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Sample records for cell-based fluorescence resonance

  1. Development of a Cell-Based Fluorescence Resonance Energy Transfer Reporter for Bacillus anthracis Lethal Factor Protease

    Energy Technology Data Exchange (ETDEWEB)

    Kimura, R H; Steenblock, E R; Camarero, J A

    2007-03-22

    We report the construction of a cell-based fluorescent reporter for anthrax lethal factor (LF) protease activity using the principle of fluorescence resonance energy transfer (FRET). This was accomplished by engineering an Escherichia coli cell line to express a genetically encoded FRET reporter and LF protease. Both proteins were encoded in two different expression plasmids under the control of different tightly controlled inducible promoters. The FRET-based reporter was designed to contain a LF recognition sequence flanked by the FRET pair formed by CyPet and YPet fluorescent proteins. The length of the linker between both fluorescent proteins was optimized using a flexible peptide linker containing several Gly-Gly-Ser repeats. Our results indicate that this FRET-based LF reporter was readily expressed in E. coli cells showing high levels of FRET in vivo in the absence of LF. The FRET signal, however, decreased 5 times after inducing LF expression in the same cell. These results suggest that this cell-based LF FRET reporter may be used to screen genetically encoded libraries in vivo against LF.

  2. Magnetic resonance imaging and cell-based neurorestorative therapy after brain injury

    Directory of Open Access Journals (Sweden)

    Quan Jiang

    2016-01-01

    Full Text Available Restorative cell-based therapies for experimental brain injury, such as stroke and traumatic brain injury, substantially improve functional outcome. We discuss and review state of the art magnetic resonance imaging methodologies and their applications related to cell-based treatment after brain injury. We focus on the potential of magnetic resonance imaging technique and its associated challenges to obtain useful new information related to cell migration, distribution, and quantitation, as well as vascular and neuronal remodeling in response to cell-based therapy after brain injury. The noninvasive nature of imaging might more readily help with translation of cell-based therapy from the laboratory to the clinic.

  3. Magnetic resonance imaging and cell-based neurorestorative therapy after brain injury

    Institute of Scientific and Technical Information of China (English)

    Quan Jiang

    2016-01-01

    Restorative cell-based therapies for experimental brain injury, such as stroke and traumatic brain injury, substantially improve functional outcome. We discuss and review state of the art magnetic resonance im-aging methodologies and their applications related to cell-based treatment after brain injury. We focus on the potential of magnetic resonance imaging technique and its associated challenges to obtain useful new information related to cell migration, distribution, and quantitation, as well as vascular and neuronal remodeling in response to cell-based therapy after brain injury. The noninvasive nature of imaging might more readily help with translation of cell-based therapy from the laboratory to the clinic.

  4. Fluorescence Resonance Energy Transfer (FRET) sensor

    CERN Document Server

    Hussain, Syed Arshad; Chakraborty, Sekhar; Saha, Jaba; Roy, Arpan Datta; Chakraborty, Santanu; Debnath, Pintu; Bhattacharjee, D

    2014-01-01

    The applications of Fluorescence resonance energy transfer (FRET) have expanded tremendously in the last 25 years, and the technique has become a staple technique in many biological and biophysical fields. FRET can be used as spectroscopic ruler in various areas such as structural elucidation of biological molecules and their interactions, in vitro assays, in vivo monitoring in cellular research, nucleic acid analysis, signal transduction, light harvesting, and metallic nanomaterials etc. Based on the mechanism of FRET a variety of novel chemical sensors and Biosensors have been developed. This review highlights the recent applications of sensitive and selective ratiometric FRET based sensors.

  5. Surface Plasmon Resonance for Cell-Based Clinical Diagnosis

    Directory of Open Access Journals (Sweden)

    Yuhki Yanase

    2014-03-01

    Full Text Available Non-invasive real-time observations and the evaluation of living cell conditions and functions are increasingly demanded in life sciences. Surface plasmon resonance (SPR sensors detect the refractive index (RI changes on the surface of sensor chips in label-free and on a real-time basis. Using SPR sensors, we and other groups have developed techniques to evaluate living cells’ reactions in response to stimuli without any labeling in a real-time manner. The SPR imaging (SPRI system for living cells may visualize single cell reactions and has the potential to expand application of SPR cell sensing for clinical diagnosis, such as multi-array cell diagnostic systems and detection of malignant cells among normal cells in combination with rapid cell isolation techniques.

  6. Magnetic resonance tracking of fluorescent nanodiamond fabrication

    Science.gov (United States)

    Shames, A. I.; Osipov, V. Yu; Boudou, J. P.; Panich, A. M.; von Bardeleben, H. J.; Treussart, F.; Vul', A. Ya

    2015-04-01

    Magnetic resonance techniques (electron paramagnetic resonance (EPR) and nuclear magnetic resonance (NMR)) are used for tracking the multi-stage process of the fabrication of fluorescent nanodiamonds (NDs) produced by high-energy electron irradiation, annealing, and subsequent nano-milling. Pristine commercial high pressure and high temperature microdiamonds (MDs) with mean size 150 μm contain ~5  ×  1018 spins/g of singlet (S = 1/2) substitutional nitrogen defects P1, as well as sp3 C-C dangling bonds in the crystalline lattice. The half-field X-band EPR clearly shows (by the appearance of the intense ‘forbidden’ g = 4.26 line) that high-energy electron irradiation and annealing of MDs induce a large amount (~5  ×  1017 spins/g) of triplet (S = 1) magnetic centers, which are identified as negatively charged nitrogen vacancy defects (NV-). This is supported by EPR observations of the ‘allowed’ transitions between Zeeman sublevels of the triplet state. After progressive milling of the fluorescent MDs down to an ultrasubmicron scale (≤100 nm), the relative abundance of EPR active NV- defects in the resulting fluorescent NDs (FND) substantially decreases and, vice versa, the content of C-inherited singlet defects correlatively increases. In the fraction of the finest FNDs (mean particle size dried supernatant of ultracentrifuged aqueous dispersion of FNDs, the NV- content is found to be reduced by one order of magnitude whereas the singlet defects content increases up to ~2  ×  1019 spins/g. In addition, another triplet-type defect, which is characterized by the g = 4.00 ‘forbidden’ line, appears. On reduction of the particle size below the 20 nm limit, the ‘allowed’ EPR lines become practically unobservable, whereas the ‘forbidden’ lines remain as a reliable fingerprint of the presence of NV- centers in small ND systems. The same size reduction causes the disappearance of the characteristic hyperfine satellites in the

  7. Distance dependence of fluorescence resonance energy transfer

    Indian Academy of Sciences (India)

    R S Swathi; K L Sebastian

    2009-09-01

    Deviations from the usual -6 dependence of the rate of fluorescence resonance energy transfer (FRET) on the distance between the donor and the acceptor have been a common scenario in the recent times. In this paper, we present a critical analysis of the distance dependence of FRET, and try to illustrate the non--6 type behaviour of the rate for the case of transfer from a localized electronic excitation on the donor, a dye molecule to three different energy acceptors with delocalized electronic excitations namely, graphene, a two-dimensional semiconducting sheet and the case of such a semiconducting sheet rolled to obtain a nanotube. We use simple analytic models to understand the distance dependence in each case.

  8. Nuclear Resonance Fluorescence for Safeguards Applications

    Energy Technology Data Exchange (ETDEWEB)

    Ludewigt, Bernhard A; Quiter, Brian J; Ambers, Scott D

    2011-02-04

    In nuclear resonance fluorescence (NRF) measurements, resonances are excited by an external photon beam leading to the emission of {gamma} rays with specific energies that are characteristic of the emitting isotope. The promise of NRF as a non-destructive analysis technique (NDA) in safeguards applications lies in its potential to directly quantify a specific isotope in an assay target without the need for unfolding the combined responses of several fissile isotopes as often required by other NDA methods. The use of NRF for detection of sensitive nuclear materials and other contraband has been researched in the past. In the safeguards applications considered here one has to go beyond mere detection and precisely quantify the isotopic content, a challenge that is discussed throughout this report. Basic NRF measurement methods, instrumentation, and the analytical calculation of NRF signal strengths are described in Section 2. Well understood modeling and simulation tools are needed for assessing the potential of NRF for safeguards and for designing measurement systems. All our simulations were performed with the radiation transport code MCNPX, a code that is widely used in the safeguards community. Our initial studies showed that MCNPX grossly underestimated the elastically scattered background at backwards angles due to an incorrect treatment of Rayleigh scattering. While new, corrected calculations based on ENDF form factors showed much better agreement with experimental data for the elastic scattering of photons on an uranium target, the elastic backscatter is still not rigorously treated. Photonuclear scattering processes (nuclear Thomson, Delbruck and Giant Dipole Resonance scattering), which are expected to play an important role at higher energies, are not yet included. These missing elastic scattering contributions were studied and their importance evaluated evaluated against data found in the literature as discussed in Section 3. A transmission experiment

  9. Resonance fluorescence from a telecom-wavelength quantum dot

    CERN Document Server

    Al-Khuzheyri, R; Huwer, J; Santana, T S; Szymanska, J Skiba-; Felle, M; Ward, M B; Stevenson, R M; Farrer, I; Tanner, M G; Hadfield, R H; Ritchie, D A; Shields, A J; Gerardot, B D

    2016-01-01

    We report on resonance fluorescence from a single quantum dot emitting at telecom wavelengths. We perform high-resolution spectroscopy and observe the Mollow triplet in the Rabi regime--a hallmark of resonance fluorescence. The measured resonance-fluorescence spectra allow us to rule out pure dephasing as a significant decoherence mechanism in these quantum dots. Combined with numerical simulations, the experimental results provide robust characterisation of charge noise in the environment of the quantum dot. Resonant control of the quantum dot opens up new possibilities for on-demand generation of indistinguishable single photons at telecom wavelengths as well as quantum optics experiments and direct manipulation of solid-state qubits in telecom-wavelength quantum dots.

  10. Resonance fluorescence and electron spin in semiconductor quantum dots

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Yong

    2009-11-18

    The work presented in this dissertation contains the first observation of spin-resolved resonance fluorescence from a single quantum dot and its application of direct measurement of electron spin dynamics. The Mollow triplet and the Mollow quintuplet, which are the hallmarks of resonance fluorescence, are presented as the non-spin-resolved and spin-resolved resonance fluorescence spectrum, respectively. The negligible laser background contribution, the near pure radiative broadened spectrum and the anti-bunching photon statistics imply the sideband photons are background-free and near transform-limited single photons. This demonstration is a promising step towards the heralded single photon generation and electron spin readout. Instead of resolving spectrum, an alternative spin-readout scheme by counting resonance fluorescence photons under moderate laser power is demonstrated. The measurements of n-shot time-resolved resonance fluorescence readout are carried out to reveal electron spin dynamics of the measurement induced back action and the spin relaxation. Hyperfine interaction and heavy-light hole mixing are identified as the relevant mechanisms for the back action and phonon-assistant spin-orbit interaction dominates the spin relaxation. After a detailed discussion on charge-spin configurations in coupled quantum dots system, the single-shot readout on electron spin are proposed. (orig.)

  11. Fluorescence Resonance Energy Transfer Using Spiropyran and Diarylethene Photochromic Acceptors

    Directory of Open Access Journals (Sweden)

    E. A. Jares-Erijman

    2000-03-01

    Full Text Available We describe the preparation and photophysical characterization of two model compounds designed to test a new approach for the quantitative determination of Fluorescence Resonance Energy Transfer (FRET in biological systems. The method enables modulation of FRET by exploiting the unique reversible spectral properties of photochromic diarylethenes and spiropyrans to create switchable energy acceptors.

  12. Resonance fluorescence of a cold atom in a high-finesse resonator

    CERN Document Server

    Bienert, M; Torres, J M; Zippilli, S; Bienert, Marc; Morigi, Giovanna; Zippilli, Stefano

    2007-01-01

    We study the spectra of emission of a system composed by an atom, tightly confined inside a high-finesse resonator, when the atom is driven by a laser and is at steady state of the cooling dynamics induced by laser and cavity field. In general, the spectrum of resonance fluorescence and the spectrum at the cavity output contain complementary information about the dynamics undergone by the system. In certain parameter regimes, quantum interference effects between the scattering processes induced by cavity and laser field lead to the selective suppression of features of the resonance fluorescence spectrum, which are otherwise visible in the spectrum of laser-cooled atoms in free space.

  13. Fluorescence resonance energy transfer between fluorescent proteins as powerful toolkits for in vivo studies

    Science.gov (United States)

    Rusanov, A. L.; Savitsky, A. P.

    2011-02-01

    To expand the field of research in biological systems development of extra-sensitive analytical methods is highly desirable. In this review, the latest advances in technologies relying on the fluorescence resonance energy transfer between fluorescent proteins (FP's) to visualize numerous molecular processes in living cells are discussed. Variety of FP's as well as of novel experimental techniques allows one to choose the most appropriate tools to attack concrete problems.

  14. Resonant soft X-ray fluorescence spectra of molecules

    Energy Technology Data Exchange (ETDEWEB)

    Nordgren, J.; Glans, P.; Gunnelin, K.; Guo, J.; Skytt, P.; Saathe, C.; Wassdahl, N. [Uppsala Univ. (Sweden). Dept. of Phys.

    1997-08-01

    A review is presented of some recent advancements in resonant soft X-ray fluorescence spectroscopy applied to molecules. The concept of inelastic scattering is discussed for molecules with an element of symmetry, which introduces degeneracy or near-degeneracy of core excited states. The potential of the symmetry selection rules associated with resonant X-ray fluorescence is illustrated in studies of electronic structure as well as molecular dynamics. The use of polarized photons for exciting X-ray emission is discussed, and examples are presented where anisotropy of X-ray emission provides more detailed information. A brief section discusses the experimental considerations relevant for the spectroscopic technique. (orig.) With 013 figs., 28 refs.

  15. Far-red fluorescence gene reporter tomography for determination of placement and viability of cell-based gene therapies.

    Science.gov (United States)

    Lu, Yujie; Darne, Chinmay D; Tan, I-Chih; Zhu, Banghe; Hall, Mary A; Lazard, Zawaunyka W; Davis, Alan R; Simpson, Lashan; Sevick-Muraca, Eva M; Olmsted-Davis, Elizabeth A

    2013-10-01

    Non-invasive injectable cellular therapeutic strategies based on sustained delivery of physiological levels of BMP-2 for spinal fusion are emerging as promising alternatives, which could provide sufficient fusion without the associated surgical risks. However, these injectable therapies are dependent on bone formation occurring only at the specific target region. In this study, we developed and deployed fluorescence gene reporter tomography (FGRT) to provide information on in vivo cell localization and viability. This information is sought to confirm the ideal placement of the materials with respect to the area where early bone reaction is required, ultimately providing three dimensional data about the future fusion. However, because almost all conventional fluorescence gene reporters require visible excitation wavelengths, current in vivo imaging of fluorescent proteins is limited by high tissue absorption and confounding autofluorescence. We previously administered fibroblasts engineered to produce BMP-2, but is difficult to determine 3-D information of placement prior to bone formation. Herein we used the far-red fluorescence gene reporter, IFP1.4 to report the position and viability of fibroblasts and developed 3-D tomography to provide placement information. A custom small animal, far-red fluorescence tomography system integrated into a commercial CT scanner was used to assess IFP1.4 fluorescence and to demark 3-D placement of encapsulated fibroblasts with respect to the vertebrae and early bone formation as assessed from CT. The results from three experiments showed that the placement of the materials within the spine could be detected. This work shows that in vivo fluorescence gene reporter tomography of cell-based gene therapy is feasible and could help guide cell-based therapies in preclinical models.

  16. Contraband Detection with Nuclear Resonance Fluorescence: Feasibility and Impact

    Energy Technology Data Exchange (ETDEWEB)

    Pruet, J; Lange, D

    2007-01-03

    In this report they show that cargo interrogation systems developed to thwart trafficking of illicit nuclear materials could also be powerful tools in the larger fight against contraband smuggling. In particular, in addition to detecting special nuclear materials, cargo scanning systems that exploit nuclear resonance fluorescence to detect specific isotopes can be used to help find: chemical weapons; some drugs as well as some chemicals regulated under the controlled substances act; precious metals; materials regulated under export control laws; and commonly trafficked fluorocarbons.

  17. A Theoretical Distinction Between Time-Resolved Resonance Raman andResonance Fluorescence

    Institute of Scientific and Technical Information of China (English)

    LU Jing; DU Si-De; FAN Kang-Nian; Lee Soo-Ying

    2000-01-01

    Based on the time-dependent theory, an analysis of the distinction between resonance Raman (RR) and resonance fluorescence (RF) with pulse excitation was presented. The real population of the intermediate state gives two optical components-the independent time evolution of intermediate ket and bra states generates RR while RF originates from the phase coherent between ket and bra states. In cw limit, the transition probability of spontaneous emission with pulse excitation can be reduced to the classical theory.

  18. Using magnetic resonance imaging to evaluate dendritic cell-based vaccination.

    Directory of Open Access Journals (Sweden)

    Peter M Ferguson

    Full Text Available Cancer immunotherapy with antigen-loaded dendritic cell-based vaccines can induce clinical responses in some patients, but further optimization is required to unlock the full potential of this strategy in the clinic. Optimization is dependent on being able to monitor the cellular events that take place once the dendritic cells have been injected in vivo, and to establish whether antigen-specific immune responses to the tumour have been induced. Here we describe the use of magnetic resonance imaging (MRI as a simple, non-invasive approach to evaluate vaccine success. By loading the dendritic cells with highly magnetic iron nanoparticles it is possible to assess whether the injected cells drain to the lymph nodes. It is also possible to establish whether an antigen-specific response is initiated by assessing migration of successive rounds of antigen-loaded dendritic cells; in the face of a successfully primed cytotoxic response, the bulk of antigen-loaded cells are eradicated on-route to the node, whereas cells without antigen can reach the node unchecked. It is also possible to verify the induction of a vaccine-induced response by simply monitoring increases in draining lymph node size as a consequence of vaccine-induced lymphocyte trapping, which is an antigen-specific response that becomes more pronounced with repeated vaccination. Overall, these MRI techniques can provide useful early feedback on vaccination strategies, and could also be used in decision making to select responders from non-responders early in therapy.

  19. Resonance fluorescence of a trapped three-level atom

    CERN Document Server

    Bienert, M; Morigi, G; Bienert, Marc; Merkel, Wolfgang; Morigi, Giovanna

    2003-01-01

    We investigate theoretically the spectrum of resonance fluorescence of a harmonically trapped atom, whose internal transitions are $\\Lambda$--shaped and driven at two-photon resonance by a pair of lasers, which cool the center--of--mass motion. For this configuration, photons are scattered only due to the mechanical effects of the quantum interaction between light and atom. We study the spectrum of emission in the final stage of laser--cooling, when the atomic center-of-mass dynamics is quantum mechanical and the size of the wave packet is much smaller than the laser wavelength (Lamb--Dicke limit). We use the spectral decomposition of the Liouville operator of the master equation for the atomic density matrix and apply second order perturbation theory. We find that the spectrum of resonance fluorescence is composed by two narrow sidebands -- the Stokes and anti-Stokes components of the scattered light -- while all other signals are in general orders of magnitude smaller. For very low temperatures, however, th...

  20. Development and validation of a simple cell-based fluorescence assay for dipeptidyl peptidase 1 (DPP1) activity.

    Science.gov (United States)

    Thong, Bob; Pilling, James; Ainscow, Edward; Beri, Raj; Unitt, John

    2011-01-01

    Dipeptidyl peptidase 1 (DPP1) (EC 3.4.14.1; also known as cathepsin C, cathepsin J, dipeptidyl aminopeptidase, and dipeptidyl aminotransferase) is a lysosomal cysteinyl protease of the papain family involved in the intracellular degradation of proteins. Isolated enzyme assays for DPP1 activity using a variety of synthetic substrates such as dipeptide or peptide linked to amino-methyl-coumarin (AMC) or other fluorophores are well established. There is, however, no report of a simple whole-cell-based assay for measuring lysosomal DPP1 activity other than the use of flow cytometry (fluorescence-activated cell sorting) or the use of invasive activity-based probes or the production of physiological products such as neutrophil elastase. The authors investigated a number of DPP1 fluorogenic substrates that have the potential to access the lysosome and enable the measurement of DPP1 enzyme activity in situ. They describe the development and evaluation of a simple noninvasive fluorescence assay for measuring DPP1 activity in fresh or cryopreserved human THP-1 cells using the substrate H-Gly-Phe-AFC (amino-fluoro-coumarin). This cell-based fluorescence assay can be performed in a 96-well plate format and is ideally suited for determining the cell potency of potential DPP1 enzyme inhibitors.

  1. Highly sensitive turn-on fluorescence detection of thrombomodulin based on fluorescence resonance energy transfer

    Science.gov (United States)

    Kong, Liyan; Zhu, Jiaming; Wang, Wen; Jin, Lehe; Fu, Yanjiao; Duan, Bohui; Tan, Liang

    2017-02-01

    As an integral glycoprotein on the surface of endothelial cells, thrombomodulin (TM) has very high affinity for thrombin. TM has been regarded to be a marker of endothelial damage since it can be released during endothelial cell injury. In this work, a highly sensitive fluorescence method for the quantitative detection of TM was developed. TM antibody (Ab) and bovine serum albumin (BSA) were bound on gold nanoparticles (AuNPs) to construct BSA-AuNPs-Ab nanocomposites and they were characterized by transmission electron microscope and UV-vis spectrophotometry. The fluorescence of acridine orange (AO) was quenched by the prepared gold nanocomposites based on fluorescence resonance energy transfer (FRET). In the presence of TM, the fluorescence was turned on due to the effective separation of AO from the surface of gold nanocomposites. Under optimum conditions, the enhanced fluorescence intensity displayed a linear relationship with the logarithm of the TM concentration from 0.1 pg mL- 1 to 5 ng mL- 1 with a low detection limit of 12 fg mL- 1. The release of soluble thrombomodulin (sTM) by the injured HUVEC-C cells in the presence of H2O2 was investigated using the proposed method. The released sTM content in the growth medium was found to be increased with the enhancement of contact time of the cells with H2O2.

  2. Effects of external fields, dimension and polarization on the resonance fluorescence of quantum dots

    Science.gov (United States)

    Vaseghi, B.; Razavi, S. M.

    2017-02-01

    In this paper simultaneous effects of external electric and magnetic fields, dimension and polarization on the resonance fluorescence spectrum and photon statistics of a spherical quantum dot with parabolic confinement are investigated. With special attention to the optical scattering processes resonance fluorescence spectrum and second-order correlation function are calculated and plotted for different external parameters. Our results show the occurrence of resonance fluorescence similar to atomic systems and considerable effects of external fields, quantum confinement and light polarization on the resonance fluorescence spectrum and second-order correlation function in the quantum dot systems. The existence of Mollow triplets and photon antibunching are strongly depend on these external agents.

  3. Fluorescence assay for glycan expression on living cancer cells based on competitive strategy coupled with dual-functionalized nanobiocomposites.

    Science.gov (United States)

    Fu, Ying; Lu, Danqin; Lin, Bin; Sun, Qianqian; Liu, Kai; Xu, Lili; Zhang, Shengping; Hu, Chen; Wang, Chuangui; Xu, Zhiai; Zhang, Wen

    2013-11-21

    Cell surface glycans are a class of sophisticated biomolecules related to cancer development and progression, and their analysis is of great significance for early cancer diagnosis and treatment. In this paper, we proposed a fluorescence assay to evaluate glycan expression on living cancer cells based on a competitive strategy coupled with dual-functionalized nanobiocomposites. The competitive assay was conducted between living cancer cells and thiomannosyl derivatives using concanavalin A (Con A)-modified electrode as the interaction platform. To impart fluorescence signaling ability to competitive derivatives, quantum dots (QDs) were anchored on BSA-protected Au nanoparticles, and thiomannosyl derivatives were further immobilized on the nanoparticle surface through Au-S binding. Due to the spacing between QDs and Au nanoparticles by BSA, the {QDs-Au-BSA-mannose} nanobiocomposites maintained the fluorescence of QDs and showed binding ability with the Con A-modified electrode. Au nanorods (AuNRs)-modified electrode was used as an effective substrate to immobilize Con A. This assay was successfully applied to the analysis of two cancer cells lines (A549 and QGY-7701). The method is simple and shows promise for the study of glycan expression on living cancer cells.

  4. Performance of a Cyanobacteria Whole Cell-Based Fluorescence Biosensor for Heavy Metal and Pesticide Detection

    Directory of Open Access Journals (Sweden)

    Salmijah Surif

    2013-05-01

    Full Text Available Whole cell biosensors always face the challenge of low stability of biological components and short storage life. This paper reports the effects of poly(2-hydroxyethyl methacrylate (pHEMA immobilization on a whole cell fluorescence biosensor for the detection of heavy metals (Cu, Pb, Cd, and pesticides (dichlorophenoxyacetic acid (2,4-D, and chlorpyrifos. The biosensor was produced by entrapping the cyanobacterium Anabaena torulosa on a cellulose membrane, followed by applying a layer of pHEMA, and attaching it to a well. The well was then fixed to an optical probe which was connected to a fluorescence spectrophotometer and an electronic reader. The optimization of the biosensor using several factors such as amount of HEMA and drying temperature were undertaken. The detection limits of biosensor without pHEMA for Cu, Cd, Pb, 2,4-D and chlorpyrifos were 1.195, 0.027, 0.0100, 0.025 and 0.025 µg/L respectively. The presence of pHEMA increased the limits of detection to 1.410, 0.250, 0.500, 0.235 and 0.117 µg/L respectively. pHEMA is known to enhance the reproducibility of the biosensor with average relative standard deviation (RSD of ±1.76% for all the pollutants tested, 48% better than the biosensor without pHEMA (RSD = ±3.73%. In storability test with Cu 5 µg/L, the biosensor with pHEMA performed 11.5% better than the test without pHEMA on day-10 and 5.2% better on day-25. pHEMA is therefore a good candidate to be used in whole cell biosensors as it increases reproducibility and enhances biosensor storability.

  5. A Label-Free Microfluidic Biosensor for Activity Detection of Single Microalgae Cells Based on Chlorophyll Fluorescence

    Directory of Open Access Journals (Sweden)

    Junsheng Wang

    2013-11-01

    Full Text Available Detection of living microalgae cells is very important for ballast water treatment and analysis. Chlorophyll fluorescence is an indicator of photosynthetic activity and hence the living status of plant cells. In this paper, we developed a novel microfluidic biosensor system that can quickly and accurately detect the viability of single microalgae cells based on chlorophyll fluorescence. The system is composed of a laser diode as an excitation light source, a photodiode detector, a signal analysis circuit, and a microfluidic chip as a microalgae cell transportation platform. To demonstrate the utility of this system, six different living and dead algae samples (Karenia mikimotoi Hansen, Chlorella vulgaris, Nitzschia closterium, Platymonas subcordiformis, Pyramidomonas delicatula and Dunaliella salina were tested. The developed biosensor can distinguish clearly between the living microalgae cells and the dead microalgae cells. The smallest microalgae cells that can be detected by using this biosensor are 3 μm ones. Even smaller microalgae cells could be detected by increasing the excitation light power. The developed microfluidic biosensor has great potential for in situ ballast water analysis.

  6. Indicators based on fluorescence resonance energy transfer (FRET).

    Science.gov (United States)

    Tsien, Roger Y

    2009-07-01

    One of the major new trends in the design of indicators for optically imaging biochemical and physiological functions of living cells has been the exploitation of fluorescence resonance energy transfer (FRET). FRET is a well-known spectroscopic technique for monitoring changes in the proximity and mutual orientation of pairs of chromophores. It has long been used in biochemistry and cell biology to assess distances and orientations between specific labeling sites within a single macromolecule or between two separate molecules. More recently, macromolecules or molecular pairs have been engineered to change their FRET in response to biochemical and physiological signals such as membrane potential, cyclic AMP (cAMP), protease activity, free Ca(2+) and Ca(2+)-calmodulin (CaM) concentrations, protein-protein heterodimerization, phosphorylation, and reporter-gene expression. Because FRET is general, nondestructive, and easily imaged, it has proven to be one of the most versatile spectroscopic readouts available to the designer of new probes. FRET is particularly amenable to emission ratioing, which is more reliably quantifiable than single-wavelength monitoring and better suited than excitation ratioing to high-speed and laser-excited imaging. This article summarizes the photophysical principles of FRET and the types of indicators used.

  7. Biomolecular interactions probed by fluorescence resonance energy transfer

    Science.gov (United States)

    Lange, Daniela Charlotte

    2000-09-01

    This thesis describes how a physical phenomenon, Fluorescence Resonance Energy Transfer (FRET), can be exploited for the study of interactions between biomolecules. The physical basis of this phenomenon is discussed and it is described how some of its characteristics can be exploited in measurement. A recently introduced method, photobleaching FRET microscopy, was implemented and its image analysis refined to suit our biological context. Further, a new technique is proposed, which combines FRET with confocal laser scanning microscopy to optimize resolution and to allow for 3D-studies in living cells. The first part of this thesis presents the application of FRET to the study of oligomerization of G-protein coupled receptors (GPCRs), which was performed at the Fraser Laboratories at McGill University in Montreal. It is demonstrated how FRET microscopy allowed us to circumvent problems of traditional biochemical approaches and provided the first direct evidence for GPCR oligomerization in intact cells. We found that somatostatin receptors (SSTRs) functionally interact by forming oligomers with their own kind, with different SSTR isoforms, and even with distantly related GPCRs, such as dopamine receptors, the latter of which is breaking with the dogma that GPCRs would only pair up with their own kind. The high sensitivity of the FRET technique allowed us to characterize these interactions under more physiological conditions, which lead to the observation that oligomerization is induced by receptor agonist. We further studied the differential effects of agonists and antagonists on receptor oligomerization, leading to a model for the molecular mechanism underlying agonist/antagonist function and receptor activation. The second part was carried out at the Neurobiology Laboratory of the VA Medical Center in Newington, CT. The objective was to further our understanding of Niemann- Pick type C disease, which is characterized by a defect in intracellular cholesterol

  8. Development of cell-based quantitative evaluation method for cell cycle-arrest type cancer drugs for apoptosis by high precision surface plasmon resonance sensor

    Science.gov (United States)

    Ona, Toshihiro; Nishijima, Hiroshi; Kosaihira, Atsushi; Shibata, Junko

    2008-04-01

    In vitro rapid and quantitative cell-based assay is demanded to verify the efficacy prediction of cancer drugs since a cancer patient may have unconventional aspects of tumor development. Here, we show the rapid and non-label quantitative verifying method and instrumentation of apoptosis for cell cycle-arrest type cancer drugs (Roscovitine and D-allose) by reaction analysis of living liver cancer cells cultured on a sensor chip with a newly developed high precision (50 ndeg s -1 average fluctuation) surface plasmon resonance (SPR) sensor. The time-course cell reaction as the SPR angle change rate for 10 min from 30 min cell culture with a drug was significantly related to cell viability. By the simultaneous detection of differential SPR angle change and fluorescence by specific probes using the new instrument, the SPR angle was related to the nano-order potential decrease in inner mitochondrial membrane potential. The results obtained are universally valid for the cell cycle-arrest type cancer drugs, which mediate apoptosis through different cell-signaling pathways, by a liver cancer cell line of Hep G2 (Pcancer cells from patients in clinical use.

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

    Science.gov (United States)

    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.

  10. Dependence of fluorescence intensity on the spectral overlap between fluorophores and plasmon resonant single silver nanoparticles.

    Science.gov (United States)

    Chen, Yeechi; Munechika, Keiko; Ginger, David S

    2007-03-01

    We investigate the fluorescence from dyes coupled to individual DNA-functionalized metal nanoparticles. We use single-particle darkfield scattering and fluorescence microscopy to correlate the fluorescence intensity of the dyes with the localized surface plasmon resonance (LSPR) spectra of the individual metal nanoparticles to which they are attached. For each of three different dyes, we observe a strong correlation between the fluorescence intensity of the dye and the degree of spectral overlap with the plasmon resonance of the nanoparticle. On average, we observe the brightest fluorescence from dyes attached to metal nanoparticles that have a LSPR scattering peak approximately 40-120 meV higher in energy than the emission peak of the fluorophore. These results should prove useful for understanding and optimizing metal-enhanced fluorescence.

  11. Emission wavelength tuning of fluorescence by fine structural control of optical metamaterials with Fano resonance

    Science.gov (United States)

    Moritake, Y.; Kanamori, Y.; Hane, K.

    2016-09-01

    We demonstrated fine emission wavelength tuning of quantum dot (QD) fluorescence by fine structural control of optical metamaterials with Fano resonance. An asymmetric-double-bar (ADB), which was composed of only two bars with slightly different bar lengths, was used to obtain Fano resonance in the optical region. By changing the short bar length of ADB structures with high dimensional accuracy in the order of 10 nm, resonant wavelengths of Fano resonance were controlled from 1296 to 1416 nm. Fluorescence of QDs embedded in a polymer layer on ADB metamaterials were modified due to coupling to Fano resonance and fine tuning from 1350 to 1376 nm was observed. Wavelength tuning of modified fluorescence was reproduced by analysis using absorption peaks of Fano resonance. Tuning range of modified fluorescence became narrow, which was interpreted by a simple Gaussian model and resulted from comparable FWHM in QD fluorescence and Fano resonant peaks. The results will help the design and fabrication of metamaterial devices with fluorophores such as light sources and biomarkers.

  12. Emission wavelength tuning of fluorescence by fine structural control of optical metamaterials with Fano resonance.

    Science.gov (United States)

    Moritake, Y; Kanamori, Y; Hane, K

    2016-09-13

    We demonstrated fine emission wavelength tuning of quantum dot (QD) fluorescence by fine structural control of optical metamaterials with Fano resonance. An asymmetric-double-bar (ADB), which was composed of only two bars with slightly different bar lengths, was used to obtain Fano resonance in the optical region. By changing the short bar length of ADB structures with high dimensional accuracy in the order of 10 nm, resonant wavelengths of Fano resonance were controlled from 1296 to 1416 nm. Fluorescence of QDs embedded in a polymer layer on ADB metamaterials were modified due to coupling to Fano resonance and fine tuning from 1350 to 1376 nm was observed. Wavelength tuning of modified fluorescence was reproduced by analysis using absorption peaks of Fano resonance. Tuning range of modified fluorescence became narrow, which was interpreted by a simple Gaussian model and resulted from comparable FWHM in QD fluorescence and Fano resonant peaks. The results will help the design and fabrication of metamaterial devices with fluorophores such as light sources and biomarkers.

  13. Resonance Fluorescence of Fused Silica by the Depopulation of the Ground State

    Directory of Open Access Journals (Sweden)

    Fuat Bayrakceken

    2012-01-01

    Full Text Available Spectroscopically pure fused silica has been used in many applications ranging from optoelectronics and optical fibers to laser flash spectroscopy. Although ultraviolet light irradiated optical absorption spectra and coherence fluorescence of silicon dioxide have been studied in the past, we present discrete absorption and resonance coherent fluorescence line of silicon dioxide which were recorded photographically at 288.2 nm. This discrete fluorescence is observed at room temperature using high photon flux (1024 photon/pulse excitation spectroscopy.

  14. Magnification of photonic crystal fluorescence enhancement via TM resonance excitation and TE resonance extraction on a dielectric nanorod surface

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Hsin-Yu; Cunningham, Brian T [Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, 1406 W Green Street, Urbana, IL 61801 (United States); Zhang Wei [Department of Material Science and Engineering, University of Illinois at Urbana-Champaign, 1304 W Green Street, Urbana, IL 61801 (United States); Mathias, Patrick C, E-mail: bcunning@illinois.edu [Department of Bioengineering, University of Illinois at Urbana-Champaign, 1304 W Springfield Avenue, Urbana, IL 61801 (United States)

    2010-03-26

    Using a one-dimensional grating surface photonic crystal (PC), we experimentally demonstrate that the detection of fluorescent molecules on a PC surface can be substantially magnified through the combined effects of resonance-enhanced excitation of the fluorescent dye, resonance-enhanced extraction of the fluorescence emission and a dielectric nanorod surface coating increasing the surface area available for fluorophore-PC interaction. Enhanced excitation is obtained by engineering a high-Q TM resonant mode to efficiently couple with an incident TM-polarized {lambda} = 633 nm laser for exciting Cyanine-5 (Cy5). Enhanced extraction results from a low-Q TE resonance designed to spectrally overlap the Cy5 emission spectrum for channeling TE-polarized emission towards the detection instrument. The entire PC surface is coated with a porous film of TiO{sub 2} nanorods that allows more fluorophores to penetrate into the region of enhanced near-electric fields. Experimental results reveal a 588-fold enhancement in fluorescence intensity relative to an unpatterned glass surface.

  15. Experimental models of brain ischemia: a review of techniques, magnetic resonance imaging and investigational cell-based therapies

    Directory of Open Access Journals (Sweden)

    Alessandra eCanazza

    2014-02-01

    Full Text Available Stroke continues to be a significant cause of death and disability worldwide. Although major advances have been made in the past decades in prevention, treatment and rehabilitation, enormous challenges remain in the way of translating new therapeutic approaches from bench to bedside. Thrombolysis, while routinely used for ischemic stroke, is only a viable option within a narrow time window. Recently, progress in stem cell biology has opened up avenues to therapeutic strategies aimed at supporting and replacing neural cells in infarcted areas. Realistic experimental animal models are crucial to understand the mechanisms of neuronal survival following ischemic brain injury and to develop therapeutic interventions. Current studies on experimental stroke therapies evaluate the efficiency of neuroprotective agents and cell-based approaches using primarily rodent models of permanent or transient focal cerebral ischemia. In parallel, advancements in imaging techniques permit better mapping of the spatial-temporal evolution of the lesioned cortex and its functional responses. This review provides a condensed conceptual review of the state of the art of this field, from models and magnetic resonance imaging techniques through to stem cell therapies.

  16. Near-infrared emitting fluorescent nanocrystals-labeled natural killer cells as a platform technology for the optical imaging of immunotherapeutic cells-based cancer therapy

    Science.gov (United States)

    Taik Lim, Yong; Cho, Mi Young; Noh, Young-Woock; Chung, Jin Woong; Chung, Bong Hyun

    2009-11-01

    This study describes the development of near-infrared optical imaging technology for the monitoring of immunotherapeutic cell-based cancer therapy using natural killer (NK) cells labeled with fluorescent nanocrystals. Although NK cell-based immunotherapeutic strategies have drawn interest as potent preclinical or clinical methods of cancer therapy, there are few reports documenting the molecular imaging of NK cell-based cancer therapy, primarily due to the difficulty of labeling of NK cells with imaging probes. Human natural killer cells (NK92MI) were labeled with anti-human CD56 antibody-coated quantum dots (QD705) for fluorescence imaging. FACS analysis showed that the NK92MI cells labeled with anti-human CD56 antibody-coated QD705 have no effect on the cell viability. The effect of anti-human CD56 antibody-coated QD705 labeling on the NK92MI cell function was investigated by measuring interferon gamma (IFN- γ) production and cytolytic activity. Finally, the NK92MI cells labeled with anti-human CD56 antibody-coated QD705 showed a therapeutic effect similar to that of unlabeled NK92MI cells. Images of intratumorally injected NK92MI cells labeled with anti-human CD56 antibody-coated could be acquired using near-infrared optical imaging both in vivo and in vitro. This result demonstrates that the immunotherapeutic cells labeled with fluorescent nanocrystals can be a versatile platform for the effective tracking of injected therapeutic cells using optical imaging technology, which is very important in cell-based cancer therapies.

  17. Sensitive turn-on fluorescent detection of tartrazine based on fluorescence resonance energy transfer.

    Science.gov (United States)

    Huang, Sheng Tian; Shi, Yan; Li, Nian Bing; Luo, Hong Qun

    2012-01-18

    We introduce a sensitive, rapid, label-free and general fluorescent method for the determination of tartrazine by competitive binding to reduced graphene oxide (rGO) against fluorescein, and the fluorescence recovery upon fluorescein desorption from rGO provides a quantitative readout for tartrazine, giving a detection limit of 0.53 ng mL(-1).

  18. Resonance fluorescence from an asymmetric quantum dot dressed by a bichromatic electromagnetic field

    Science.gov (United States)

    Kryuchkyan, G. Yu.; Shahnazaryan, V.; Kibis, O. V.; Shelykh, I. A.

    2017-01-01

    We present the theory of resonance fluorescence from an asymmetric quantum dot driven by a two-component electromagnetic field with two different frequencies, polarizations, and amplitudes (bichromatic field) in the regime of strong light-matter coupling. It follows from the elaborated theory that the broken inversion symmetry of the driven quantum system and the bichromatic structure of the driving field result in unexpected features of the resonance fluorescence, including the infinite set of Mollow triplets, the quench of fluorescence peaks induced by the dressing field, and the oscillating behavior of the fluorescence intensity as a function of the dressing field amplitude. These quantum phenomena are of general physical nature and, therefore, can take place in various double-driven quantum systems with broken inversion symmetry.

  19. Charge-driven feedback loop in the resonance fluorescence of a single quantum dot

    Science.gov (United States)

    Merkel, B.; Kurzmann, A.; Schulze, J.-H.; Strittmatter, A.; Geller, M.; Lorke, A.

    2017-03-01

    We demonstrate a feedback loop that manifests itself in a strong hysteresis and bistability of the exciton resonance fluorescence signal. Field ionization of photogenerated quantum dot excitons leads to the formation of a charged interface layer that drags the emission line along over a frequency range of more than 30 GHz . These measurements are well described by a rate equation model. With a time-resolved resonance fluorescence measurement we determined the buildup times for the hole gas in the orders of milliseconds. This internal charge-driven feedback loop could be used to reduce the spectral wandering in the emission spectra of single self-assembled quantum dots.

  20. Determination of level widths in 15N using nuclear resonance fluorescence

    Directory of Open Access Journals (Sweden)

    Szücs T.

    2015-01-01

    Full Text Available Level widths in 15N have been measured with the nuclear resonance fluorescence (NRF technique. Solid nitrogen compounds, bremsstrahlung, and HPGe detectors have been used as target, beam, and detectors, respectively. The preliminarily level widths are in agreement with the literature values, but more precise.

  1. A fluorescence resonance energy transfer-based method for histone methyltransferases

    DEFF Research Database (Denmark)

    Devkota, Kanchan; Lohse, Brian; Nyby Jakobsen, Camilla;

    2015-01-01

    A simple dye–quencher fluorescence resonance energy transfer (FRET)-based assay for methyltransferases was developed and used to determine kinetic parameters and inhibitory activity at EHMT1 and EHMT2. Peptides mimicking the truncated histone H3 tail were functionalized in each end with a dye...

  2. DEVELOPMENT OF A REAL-TIME FLUORESCENCE RESONANCE ENERGY TRANSFER (FRET) PCR TO DETECT ARCOBACTER SPECIES

    Science.gov (United States)

    A real-time PCR targeting the gyrase A subunit gene outside the quinolone resistance-determining region has been developed to detect Arcobacter species. The species identification was made by probe hybridization and melting curve analysis, using the Fluorescence Resonance Energy Transfer technology...

  3. DEVELOPMENT OF A REAL-TIME FLUORESCENCE RESONANCE ENERGY TRANSFER PCR TO DETECT ARCOBACTER SPECIES

    Science.gov (United States)

    A real-time PCR targeting the gyrase A subunit gene outside the quinolone resistance-determining region has been developed to detect Arcobacter species. The species identification was made by probe hybridization and melting curve analysis, using Fluorescence Resonance Energy Transfer technology. D...

  4. Spectrophotometry of Bowen resonance fluorescence lines in three planetary nebulae

    Science.gov (United States)

    O'Dell, C. R.; Miller, Christopher O.

    1992-01-01

    The results are presented of a uniquely complete, carefully reduced set of observations of the O III Bowen fluorescence lines in the planetary nebulae NGC 6210, NGC 7027, and NGC 7662. A detailed comparison with the predictions of radiative excitation verify that some secondary lines are enhanced by selective population by the charge exchange mechanism involving O IV. Charge exchange is most important in NGC 6210, which is of significantly lower ionization than the other nebulae. In addition to the principal Bowen lines arising from Ly-alpha pumping of the O III O1 line, lines arising from pumping of the O3 line are also observed. Comparison of lines produced by O1 and O3 with the theoretical predictions of Neufeld indicate poor agreement; comparison with the theoretical predictions of Harrington show agreement with NGC 7027 and NGC 7662.

  5. 基于荧光检测的新型细胞传感器%Novel Cell-based Biosensors Based on Measurement of Fluorescence

    Institute of Scientific and Technical Information of China (English)

    辛文文; 王景林

    2011-01-01

    The diagnosis of infectious diseases, the monitoring of environment and detection of potential bioterrorism agents greatly require a pathogen identification method with better combined speed, accuracy and sensitivity. Here, a kind of novel cell-based biosensors based on measurement of fluorescence is introduced, which could detect pathogens and other antigens by measuring fluorescent signals within minutes. It is based on the specifically bind character of antigens and antibodies and the theory of bioluminescence or chemiluminescence.The cell could emit light via calcium chemically fluorescent indicators such as Fluo-4, or calcium fluorescent proteins such as aequorin, green fluorescent protein. B cell-based biosensors and mast cell-based biosensors have been applied in some fields. This kind of biosensors has advantages in combined sensitivity, accuracy and speed,while it also has disadvantages such as cross reactivity and problems with cellular storage and maintenance. This is a promising kind of biosensors applied in the diagnosis of infectious diseases, the monitoring of environment and detection of potential bioterrorism agents.%主要介绍了一类基于荧光检测的新型细胞传感器,这类传感器利用免疫细胞表面分子特异性识别、结合抗原的特性和生物(或化学)发光技术,通过检测荧光信号在数分钟内达到检测病原体或其他抗原的目的.这类传感器的发光原理主要是利用钙离子敏感型化学荧光探针发光,如Fluo-4等,或钙离子敏感型发光蛋自发光,如水母发光蛋白、绿色荧光蛋白等.现在已经应用的主要是B细胞传感器和肥大细胞传感器.这类传感器具有灵敏度高、检测准确、反应速度快的优点.同时又存在交叉反应、细胞不易保存等不足之处.这类传感器在疾病诊断、环境监测、生物战剂检测等领域具有较大的应用前景.

  6. Study on the interaction between diphenhydramine and erythrosin by absorption, fluorescence and resonance Rayleigh scattering spectra

    Institute of Scientific and Technical Information of China (English)

    TANG XiaoLing; LIU ZhongFang; LIU ShaoPu; HU XiaoLi

    2007-01-01

    In pH 4.5 Britton-Robinson (BR) buffer solution, erythrosin (ET) can react with diphenhydramine (DP) to form a 1:1 ion-association complex, which not only results in the change of the absorption spectra, but also results in the great enhancement of resonance Rayleigh scattering (RRS) and the quenching of fluorescence. Furthermore, a new RRS spectrum will appear, and the maximum RRS wavelength was located at about 580 nm.In this work, the spectral characteristics of the absorption, fluorescence and RRS, the optimum conditions of the reaction and the properties of an analytical chemistry were investigated. A sensitive, simple and new method for the determination of DP by using erythrosin as a probe has been developed. The detection limits for DP were 0.0020 μg/mL for RRS method, 0.088 μg/mL for absorption method and 0.094 μg/mL for fluorophotometry. There was a linear relationship between the absorbance, RRS and fluorescence intensities and the drug concentration in the range of 0.0067-2.0, 0.29-6.4 and 0.31-3.2 μg/mL, respectively. The effects of the interaction of diphenhydramine and erythrosin on the absorption, fluorescence and resonance Rayleigh scattering spectra were discussed. In light polarization experiment, the polarization of RRS at maximum wavelength was measured to be P = 0.9779, and it revealed that the RRS spectrum of DP-ET complex consists mostly of resonance scattering and few resonance fluorescence. In this study, enthalpy of formation and mean polarizability were calculated by AM1 quantum chemistry method. In addition, the reaction mechanism and the reasons for the enhancement of scattering spectra and the energy transfer between absorption, fluorescence and RRS were discussed.

  7. Monitoring p21 mRNA expression in living cell based on molecular beacon fluorescence increasing rate

    Institute of Scientific and Technical Information of China (English)

    TANG HongXing; YANG XiaoHai; WANG KeMin; TAN WeiHong; LIU Bin; HE LiFang; WANG Wei

    2008-01-01

    Studying the expression level of mRNA in living cells will offer tremendous opportunities for ad-vancement in cell biology research, disease diagnostics, and drug discovery. In this paper, a molecular beacon (MB) specific for the important tumor suppressor gene p21 has been designed and synthesized. The fluorescence signal was detected in real-time after the MB entered the cytoplasm of nasopharyn-geal carcinoma cells. After injecting the p21MB into nasopharyngeal carcinoma cell and p33-trans-fected nasopharyngeal carcinoma cell, the consistent increase of fluorescent signal intensity was de-tected in both cell lines, and maximum fluorescence intensity achieved in about 15 min. In about 4 min following microinjection, the fluorescence increasing rate was significantly different between these two cell lines, which indicate the different p21 mRNA expression levels. The results obtained in the real-time detection were also validated by RT-PCR. Analysis of the initial fluorescence increasing rate can effi-ciently reduce the side effect of enzyme and improve the accuracy in living cell mRNA detection.

  8. Fluorescence resonance energy transfer from tryptophan in human serum albumin to a bioactive indoloquinolizine system

    Indian Academy of Sciences (India)

    Paramita Das; Arabinda Mallick; Basudeb Haldar; Alok Chakrabarty; Nitin Chattopadhyay

    2007-03-01

    The interaction between a bioactive molecule, 3-acetyl-4-oxo-6,7-dihydro-12H indolo-[2,3-a] quinolizine (AODIQ), with human serum albumin (HSA) has been studied using steady-state absorption and fluorescence techniques. A 1 : 1 complex formation has been established and the binding constant () and free energy change for the process have been reported. The AODIQ-HSA complex results in fluorescence resonance energy transfer (FRET) from the tryptophan moiety of HSA to the probe. The critical energy-transfer distance (0) for FRET and the Stern-Volmer constant (sv) for the fluorescence quenching of the donor in the presence of the acceptor have been determined. Importantly, SV has been shown to be equal to the binding constant itself, implying that the fluorescence quenching arises only from the FRET process. The study suggests that the donor and the acceptor are bound to the same protein at different locations but within the quenching distance.

  9. The fluorescence quenching and resonance Rayleigh scattering enhancement of ET-CPM system

    Institute of Scientific and Technical Information of China (English)

    YANG JiDong; ZHOU Shang

    2009-01-01

    In pH 4.4 BR buffer medium, erythrosine (ET) and Chlorphensmine Maleate (CPM) could form ion-as-sociation complex, which led to the quenching of fluorescence and synchronous fluorescence, and the significant enhancement of resonance Rayleigh scattering(RRS) of erythrosine. Furthermore, a new RRS spectrum would appear, and the maximum RRS wavelength was located at about 578 rim. The quenched fluorescence and enhanced RRS intensity was directly proportional to the concentration of CPM in the ranges of 0.24-8.0 μg/mL, and 0.008-3.6 μg/mL, respectively. The method has been ap-plied to determine CPM in urine samples with satisfactory results. The mechanisms of the RRS en-hancement and fluorescence quenching were discussed as well.

  10. Measuring distances within unfolded biopolymers using fluorescence resonance energy transfer: The effect of polymer chain dynamics on the observed fluorescence resonance energy transfer efficiency

    Science.gov (United States)

    Makarov, Dmitrii E.; Plaxco, Kevin W.

    2009-01-01

    Recent years have seen a number of investigations in which distances within unfolded proteins, polypeptides, and other biopolymers are probed via fluorescence resonance energy transfer, a method that relies on the strong distance dependence of energy transfer between a pair of dyes attached to the molecule of interest. In order to interpret the results of such experiments it is commonly assumed that intramolecular diffusion is negligible during the excited state lifetime. Here we explore the conditions under which this “frozen chain” approximation fails, leading to significantly underestimated donor-acceptor distances, and describe a means of correcting for polymer dynamics in order to estimate these distances more accurately. PMID:19725638

  11. Firefly Luciferase-Based Sequential Bioluminescence Resonance Energy Transfer (BRET)-Fluorescence Resonance Energy Transfer (FRET) Protease Assays.

    Science.gov (United States)

    Branchini, Bruce

    2016-01-01

    We describe here the preparation of ratiometric luminescent probes that contain two well-separated emission peaks produced by a sequential bioluminescence resonance energy transfer (BRET)-fluorescence resonance energy transfer (FRET) process. The probes are single soluble fusion proteins consisting of a thermostable firefly luciferase variant that catalyzes yellow-green (560 nm maximum) bioluminescence and a red fluorescent protein covalently labeled with a near-Infrared fluorescent dye. The two proteins are connected by a decapeptide containing a protease recognition site specific for factor Xa, thrombin, or caspase 3. The rates of protease cleavage of the fusion protein substrates were monitored by recording emission spectra and plotting the change in peak ratios over time. Detection limits of 0.41 nM for caspase 3, 1.0 nM for thrombin, and 58 nM for factor Xa were realized with a scanning fluorometer. This method successfully employs an efficient sequential BRET-FRET energy transfer process based on firefly luciferase bioluminescence to assay physiologically important protease activities and should be generally applicable to the measurement of any endoprotease lacking accessible cysteine residues.

  12. Discrete deexcitations in 235U from Nuclear Resonance Fluorescence

    Science.gov (United States)

    Kwan, E.; Howell, C. R.; Raut, R.; Rusev, G.; Tonchev, A. P.; Tornow, W.; Adekola, A. S.; Hammond, S. L.; Karwowski, H. J.; Pedroni, R.; Kelley, J. H.

    2010-11-01

    Systematics of the even-even rare-earth nuclei suggest a concentration of M1 excitations peaking around 3 MeV with a ∑B(M1) strength of ˜3μN^2. In addition, a linear dependence on the square of the ground-state deformation was observed in the systematics of the ∑B(M1) strengths. The actinide region is interesting for investigation of the ``scissors'' mode of M1 excitations because it has neutron-rich nuclei with large deformations. Evidence of M1 resonances concentrated around 2.0-2.5 MeV were found in ^238U & ^232Th. A research program has been initiated at TUNL to measure dipole transitions in the actinide using HIγS. Nearly monoenergic & circular polarized γ-ray beams below 3.0 MeV was used to measure transitions in ^235U. More than 20 transitions were observed. The integrated cross sections, B(M1) strengths & branching transitions intensities will be presented and compared with previous measurements.

  13. Interaction between fluorescein isothiocyanate and carbon dots: Inner filter effect and fluorescence resonance energy transfer

    Science.gov (United States)

    Liu, Huabing; Xu, Chaoyong; Bai, Yanli; Liu, Lin; Liao, Dongmei; Liang, Jiangong; Liu, Lingzhi; Han, Heyou

    2017-01-01

    Carbon dots (CDs) have been widely used for the preparation of multifunctional probes by conjugation with organic fluorescent dyes. However, the effect of organic fluorescent dyes on CDs still remains poorly understood. Herein, the effect of fluorescein isothiocyanate (FITC) on CDs was explored by spectroscopic techniques at pH 5.1, 7.0 and 9.0. The fluorescent intensity of CDs was found to be quenched gradually after mixing directly with different concentrations of FITC, but the fluorescent lifetime of CDs remained unchanged. According to the results of UV-vis absorption spectra and fluorescent lifetime measurements, a pH-dependent inner filter effect (IFE) between CDs and FITC was proposed. However, the fluorescent lifetime of CDs deceased after their conjugation with FITC, implying the fluorescence resonance energy transfer (FRET) between CDs and FITC. This study has revealed two different effects of FITC on CDs with varying pH values and provided useful theoretical guidelines for further research on the interaction between other nanoparticles and fluorophores.

  14. Mammalian cell-based optimization of the biarsenical-binding tetracysteine motif for improved fluorescence and affinity

    NARCIS (Netherlands)

    Martin, Brent R; Giepmans, Ben N G; Adams, Stephen R; Tsien, Roger Y

    2005-01-01

    Membrane-permeant biarsenical dyes such as FlAsH and ReAsH fluoresce upon binding to genetically encoded tetracysteine motifs expressed in living cells, yet spontaneous nonspecific background staining can prevent detection of weakly expressed or dilute proteins. If the affinity of the tetracysteine

  15. Fluorescence resonance energy transfer induced by conjugation of metalloproteins to nanoparticles

    Science.gov (United States)

    Pompa, P. P.; Chiuri, R.; Manna, L.; Pellegrino, T.; del Mercato, L. L.; Parak, W. J.; Calabi, F.; Cingolani, R.; Rinaldi, R.

    2006-01-01

    We show the possibility of realizing a hybrid system composed of a semiconductor nanoparticle (NP) and a metalloprotein, in which the photophysical properties of the two species can be exploited to elicit fluorescence resonance energy transfer mechanisms from the biomolecule to the NP. A specific conjugation process between CdSe/ZnS core/shell water soluble NPs, functionalized with surface exposed thiol groups (-SH), and the apo form of the metalloprotein azurin (Az) has been achieved, resulting in a fixed distance in the donor-acceptor pairs. The increase in the NP fluorescence intensity was found to be dependent on the Az to NP molar ratio.

  16. High-precision metrology of highly charged ions via relativistic resonance fluorescence.

    Science.gov (United States)

    Postavaru, O; Harman, Z; Keitel, C H

    2011-01-21

    Resonance fluorescence of laser-driven highly charged ions is investigated with regard to precisely measuring atomic properties. For this purpose an ab initio approach based on the Dirac equation is employed that allows for studying relativistic ions. These systems provide a sensitive means to test correlated relativistic dynamics, quantum electrodynamic phenomena and nuclear effects by applying x-ray lasers. We show how the narrowing of sidebands in the x-ray fluorescence spectrum by interference due to an additional optical driving can be exploited to determine atomic dipole or multipole moments to unprecedented accuracy.

  17. Correlation functions in resonance fluorescence with spectral resolution: Signal-processing approach

    Science.gov (United States)

    Shatokhin, Vyacheslav N.; Kilin, Sergei Ya.

    2016-09-01

    In the framework of the signal processing approach to single-atom resonance fluorescence with spectral resolution, we diagrammatically derive an analytical formula for arbitrary-order spectral correlation functions of the scattered fields that pass through Fabry-Perot interferometers. Our general expression is then applied to study correlation signals in the limit of well separated spectral lines of the resonance fluorescence spectrum. In particular, we study the normalized second-order temporal intensity correlation functions in the case of the interferometers tuned to the components of the spectrum and obtain interferential corrections to the approximate results derived in the secular limit. In addition, we explore purely spectral correlations and show that they can fully be understood in terms of the two-photon cascades down the dressed state ladder.

  18. Decay of the resonance fluorescence following pulsed excitation of a weakly disordered excitonic system

    Science.gov (United States)

    Boukahil, A.; Huber, D. L.

    1993-12-01

    A study is made of the decay of the resonance fluorescence following pulsed excitation of a weakly disordered system whose optical excitations are Frenkel excitons. The disorder is characterized by a Gaussian distribution of optical transition frequencies with no correlation between different sites. The duration of the resonant pulse is taken to be short in comparison with the reciprocal of the optical linewidth, and the wavelength of the light is assumed to be large in comparison with either the size of the array or the exciton mean free path associated with the disorder. In the limit where σ, the standard deviation of the Gaussian distribution, is much less than the exciton bandwidth, the integrated intensity of the fluorescence decays non-exponentially and is characterized by universal functions of σ xt, where x= 4/3, 2, and 4 in one, two, and three dimensions, respectively. Analytic approximations to the scaling functions in two and three dimensions are presented.

  19. Fluorescence resonance energy transfer between conjugated molecules infiltrated in three-dimensional opal photonic crystals

    Energy Technology Data Exchange (ETDEWEB)

    Zou, Lu; Sui, Ning; Wang, Ying-Hui, E-mail: yinghui_wang@jlu.edu.cn; Qian, Cheng; Ma, Yu-Guang; Zhang, Han-Zhuang, E-mail: zhanghz@jlu.edu.cn

    2015-02-15

    Fluorescence resonance energy transfer (FRET) from Coumarin 6 (C-6) to Sulforhodamine B (S-B) infiltrated into opal PMMA (poly-methyl-methacrylate) photonic crystals (PCs) has been studied in detail. The intrinsic mesh micro-porous structure of opal PCs could increase the luminescent efficiency through inhibiting the intermolecular interaction. Meanwhile, its structure of periodically varying refractive indices could also modify the FRET through affecting the luminescence characteristics of energy donor or energy acceptor. The results demonstrate that the FRET efficiency between conjugated dyes was easily modified by opal PCs. - Highlights: • We investigate the fluorescence resonance energy transfer between two kinds of dyes. • These two kinds of dyes are infiltrated in PMMA opal photonic crystals. • The structure of opal PCs could improve the luminescent characteristics. • The structure of opal PCs could improve the energy transfer characteristics.

  20. Investigation of Fluorescence Resonance Energy Transfer between Fluorescein and Rhodamine 6G

    Science.gov (United States)

    Saha, Jaba; Datta Roy, Arpan; Dey, Dibyendu; Chakraborty, Santanu; Bhattacharjee, D.; Paul, P. K.; Hussain, Syed Arshad

    2015-10-01

    Fluorescence Resonance Energy Transfer between two organic dyes Fluorescein and Rhodamine 6G was investigated in aqueous solution in presence and absence of synthetic clay laponite. Spectroscopic studies suggest that both the dyes were present mainly as monomer in solution. Fluorescence Resonance Energy Transfer occurred from Fluorescein to Rhodamine 6G in solutions. Energy transfer efficiency increases in presence of laponite and the maximum efficiency was 72.00% in aqueous laponite dispersion. Energy transfer efficiency was found to be pH sensitive. It has been demonstrated that with proper calibration it is possible to use the present system under investigation to sense pH over a wide range from 1.5 to 8.0.

  1. Fluorescence resonance energy transfer sensors for quantitative monitoring of pentose and disaccharide accumulation in bacteria

    Directory of Open Access Journals (Sweden)

    Looger Loren L

    2008-06-01

    Full Text Available Abstract Background Engineering microorganisms to improve metabolite flux requires detailed knowledge of the concentrations and flux rates of metabolites and metabolic intermediates in vivo. Fluorescence resonance energy transfer sensors represent a promising technology for measuring metabolite levels and corresponding rate changes in live cells. These sensors have been applied successfully in mammalian and plant cells but potentially could also be used to monitor steady-state levels of metabolites in microorganisms using fluorimetric assays. Sensors for hexose and pentose carbohydrates could help in the development of fermentative microorganisms, for example, for biofuels applications. Arabinose is one of the carbohydrates to be monitored during biofuels production from lignocellulose, while maltose is an important degradation product of starch that is relevant for starch-derived biofuels production. Results An Escherichia coli expression vector compatible with phage λ recombination technology was constructed to facilitate sensor construction and was used to generate a novel fluorescence resonance energy transfer sensor for arabinose. In parallel, a strategy for improving the sensor signal was applied to construct an improved maltose sensor. Both sensors were expressed in the cytosol of E. coli and sugar accumulation was monitored using a simple fluorimetric assay of E. coli cultures in microtiter plates. In the case of both nanosensors, the addition of the respective ligand led to concentration-dependent fluorescence resonance energy transfer responses allowing quantitative analysis of the intracellular sugar levels at given extracellular supply levels as well as accumulation rates. Conclusion The nanosensor destination vector combined with the optimization strategy for sensor responses should help to accelerate the development of metabolite sensors. The new carbohydrate fluorescence resonance energy transfer sensors can be used for in vivo

  2. Determination of gamma-ray widths in $^{15}$N using nuclear resonance fluorescence

    OpenAIRE

    Szücs, T.; Bemmerer, D.; Caciolli, A.; Fülöp, Zs.; Massarczyk, R.; Michelagnoli, C.; Reinhardt, T. P.; Schwengner, R.; Takács, M. P.; Ur, C. A.; Wagner, A.; Wagner, L.

    2015-01-01

    The stable nucleus $^{15}$N is the mirror of $^{15}$O, the bottleneck in the hydrogen burning CNO cycle. Most of the $^{15}$N level widths below the proton emission threshold are known from just one nuclear resonance fluorescence (NRF) measurement, with limited precision in some cases. A recent experiment with the AGATA demonstrator array determined level lifetimes using the Doppler Shift Attenuation Method (DSAM) in $^{15}$O. As a reference and for testing the method, level lifetimes in $^{1...

  3. Development of a Real-Time Fluorescence Resonance Energy Transfer PCR To Detect Arcobacter Species▿

    OpenAIRE

    Abdelbaqi, Khalil; Buissonnière, Alice; Prouzet-Mauleon, Valérie; Gresser, Jessica; Wesley, Irene; Mégraud, Francis; Ménard, Armelle

    2007-01-01

    A real-time PCR targeting the gyrase A subunit gene outside the quinolone resistance-determining region has been developed to detect Arcobacter species. The species identification was done by probe hybridization and melting curve analysis, using fluorescence resonance energy transfer technology. Discrimination between Arcobacter species was straightforward, as the corresponding melting points showed significant differences with the characteristic melting temperatures of 63.5°C, 58.4°C, 60.6°C...

  4. Fluorescent resonance energy transfer based detection of biological contaminants through hybrid quantum dot-quencher interactions.

    Science.gov (United States)

    Ramadurai, D; Norton, E; Hale, J; Garland, J W; Stephenson, L D; Stroscio, M A; Sivananthan, S; Kumar, A

    2008-06-01

    A nanoscale sensor employing fluorescent resonance energy transfer interactions between fluorescent quantum dots (QDs) and organic quencher molecules can be used for the multiplexed detection of biological antigens in solution. Detection occurs when the antigens to be detected displace quencher-labelled inactivated (or dead) antigens of the same type attached to QD-antibody complexes through equilibrium reactions. This unquenches the QDs, allowing detection to take place through the observation of photoluminescence in solution or through the fluorescence imaging of unquenched QD complexes trapped on filter surfaces. Multiplexing can be accomplished by using several different sizes of QDs, with each size QD labelled with an antibody for a different antigen, providing the ability to detect several types of antigens or biological contaminants simultaneously in near real-time with high specificity and sensitivity.

  5. Resonance fluorescence as a probe to elucidate mechanisms of intramolecular relaxation processes

    Energy Technology Data Exchange (ETDEWEB)

    Chen, K.; Yeung, E.S.

    1979-12-15

    Density matrix formalism is used to study resonance fluorescence processes of isolated gaseous molecules. The Wangsness--Bloch--Fano theory is applied to the irreversible intramolecular relaxation process. In the present treatment, it is assumed that the rotational angular momentum J is a constant of motion in the zeroth order approximation. Thus intramolecular interactions can be classified as tensor operators with respect to J. The fluorescence decay patterns depend on excitation bandwidths and monitoring conditions. Under narrow band excitation, the population (monopole), orientation (dipole), and alignment (quadrupole) of the excited state density matrix rho/sub e/ can be prepared. For a sharply prepared angular momentum state under pulsed excitation, the population does not undergo radiationless decay and the resonance fluorescence intensity is a sum of three exponentials in general. For cw experiments, where lifetime limited linewidth or polarized fluorescence intensities are measured, the population can decay via an intramolecular relaxation channel providing the orientation component is initially prepared. Explicit J dependences of lifetimes are given for pulsed experiments. Under broad band excitation, quantum beats due to the coherence of rho/sub e/ are derived from the Liouville equation. Most important of all, the population can decay under pulsed, broad-band excitation conditions. The scalar part of the nonadiabatic coupling becomes an additional contributor to the intramolecular relaxation process in this last case.

  6. Resonance fluorescence of strongly driven two-level system coupled to multiple dissipative reservoirs

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Yiying, E-mail: yiyingyan@sjtu.edu.cn; Lü, Zhiguo, E-mail: zglv@sjtu.edu.cn; Zheng, Hang, E-mail: hzheng@sjtu.edu.cn

    2016-08-15

    We present a theoretical formalism for resonance fluorescence radiating from a two-level system (TLS) driven by any periodic driving and coupled to multiple reservoirs. The formalism is derived analytically based on the combination of Floquet theory and Born–Markov master equation. The formalism allows us to calculate the spectrum when the Floquet states and quasienergies are analytically or numerically solved for simple or complicated driving fields. We can systematically explore the spectral features by implementing the present formalism. To exemplify this theory, we apply the unified formalism to comprehensively study a generic model that a harmonically driven TLS is simultaneously coupled to a radiative reservoir and a dephasing reservoir. We demonstrate that the significant features of the fluorescence spectra, the driving-induced asymmetry and the dephasing-induced asymmetry, can be attributed to the violation of detailed balance condition, and explained in terms of the driving-related transition quantities between Floquet-states and their steady populations. In addition, we find the distinguished features of the fluorescence spectra under the biharmonic and multiharmonic driving fields in contrast with that of the harmonic driving case. In the case of the biharmonic driving, we find that the spectra are significantly different from the result of the RWA under the multiple resonance conditions. By the three concrete applications, we illustrate that the present formalism provides a routine tool for comprehensively exploring the fluorescence spectrum of periodically strongly driven TLSs.

  7. Paper-based fluorescence resonance energy transfer assay for directly detecting nucleic acids and proteins.

    Science.gov (United States)

    Li, Hua; Fang, Xueen; Cao, Hongmei; Kong, Jilie

    2016-06-15

    Paper-based fluorescence resonance energy transfer assay (FRET) is gaining great interest in detecting macro-biological molecule. It is difficult to achieve conveniently and fast detection for macro-biological molecule. Herein, a graphene oxide (GO)-based paper chip (glass fiber) integrated with fluorescence labeled single-stranded DNA (ssDNA) for fast, inexpensive and direct detection of biological macromolecules (proteins and nucleic acids) has been developed. In this paper, we employed the Cy3/FAM-labeled ssDNA as the reporter and the GO as quencher and the original glass fiber paper as data acquisition substrates. The chip which was designed and fabricated by a cutting machine is a miniature biosensor that monitors fluorescence recovery from resonance energy transfer. The hybridization assays and fluorescence detection were all simplified, and the surface of the chip did not require immobilization or washing. A Nikon Eclipse was employed as excited resource and a commercial digital camera was employed for capturing digital images. This paper-based microfluidics chip has been applied in the detection of proteins and nucleic acids. The biosensing capability meets many potential requirements for disease diagnosis and biological analysis.

  8. Resonance fluorescence of strongly driven two-level system coupled to multiple dissipative reservoirs

    Science.gov (United States)

    Yan, Yiying; Lü, Zhiguo; Zheng, Hang

    2016-08-01

    We present a theoretical formalism for resonance fluorescence radiating from a two-level system (TLS) driven by any periodic driving and coupled to multiple reservoirs. The formalism is derived analytically based on the combination of Floquet theory and Born-Markov master equation. The formalism allows us to calculate the spectrum when the Floquet states and quasienergies are analytically or numerically solved for simple or complicated driving fields. We can systematically explore the spectral features by implementing the present formalism. To exemplify this theory, we apply the unified formalism to comprehensively study a generic model that a harmonically driven TLS is simultaneously coupled to a radiative reservoir and a dephasing reservoir. We demonstrate that the significant features of the fluorescence spectra, the driving-induced asymmetry and the dephasing-induced asymmetry, can be attributed to the violation of detailed balance condition, and explained in terms of the driving-related transition quantities between Floquet-states and their steady populations. In addition, we find the distinguished features of the fluorescence spectra under the biharmonic and multiharmonic driving fields in contrast with that of the harmonic driving case. In the case of the biharmonic driving, we find that the spectra are significantly different from the result of the RWA under the multiple resonance conditions. By the three concrete applications, we illustrate that the present formalism provides a routine tool for comprehensively exploring the fluorescence spectrum of periodically strongly driven TLSs.

  9. Magnetic Separation-Assistant Fluorescence Resonance Energy Transfer Inhibition for Highly Sensitive Probing of Nucleolin.

    Science.gov (United States)

    Li, Yan-Ran; Liu, Qian; Hong, Zhangyong; Wang, He-Fang

    2015-12-15

    For the widely used "off-on" fluorescence (or phosphorescence) resonance energy transfer (FRET or PRET) system, the separation of donors and acceptors species was vital for enhancing the sensitivity. To date, separation of free donors from FRET/PRET inhibition systems was somewhat not convenient, whereas separation of the target-induced far-between acceptors has hardly been reported yet. We presented here a novel magnetic separation-assistant fluorescence resonance energy transfer (MS-FRET) inhibition strategy for highly sensitive detection of nucleolin using Cy5.5-AS1411 as the donor and Fe3O4-polypyrrole core-shell (Fe3O4@PPY) nanoparticles as the NIR quenching acceptor. Due to hydrophobic interaction and π-π stacking of AS1411 and PPY, Cy5.5-AS1411 was bound onto the surface of Fe3O4@PPY, resulting in 90% of fluorescence quenching of Cy5.5-AS1411. Owing to the much stronger specific interaction of AS1411 and nucleolin, the presence of nucleolin could take Cy5.5-AS1411 apart from Fe3O4@PPY and restore the fluorescence of Cy5.5-AS1411. The superparamagnetism of Fe3O4@PPY enabled all separations and fluorescence measurements complete in the same quartz cell, and thus allowed the convenient but accurate comparison of the sensitivity and fluorescence recovery in the cases of separation or nonseparation. Compared to nonseparation FRET inhibition, the separation of free Cy5.5-AS1411 from Cy5.5-AS1411-Fe3O4@PPY solution (the first magnetic separation, MS-1) had as high as 25-fold enhancement of the sensitivity, whereas further separation of the nucleolin-inducing far-between Fe3O4@PPY from the FRET inhibition solution (the second magnetic separation, MS-2) could further enhance the sensitivity to 35-fold. Finally, the MS-FRET inhibition assay displayed the linear range of 0.625-27.5 μg L(-1) (8.1-359 pM) and detection limit of 0.04 μg L(-1) (0.05 pM) of nucleolin. The fluorescence intensity recovery (the percentage ratio of the final restoring fluorescence intensity

  10. The Effect of Incoherent Population Pumping on Squeezing in Resonance Fluorescence

    Institute of Scientific and Technical Information of China (English)

    CHEN Zhaoyang; ZHANG Jingtao; XU Zhizhan

    2000-01-01

    The effect of incoherent population pumping on the steady-state population inversion and the quadrature squeezing spectra produced in the resonance fluorescence of a two-level atom is investigated. In the presence of incoherent population pumping, the steady-state population inversion is increased for small frequency detuning but is not changed for large frequency detuning. For resonant excitation at low intensities, the weak incoherent pumping degrades the degree of the squeezing and shifts the position of the maximum squeezing; for off-resonant excitation at strong intensities, the weak incoherent pumping hardly changes the squeezing spectra. But when the incoherent pumping is strong the squeezing may be completely destroyed for both cases.

  11. Carbon quantum dots as fluorescence resonance energy transfer sensors for organophosphate pesticides determination.

    Science.gov (United States)

    Wu, Xiaoli; Song, Yang; Yan, Xu; Zhu, Chengzhou; Ma, Yongqiang; Du, Dan; Lin, Yuehe

    2017-03-07

    Carbon quantum dots (CQDs) obtained from natural organics attract significant attention due to the abundance of carbon sources, varieties of heteroatom doping (such as N, S, P) and good biocompatibility of precursor. In this study, tunable fluorescence emission CQDs originated from chlorophyll were synthesized and characterized. The fluorescence emission can be effectively quenched by gold nanoparticles (Au NPs) via fluorescence resonance energy transfer (FRET). Thiocholine, which was produced from acetylthiocholine (ATC) by the hydrolysis of butyrylcholinesterase (BChE), could cause the aggregation of Au NPs and the corresponding recovery of FRET-quenched fluorescence emission. The catalytic activity of BChE could be irreversibly inhibited by organophosphorus pesticides (OPs), thus, the recovery effect was reduced. By evaluating the fluorescence emission intensity of CQDs, a FRET-based sensing platform for OPs determination was established. Paraoxon was studied as an example of OPs. The sensing platform displayed a linear relationship with the logarithm of the paraoxon concentrations in the range of 0.05-50μgL(-1) and the limit of detection (LOD) was 0.05μgL(-1). Real sample study in tap and river water revealed that this sensing platform was repeatable and accurate. The results indicate that the OP sensor is promising for applications in food safety and environmental monitoring.

  12. Amplification of the Signal Intensity of Fluorescence-Based Fiber-Optic Biosensors Using a Fabry-Perot Resonator Structure

    Directory of Open Access Journals (Sweden)

    Meng-Chang Hsieh

    2015-02-01

    Full Text Available Fluorescent biosensors have been widely used in biomedical applications. To amplify the intensity of fluorescence signals, this study developed a novel structure for an evanescent wave fiber-optic biosensor by using a Fabry-Perot resonator structure. An excitation light was coupled into the optical fiber through a laser-drilled hole on the proximal end of the resonator. After entering the resonator, the excitation light was reflected back and forth inside the resonator, thereby amplifying the intensity of the light in the fiber. Subsequently, the light was used to excite the fluorescent molecules in the reactive region of the sensor. The experimental results showed that the biosensor signal was amplified eight-fold when the resonator reflector was formed using a 92% reflective coating. Furthermore, in a simulation, the biosensor signal could be amplified 20-fold by using a 99% reflector.

  13. Fluorescence resonance energy transfer biosensor between upconverting nanoparticles and palladium nanoparticles for ultrasensitive CEA detection.

    Science.gov (United States)

    Li, Hui; Shi, Liang; Sun, De-En; Li, Peiwu; Liu, Zhihong

    2016-12-15

    An ultrasensitive biosensor for carcinoembryonic antigen (CEA) was constructed based on fluorescence resonance energy transfer (FRET) between upconverting nanoparticles (UCPs) and palladium nanoparticles (PdNPs). PdNPs was synthesized by the addition of a solution of Na2PdCl4 into a mixture of N2H4·H2O as the reducing agent and 11-mercaptoundecanoic acid (MUDA) as the stabilizer. The CEA aptamer (5'-NH2-ATACCAGCTTATTCAATT-3') was conjugated to hexanedioic acid (HDA) modified UCPs (HDA-UCPs) through an EDC-NHS coupling protocol. The coordination interaction between nitrogen functional groups of the CEA aptamer and PdNPs brought UCPs and PdNPs in close proximity, which resulted in the fluorescence quenching of UCPs to an extent of 85%. And the non-specific fluorescence quenching caused by PdNPs towards HDA-UCPs was negligible. After the introduction of CEA into the UCPs-CEA aptamer-PdNPs fluorescence quenching system, the CEA aptamer preferentially combined with CEA accompanied by the conformational change which weakened the coordination interaction between the CEA aptamer and PdNPs. So fluorescence recovery of UCPs was observed and a linear relationship between the fluorescence recovery of UCPs and the concentration of CEA was obtained in the range from 2pg/mL to 100pg/mL in the aqueous buffer with the detection limit of 0.8pg/mL. The ultrasensitive detection of CEA was also realized in diluted human serum with a linear range from 4pg/mL to 100pg/mL and a detection limit of 1.7pg/mL. This biosensor makes the most of the high quenching ability of PdNPs towards UCPs with negligible non-specific fluorescence quenching and has broad application prospects in biochemistry.

  14. Intrinsic Tryptophan Fluorescence in the Detection and Analysis of Proteins: A Focus on Förster Resonance Energy Transfer Techniques

    Directory of Open Access Journals (Sweden)

    Amar B. T. Ghisaidoobe

    2014-12-01

    Full Text Available F resonance energy transfer (FRET occurs when the distance between a donor fluorophore and an acceptor is within 10 nm, and its application often necessitates fluorescent labeling of biological targets. However, covalent modification of biomolecules can inadvertently give rise to conformational and/or functional changes. This review describes the application of intrinsic protein fluorescence, predominantly derived from tryptophan (\\(\\uplambda_{\\textsc{ex}}\\sim\\ nm, \\(\\uplambda_{\\textsc{em}}\\sim\\ 350 nm, in protein-related research and mainly focuses on label-free FRET techniques. In terms of wavelength and intensity, tryptophan fluorescence is strongly influenced by its (or the proteinlocal environment, which, in addition to fluorescence quenching, has been applied to study protein conformational changes. Intrinsic F resonance energy transfer (iFRET, a recently developed technique, utilizes the intrinsic fluorescence of tryptophan in conjunction with target-specific fluorescent probes as FRET donors and acceptors, respectively, for real time detection of native proteins.

  15. Specific survivin dual fluorescence resonance energy transfer molecular beacons for detection of human bladder cancer cells

    Institute of Scientific and Technical Information of China (English)

    Zhi-qiang WANG; Jun ZHAO; Jin ZENG; Kai-jie WU; Yu-le CHEN; Xin-ya ng WANG; Luke S CHANG; Da-lin HE

    2011-01-01

    Survivin molecular beacons can be used to detectbladder cancer cells in urine samples non-invasively.The aim of this study is to improve the specificity of detection of bladder cancer cells using survivin dual fluorescence resonance energy transfer molecular beacons (FRET MBs) that have fluorophores forming one donor-acceptor pair.Methods:Survivin-targeting dual fluorescence resonance energy transfer molecular beacons with unique target sequences were designed,which had no overlap with the other genes in the apoptosis inhibitor protein family.Human bladder cancer cell lines 5637,253J and T24,as well as the exfoliated cells in the urine of healthy adults and patients with bladder cancer were examined.Images of cells were taken using a laser scanning confocal fluorescence microscope.For assays using dual FRET MBs,the excitation wavelength was 488 nm,and the emission detection wavelengths were 520+20 nm and 560+20 nm,respectively.Results:The human bladder cancer cell lines and exfoliated cells in the urine of patients with bladder cancer incubated with the survivin dual FRET MBs exhibited strong fluorescence signals.In contrast,no fluorescence was detected in the survivin-negative human dermal fibroblasts-adult (HDF-a) cells or exfoliated cells in the urine of healthy adults incubated with the survivin dual FRET MBs.Conclusion:The results suggest that the survivin dual FRET MBs may be used as a specific and non-invasive method for early detection and follow-up of patients with bladder cancer.

  16. Folic acid-targeted magnetic Tb-doped CeF3 fluorescent nanoparticles as bimodal probes for cellular fluorescence and magnetic resonance imaging.

    Science.gov (United States)

    Ma, Zhi-Ya; Liu, Yu-Ping; Bai, Ling-Yu; An, Jie; Zhang, Lin; Xuan, Yang; Zhang, Xiao-Shuai; Zhao, Yuan-Di

    2015-10-07

    Magnetic fluorescent nanoparticles (NPs) have great potential applications for diagnostics, imaging and therapy. We developed a facile polyol method to synthesize multifunctional Fe3O4@CeF3:Tb@CeF3 NPs with small size (magnetic resonance imaging (MRI) of HeLa cells with overexpressed folate receptors (FR). The results indicated that these NPs had strong luminescence and enhanced T2-weighted MR contrast and would be promising candidates as multimodal probes for both fluorescence and MRI imaging.

  17. Probing symmetry and symmetry breaking in resonant soft-x-ray fluorescence spectra of molecules

    Energy Technology Data Exchange (ETDEWEB)

    Glans, P.; Gunnelin, K.; Guo, J. [Uppsala Univ. (Sweden)] [and others

    1997-04-01

    Conventional non-resonant soft X-ray emission brings about information about electronic structure through its symmetry and polarization selectivity, the character of which is governed by simple dipole rules. For centro-symmetric molecules with the emitting atom at the inversion center these rules lead to selective emission through the required parity change. For the more common classes of molecules which have lower symmetry or for systems with degenerate core orbitals (delocalized over identical sites), it is merely the local symmetry selectivity that provides a probe of the local atomic orbital contribution to the molecular orbital. For instance, in X-ray spectra of first row species the intensities essentially map the p-density at each particular atomic site, and, in a molecular orbital picture, the contribution of the local p-type atomic orbitals in the LCAO description of the molecular orbitals. The situation is different for resonant X-ray fluorescence spectra. Here strict parity and symmetry selectivity gives rise to a strong frequency dependence for all molecules with an element of symmetry. In addition to symmetry selectivity the strong frequency dependence of resonant X-ray emission is caused by the interplay between the shape of a narrow X-ray excitation energy function and the lifetime and vibrational broadenings of the resonantly excited core states. This interplay leads to various observable effects, such as linear dispersion, resonance narrowing and emission line (Stokes) doubling. Also from the point of view of polarization selectivity, the resonantly excited X-ray spectra are much more informative than the corresponding non-resonant spectra. Examples are presented for nitrogen, oxygen, and carbon dioxide molecules.

  18. Efficient HOMO-LUMO separation by multiple resonance effect toward ultrapure blue thermally activated delayed fluorescence

    Science.gov (United States)

    Hatakeyama, Takuji; Ikuta, Toshiaki; Shiren, Kazushi; Nakajima, Kiichi; Nomura, Shintaro; Ni, Jingping

    2016-09-01

    Organic light-emitting diodes (OLEDs) play an important role in the new generation of flat-panel displays. Conventional OLEDs employing fluorescent materials together with triplet-triplet annihilation suffer from a relatively low internal quantum efficiency (IQE) of 62.5%. On the other hand, the IQE of OLEDs employing phosphorescent or thermally activated delayed fluorescence (TADF) materials can reach 100%. However, these materials exhibit very broad peaks with a full-width at half-maximum (FWHM) of 70-100 nm and cannot satisfy the color-purity requirements for displays. Therefore, the latest commercial OLED displays employ blue fluorescent materials with a relatively low IQE, and efficient blue emitters with a small FWHM are highly needed. In our manuscript, we present organic molecules that exhibit ultrapure blue fluorescence based on TADF. These molecules consist of three benzene rings connected by one boron and two nitrogen atoms, which establish a rigid polycyclic framework and significant localization of the highest occupied and lowest unoccupied molecular orbitals by a multiple resonance effect. An OLED device based on the new emitter exhibits ultrapure blue emission at 467 nm with an FWHM of 28 nm, Commission Internationale de l'Eclairage (CIE) coordinates of (0.12, 0.13), and an IQE of 100%, which represent record-setting performance for blue OLED devices.

  19. Novel fluorescent silica nanoparticle probe for Förster Resonance Energy Transfer

    Directory of Open Access Journals (Sweden)

    Mouhamad Khalil

    2014-04-01

    Full Text Available The preparation and utilization of a novel particulate label based on fluorescent hybrid silica nanoparticles are reported in this article. These fluorescent nanoparticles have shown several unique advantages over existing dye molecules, quantum dots, and latex-based fluorescent particles in easy preparation, good photostability and high sensitivity. Alexa Fluor® dyes, the leading and most trusted fluorescent dyes available today were used to make the grafting onto silica nanoparticles. Alexa Fluor® was modified with 3-aminopropyl-triethoxysilane (APS to provide reactive groups Alexa-APS. Alexa-APS can be grafted onto silica nanoparticles via silylation process. The silica nanoparticles were prepared by Stöber methods and treated by Etching to functionalized their surface. A mixture of Alexafluoro donor A488 and Alexafluoro acceptor A568 coated silica nanoparticles leads to the Förster Resonance Energy Transfer (FRET by excitation of the acceptor when the particles are distant by less than 10nm.

  20. Nicking enzyme-assisted biosensor for Salmonella enteritidis detection based on fluorescence resonance energy transfer.

    Science.gov (United States)

    Song, Yang; Li, Wenkai; Duan, Yingfen; Li, Zhongjie; Deng, Le

    2014-05-15

    Salmonella enteritidis (S. enteritidis) outbreaks continue to occur, and have increased public awareness of this pathogen. Nicking endonuclease Nb.BbvC I is widely used for the detection of biomolecules and displays activity for specific double-stranded DNA (dsDNA). In this study, we developed a biosensor to detect S. enteritidis based on fluorescence resonance energy transfer (FRET) using nicking enzyme and carbon nanoparticles (CNPs). Because of the quenching effect of black hole quencher 1 (BHQ 1), the CNPs do not fluoresce in the reaction system. When the target bacteria are added, the nicking enzyme recognizes and cleaves the dsDNA fabricated by the interaction between probe and target. As a result, the CNPs dissociate from BHQ 1 and emit strong fluorescence. Using the nicking enzyme, the fluorescence signals of the biosensor are greatly amplified. The biosensor exhibited a linear relationship with the concentration of S. enteritidis ranging from 10(2) to 3 × 10(3)CFU/mL in water and from 1.5 × 10(2) to 3 × 10(3)CFU/mL in milk. The present results indicate that our FRET-based detection system can be widely employed for the effective detection of pathogens.

  1. Highly indistinguishable on-demand resonance fluorescence photons from a deterministic quantum dot micropillar device with 74% extraction efficiency

    DEFF Research Database (Denmark)

    Gregersen, Niels

    2016-01-01

    device is a major challenge. Here, we report on the observation of bright single photon emission generated via pulsed, resonance fluorescence conditions from a single quantum dot (QD) deterministically centered in a micropillar cavity device via cryogenic optical lithography. The brightness of the QD...... fluorescence is greatly enhanced on resonance with the fundamental mode of the pillar, leading to an overall device efficiency of η = (74 ± 4) % for a single photon emission as pure as g (2) (0) = 0.0092 ± 0.0004. The combination of large Purcell enhancement and resonant pumping conditions allows us to observe...... a two-photon wave packet overlap up to ν = (88±3) %....

  2. Measuring evaporation rates of laser-trapped droplets by use of fluorescent morphology-dependent resonances.

    Science.gov (United States)

    Pastel, R; Struthers, A

    2001-05-20

    Morphology-dependent resonances (MDRs) are used to measure accurately the evaporation rates of laser-trapped 1- to 2-mum droplets of ethylene glycol. Droplets containing 3 x 10(-5) M Rhodamine-590 laser dye are optically trapped in a 20-mum hollow fiber by two counterpropagating 150-mW, 800-nm laser beams. A weaker 532-nm laser excites the dye, and fluorescence emission is observed near 560 nm as the droplet evaporates. A complete series of first-order TE and TM MDRs dominates the fluorescent output. MDR mode identification sizes the droplets and provides accurate evaporation rates. We verify the automated MDR mode identification by counting fringes in a videotape of the experiment. The longitudinal spring constant of the trap, measured by analysis of the videotaped motion of droplets perturbed from the trap center, provides independent verification of the laser's intensity within the trap.

  3. RecQ Helicase-catalyzed DNA Unwinding Detected by Fluorescence Resonance Energy Transfer

    Institute of Scientific and Technical Information of China (English)

    Xing-Dong ZHANG; Shuo-Xing DOU; Ping XIE; Peng-Ye WANG; Xu Guang XI

    2005-01-01

    A fluorometric assay was used to study the DNA unwinding kinetics induced by Escherichia coli RecQ helicase. This assay was based on fluorescence resonance energy transfer and carried out on stopped-flow, in which DNA unwinding was monitored by fluorescence emission enhancement of fluorescein resulting from helicase-catalyzed DNA unwinding. By this method, we determined the DNA unwinding rate of RecQ at different enzyme concentrations. We also studied the dependences of DNA unwinding magnitude and rate on magnesium ion concentration. We showed that this method could be used to determine the polarity of DNA unwinding. This assay should greatly facilitate further study of the mechanism for RecQcatalyzed DNA unwinding.

  4. The Photochemical Study of HSA and BSA with Resonance Light-Scattering and Fluorescence Spectra

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The resonance light-scattering (RLS) of human serum albumin (HSA) and bovine serum albumin (BSA) is reported for the first time, and applied to study photochemical reaction of HSA and BSA. The fact of photocrosslinking self-association effect in HSA and BSA solutions is identified by the enhancement of RLS. The fluorescence quenching at about 350 nm and 700 nm proves that tryptophan (Trp) residues are one of the photochemical activity sites in HSA and BSA molecules. The Rayleigh scattering (RS) spectra of HSA and BSA that were neglected in fluorescence spectra before are found at about 296 nm, 592 nm and 888 nm for the first time, and are of adventageous to studying the aggregation of HSA or BSA. The possible photochemical reaction mechanism is also proposed.

  5. Upconversion nanoparticle-based fluorescence resonance energy transfer assay for organophosphorus pesticides.

    Science.gov (United States)

    Long, Qian; Li, Haitao; Zhang, Youyu; Yao, Shouzhuo

    2015-06-15

    This paper reports a novel nanosensor for organophosphorus pesticides based on the fluorescence resonance energy transfer (FRET) between NaYF4:Yb,Er upconversion nanoparticles (UCNPs) and gold nanoparticles (AuNPs). The detection mechanism is based on the facts that AuNPs quench the fluorescence of UCNPs and organophosphorus pesticides (OPs) inhibit the activity of acetylcholinesterase (AChE) which catalyzes the hydrolysis of acetylthiocholine (ATC) into thiocholine. Under the optimized conditions, the logarithm of the pesticides concentration was proportional to the inhibition efficiency. The detection limits of parathion-methyl, monocrotophos and dimethoate reached 0.67, 23, and 67 ng/L, respectively. Meanwhile, the biosensor shows good sensitivity, stability, and could be successfully applied to detection of OPs in real food samples, suggesting the biosensor has potentially extensive application clinic diagnoses assays.

  6. Far-field resonance fluorescence from a dipole-interacting laser-driven cold atomic gas

    Science.gov (United States)

    Jones, Ryan; Saint, Reece; Olmos, Beatriz

    2017-01-01

    We analyze the temporal response of the fluorescence light that is emitted from a dense gas of cold atoms driven by a laser. When the average interatomic distance is comparable to the wavelength of the photons scattered by the atoms, the system exhibits strong dipolar interactions and collective dissipation. We solve the exact dynamics of small systems with different geometries and show how these collective features are manifest in the scattered light properties such as the photon emission rate, the power spectrum and the second-order correlation function. By calculating these quantities beyond the weak (linear) driving limit, we make progress in understanding the signatures of collective behavior in these many-body systems. Furthermore, we shed light on the role of disorder and averaging on the resonance fluorescence, of direct relevance for recent experimental efforts that aim at the exploration of many-body effects in dipole-dipole interacting gases of atoms.

  7. Multiway study of hybridization in nanoscale semiconductor labeled DNA based on fluorescence resonance energy transfer

    DEFF Research Database (Denmark)

    Gholami, Somayeh; Kompany Zare, Mohsen

    2013-01-01

    The resolution of the ternary-binary complex competition of a target sequence and of its two complementary probes in sandwich DNA hybridization is reported. To achieve this goal, Fluorescence Resonance Energy Transfer (FRET) between oligonucleotide-functionalized quantum dot (QD) nanoprobes (QD...... in the photoluminescence excitation (PLE) plot. From the obtained data, energy transfer efficiency and Forster radius (R-0) were calculated. In particular, our results demonstrated that energy transfer by using QD donor-QD acceptor FRET pairs is more efficient in comparison with QD donor-organic dye acceptor pairs. Soft...

  8. Quantum dot-DNA bioconjugates for fluorescence-resonance-energy-transfer-based biosensing

    Science.gov (United States)

    Medintz, Igor L.; Berti, Lorenzo; Pons, Thomas; Mattoussi, Hedi

    2007-02-01

    Semiconductor quantum dots (QDs) have unique photophysical properties which make them excellent fluorescence resonance energy transfer donors. However, lack of facile methods for conjugating biomolecules such as DNA, proteins and peptides to QDs have limited their applications. In this report, we describe a general procedure for the preparation of a synthetic peptide that can be covalently attached to DNA segments and used to facilitate the self-assembly of the modified DNA onto water soluble QDs. To characterize this conjugation strategy, dye-labeled DNA is first reacted with the synthetic peptide and the resulting peptide-DNA then self-assembled onto QDs. QD attachment is verified by monitoring resonance energy transfer efficiency from the QD donor to the dye-labeled DNA acceptor. QD-DNA bioconjugates assembled using this method may find applications as molecular beacons and hybridization probes.

  9. The effect of gold colloid on the fluorescence spectrum from Safranine T: a physical mechanism based on resonance light scattering.

    Science.gov (United States)

    Li, Jianjun; Zhu, Jian; Zhao, Junwu

    2011-05-01

    The non-monotonic change of fluorescence intensity from Safranine T was observed experimentally by adding gold colloids to the Safranine T aqueous solution. Increasing the content of gold colloids leads to the fluorescence emission peak red shift and increases first and then decreases. We believe that one of the important reasons of the fluorescence changing and shifting is due to the resonance light scattering from gold nanoparticles which is stimulated by the fluorescence of Safranine T and is tunable by the shell thickness of the ST that coated around the gold particles.

  10. A cell-based time-resolved fluorescence assay for selection of antibody reagents for G protein-coupled receptor immunohistochemistry.

    Science.gov (United States)

    Su, Jui-Lan; Fornwald, Jim; Rivers, Philip; Goldsworthy, Susan; Looney, Noeleen A; Hanvey, Jeff; Plumpton, Chris; Parham, Janet; Romanos, Michael; Kost, Thomas A; Kull, Frederick C

    2004-08-01

    A cell-based time-resolved fluorescence (celTRF) immunoassay is described for pre-screening antibodies to G protein-coupled receptor (GPCR) peptides that predicts suitability for immunohistochemistry (IHC). Rat GPCRs were expressed in Saos-2 human osteosarcoma cells via recombinant baculoviruses designed for mammalian cell expression, i.e., the transduced cells were used as a "screening lawn". The lawn was fixed and permeabilized similarly to IHC tissue. The celTRF, a dissociation-enhanced lanthanide fluorescence immunoassay (DELFIA), employed Eu-labelled goat anti-rabbit IgG. It exhibited a broad dynamic range upon which enzyme-linked immunosorbant assay (ELISA)-positive affinity-purified anti-peptide antibody reagents were examined for specificity and potency. Over 150 anti-peptide reagents to 27 GPCRs were characterized. All celTRF-positive antibodies were found to be suitable for IHC, whereas ELISA alone did not predict IHC utility. Examples are illustrated with five rabbit anti-neuropeptide FF receptor 1 (NPFF1) antibodies, where a strong correlation between celTRF potency and IHC utility was observed in both applications. In contrast, two high anti-peptide ELISA titer but celTRF-negative antibodies failed to recognize the NPFF1 receptor in IHC. The celTRF assay was performed manually and in an automated fashion, in our case, using a Biomek FX station and Sami scheduling software. The celTRF is the first in vitro automated assay that offers confident pre-selection of antibodies for IHC and the versatility to accommodate the rapid screening of large numbers of GPCRs. The celTRF is readily applicable to other protein target classes.

  11. High Repetition Rate, LINAC-based Nuclear Resonance Fluorescence FY 2009 Final Report

    Energy Technology Data Exchange (ETDEWEB)

    Mathew Kinlaw; Scott Watson; James Johnson; Alan Hunt; Heather Seipel; Edward Reedy

    2009-10-01

    Nuclear Resonance Fluorescence (NRF), which is possible for nuclei with atomic numbers greater than helium (Z=2), occurs when a nuclear level is excited by resonant absorption of a photon and subsequently decays by reemission of a photon. The excited nuclear states can become readily populated, provided the incident photon’s energy is within the Doppler-broadened width of the energy level being excited. Utilizing continuous energy photon spectra, as is characteristic of a bremsstrahlung photon beam, as the inspection source, ensures that at least some fraction of the impinging beam will contribute to the population of the excited energy levels in the material of interest. Upon de-excitation, either to the ground state or to a lower-energy excited state, the emitted fluorescence photon’s energy will correspond to the energy difference between the excited state and the state to which it decays. As each isotope inherently contains unique nuclear energy levels, the NRF states for each isotope are also unique. By exploiting this phenomenon, NRF photon detection provides a well-defined signature for identifying the presence of individual nuclear species. This report summarizes the second year (Fiscal Year [FY] 2009) of a collaborative research effort between Idaho National Laboratory, Idaho State University’s Idaho Accelerator Center, and Pacific Northwest National Laboratory. This effort focused on continuing to assess and optimize NRF-based detection techniques utilizing a slightly modified, commercially available, pulsed medical electron accelerator.

  12. Fluorescent material concentration dependency: Förster resonance energy transfer in quasi-solid state DSSCs

    Science.gov (United States)

    Kim, Dong Woo; Jo, Hyun-Jun; Thogiti, Suresh; Yang, Weon Ki; Cheruku, Rajesh; Kim, Jae Hong

    2017-03-01

    Förster resonance energy transfer (FRET) is critical for wide spectral absorption, an increased dye loading, and photocurrent generation of dye-sensitized solar cells (DSSCs). This process consists of organic fluorescent materials (as an energy donor), and an organic dye (as an energy acceptor on TiO2 surfaces) with quasi-solid electrolyte. The judicious choice of the energy donor and acceptor facilitates a strong spectral overlap between the emission and absorption regions of the fluorescent materials and dye. This FRET process enhances the light-harvesting characteristics of quasi-solid state DSSCs. In this study, DSSCs containing different concentrations (0, 1, and 1.5 wt%) of a fluorescent material (FM) as the energy donor are investigated using FRET. The power conversion efficiency of DSSCs containing FMs in a quasi-solid electrolyte increased by 33% over a pristine cell. The optimized cell fabricated with the quasi-solid state DSSC containing 1.0 wt% FM shows a maximum efficiency of 3.38%, with a short-circuit current density (J SC ) of 4.32 mA/cm-2, and an open-circuit voltage (V OC ) of 0.68 V under illumination of simulated solar light (AM 1.5G, 100 mW/cm-2). [Figure not available: see fulltext.

  13. Absorption, fluorescence and resonance Rayleigh scattering spectral characteristics of interaction of gold nanoparticle with safranine T

    Institute of Scientific and Technical Information of China (English)

    HE Youqiu; LIU Shaopu; LIU Qin; LIU Zhongfang; HU Xiaoli

    2005-01-01

    The interaction between gold nanoparticle and safranine T (ST) has been studied with resonance Rayleigh scattering (RRS) spectra, absorption and fluorescence spectra. In the pH 5 solution, citrate [(H2L)2-] self-assembles on the surface of positively-charged gold nanoparticle, which results in the [(Au)n(H2L)m]x- complex. In other words, one of carboxylate oxygens in (H2L)2- moves inward and combines with gold nanoparticle. The other carboxylate oxygens moves outward to form a supermolecular complex anion with x negative charges. Then by virtue of electrostatic attraction, hydrophobic force and charge transfer action, the complex anion binds with ST cation to form a new ion-association complex. Here (H2L)2- acts as a bridge. The formation of the complex results in the significant enhancement of RRS intensity, the appearance of new RRS spectrum, the red shift of plasma absorption band of gold nanoparticle as well as the decrease in the absorbance and fluorescence quenching for safranine T. In this work, the interaction between gold nanoparticle and ST on the RRS, absorption and fluorescence spectra has been investigated. The reason why RRS intensity increases greatly and the reaction mechanism have been inquired. The results show that RRS spectra can not only be used to study nanoparticle and reaction product, but also are a sensitive means to characterize and detect nanoparticles.

  14. Effect of Zinc oxide nanoparticle on Fluorescence Resonance Energy transfer between Fluorescein and Rhodamine 6G

    Science.gov (United States)

    Saha, Jaba; Roy, Arpan Datta; Dey, Dibyendu; Bhattacharjee, D.; Paul, Pabitra Kumar; Das, R.; Hussain, Syed Arshad

    2017-03-01

    Fluorescence Resonance Energy Transfer between two dyes Fluorescein and Rhodamine 6G were investigated in solution in the presence and absence of Zinc oxide nanoparticle. Zinc oxide nanostructure is used as the fluorescence enhancing agent for the present study since donor (Fluorescein) fluorescence increase significantly in presence of nanoparticle. Accordingly, the energy transfer efficiency in the presence of nanoparticle increases. The maximum efficiency was 69% for acceptor (Rhodamine 6G) concentration of 0.75 × 10- 5 M. The energy transfer efficiency was found to be pH sensitive and it varies from 4.15% to 90.00% in mixed dye solution for a change in pH from 1.5 to 10.0. With proper calibration it is possible to use the present system under investigation to sense pH which is better with respect to our previous reported results [Spectrochim. Acta Part A. 149 (2015) 143-149] as it can sense a wide range of pH and with better sensitivity.

  15. Intramolecular fluorescence resonance energy transfer and living cell imaging of novel pyridyltriphenylamine dye

    Science.gov (United States)

    Cao, Duojun; Qian, Ying

    2016-07-01

    A novel pyridyltriphenylamine-rhodamine dye PTRh and a pyridyltriphenylamine derivative PTO were synthesized and characterized by 1H NMR and HRMS-MALDI-TOF. PTRh performed typical fluorescence resonance energy transfer (FRET) signal from pyridyltriphenylamine to rhodamine along with notable color change from green to rose when interacting with Hg2+ in EtOH/H2O. And PTRh as a ratiometric probe for Hg2+ based on FRET could achieve a very low detection limit of 32 nM and energy transfer efficiency of 83.7% in aqueous organic system. On the other hand, spectra properties of PTO in its aggregates, THF/H2O mixed solution and silica nanoparticles (Si-NPs) dispersed in water were investigated. And the results indicated PTO exhibited bright green fluorescence in solid state, and PTO was successfully encapsulated in silica matrix (30-40 nm), emitting bright blue fluorescence with 11.7% quantum yield. Additionally, living cell imaging experiments demonstrated that PTRh could effectively response to intracellular Hg2+ and PTO-doped Si-NPs were well uptaken by MCF-7 breast cancer cells. It could be concluded that the chromophores are promising materials used as biosensors.

  16. Synthesis and characterization of a combined fluorescence, phosphorescence, and electron paramagnetic resonance probe

    Science.gov (United States)

    Beth, Albert H.; Cobb, Charles E.; Beechem, Joseph M.

    1992-04-01

    A spin-labeled derivative of eosin was chemically synthesized from 5-aminoeosin and the nitroxide spin label 2,2,5,5-tetramethylpyrrolin-1-oxyl-3-carboxylic acid. Following determination of the chemical identity of the spin-labeled eosin (5-SLE) by FAB mass spectroscopy, its optical and magnetic resonance spectroscopic properties were characterized in aqueous solution and compared to a diamagnetic eosin derivative, 5-acetamido eosin (5- AcE). The visible light absorption maximum of 5-SLE was 518 nm, the same as for 5-AcE. The fluorescence quantum yield of 5-SLE was only reduced by approximately 10% relative to 5-AcE, and the fluorescence lifetime was marginally reduced relative to 5-AcE. The phosphorescence lifetime and yield for 5-SLE were very similar to those for 5-AcE. The phosphorescence yield of 5-SLE bound noncovalently to BSA was reduced by approximately 60% relative to 5-AcE, and the phosphorescence lifetime reduced from approximately 2.4 msec (5-AcE) to 1.6 msec (5-SLE). Reduction of the nitroxide moiety of the 5-SLE with sodium ascorbate resulted in minimal changes in the fluorescence and phosphorescence quantum yields and lifetimes. This indicated that the unpaired electron of the nitroxide spin label did not seriously affect the optical spectroscopic characteristics of the spin-labeled eosin molecule. The quantum yields and lifetimes of 5-SLE were still quite acceptable for time- resolved fluorescence and phosphorescence studies. The electron paramagnetic resonance (EPR) spectrum of 5-SLE in aqueous solution has a lineshape consistent with a molecule the size of 5-SLE undergoing rapid rotational reorientation. When bound to BSA, the EPR spectrum of 5-SLE was broadened to a near slow motion limit for EPR, as expected for the relatively slowly rotating protein-5-SLE complex. Time-resolved phosphorescence anisotropy and saturation transfer EPR (ST-EPR) experiments with samples of 5-SLE bound to BSA in solutions of varying glycerol concentrations at 2

  17. Probing the graphite band structure with resonant soft-x-ray fluorescence

    Energy Technology Data Exchange (ETDEWEB)

    Carlisle, J.A.; Shirley, E.L.; Hudson, E.A. [Lawrence Berkeley National Lab., CA (United States)] [and others

    1997-04-01

    Soft x-ray fluorescence (SXF) spectroscopy using synchrotron radiation offers several advantages over surface sensitive spectroscopies for probing the electronic structure of complex multi-elemental materials. Due to the long mean free path of photons in solids ({approximately}1000 {angstrom}), SXF is a bulk-sensitive probe. Also, since core levels are involved in absorption and emission, SXF is both element- and angular-momentum-selective. SXF measures the local partial density of states (DOS) projected onto each constituent element of the material. The chief limitation of SXF has been the low fluorescence yield for photon emission, particularly for light elements. However, third generation light sources, such as the Advanced Light Source (ALS), offer the high brightness that makes high-resolution SXF experiments practical. In the following the authors utilize this high brightness to demonstrate the capability of SXF to probe the band structure of a polycrystalline sample. In SXF, a valence emission spectrum results from transitions from valence band states to the core hole produced by the incident photons. In the non-resonant energy regime, the excitation energy is far above the core binding energy, and the absorption and emission events are uncoupled. The fluorescence spectrum resembles emission spectra acquired using energetic electrons, and is insensitive to the incident photon`s energy. In the resonant excitation energy regime, core electrons are excited by photons to unoccupied states just above the Fermi level (EF). The absorption and emission events are coupled, and this coupling manifests itself in several ways, depending in part on the localization of the empty electronic states in the material. Here the authors report spectral measurements from highly oriented pyrolytic graphite.

  18. Vibrational techniques applied to photosynthesis: Resonance Raman and fluorescence line-narrowing.

    Science.gov (United States)

    Gall, Andrew; Pascal, Andrew A; Robert, Bruno

    2015-01-01

    Resonance Raman spectroscopy may yield precise information on the conformation of, and the interactions assumed by, the chromophores involved in the first steps of the photosynthetic process. Selectivity is achieved via resonance with the absorption transition of the chromophore of interest. Fluorescence line-narrowing spectroscopy is a complementary technique, in that it provides the same level of information (structure, conformation, interactions), but in this case for the emitting pigment(s) only (whether isolated or in an ensemble of interacting chromophores). The selectivity provided by these vibrational techniques allows for the analysis of pigment molecules not only when they are isolated in solvents, but also when embedded in soluble or membrane proteins and even, as shown recently, in vivo. They can be used, for instance, to relate the electronic properties of these pigment molecules to their structure and/or the physical properties of their environment. These techniques are even able to follow subtle changes in chromophore conformation associated with regulatory processes. After a short introduction to the physical principles that govern resonance Raman and fluorescence line-narrowing spectroscopies, the information content of the vibrational spectra of chlorophyll and carotenoid molecules is described in this article, together with the experiments which helped in determining which structural parameter(s) each vibrational band is sensitive to. A selection of applications is then presented, in order to illustrate how these techniques have been used in the field of photosynthesis, and what type of information has been obtained. This article is part of a Special Issue entitled: Vibrational spectroscopies and bioenergetic systems.

  19. Upconversion fluorescence resonance energy transfer based biosensor for ultrasensitive detection of matrix metalloproteinase-2 in blood.

    Science.gov (United States)

    Wang, Yuhui; Shen, Pei; Li, Chunya; Wang, Yanying; Liu, Zhihong

    2012-02-01

    Matrix metalloproteinase-2 (MMP-2) is a very important biomarker in blood. Presently, sensitive and selective determination of MMP-2 directly in blood samples is still a challenging job because of the high complexity of the sample matrix. In this work, we reported a new homogeneous biosensor for MMP-2 based on fluorescence resonance energy transfer (FRET) from upconversion phosphors (UCPs) to carbon nanoparticles (CNPs). A polypeptide chain (NH(2)-GHHYYGPLGVRGC-COOH) comprising both the specific MMP-2 substrate domain (PLGVR) and a π-rich motif (HHYY) was designed and linked to the surface of UCPs at the C terminus. The FRET process was initiated by the π-π interaction between the peptide and CNPs, which thus quenched the fluorescence of the donor. Upon the cleavage of the substrate by the protease at the amide bond between Gly and Val, the donor was separated from the acceptor while the π-rich motif stayed on the acceptor. As a result, the fluorescence of the donor was restored. The fluorescence recovery was found to be proportional to the concentration of MMP-2 within the range from 10-500 pg/mL in an aqueous solution. The quantification limit of this sensor was at least 1 order of magnitude lower than that of other reported assays for MMP-2. The sensor was used to determine the MMP-2 level directly in human plasma and whole blood samples with satisfactory results obtained. Owing to the hypersensitivity of the method, clinical samples of only less than 1 μL were needed for accurate quantification, which can be meaningful in MMP-2-related clinical and bioanalytical applications.

  20. Transmission Nuclear Resonance Fluorescence Measurements of 238U in Thick Targets

    Energy Technology Data Exchange (ETDEWEB)

    Quiter, Brian J.; Ludewigt, Bernhard A.; Mozin, Vladimir V.; Wilson, Cody; Korbly, Steve

    2010-08-31

    Transmission nuclear resonance fluorescence measurements were made on targets consisting of Pb and depleted U with total areal densities near 86 g/cm2. The 238U content n the targets varied from 0 to 8.5percent (atom fraction). The experiment demonstrates the capability of using transmission measurements as a non-destructive technique to identify and quantify the presence of an isotope in samples with thicknesses comparable to he average thickness of a nuclear fuel assembly. The experimental data also appear to demonstrate the process of notch refilling with a predictable intensity. Comparison of measured spectra to previous backscatter 238U measurements indicates general agreement in observed excited states. Two new 238U excited states and possibly a third state have also been observed.

  1. Hybrid aptamer-antibody linked fluorescence resonance energy transfer based detection of trinitrotoluene.

    Science.gov (United States)

    Sabherwal, Priyanka; Shorie, Munish; Pathania, Preeti; Chaudhary, Shilpa; Bhasin, K K; Bhalla, Vijayender; Suri, C Raman

    2014-08-05

    Combining synthetic macromolecules and biomolecular recognition units are promising in developing novel diagnostic and analysis techniques for detecting environmental and/or clinically important substances. Fluorescence resonance energy transfer (FRET) apta-immunosensor for explosive detection is reported using 2,4,6-trinitrotoluene (TNT) specific aptamer and antibodies tagged with respective FRET pair dyes in a sandwich immunoassay format. FITC-labeled aptamer was used as a binder molecule in the newly developed apta-immunoassay format where the recognition element was specific anti-TNT antibody labeled with rhodamine isothiocyanate. The newly developed sensing platform showed excellent sensitivity with a detection limit of the order of 0.4 nM presenting a promising candidate for routine screening of TNT in samples.

  2. Gamma-ray width measurements in {sup 15}N at the ELBE nuclear resonance fluorescence setup

    Energy Technology Data Exchange (ETDEWEB)

    Szuecs, Tamas [Helmholtz-Zentrum Dresden-Rossendorf (HZDR), Dresden (Germany); MTA ATOMKI, Debrecen/Hungary (Hungary); Bemmerer, Daniel; Schwengner, Ronald [Helmholtz-Zentrum Dresden-Rossendorf (HZDR), Dresden (Germany); Massarczyk, Ralph; Takacs, Marcell; Wagner, Louis [Helmholtz-Zentrum Dresden-Rossendorf (HZDR), Dresden (Germany); TU Dresden (Germany)

    2014-07-01

    The stable nucleus {sup 15}N is the mirror of the astrophysically important {sup 15}O, compound nucleus of the leading reaction of the Bethe-Weizsaecker cycle of hydrogen burning. Most of the {sup 15}N level widths below the neutron and proton emission thresholds are known from just one nuclear resonance fluorescence (NRF) measurement published more than 30 years ago, with unsatisfactory precision on some cases. A recent experiment with the AGATA demonstrator array aimed to determine level widths with the Doppler Shift Attenuation Method (DSAM) in {sup 15}O and {sup 15}N populated in {sup 14}N + {sup 2}H reaction. In order to set a benchmark value for the upcoming AGATA demonstrator data, the widths of several {sup 15}N levels are being studied using the bremsstrahlung facility γELBE at the electron accelerator of Helmholtz-Zentrum Dresden-Rossendorf (HZDR). The γELBE experiment and its preliminary results are presented.

  3. Fluorescence resonance energy transfer of gas-phase ions under ultra high vacuum and ambient conditions.

    Science.gov (United States)

    Frankevich, Vladimir; Chagovets, Vitaliy; Widjaja, Fanny; Barylyuk, Konstantin; Yang, Zhiyi; Zenobi, Renato

    2014-05-21

    We report evidence for fluorescence resonance energy transfer (FRET) of gas-phase ions under ultra high vacuum conditions (10(-9) mbar) inside a mass spectrometer as well as under ambient conditions inside an electrospray plume. Two different FRET pairs based on carboxyrhodamine 6G (donor) and ATTO590 or Bodipy TR (acceptor) dyes were examined and their gas-phase optical properties were studied. Our measurements indicate a different behavior for the two FRET pairs, which can be attributed to their different conformations in the gas phase. Upon desolvation via electrospray ionization, one of the FRET pairs undergoes a conformational change that leads to disappearance of FRET. This study shows the promise of FRET to obtain a direct correlation between solution and gas-phase structures.

  4. Resonance energy transfer between fluorescent BSA protected Au nanoclusters and organic fluorophores.

    Science.gov (United States)

    Raut, Sangram; Rich, Ryan; Fudala, Rafal; Butler, Susan; Kokate, Rutika; Gryczynski, Zygmunt; Luchowski, Rafal; Gryczynski, Ignacy

    2014-01-01

    Bovine serum albumin (BSA) protected nanoclusters (Au and Ag) represent a group of nanomaterials that holds great promise in biophysical applications due to their unique fluorescence properties and lack of toxicity. These metal nanoclusters have utility in a variety of disciplines including catalysis, biosensing, photonics, imaging and molecular electronics. However, they suffer from several disadvantages such as low fluorescence quantum efficiency (typically near 6%) and broad emission spectrum (540 nm to 800 nm). We describe an approach to enhance the apparent brightness of BSA Au clusters by linking them with a high extinction donor organic dye pacific blue (PB). In this conjugate PB acts as a donor to BSA Au clusters and enhances its brightness by resonance energy transfer (RET). We found that the emission of BSA Au clusters can be enhanced by a magnitude of two-fold by resonance energy transfer (RET) from the high extinction donor PB, and BSA Au clusters can act as an acceptor to nanosecond lifetime organic dyes. By pumping the BSA Au clusters using a high extinction donor, one can increase the effective brightness of less bright fluorophores like BSA Au clusters. Moreover, we prepared another conjugate of BSA Au clusters with the near infrared (NIR) dye Dylight 750 (Dy750), where BSA Au clusters act as a donor to Dy750. We observed that BSA Au clusters can function as a donor, showing 46% transfer efficiency to the NIR dye Dy750 with a long lifetime component in the acceptor decay through RET. Such RET-based probes can be used to prevent the problems of a broad emission spectrum associated with the BSA Au clusters. Moreover, transferring energy from BSA Au clusters to Dy750 will result in a RET probe with a narrow emission spectrum and long lifetime component which can be utilized in imaging applications.

  5. Study on the fluorescence resonance energy transfer between CdS quantum dots and Eosin Y.

    Science.gov (United States)

    Yan, Zhengyu; Zhang, Zhengwei; Yu, Yan; Chen, Jianqiu

    2015-03-01

    Water-soluble CdS quantum dots (QDs) were prepared using mercaptoacetic acid (TGA) as the stabilizer in an aqueous system. A fluorescence resonance energy transfer (FRET) system was constructed between water-soluble CdS QDs (donor) and Eosin Y (acceptor). Several factors that impacted the fluorescence spectra of the FRET system, such as pH (3.05-10.10), concentration of Eosin Y (2-80 mg/L) and concentration of CdS QDs (2-80 mg/L), were investigated and refined. Donor-to-acceptor ratios, the energy transfer efficiency (E) and the distance (r) between CdS QDs and Eosin Y were obtained. The results showed that a FRET system could be established between water-soluble CdS QDs and Eosin Y at pH 5.0; donor-to-acceptor ratios demonstrated a 1: 8 proportion of complexes; the energy transfer efficiency (E) and the distance (r) between the QDs and Eosin Y were 20.07% and 4.36 nm,respectively.

  6. Drug transport mechanism of P-glycoprotein monitored by single molecule fluorescence resonance energy transfer

    Science.gov (United States)

    Ernst, S.; Verhalen, B.; Zarrabi, N.; Wilkens, S.; Börsch, M.

    2011-03-01

    In this work we monitor the catalytic mechanism of P-glycoprotein (Pgp) using single-molecule fluorescence resonance energy transfer (FRET). Pgp, a member of the ATP binding cassette family of transport proteins, is found in the plasma membrane of animal cells where it is involved in the ATP hydrolysis driven export of hydrophobic molecules. When expressed in the plasma membrane of cancer cells, the transport activity of Pgp can lead to the failure of chemotherapy by excluding the mostly hydrophobic drugs from the interior of the cell. Despite ongoing effort, the catalytic mechanism by which Pgp couples MgATP binding and hydrolysis to translocation of drug molecules across the lipid bilayer is poorly understood. Using site directed mutagenesis, we have introduced cysteine residues for fluorescence labeling into different regions of the nucleotide binding domains (NBDs) of Pgp. Double-labeled single Pgp molecules showed fluctuating FRET efficiencies during drug stimulated ATP hydrolysis suggesting that the NBDs undergo significant movements during catalysis. Duty cycle-optimized alternating laser excitation (DCO-ALEX) is applied to minimize FRET artifacts and to select the appropriate molecules. The data show that Pgp is a highly dynamic enzyme that appears to fluctuate between at least two major conformations during steady state turnover.

  7. Development of L-lactate dehydrogenase biosensor based on porous silicon resonant microcavities as fluorescence enhancers.

    Science.gov (United States)

    Jenie, S N Aisyiyah; Prieto-Simon, Beatriz; Voelcker, Nicolas H

    2015-12-15

    The up-regulation of L-lactate dehydrogenase (LDH), an intracellular enzyme present in most of all body tissues, is indicative of several pathological conditions and cellular death. Herein, we demonstrate LDH detection using porous silicon (pSi) microcavities as a luminescence-enhancing optical biosensing platform. Non-fluorescent resazurin was covalently attached onto the pSi surface via thermal hydrocarbonisation, thermal hydrosylilation and acylation. Each surface modification step was confirmed by means of FTIR and the optical shifts of the resonance wavelength of the microcavity. Thermal hydrocarbonisation also afforded excellent surface stability, ensuring that the resazurin was not reduced on the pSi surface. Using a pSi microcavity biosensor, the fluorescence signal upon detection of LDH was amplified by 10 and 5-fold compared to that of a single layer and a detuned microcavity, respectively, giving a limit of detection of 0.08 U/ml. The biosensor showed a linear response between 0.16 and 6.5 U/ml, covering the concentration range of LDH in normal as well as damaged tissues. The biosensor was selective for LDH and did not produce a signal upon incubation with another NAD-dependant enzyme L-glutamic dehydrogenase. The use of the pSi microcavity as a sensing platform reduced reagent usage by 30% and analysis time threefold compared to the standard LDH assay in solution.

  8. Fluorescence Resonance Energy Transfer Analysis of Bid Activation in Living Cells during Ultraviolet-induced Apoptosis

    Institute of Scientific and Technical Information of China (English)

    Yinyuan WU; Da XING; Lei LIU; Tongsheng CHEN; Wei R. CHEN

    2007-01-01

    Ultraviolet (UV) irradiation is a DNA-damaging agent that triggers apoptosis through both the membrane death receptor and mitochondrial apoptotic signaling pathways. Bid, a pro-apoptotic Bcl-2family member, is important in most cell types to apoptosis in response to DNA damage. In this study, a recombinant plasmid, YFP-Bid-CFP, comprised of yellow and cyan fluorescent protein and a full length Bid,was used as a fluorescence resonance energy transfer analysis (FRET) probe. Using the FRET technique based on YFP-Bid-CFP, we found that Bid activation was initiated at 9±1 h after UV irradiation, and the average duration of the activation was 75± 10 min. Bid activation coincided with a collapse of the mitochondrial membrane potential with an average duration of 50±10 min. When cells were pretreated with Z-IETD-fmk(caspase-8 specific inhibitor) the process of Bid activation was completely inhibited, but the apoptosis was only partially affected. Z-DEVD-fmk (caspase-3 inhibitor) and Z-FA-fmk (non asp specific inhibitor) did not block Bid activation. Furthermore, the endogenous Bid activation with or without Z-IETD-fmk in response to UV irradiation was confirmed by Western blotting. In summary, using the FRET technique, we observed the dynamics of Bid activation during UV-induced apoptosis and found that it was a caspase-8 dependent event.

  9. Integrated Lymphography using Fluorescence Imaging and Magnetic Resonance Imaging in Intact Mice

    Directory of Open Access Journals (Sweden)

    Yusuke Inoue

    2011-09-01

    Full Text Available We assessed lymph drainage in living mice by an integrated imaging method using fluorescence imaging (FLI and magnetic resonance imaging (MRI. Mice were subcutaneously injected with quantum dots and gadofluorine 8 into the right rear footpad. They were fixed on a transparent flat plate and underwent FLI and MRI successively. Small markers were attached to the mouse surface for spatial coregistration, and image fusion of FLIs and MRIs was performed. Two-dimensional fluorescence reflectance imaging was used for FLI. FLI and MRI provided generally consistent results and demonstrated lymphatic flow to the popliteal, sacral, and iliac lymph nodes in most mice and to the renal, inguinal, and lumbar-aortic lymph nodes in some mice. On the fusion images, the locations of the lymph nodes in the mouse trunk were in good agreement between FLI and MRI, indicating successful spatial registration even for the deep structures. The popliteal node tended to be visualized a little farther caudally in FLI than in MRI, presumably because the overlying tissues were thicker in the cranial portion. Integrated FLI/MRI lymphography with image fusion appears to be a useful tool for analysis of the murine lymphatic system.

  10. Nuclear Resonance Fluorescence Measurements on ^237Np for Security and Safeguards Applications

    Science.gov (United States)

    Angell, C. T.; Joshi, T.; Yee, Ryan; Norman, E. B.; Kulp, W. D.; Warren, G. A.; Korbly, S.; Klimenko, A.; Wilson, C.; Copping, R.; Shuh, D. K.

    2009-10-01

    The smuggling of nuclear material and the diversion of fissile material for covert weapon programs both present grave risks to world security. Methods are needed to detect nuclear material smuggled in cargo, and for proper material accountability in civilian fuel re-processing facilities. Nuclear resonance fluorescence (NRF) is a technique that can address both needs. It is a non-destructive active interrogation method that provides isotope-specific information. It works by using a γ-ray beam to resonantly excite levels in a nucleus and observing the γ-rays emitted whose energy and intensity are characteristic of that isotope. ^237Np presents significant safeguard challenges; it is fissile yet currently has fewer safeguard restrictions. NRF measurements on ^237Np will expand the nuclear database and will permit designing interrogation and assay systems. Measurements were made using the bremsstrahlung beam at the HVRL at MIT on a 7 g target of ^237Np with two incident electron energies of 2.8 and 3.1 MeV. Results will be presented with discussion of the relevant nuclear structure necessary to predict levels in other actinides.

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

    Science.gov (United States)

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

    2015-02-01

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

  12. Spectroscopy and photophysics of self-organized zinc porphyrin nanolayers. 3. Fluorescence detected magnetic resonance of triplet states

    NARCIS (Netherlands)

    Schaafsma, T.J.; Dag, I.; Sitters, R.; Glasbeek, M.; Lifshitz, E.

    2005-01-01

    Fluorescence detected magnetic resonance (FDMR) has been applied to ~25-nm-thick porphyrin films, containing ordered domains of zinc tetra-(p-octylphenyl)-porphyrin (ZnTOPP) spin-coated onto quartz slides. Illuminating the films at 1.4 K with 457.9-nm light from a continuous wave Ar+ laser produces

  13. Peptide-based fluorescence resonance energy transfer protease substrates for the detection and diagnosis of Bacillus species

    NARCIS (Netherlands)

    Kaman, W.E.; Hulst, A.G.; Alphen, P.T.W. van; Roffel, S.; Schans, M.J. van der; Merkel, T.; Belkum, A. van; Bikker, F.J.

    2011-01-01

    We describe the development of a highly specific enzyme-based fluorescence resonance energy transfer (FRET) assay for easy and rapid detection both in vitro and in vivo of Bacillus spp., among which are the members of the B. cereus group. Synthetic substrates for B. anthracis proteases were designed

  14. Resonant inelastic scattering in dilute magnetic semiconductors by x-ray fluorescence spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Lawniczak-Jablonska, K. [Lawrence Berkeley National Lab., CA (United States)]|[Institute of Physics, Warsaw (Poland); Jia, J.J.; Underwood, J.H. [Lawrence Berkeley National Lab., CA (United States)] [and others

    1997-04-01

    As modern, technologically important materials have become more complex, element specific techniques have become invaluable in studying the electronic structure of individual components from the system. Soft x-ray fluorescence (SXF) and absorption (SXA) spectroscopies provide a unique means of measuring element and angular momentum density of electron states, respectively, for the valence and conducting bands in complex materials. X-ray absorption and the decay through x-ray emission are generally assumed to be two independent one-photon processes. Recent studies, however have demonstrated that SXF excited near the absorption threshold generate an array of spectral features that depend on nature of materials, particularly on the localization of excited states in s and d-band solids and that these two processes can no be longer treated as independent. Resonant SXF offers thus the new way to study the dynamics of the distribution of electronic valence states in the presence of a hole which is bound to the electron low lying in the conduction band. This process can simulate the interaction between hole-electron pair in wide gap semiconductors. Therefore such studies can help in understanding of transport and optics phenomena in the wide gap semiconductors. The authors report the result of Mn and S L-resonant emission in Zn{sub 1{minus}x}Mn{sub x}S (with x=0.2 and 0.3) and MnS as the energy of exciting radiation is tuned across the Mn and S L{sub 3,2} absorption edge, along with the resonant excited spectra from elemental Mn as a reference.

  15. Nuclear Resonance Fluorescence to Measure Plutonium Mass in Spent Nuclear Fuel

    Energy Technology Data Exchange (ETDEWEB)

    Ludewigt, Bernhard A; Quiter, Brian J.; Ambers, Scott D.

    2011-01-14

    The Next Generation Safeguard Initiative (NGSI) of the U.S Department of Energy is supporting a multi-lab/university collaboration to quantify the plutonium (Pu) mass in spent nuclear fuel (SNF) assemblies and to detect the diversion of pins with non-destructive assay (NDA) methods. The following 14 NDA techniques are being studied: Delayed Neutrons, Differential Die-Away, Differential Die-Away Self-Interrogation, Lead Slowing Down Spectrometer, Neutron Multiplicity, Passive Neutron Albedo Reactivity, Total Neutron (Gross Neutron), X-Ray Fluorescence, {sup 252}Cf Interrogation with Prompt Neutron Detection, Delayed Gamma, Nuclear Resonance Fluorescence, Passive Prompt Gamma, Self-integration Neutron Resonance Densitometry, and Neutron Resonance Transmission Analysis. Understanding and maturity of the techniques vary greatly, ranging from decades old, well-understood methods to new approaches. Nuclear Resonance Fluorescence (NRF) is a technique that had not previously been studied for SNF assay or similar applications. Since NRF generates isotope-specific signals, the promise and appeal of the technique lies in its potential to directly measure the amount of a specific isotope in an SNF assay target. The objectives of this study were to design and model suitable NRF measurement methods, to quantify capabilities and corresponding instrumentation requirements, and to evaluate prospects and the potential of NRF for SNF assay. The main challenge of the technique is to achieve the sensitivity and precision, i.e., to accumulate sufficient counting statistics, required for quantifying the mass of Pu isotopes in SNF assemblies. Systematic errors, considered a lesser problem for a direct measurement and only briefly discussed in this report, need to be evaluated for specific instrument designs in the future. Also, since the technical capability of using NRF to measure Pu in SNF has not been established, this report does not directly address issues such as cost, size

  16. A reduced graphene oxide-based fluorescence resonance energy transfer sensor for highly sensitive detection of matrix metalloproteinase 2.

    Science.gov (United States)

    Xi, Gaina; Wang, Xiaoping; Chen, Tongsheng

    2016-01-01

    A novel fluorescence nanoprobe (reduced nano-graphene oxide [nrGO]/fluorescein isothiocyanate-labeled peptide [Pep-FITC]) for ultrasensitive detection of matrix metalloproteinase 2 (MMP2) has been developed by engineering the Pep-FITC comprising the specific MMP2 substrate domain (PLGVR) onto the surface of nrGO particles through non-covalent linkage. The nrGO was obtained by water bathing nano-graphene oxide under 90°C for 4 hours. After mixing the nrGO and Pep-FITC for 30 seconds, the fluorescence from Pep-FITC was almost completely quenched due to the fluorescence resonance energy transfer between fluorescein isothiocyanate (FITC) and nrGO. Upon cleavage of the amide bond between Leu and Gly in the Pep-FITC by protease-MMP2, the FITC bound to nrGO was separated from nrGO surface, disrupting the fluorescence resonance energy transfer process and resulting in fluorescence recovery of FITC. Under optimal conditions, the fluorescence recovery of nrGO/Pep-FITC was found to be directly proportional to the concentration of MMP2 within 0.02-0.1 nM. The detection limit of the nrGO/Pep-FITC was determined to be 3 pM, which is approximately tenfold lower than that of the unreduced carboxylated nano-graphene oxide/Pep-FITC probe.

  17. Highly indistinguishable on-demand resonance fluorescence photons from a deterministic quantum dot micropillar device with 74% extraction efficiency

    DEFF Research Database (Denmark)

    Gregersen, Niels

    2016-01-01

    The implementation and engineering of bright and coherent solid state quantum light sources is key for the realization of both on chip and remote quantum networks. Despite tremendous efforts for more than 15 years, the combination of these two key prerequisites in a single, potentially scalable...... device is a major challenge. Here, we report on the observation of bright single photon emission generated via pulsed, resonance fluorescence conditions from a single quantum dot (QD) deterministically centered in a micropillar cavity device via cryogenic optical lithography. The brightness of the QD...... fluorescence is greatly enhanced on resonance with the fundamental mode of the pillar, leading to an overall device efficiency of η = (74 ± 4) % for a single photon emission as pure as g (2) (0) = 0.0092 ± 0.0004. The combination of large Purcell enhancement and resonant pumping conditions allows us to observe...

  18. The Science of Nuclear Materials Detection using gamma-ray beams: Nuclear Resonance Fluorescence

    Science.gov (United States)

    Ohgaki, Hideaki

    2014-09-01

    An atomic nucleus is excited by absorption of incident photons with an energy the same as the excitation energy of the level, and subsequently a gamma-ray is emitted as it de-excites. This phenomenon is called Nuclear Resonance Fluorescence and mostly used for studies on Nuclear Physics field. By measuring the NRF gamma-rays, we can identify nuclear species in any materials because the energies of the NRF gamma-rays uniquely depend on the nuclear species. For example, 235U has an excitation level at 1733 keV. If we irradiate a material including 235U with a gamma-ray tuned at this excitation level, the material absorbs the gamma-ray and re-emits another gamma-ray immediately to move back towards the ground state. Therefore we can detect the 235U by measuring the re-emitted (NRF) gamma-rays. Several inspection methods using gamma-rays, which can penetrate a thick shielding have been proposed and examined. Bertozzi and Ledoux have proposed an application of nuclear resonance fluorescence (NRF) by using bremsstrahlung radiations. However the signal-to-noise (SN) ratio of the NRF measurement with the bremsstrahlung radiation is, in general, low. Only a part of the incident photons makes NRF with a narrow resonant band (meV-eV) whereas most of incident radiation is scattered by atomic processes in which the reaction rate is higher than that of NRF by several orders of magnitudes and causes a background. Thus, the NRF with a gamma-ray quasi-monochromatic radiation beam is proposed. The monochromatic gamma-rays are generated by using laser Compton scattering (LCS) of electrons and intense laser photons by putting a collimator to restrict the gamma-ray divergence downstream. The LCS gamma-ray, which is energy-tunable and monochromatic, is an optimum apparatus for NRF measurements We have been conducted NRF experiment for nuclear research, especially with high linear polarized gamma-ray generated by LCS, to survey the distribution of M1 strength in MeV region in LCS

  19. Micro-RNA detection based on fluorescence resonance energy transfer of DNA-carbon quantum dots probes.

    Science.gov (United States)

    Khakbaz, Faeze; Mahani, Mohamad

    2017-04-15

    Carbon quantum dots have been proposed as an effective platform for miRNA detection. Carbon dots were synthesized by citric acid. The synthesized dots were characterized by dynamic light scattering, UV-Vis spectrophotometry, spectrofluorimetry, transmission electron microscopy and FT-IR spectrophotometry. The fluorescence quantum yield of the synthesized dots was determined using quinine sulfate as the standard. The FAM-labeled single stranded DNA, as sensing element, was adsorbed on dots by π-π interaction. The quenching of the dots fluorescence due to fluorescence resonance energy transfer (FRET) was used for mir 9-1 detection. In the presence of the complementary miRNA, the FRET did not take place and the fluorescence was recovered.

  20. Fluorescence resonance energy transfer in AOT/4-chlorophenol/m-xylene organogels

    Energy Technology Data Exchange (ETDEWEB)

    Dandapat, Manika; Mandal, Debabrata, E-mail: dmandal.chemistry@gmail.com

    2015-06-15

    Fluorescence Resonance Energy Transfer (FRET) between donor coumarins (C102 and C153) and acceptor Rhodamine 6G were studied in AOT/4-chlorophenol/m-xylene organogels. The gel comprises a three-dimensional network of fiber bundles trapping the m-xylene solvent. Each fiber is an aggregate of several strands, and each strand consists of a central columnar stack of the phenols, surrounded by AOT headgroups. Our acceptor is ionic so that it was concentrated near the polar center of the strand, while the neutral donors were likely distributed over a wider region. With C153 as donor, clear evidence of FRET (time-constant~100 ps) was found, which indicated that the donor and acceptor may reside in neighboring strands within the same fiber. However, with C102 as donor, FRET probably occurred over an ultrashort, sub-picosecond time-scale suggesting that the donor and acceptor in this case resided in close vicinity. Thus, C102 tends to localize near the polar centre of the strands, compared to the more hydrophobic C153, which prefers to occupy the relatively non-polar peripheral regions of the strands and fibers. - Highlights: • FRET between coumarin donors and Rhodamine 6G acceptor studied in AOT organogels. • With Coumarin 153 donor, a ~100 ps FRET component detected in both donor and acceptor fluorescence. • With Coumarin 102 donor, FRET component too short to be detected with a time-resolution of ~70 ps. • The FRET rates reveal crucial differences in donor–acceptor distances for the two coumarin donors.

  1. Study on the interaction between diphenhydramine and erythrosin by absorption,fluorescence and resonance Rayleigh scattering spectra

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    In pH 4.5 Britton-Robinson(BR)buffer solution,erythrosin(ET)can react with diphenhydramine(DP)to form a 1:1 ion-association complex,which not only results in the change of the absorption spectra,but also results in the great enhancement of resonance Rayleigh scattering(RRS)and the quenching of fluorescence.Furthermore,a new RRS spectrum will appear,and the maximum RRS wavelength was located at about 580 nm.In this work,the spectral characteristics of the absorption,fluorescence and RRS,the optimum conditions of the reaction and the properties of an analytical chemistry were inves- tigated.A sensitive,simple and new method for the determination of DP by using erythrosin as a probe has been developed.The detection limits for DP were 0.0020μg/mL for RRS method,0.088μg/mL for absorption method and 0.094μg/mL for fluorophotometry.There was a linear relationship between the absorbance,RRS and fluorescence intensities and the drug concentration in the range of 0.0067-2.0, 0.29-6.4 and 0.31-3.2μg/mL,respectively.The effects of the interaction of diphenhydramine and erythrosin on the absorption,fluorescence and resonance Rayleigh scattering spectra were discussed. In light polarization experiment,the polarization of RRS at maximum wavelength was measured to be P =0.9779,and it revealed that the RRS spectrum of DP-ET complex consists mostly of resonance scat- tering and few resonance fluorescence.In this study,enthalpy of formation and mean polarizability were calculated by AM1 quantum chemistry method.In addition,the reaction mechanism and the rea- sons for the enhancement of scattering spectra and the energy transfer between absorption,fluores- cence and RRS were discussed.

  2. The effect of pressure on the nuclear quadrupole resonance and ruby fluorescence in a NiCrAl pressure cell

    Energy Technology Data Exchange (ETDEWEB)

    Fujiwara, N [Graduate School of Human and Environmental Studies, Kyoto University, Yoshida-nihonmatsu-cho, Sakyo-ku, Kyoto 606-8501 (Japan); Koyama-Nakazawa, K [Institute for Solid State Physics, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8581 (Japan); Matsumoto, T [National Institute for Materials Science, Tsukuba 305-0047 (Japan); Hisada, A [Graduate School of Human and Environmental Studies, Kyoto University, Yoshida-nihonmatsu-cho, Sakyo-ku, Kyoto 606-8501 (Japan); Uwatoko, Y [Institute for Solid State Physics, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8581 (Japan)

    2007-10-24

    A pressure of 4.0 GPa was achieved using a piston-cylinder-type NiCrAl pressure cell which possesses a large sample space of 4.4 mm diameter x 15 mm length at ambient pressure. As monitors of the pressure, the {sup 63}Cu nuclear quadrupole resonance (NQR) of Cu{sub 2}O and ruby fluorescence were studied at the same time. The relation between the NQR frequency and the R1 line of the ruby fluorescence was investigated at pressures below 3.2 GPa.

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

    Directory of Open Access Journals (Sweden)

    Paul J. Endres

    2006-10-01

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

  4. Forster Resonance Energy Transfer and Laser Fluorescent Analysis of Defects in DNA Double Helix

    CERN Document Server

    Bregadze, Vasil G; Giorgadze, Tamar G; Jaliashvili, Zaza V; Chkhaberidze, Jemal G; Monaselidze, Jamlet R; Khuskivadze, Temur B

    2013-01-01

    Real time laser induced fluorescence spectroscopy usage for microanalysis of DNA double helix defects is shown. The method is based on Forster resonance energy transfer (FRET) in intercalator-donor pair (acridine orange as a donor and ethidium bromide as an acceptor). Transition metal ions such as Cu(II), Cu(I), Ag(I), silver nanoparticles (AgNPs), photo- and thermo effects were used to cause double helix defects in DNA. FRET radii were experimentally estimated in background electrolyte solution (0.01 M NaNO3) and proved to be 3.9 +- 0.3 nm and the data are in satisfactory agreement with the theoretically calculated value Ro = 3.5 +- 0.3 nm. Concentration of DNA sites, exposed to Cu(II), Cu(I), Ag(I) ions, AgNPs impact as well as laser irradiation ({\\lambda} = 457 nm) and temperature, which are applicable for intercalation, were estimated in relative units. FRET method allows to estimate the concentration of double helix areas with high quality stability applicable for intercalation in DNA after it was subjec...

  5. Determination of gamma-ray widths in $^{15}$N using nuclear resonance fluorescence

    CERN Document Server

    Szücs, T; Caciolli, A; Fülöp, Zs; Massarczyk, R; Michelagnoli, C; Reinhardt, T P; Schwengner, R; Takács, M P; Ur, C A; Wagner, A; Wagner, L

    2015-01-01

    The stable nucleus $^{15}$N is the mirror of $^{15}$O, the bottleneck in the hydrogen burning CNO cycle. Most of the $^{15}$N level widths below the proton emission threshold are known from just one nuclear resonance fluorescence (NRF) measurement, with limited precision in some cases. A recent experiment with the AGATA demonstrator array determined level lifetimes using the Doppler Shift Attenuation Method (DSAM) in $^{15}$O. As a reference and for testing the method, level lifetimes in $^{15}$N have also been determined in the same experiment. The latest compilation of $^{15}$N level properties dates back to 1991. The limited precision in some cases in the compilation calls for a new measurement in order to enable a comparison to the AGATA demonstrator data. The widths of several $^{15}$N levels have been studied with the NRF method. The solid nitrogen compounds enriched in $^{15}$N have been irradiated with bremsstrahlung. The $\\gamma$-rays following the deexcitation of the excited nuclear levels were dete...

  6. Investigation of Prolactin Receptor Activation and Blockade Using Time-Resolved Fluorescence Resonance Energy Transfer

    Directory of Open Access Journals (Sweden)

    Estelle eTallet

    2011-09-01

    Full Text Available The prolactin receptor (PRLR is emerging as a therapeutic target in oncology. Knowledge-based drug design led to the development of a pure PRLR antagonist (Del1-9-G129R-hPRL that was recently shown to prevent PRL-induced mouse prostate tumorogenesis. In humans, the first gain-of-function mutation of the PRLR (PRLRI146L was recently identified in breast tumor patients. At the molecular level, the actual mechanism of action of these two novel players in the PRL system remains elusive. In this study, we addressed whether constitutive PRLR activation (PRLRI146L or PRLR blockade (antagonist involved alteration of receptor oligomerization and/or of inter-chain distances compared to unstimulated and PRL-stimulated PRLR. Using a combination of various biochemical and spectroscopic approaches (co-IP, blue-native electrophoresis, BRET1, we demonstrated that preformed PRLR homodimers are altered neither by PRL- or I146L-induced receptor triggering, nor by antagonist-mediated blockade. These findings were confirmed using a novel time-resolved fluorescence resonance energy transfer (TR-FRET technology that allows monitoring distance changes between cell-surface tagged receptors. This technology revealed that PRLR blockade or activation did not involve detectable distance changes between extracellular domains of receptor chains within the dimer. This study merges with our previous structural investigations suggesting that the mechanism of PRLR activation solely involves intermolecular contact adaptations leading to subtle intramolecular rearrangements.

  7. Association of a novel preribosomal complex in Trypanosoma brucei determined by fluorescence resonance energy transfer.

    Science.gov (United States)

    Wang, Lei; Ciganda, Martin; Williams, Noreen

    2013-02-01

    We have previously reported that the trypanosome-specific proteins P34 and P37 form a unique preribosomal complex with ribosomal protein L5 and 5S rRNA in the nucleoplasm. We hypothesize that this novel trimolecular complex is necessary for stabilizing 5S rRNA in Trypanosoma brucei and is essential for the survival of the parasite. In vitro quantitative analysis of the association between the proteins L5 and P34 is fundamental to our understanding of this novel complex and thus our ability to exploit its unique characteristics. Here we used in vitro fluorescence resonance energy transfer (FRET) to analyze the association between L5 and P34. First, we demonstrated that FRET can be used to confirm the association between L5 and P34. We then determined that the binding constant for L5 and P34 is 0.60 ± 0.03 μM, which is in the range of protein-protein binding constants for RNA binding proteins. In addition, we used FRET to identify the critical regions of L5 and P34 involved in the protein-protein association. We found that the N-terminal APK-rich domain and RNA recognition motif (RRM) of P34 and the L18 domain of L5 are important for the association of the two proteins with each other. These results provide us with the framework for the discovery of ways to disrupt this essential complex.

  8. A high-throughput fluorescence resonance energy transfer-based assay for DNA ligase.

    Science.gov (United States)

    Shapiro, Adam B; Eakin, Ann E; Walkup, Grant K; Rivin, Olga

    2011-06-01

    DNA ligase is the enzyme that catalyzes the formation of the backbone phosphodiester bond between the 5'-PO(4) and 3'-OH of adjacent DNA nucleotides at single-stranded nicks. These nicks occur between Okazaki fragments during replication of the lagging strand of the DNA as well as during DNA repair and recombination. As essential enzymes for DNA replication, the NAD(+)-dependent DNA ligases of pathogenic bacteria are potential targets for the development of antibacterial drugs. For the purposes of drug discovery, a high-throughput assay for DNA ligase activity is invaluable. This article describes a straightforward, fluorescence resonance energy transfer-based DNA ligase assay that is well suited for high-throughput screening for DNA ligase inhibitors as well as for use in enzyme kinetics studies. Its use is demonstrated for measurement of the steady-state kinetic constants of Haemophilus influenzae NAD(+)-dependent DNA ligase and for measurement of the potency of an inhibitor of this enzyme.

  9. Single-molecule-sensitive fluorescence resonance energy transfer in freely-diffusing attoliter droplets

    Energy Technology Data Exchange (ETDEWEB)

    Rahmanseresht, Sheema; Ramos, Kieran P.; Gamari, Ben D.; Goldner, Lori S., E-mail: lgoldner@physics.umass.edu [Department of Physics, University of Massachusetts, Amherst, Massachusetts 01003 (United States); Milas, Peker [Department of Neuroscience, University of Wisconsin, Madison, Wisconsin 53705 (United States)

    2015-05-11

    Fluorescence resonance energy transfer (FRET) from individual, dye-labeled RNA molecules confined in freely-diffusing attoliter-volume aqueous droplets is carefully compared to FRET from unconfined RNA in solution. The use of freely-diffusing droplets is a remarkably simple and high-throughput technique that facilitates a substantial increase in signal-to-noise for single-molecular-pair FRET measurements. We show that there can be dramatic differences between FRET in solution and in droplets, which we attribute primarily to an altered pH in the confining environment. We also demonstrate that a sufficient concentration of a non-ionic surfactant mitigates this effect and restores FRET to its neutral-pH solution value. At low surfactant levels, even accounting for pH, we observe differences between the distribution of FRET values in solution and in droplets which remain unexplained. Our results will facilitate the use of nanoemulsion droplets as attoliter volume reactors for use in biophysical and biochemical assays, and also in applications such as protein crystallization or nanoparticle synthesis, where careful attention to the pH of the confined phase is required.

  10. Fluorescence Resonance Energy Transfer Assay for High-Throughput Screening of ADAMTS1 Inhibitors

    Directory of Open Access Journals (Sweden)

    Guanhua Du

    2011-12-01

    Full Text Available A disintegrin and metalloprotease with thrombospondin type I motifs-1 (ADAMTS1 plays a crucial role in inflammatory joint diseases and its inhibitors are potential candidates for anti-arthritis drugs. For the purposes of drug discovery, we reported the development and validation of fluorescence resonance energy transfer (FRET assay for high-throughput screening (HTS of the ADAMTS1 inhibitors. A FRET substrate was designed for a quantitative assay of ADAMTS1 activity and enzyme kinetics studies. The assay was developed into a 50-µL, 384-well assay format for high throughput screening of ADAMTS1 inhibitors with an overall Z’ factor of 0.89. ADAMTS1 inhibitors were screened against a diverse library of 40,960 total compounds with the established HTS system. Four structurally related hits, naturally occurring compounds, kuwanon P, kuwanon X, albafuran C and mulberrofuran J, extracted from the Chinese herb Morus alba L., were identified for further investigation. The results suggest that this FRET assay is an excellent tool, not only for measurement of ADAMTS1 activity but also for discovery of novel ADAMTS1 inhibitors with HTS.

  11. Detection of Citrus tristeza virus by using fluorescence resonance energy transfer-based biosensor

    Science.gov (United States)

    Shojaei, Taha Roodbar; Salleh, Mohamad Amran Mohd; Sijam, Kamaruzaman; Rahim, Raha Abdul; Mohsenifar, Afshin; Safarnejad, Reza; Tabatabaei, Meisam

    2016-12-01

    Due to the low titer or uneven distribution of Citrus tristeza virus (CTV) in field samples, detection of CTV by using conventional detection techniques may be difficult. Therefore, in the present work, the cadmium-telluride quantum dots (QDs) was conjugated with a specific antibody against coat protein (CP) of CTV, and the CP were immobilized on the surface of gold nanoparticles (AuNPs) to develop a specific and sensitive fluorescence resonance energy transfer (FRET)-based nanobiosensor for detecting CTV. The maximum FRET efficiency for the developed nano-biosensor was observed at 60% in AuNPs-CP/QDs-Ab ratio of 1:8.5. The designed system showed higher sensitivity and specificity over enzyme linked immunosorbent assay (ELISA) with a limit of detection of 0.13 μg mL- 1 and 93% and 94% sensitivity and specificity, respectively. As designed sensor is rapid, sensitive, specific and efficient in detecting CTV, this could be envisioned for diagnostic applications, surveillance and plant certification program.

  12. Intramolecular and intermolecular fluorescence resonance energy transfer in fluorescent protein-tagged Na-K-Cl cotransporter (NKCC1): sensitivity to regulatory conformational change and cell volume.

    Science.gov (United States)

    Pedersen, Meike; Carmosino, Monica; Forbush, Biff

    2008-02-01

    To examine the structure and function of the Na-K-Cl cotransporter, NKCC1, we tagged the transporter with cyan (CFP) and yellow (YFP) fluorescent proteins and measured fluorescence resonance energy transfer (FRET) in stably expressing human embryonic kidney cell lines. Fluorescent protein tags were added at the N-terminal residue between the regulatory domain and the membrane domain and within a poorly conserved region of the C terminus. Both singly and doubly tagged NKCC1s were appropriately trafficked to the cell membrane and were fully functional; regulation was normal except when YFP was inserted near the regulatory domain, in which case activation occurred only upon incubation with calyculin A. Quenching of YFP fluorescence by Cl(-) provided a ratiometric indicator of intracellular [Cl(-)]. All of the CFP/YFP NKCC pairs exhibited some level of FRET, demonstrating the presence of dimers or higher multimers in functioning NKCC1. With YFP near the regulatory domain and CFP in the C terminus, we recorded a 6% FRET change signaling the regulatory phosphorylation event. On the other hand, when the probe was placed at the extreme N terminus, such changes were not seen, presumably due to the length and predicted flexibility of the N terminus. Substantial FRET changes were observed contemporaneous with cell volume changes, possibly reflective of an increase in molecular crowding upon cell shrinkage.

  13. Fluorescence Resonance Energy Transfer Imaging Reveals that Chemokine-Binding Modulates Heterodimers of CXCR4 and CCR5 Receptors

    OpenAIRE

    2008-01-01

    BACKGROUND: Dimerization has emerged as an important feature of chemokine G-protein-coupled receptors. CXCR4 and CCR5 regulate leukocyte chemotaxis and also serve as a co-receptor for HIV entry. Both receptors are recruited to the immunological synapse during T-cell activation. However, it is not clear whether they form heterodimers and whether ligand binding modulates the dimer formation. METHODOLOGY/PRINCIPAL FINDINGS: Using a sensitive Fluorescence Resonance Energy Transfer (FRET) imaging ...

  14. Polarised bremsstrahlung nuclear resonance fluorescence set-up at the 15 MeV linac in Gent

    Science.gov (United States)

    Govaert, K.; Mondelaers, W.; Jacobs, E.; De Frenne, D.; Persyn, K.; Pommé, S.; Yoneama, M.-L.; Lindenstruth, S.; Huber, K.; Jung, A.; Starck, B.; Stock, R.; Wesselborg, C.; Heil, R.-D.; Kneissl, U.; Pitz, H. H.

    1994-01-01

    Nuclear resonance fluorescence experiments using unpolarised as well as off-axis linearly polarised bremsstrahlung represent an outstanding tool to determine in a completely model independent way transition probabilities, multipole orders and parities of electromagnetic transitions to bound states in nuclei. A new polarised bremsstrahlung facility has been constructed at the 15 MeV linac in Gent. The experimental arrangement is discussed and first results are presented.

  15. Practical use of corrected fluorescence excitation and emission spectra of fluorescent proteins in Förster Resonance Energy Transfer (FRET) studies

    NARCIS (Netherlands)

    Hink, M.A.; Visser, N.V.; Borst, J.W.; Hoek, van A.; Visser, A.J.W.G.

    2003-01-01

    Corrected fluorescence excitation and emission spectra have been obtained from several enhanced variants of the green fluorescent protein (EGFP) isolated from the jellyfish Aequorea victoria, blue fluorescence protein (EBFP), cyan fluorescent protein (ECFP), EGFP and yellow fluorescent protein (EYFP

  16. Ultrafast fluorescence resonance energy transfer in a bile salt aggregate: Excitation wavelength dependence

    Indian Academy of Sciences (India)

    Ujjwal Mandal; Subhadip Ghosh; Dibyendu Kumar Das; Aniruddha Adhikari; Shantanu Dey; Kankan Bhattacharyya

    2008-01-01

    Fluorescence resonance energy transfer (FRET) from Coumarin 153 (C153) to Rhodamine 6G (R6G) in a secondary aggregate of a bile salt (sodium deoxycholate, NaDC) is studied by femtosecond up-conversion. The emission spectrum of C153 in NaDC is analysed in terms of two spectra-one with emission maximum at 480 nm which corresponds to a non-polar and hydrophobic site and another with maximum at ∼ 530 nm which arises from a polar hydrophilic site. The time constants of FRET were obtained from the rise time of the emission of the acceptor (R6G). In the NaDC aggregate, FRET occurs in multiple time scales -4 ps and 3700 ps. The 4 ps component is assigned to FRET from a donor (D) to an acceptor (A) held at a close distance (DA ∼ 17 Å) inside the bile salt aggregate. The 3700 ps component corresponds to a donor-acceptor distance ∼ 48 Å. The long (3700 ps) component may involve diffusion of the donor. With increase in the excitation wavelength (ex) from 375 to 435 nm, the relative contribution of the ultrafast component of FRET (∼ 4 ps) increases from 3 to 40% with a concomitant decrease in the contribution of the ultraslow component (∼3700 ps) from 97 to 60%. The ex dependence is attributed to the presence of donors at different locations. At a long ex (435 nm) donors in the highly polar peripheral region are excited. A short ex (375 nm) `selects’ donor at a hydrophobic location.

  17. High-efficiency receiver architecture for resonance-fluorescence and Doppler lidars.

    Science.gov (United States)

    Smith, John A; Chu, Xinzhao

    2015-04-10

    A high-efficiency lidar receiver architecture that emphasizes boosting the receiver collection efficiency of resonance-fluorescence and Doppler lidars has opened up new avenues of study for the mesosphere and lower thermosphere-extended (MLT-X) at sites in Boulder, Colorado, USA, and Cerro Pachón, Chile. Described in this work are in-depth considerations in the design, construction, and alignment of Na Doppler lidar receivers that have yielded signal levels typically 5-10 times higher per power-aperture product than any demonstrated in the literature, to these authors' knowledge, making studies of fine-scale MLT turbulence and tenuous thermospheric layers in Na possible with temperature and vertical wind capability for the first time. A lowering of the detection threshold by higher receiver collection efficiency at Cerro Pachón has enabled this Na Doppler lidar to extend its measurement range far higher into the thermosphere, to regions with Na density less than 3  cm(-3). With renewed interest in the MLT-X region prompted by recent lidar discoveries of Fe in the thermosphere reaching 170 km at McMurdo, Antarctica, the receiver optimizations we have made now enable addressing an important need in the community. In addition, the higher spatial and temporal resolutions afforded by high signal-to-noise ratio, down to resolutions of ∼20  s and ∼20  m, promise to make the first direct measurements of eddy flux in the mesopause region possible. Results from deployment of optimized receivers at the Table Mountain Lidar Observatory in Boulder, the Andes Lidar Observatory at Cerro Pachón, and the Arecibo Observatory in Puerto Rico are presented to demonstrate the power and portability of our methods that are readily applicable to other lidar varieties, including, but not limited to, the newly developed Fe Doppler lidar and recently upgraded K Doppler lidar.

  18. Fluorescent resonance energy transfer: A tool for probing molecular cell-biomaterial interactions in three dimensions.

    Science.gov (United States)

    Huebsch, Nathaniel D; Mooney, David J

    2007-05-01

    The current paradigm in designing biomaterials is to optimize material chemical and physical parameters based on correlations between these parameters and downstream biological responses, whether in vitro or in vivo. Extensive developments in molecular design of biomaterials have facilitated identification of several biophysical and biochemical variables (e.g. adhesion peptide density, substrate elastic modulus) as being critical to cell response. However, these empirical observations do not indicate whether different parameters elicit cell responses by modulating redundant variables of the cell-material interface (e.g. number of cell-material bonds, cell-matrix mechanics). Recently, fluorescence resonance energy transfer (FRET) has been applied to quantitatively analyze parameters of the cell-material interface for both two- and three-dimensional adhesion substrates. Tools based on FRET have been utilized to quantify several parameters of the cell-material interface relevant to cell response, including molecular changes in matrix proteins induced by interactions both with surfaces and cells, the number of bonds between integrins and their adhesion ligands, and changes in the crosslink density of hydrogel synthetic extracellular matrix analogs. As such techniques allow both dynamic and 3-D analyses they will be useful to quantitatively relate downstream cellular responses (e.g. gene expression) to the composition of this interface. Such understanding will allow bioengineers to fully exploit the potential of biomaterials engineered on the molecular scale, by optimizing material chemical and physical properties to a measurable set of interfacial parameters known to elicit a predictable response in a specific cell population. This will facilitate the rational design of complex, multi-functional biomaterials used as model systems for studying diseases or for clinical applications.

  19. Noninvasive fluorescence resonance energy transfer imaging of in vivo premature drug release from polymeric nanoparticles.

    Science.gov (United States)

    Zou, Peng; Chen, Hongwei; Paholak, Hayley J; Sun, Duxin

    2013-11-01

    Understanding in vivo drug release kinetics is critical for the development of nanoparticle-based delivery systems. In this study, we developed a fluorescence resonance energy transfer (FRET) imaging approach to noninvasively monitor in vitro and in vivo cargo release from polymeric nanoparticles. The FRET donor dye (DiO or DiD) and acceptor dye (DiI or DiR) were individually encapsulated into poly(ethylene oxide)-b-polystyrene (PEO-PS) nanoparticles. When DiO (donor) nanoparticles and DiI (acceptor) nanoparticles were coincubated with cancer cells for 2 h, increased FRET signals were observed from cell membranes, suggesting rapid release of DiO and DiI to cell membranes. Similarly, increased FRET ratios were detected in nude mice after intravenous coadministration of DiD (donor) nanoparticles and DiR (acceptor) nanoparticles. In contrast, another group of nude mice i.v. administrated with DiD/DiR coloaded nanoparticles showed decreased FRET ratios. Based on the difference in FRET ratios between the two groups, in vivo DiD/DiR release half-life from PEO-PS nanoparticles was determined to be 9.2 min. In addition, it was observed that the presence of cell membranes facilitated burst release of lipophilic cargos while incorporation of oleic acid-coated iron oxide into PEO-PS nanoparticles slowed the release of DiD/DiR to cell membranes. The developed in vitro and in vivo FRET imaging techniques can be used to screening stable nanoformulations for lipophilic drug delivery.

  20. Detection of influenza A virus based on fluorescence resonance energy transfer from quantum dots to carbon nanotubes.

    Science.gov (United States)

    Tian, Junping; Zhao, Huimin; Liu, Meng; Chen, Yaqiong; Quan, Xie

    2012-04-20

    In this paper, a simple and sensitive approach for H5N1 DNA detection was described based on the fluorescence resonance energy transfer (FRET) from quantum dots (QDs) to carbon nanotubes (CNTs) in a QDs-ssDNA/oxCNTs system, in which the QDs (CdTe) modified with ssDNA were used as donors. In the initial stage, with the strong interaction between ssDNA and oxCNTs, QDs fluorescence was effectively quenched. Upon the recognition of the target, the effective competitive bindings of it to QDs-ssDNA occurred, which decreased the interactions between the QDs-ssDNA and oxCNTs, leading to the recovery of the QDs fluorescence. The recovered fluorescence of QDs was linearly proportional to the concentration of the target in the range of 0.01-20 μM with a detection limit of 9.39 nM. Moreover, even a single-base mismatched target with the same concentration of target DNA can only recover a limited low fluorescence of QDs, illustrating the good anti-interference performance of this QDs-ssDNA/oxCNTs system. This FRET platform in the QDs-ssDNA/oxCNTs system was facilitated to the simple, sensitive and quantitative detection of virus nucleic acids and could have a wide range of applications in molecular diagnosis.

  1. Detection of influenza A virus based on fluorescence resonance energy transfer from quantum dots to carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    Tian Junping [Key Laboratory of Industrial Ecology and Environmental Engineering (Ministry of Education, China), School of Environmental Science and Technology, Dalian University of Technology, Dalian 116024 (China); Zhao Huimin, E-mail: zhaohuim@dlut.edu.cn [Key Laboratory of Industrial Ecology and Environmental Engineering (Ministry of Education, China), School of Environmental Science and Technology, Dalian University of Technology, Dalian 116024 (China); Liu Meng; Chen Yaqiong; Quan Xie [Key Laboratory of Industrial Ecology and Environmental Engineering (Ministry of Education, China), School of Environmental Science and Technology, Dalian University of Technology, Dalian 116024 (China)

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer The quantum dots-ssDNA probe was designed for the determination of virus DNA. Black-Right-Pointing-Pointer The fluorescence of quantum dots was effectively quenched by carbon nanotubes. Black-Right-Pointing-Pointer The addition of target H5N1 DNA restored the quenched fluorescence of quantum dots. Black-Right-Pointing-Pointer The proposed method exhibited high sensitivity and good selectivity for H5N1 DNA. - Abstract: In this paper, a simple and sensitive approach for H5N1 DNA detection was described based on the fluorescence resonance energy transfer (FRET) from quantum dots (QDs) to carbon nanotubes (CNTs) in a QDs-ssDNA/oxCNTs system, in which the QDs (CdTe) modified with ssDNA were used as donors. In the initial stage, with the strong interaction between ssDNA and oxCNTs, QDs fluorescence was effectively quenched. Upon the recognition of the target, the effective competitive bindings of it to QDs-ssDNA occurred, which decreased the interactions between the QDs-ssDNA and oxCNTs, leading to the recovery of the QDs fluorescence. The recovered fluorescence of QDs was linearly proportional to the concentration of the target in the range of 0.01-20 {mu}M with a detection limit of 9.39 nM. Moreover, even a single-base mismatched target with the same concentration of target DNA can only recover a limited low fluorescence of QDs, illustrating the good anti-interference performance of this QDs-ssDNA/oxCNTs system. This FRET platform in the QDs-ssDNA/oxCNTs system was facilitated to the simple, sensitive and quantitative detection of virus nucleic acids and could have a wide range of applications in molecular diagnosis.

  2. Fluorescence resonance-energy-transfer in systems of Rhodamine 6G with ionic liquid showing emissions by excitation at wide wavelength areas.

    Science.gov (United States)

    Izawa, Hironori; Wakizono, Satoshi; Kadokawa, Jun-ichi

    2010-09-14

    Fluorescence resonance-energy-transfer occurred in a solution of Rhodamine 6G in an ionic liquid by excitation at wide wavelength areas owing to specific fluorescent behavior of the ionic liquid to show emissions at each excitation wavelength, which was also observed in the guar gum/ionic liquid gel material containing Rhodamine 6G.

  3. Study on the interaction between fluoroquinolones and erythrosine by absorption, fluorescence and resonance Rayleigh scattering spectra and their application

    Science.gov (United States)

    Wang, Jian; Liu, Zhongfang; Liu, Jiangtao; Liu, Shaopu; Shen, Wei

    2008-03-01

    In pH 4.4-4.5 Britton-Robinson (BR) buffer solution, fluoroquinolone antibiotics (FLQs) including ciprofloxacin (CIP), norfloxacin (NOR), levofloxacin (LEV) and lomefloxacin (LOM) could react with erythrosine (Ery) to form 1:1 ion-association complexes, which not only resulted in the changes of the absorption spectra and the quenching of fluorescence, but also resulted in the great enhancement of resonance Rayleigh scattering (RRS). These offered some indications of the determination of fluoroquinolone antibiotics by spectrophotometric, fluorescence and resonance Rayleigh scattering methods. The detection limits for fluoroquinolone antibiotics were in the range of 0.097-0.265 μg/mL for absorption methods, 0.022-0.100 μg/mL for fluorophotometry and 0.014-0.027 μg/mL for RRS method, respectively. Among them, the RRS method had the highest sensitivity. In this work, the spectral characteristics of the absorption, fluorescence and RRS, the optimum conditions of the reactions and the properties of the analytical chemistry were investigated. The methods have been successfully applied to determination of some fluoroquinolone antibiotics in human urine samples and tablets. Taking CIP-Ery system as an example, the charge distribution, the enthalpy of formation and the mean polarizability were calculated by density function theory (DFT) method. In addition, the reasons for the enhancement of scattering spectra were discussed.

  4. Laser photolysis-resonance fluorescence technique (LP-RF) applied to the study of reactions of atmospheric interest

    Science.gov (United States)

    Albaladejo, J.; Cuevas, C. A.; Notario, A.; Martínez, E.

    Atomic chlorine is highly reactive with a variety of organic and inorganic compounds so that relatively small concentrations can compete with the tropospheric oxidants (OH, O3 and NO3) in determining the tropospheric fate of such compounds [1]. Besides, there is a lot of evidence that bromine compounds play significant role in the ozone chemistry both in the troposphere and in the stratosphere [2]. In this work we show the laser photolysis-resonance fluorescence technique (LP-RF) applied to the study of gas phase reactions of halogen atoms with volatile organic compounds (VOCs) of interest in atmospheric chemistry [3]. By means of this technique is possible to measure the rate constants of theses reactions, and subsequently obtain the Arrhenius parameters. Halogens atoms are produced in a excess of the VOC and He, by photolyzing Cl2 at 308 nm to obtain Cl atoms, or CF2Br2 at 248 nm for Br atoms, both cases using a pulsed excimer laser. The radiation (135 nm) from a microwave-driven lamp, through which He containing a low concentrations of Cl2 or Br2 was flowed, was used to excite the resonance fluorescence from the corresponding halogen atom in the jacketed Pyrex reaction cell. Signal were obtained using photon-counting techniques in conjunction with multichannel scaling. The fluorescence signal from the PMT was processed by a preamplifier and sent to an multichannel scaler to collect the time-resolved signal. The multichannel scaler was coupled to a microcomputer for further kinetics analysis.

  5. 钠离子荧光探针用于检测雪卡毒素的细胞毒性%A cell-based detection of ciguatoxin using sodium fluorescence probe

    Institute of Scientific and Technical Information of China (English)

    袁建辉; 杨慧; 唐焕文; 黄薇; 徐新云; 刘建军; 柯跃斌; 程锦泉; 庄志雄

    2011-01-01

    Objective To establish a cell-based detection method of ciguatoxin using fluorescence assay. Methods Mouse neuroblastoma N-2A cells were exposed to ouabain and veratridine and different concentrations of standard ciguatoxin samples (P-CTX-1) to establish the curvilinear relationship between the toxin dosage and fluorescence intensity using the sodium fluorescence probe CoroNa? Green. The toxicity curvilinear relationship was also generated between the toxin dosage and cell survival using CCK-8 method. Based on these standard curves, the presence of ciguatoxin was detected in 33 samples of deep-sea coral fish. Results A correlation was found between the detection results of cell-based fluorescence assay and cytotoxicity assay, whose detection limit reached 10"13 g/ml and 10-12 g/ml, respectively. The cell-based fluorescent assay sensitivity showed a higher sensitivity than cytotoxicity assay with a 2-4 h reduction of the detection time. Conclusions The cell-based fluorescent assay can quickly and sensitively detect ciguatoxin and may serve as a good option for preliminary screening of the toxin.%目的 研究和建立以细胞为基础的结合钠离子荧光探针检测雪卡毒素的方法,为快速、敏感检测雪卡毒素提供科学的实验依据.方法 采用雪卡毒素标准品(p-CTX-1)结合乌苯苷和黎芦定处理小鼠脑神经瘤细胞(N-2A),用钠离子荧光探针(CoroNaTM Green)标记细胞,利用多功能酶标仪检测荧光强度,建立雪卡毒素浓度与荧光强度的标准曲线关系式;同时,采用CCK-8法和酶标仪检测毒素处理组细胞的存活率,建立细胞毒性实验的标准曲线关系式;分析比较两种方法的灵敏度.以两种方法分别对采集的33份深海珊瑚鱼样品进行毒素检测,对检测结果作分析比较.结果 以细胞为基础的荧光检测法检测样品中毒素的检出限为10-13g/ml,而细胞毒性试验检测法检测样品中毒素的检出限为10-12g/ml,其结果具有一定的

  6. Switchable and selective detection of Zn2+ or Cd2+ in living cells based on 3'-O-substituted arrangement of benzoxazole-derived fluorescent probes.

    Science.gov (United States)

    Xu, Yongqian; Xiao, Liangliang; Sun, Shiguo; Pei, Zhichao; Pei, Yuxin; Pang, Yi

    2014-07-18

    Two benzoxazole-derived ESIPT fluorescent sensors and show highly selective detection of Zn(2+) and Cd(2+), respectively, in aqueous solution and living cells. The selectivity switching from Zn(2+) to Cd(2+) is attributed to the different binding mode which is dependent on the 3'-O-substituted arrangement.

  7. Magnetically engineered Cd-free quantum dots as dual-modality probes for fluorescence/magnetic resonance imaging of tumors.

    Science.gov (United States)

    Ding, Ke; Jing, Lihong; Liu, Chunyan; Hou, Yi; Gao, Mingyuan

    2014-02-01

    Magnetically engineered Cd-free CuInS2@ZnS:Mn quantum dots (QDs) were designed, synthesized, and evaluated as potential dual-modality probes for fluorescence and magnetic resonance imaging (MRI) of tumors in vivo. The synthesis of Mn-doped core-shell structured CuInS2@ZnS mainly comprised three steps, i.e., the preparation of fluorescent CuInS2 seeds, the particle surface coating of ZnS, and the Mn-doping of the ZnS shells. Systematic spectroscopy studies were carried out to illustrate the impacts of ZnS coating and the following Mn-doping on the optical properties of the QDs. In combination with conventional fluorescence, fluorescence excitation, and time-resolved fluorescence measurements, the structure of CuInS2@ZnS:Mn QDs prepared under optimized conditions presented a Zn gradient CuInS2 core and a ZnS outer shell, while Mn ions were mainly located in the ZnS shell, which well balanced the optical and magnetic properties of the resultant QDs. For the following in vivo imaging experiments, the hydrophobic CuInS2@ZnS:Mn QDs were transferred into water upon ligand exchange reactions by replacing the 1-dodecanethiol ligand with dihydrolipoic acid-poly(ethylene glycol) (DHLA-PEG) ligand. The MTT assays based on HeLa cells were carried out to evaluate the cytotoxicity of the current Cd-free CuInS2@ZnS:Mn QDs for comparing with that of water soluble CdTe QDs. Further in vivo fluorescence and MR imaging experiments suggested that the PEGylated CuInS2@ZnS:Mn QDs could well target both subcutaneous and intraperitoneal tumors in vivo.

  8. Special characteristics of fluorescence and resonance Rayleigh scattering for cadmium telluride nanocrystal aqueous solution and its interactions with aminoglycoside antibiotics

    Institute of Scientific and Technical Information of China (English)

    LI TaiShan; LIU ShaoPu; LIU ZhongFang; HU XiaoLi; ZHANG LiPing

    2009-01-01

    CdTe nanocrystals (CdTe NCs) were achieved by reaction of CdCl2 with KHTe solution and were capped with sodium mercaptoacetate. The product was detected by transmission electron microscopy (TEM), high-resolution transmission electron microscopy (HRTEM), energy dispersive spectroscopy (EDS), fluorescence spectra, ultraviolet-visible spectra and X-ray diffraction (XRD). The CdTe NCs are of cubic structure and the average size is about 5 nm. The fluorescence quantum yield of CdTe NCs aqueous solution increased from 37% to 97% after 20 d under room light. The maximum λem of fluorescence changed from 543 nm to 510 nm and the blue shift was 33 nm. CdTe NCs aqueous solution can be steady for at least 10 months at 4℃ in a refrigerator. The resonance Rayleigh scattering (RRS) of CdTe NCs in the aqueous solution was investigated. The maximum scattering peak was located at about 554 nm. The interactions of CdTe NCs with amikacin sulfate (AS) and micronomicin sulfate (MS) were in-vestigated respectively. The effects of AS and MS on fluorescence and RRS of CdTe NCs were analyzed. It was found that AS and MS quenched the photoluminescence of CdTe NCs and enhanced RRS of CdTe NCs. Under optimum conditions, there are linear relationships between quenching intensity (F0-F), intensity of RRS (1-10) and concentration of AS and MS. The detection limits (3σ) of AS and MS are re-spectively 3.4 ng.mL-1 and 2.6 ng.mL-1 by the fluorescence quenching method, and 15.2 ng.mL-1 and 14.0 ng.mL-1 by the RRS method. The methods have high sensitivity, thus CdTe NCs may be used as fluorescence probes and RRS probes for the detection of aminoglycoside antibiotics.

  9. Graphene oxide based fluorescence resonance energy transfer and loop-mediated isothermal amplification for white spot syndrome virus detection.

    Science.gov (United States)

    Waiwijit, U; Phokaratkul, D; Kampeera, J; Lomas, T; Wisitsoraat, A; Kiatpathomchai, W; Tuantranont, A

    2015-10-20

    Graphene oxide (GO) is attractived for biological or medical applications due to its unique electrical, physical, optical and biological properties. In particular, GO can adsorb DNA via π-π stacking or non-covalent interactions, leading to fluorescence quenching phenomenon applicable for bio-molecular detection. In this work, a new method for white spot syndrome virus (WSSV)-DNA detection is developed based on loop-mediated isothermal amplification (LAMP) combined with fluorescence resonance energy transfer (FRET) between GO and fluorescein isothiocyanate-labeled probe (FITC-probe). The fluorescence quenching efficiency of FITC-probe was found to increase with increasing GO concentration and reached 98.7% at a GO concentration of 50 μg/ml. The fluorescence intensity of FITC-probe was recovered after hybridization with WSSV LAMP product with an optimal hybridization time of 10 min and increased accordingly with increasing amount of LAMP products. The detection limit was estimated to be as low as 10 copies of WSSV plasmid DNA or 0.6 fg of the total DNA extracted from shrimp infected with WSSV. In addition, no cross reaction was observed with other common shrimp viral pathogens. Therefore, the GO-FRET-LAMP technique is promising for fast, sensitive and specific detection of DNAs.

  10. Spectrophotometry near the atmospheric cutoff of the strongest Bowen resonance fluorescence lines of O III in two planetary nebulae

    Science.gov (United States)

    O'Dell, C. R.; Opal, Chet B.

    1989-01-01

    Spectrophotometric results are presented for the stronger, well-resolved Bowen O III resonance fluorescence emission lines in the planetary nebulae 7027 and NGC 7662 down to and including the intrinsically strong line at 3133 A. These data are combined with results from the IUE atlas of spectra and similar results for the longer wavelength lines by Likkel and Aller (1986) to give the first full coverage of the Bowen lines. Good agreement is found with fluorescence theory for the primary cascade lines, except for the Likkel and Aller results. The efficiency of conversion of the exciting He II Ly-alpha into O III lines is determined, and values comparable to other planetary nebulae are found.

  11. Charge-controlled assembling of bacteriorhodopsin and semiconductor quantum dots for fluorescence resonance energy transfer-based nanophotonic applications

    Science.gov (United States)

    Bouchonville, Nicolas; Molinari, Michael; Sukhanova, Alyona; Artemyev, Mikhail; Oleinikov, Vladimir A.; Troyon, Michel; Nabiev, Igor

    2011-01-01

    The fluorescence resonance energy transfer (FRET) between quantum dots (QDs) and photochromic protein bacteriorhodopsin within its natural purple membrane (PM) is explored to monitor their assembling. It is shown that the efficiency of FRET may be controlled by variation of the surface charge and thickness of QD organic coating. Atomic force microscopy imaging revealed correlation between the surface charge of QDs and degree of their ordering on the surface of PM. The most FRET-efficient QD-PM complexes have the highest level of QDs ordering, and their assembling design may be further optimized to engineer hybrid materials with advanced biophotonic and photovoltaic properties.

  12. Binomial distribution-based quantitative measurement of multiple-acceptors fluorescence resonance energy transfer by partially photobleaching acceptor

    Science.gov (United States)

    Zhang, Lili; Yu, Huaina; Zhang, Jianwei; Chen, Tongsheng

    2014-06-01

    We report that binomial distribution depending on acceptor photobleaching degree can be used to characterize the proportions of various kinds of FRET (Fluorescence Resonance Energy Transfer) constructs resulted from partial acceptor photobleaching of multiple-acceptors FRET system. On this basis, we set up a rigorous quantitation theory for multiple-acceptors FRET construct named as Mb-PbFRET which is not affected by the imaging conditions and fluorophore properties. We experimentally validate Mb-PbFRET with FRET constructs consisted of one donor and two or three acceptors inside living cells on confocal and wide-field microscopes.

  13. Characterization and inhibition of norovirus proteases of genogroups I and II using a fluorescence resonance energy transfer assay

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Kyeong-Ok [Department of Diagnostic Medicine and Pathobiology, College of Veterinary Medicine, Kansas State University, KS 66506 (United States); Takahashi, Daisuke; Prakash, Om [Department of Biochemistry, Kansas State University, Manhattan, KS 66506 (United States); Kim, Yunjeong, E-mail: ykim@vet.ksu.edu [Department of Diagnostic Medicine and Pathobiology, College of Veterinary Medicine, Kansas State University, KS 66506 (United States)

    2012-02-20

    Noroviruses are the major cause of food- or water-borne gastroenteritis outbreaks in humans. The norovirus protease that cleaves a large viral polyprotein to nonstructural proteins is essential for virus replication and an attractive target for antiviral drug development. Noroviruses show high genetic diversity with at least five genogroups, GI-GV, of which GI and GII are responsible for the majority of norovirus infections in humans. We cloned and expressed proteases of Norwalk virus (GI) and MD145 virus (GII) and characterized the enzymatic activities with fluorescence resonance energy transfer substrates. We demonstrated that the GI and GII proteases cleaved the substrates derived from the naturally occurring cleavage site in the open reading frame (ORF) 1 of G1 norovirus with similar efficiency, and that enzymatic activity of both proteases was inhibited by commercial protease inhibitors including chymostatin. The interaction of chymostatin to Norwalk virus protease was validated by nuclear magnetic resonance (NMR) spectroscopy.

  14. Full genotyping of a highly polymorphic human gene trait by time-resolved fluorescence resonance energy transfer.

    Directory of Open Access Journals (Sweden)

    Edoardo Totè

    Full Text Available The ability of detecting the subtle variations occurring, among different individuals, within specific DNA sequences encompassed in highly polymorphic genes discloses new applications in genomics and diagnostics. DQB1 is a gene of the HLA-II DQ locus of the Human Leukocyte Antigens (HLA system. The polymorphisms of the trait of the DQB1 gene including codons 52-57 modulate the susceptibility to a number of severe pathologies. Moreover, the donor-receiver tissue compatibility in bone marrow transplantations is routinely assessed through crossed genotyping of DQB and DQA. For the above reasons, the development of rapid, reliable and cost-effective typing technologies of DQB1 in general, and more specifically of the codons 52-57, is a relevant although challenging task. Quantitative assessment of the fluorescence resonance energy transfer (FRET efficiency between chromophores labelling the opposite ends of gene-specific oligonucleotide probes has proven to be a powerful tool to type DNA polymorphisms with single-nucleotide resolution. The FRET efficiency can be most conveniently quantified by applying a time-resolved fluorescence analysis methodology, i.e. time-correlated single-photon counting, which allows working on very diluted template specimens and in the presence of fluorescent contaminants. Here we present a full in-vitro characterization of the fluorescence responses of two probes when hybridized to oligonucleotide mixtures mimicking all the possible genotypes of the codons 52-57 trait of DQB1 (8 homozygous and 28 heterozygous. We show that each genotype can be effectively tagged by the combination of the fluorescence decay constants extrapolated from the data obtained with such probes.

  15. Structurally Rigid 9-Amino-benzo[c]cinnoliniums Make Up a Class of Compact and Large Stokes-Shift Fluorescent Dyes for Cell-Based Imaging Applications.

    Science.gov (United States)

    Shen, Yanming; Shang, Zhihao; Yang, Yanhong; Zhu, Shaojia; Qian, Xuhong; Shi, Ping; Zheng, Jing; Yang, Youjun

    2015-06-05

    Classic fluorescent dyes, such as coumarin, naphthalimide, fluorescein, BODIPY, rhodamine, and cyanines, are cornerstones of various spectroscopic and microscopic methods, which hold a prominent position in biological studies. We recently found that 9-amino-benzo[c]cinnoliniums make up a novel group of fluorophores that can be used in biological studies. They are featured with a succinct conjugative push-pull backbone, a broad absorption band, and a large Stokes shift. They are potentially useful as a small-molecule alternative to R-phycoerythrin to pair with fluorescein in multiplexing applications.

  16. Resonance fluorescence of a two-level quantum emitter near a plasmonic nanoparticle: role of the near-field polarization

    Science.gov (United States)

    Vladimirova, Yu V.; Chubchev, E. D.; Zadkov, V. N.

    2017-02-01

    It is demonstrated that the interaction of a two-level quantum emitter (atom, molecule, etc) with a plasmonic nanoparticle (prolate nanospheroid) in an external laser field features either an essential increase (up to a few orders of magnitude) or reduction (up to a few times) of the total decay rate of the emitter in specific areas around the nanoparticle in contrast to its decay rate in a vacuum. It is also shown that the resonance fluorescence spectrum of the emitter in close proximity to a plasmonic nanoparticle is very sensitive to both the location of the emitter around the nanoparticle and to polarization of the near-field, which depends in turn on the polarization of the incident laser field. This can be used in engineering potential quantum optics experiments with quantum emitters in the near-field, as well as for 3D nanoscopy of the near-field by registering the resonance fluorescence spectra of quantum emitters scattered in the vicinity of a plasmonic nanoparticle.

  17. Collective resonance fluorescence in small and dense atom clouds: Comparison between theory and experiment

    CERN Document Server

    Jenkins, S D; Javanainen, J; Jennewein, S; Bourgain, R; Pellegrino, J; Sortais, Y R P; Browaeys, A

    2016-01-01

    We study the emergence of a collective optical response of a cold and dense $^{87}$Rb atomic cloud to a near-resonant low-intensity light when the atom number is gradually increased. Experimental observations are compared with microscopic stochastic simulations of recurrent scattering processes between the atoms that incorporate the atomic multilevel structure and the optical measurement setup. We analyze the optical response of an inhomogeneously-broadened gas and find that the experimental observations of the resonance line shifts and the total collected scattered light intensity in cold atom clouds substantially deviate from those of thermal atomic ensembles, indicating strong light-induced resonant dipole-dipole interactions between the atoms. At high densities, the simulations also predict a significantly slower decay of light-induced excitations in cold than in thermal atom clouds. The role of dipole-dipole interactions is discussed in terms of resonant coupling examples and the collective radiative exc...

  18. A homogeneous europium cryptate-based assay for the diagnosis of mutations by time-resolved fluorescence resonance energy transfer.

    Science.gov (United States)

    Lopez-Crapez, E; Bazin, H; Andre, E; Noletti, J; Grenier, J; Mathis, G

    2001-07-15

    Oligonucleotide ligation assay (OLA) is considered to be a very useful methodology for the detection and characterization of mutations, particularly for clinical purposes. The fluorescence resonance energy transfer between a fluorescent donor and a suitable fluorophore as acceptor has been applied in the past to several scientific fields. This technique is well adapted to nucleic acid analysis such as DNA sequencing, DNA hybridization and polymerase chain reaction. We describe here a homogeneous format based on the use of a rare earth cryptate label as donor: tris-bipyridine-Eu(3+). The long-lived fluorescence of this label makes it possible to reach a high sensitivity by using a time-resolved detection mode. A non-radiative energy transfer technology, known as time-resolved amplification of cryptate emission (TRACE((R))) characterized by a temporal and spectral selectivity has been developed. The TRACE((R)) detection of characterized single nucleotide polymorphism using the OLA for allelic discrimination is proposed. We demonstrate the potentialities of this OLA-TRACE((R)) methodology through the analysis of K-ras oncogene point mutations.

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

    Science.gov (United States)

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

    2014-01-01

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

  20. Establishment and validation of whole-cell based fluorescence assays to identify anti-mycobacterial compounds using the Acanthamoeba castellanii-Mycobacterium marinum host-pathogen system.

    Directory of Open Access Journals (Sweden)

    Sébastien Kicka

    Full Text Available Tuberculosis is considered to be one of the world's deadliest disease with 2 million deaths each year. The need for new antitubercular drugs is further exacerbated by the emergence of drug-resistance strains. Despite multiple recent efforts, the majority of the hits discovered by traditional target-based screening showed low efficiency in vivo. Therefore, there is heightened demand for whole-cell based approaches directly using host-pathogen systems. The phenotypic host-pathogen assay described here is based on the monitoring of GFP-expressing Mycobacterium marinum during infection of the amoeba Acanthamoeba castellanii. The assay showed straight-forward medium-throughput scalability, robustness and ease of manipulation, demonstrating its qualities as an efficient compound screening system. Validation with a series of known antitubercular compounds highlighted the advantages of the assay in comparison to previously published macrophage-Mycobacterium tuberculosis-based screening systems. Combination with secondary growth assays based on either GFP-expressing D. discoideum or M. marinum allowed us to further fine-tune compound characterization by distinguishing and quantifying growth inhibition, cytotoxic properties and antibiotic activities of the compounds. The simple and relatively low cost system described here is most suitable to detect anti-infective compounds, whether they present antibiotic activities or not, in which case they might exert anti-virulence or host defense boosting activities, both of which are largely overlooked by classical screening approaches.

  1. Establishment and validation of whole-cell based fluorescence assays to identify anti-mycobacterial compounds using the Acanthamoeba castellanii-Mycobacterium marinum host-pathogen system.

    Science.gov (United States)

    Kicka, Sébastien; Trofimov, Valentin; Harrison, Christopher; Ouertatani-Sakouhi, Hajer; McKinney, John; Scapozza, Leonardo; Hilbi, Hubert; Cosson, Pierre; Soldati, Thierry

    2014-01-01

    Tuberculosis is considered to be one of the world's deadliest disease with 2 million deaths each year. The need for new antitubercular drugs is further exacerbated by the emergence of drug-resistance strains. Despite multiple recent efforts, the majority of the hits discovered by traditional target-based screening showed low efficiency in vivo. Therefore, there is heightened demand for whole-cell based approaches directly using host-pathogen systems. The phenotypic host-pathogen assay described here is based on the monitoring of GFP-expressing Mycobacterium marinum during infection of the amoeba Acanthamoeba castellanii. The assay showed straight-forward medium-throughput scalability, robustness and ease of manipulation, demonstrating its qualities as an efficient compound screening system. Validation with a series of known antitubercular compounds highlighted the advantages of the assay in comparison to previously published macrophage-Mycobacterium tuberculosis-based screening systems. Combination with secondary growth assays based on either GFP-expressing D. discoideum or M. marinum allowed us to further fine-tune compound characterization by distinguishing and quantifying growth inhibition, cytotoxic properties and antibiotic activities of the compounds. The simple and relatively low cost system described here is most suitable to detect anti-infective compounds, whether they present antibiotic activities or not, in which case they might exert anti-virulence or host defense boosting activities, both of which are largely overlooked by classical screening approaches.

  2. Label-free distinguishing between neurons and glial cells based on two-photon excited fluorescence signal of neuron perinuclear granules

    Science.gov (United States)

    Du, Huiping; Jiang, Liwei; Wang, Xingfu; Liu, Gaoqiang; Wang, Shu; Zheng, Liqin; Li, Lianhuang; Zhuo, Shuangmu; Zhu, Xiaoqin; Chen, Jianxin

    2016-08-01

    Neurons and glial cells are two critical cell types of brain tissue. Their accurate identification is important for the diagnosis of psychiatric disorders such as depression and schizophrenia. In this paper, distinguishing between neurons and glial cells by using the two-photon excited fluorescence (TPEF) signals of intracellular intrinsic sources was performed. TPEF microscopy combined with TUJ-1 and GFAP immunostaining and quantitative image analysis demonstrated that the perinuclear granules of neurons in the TPEF images of brain tissue and the primary cultured cortical cells were a unique characteristic of neurons compared to glial cells which can become a quantitative feature to distinguish neurons from glial cells. With the development of miniaturized TPEF microscope (‘two-photon fiberscopes’) imaging devices, TPEF microscopy can be developed into an effective diagnostic and monitoring tool for psychiatric disorders such as depression and schizophrenia.

  3. Development of homogeneous binding assays based on fluorescence resonance energy transfer between quantum dots and Alexa Fluor fluorophores.

    Science.gov (United States)

    Nikiforov, Theo T; Beechem, Joseph M

    2006-10-01

    We studied the fluorescence resonance energy transfer (FRET) between quantum dots emitting at 565, 605, and 655 nm as energy donors and Alexa Fluor fluorophores with absorbance maxima at 594, 633, 647, and 680 nm as energy acceptors. As a first step, we prepared covalent conjugates between all three types of quantum dots and each of the Alexa Fluor fluorophores that could act as an energy acceptor. All of these conjugates displayed efficient resonance energy transfer. Then we prepared covalent conjugates of these quantum dots with biotin, fluorescein, and cortisol and established that the binding of these conjugates to suitable Alexa Fluor-labeled antibodies and streptavidin (in the case of biotin) can be efficiently detected by measuring the resonance energy transfer in homogeneous solutions. Finally, based on these observations, competitive binding assays for these three small analytes were developed. The performance of these assays as a function of the degree of labeling of the quantum dots was evaluated. It was found that decreasing the degree of loading of the quantum dots leads to decreases of the limits of detection. The results show the great potential of this FRET system for the development of new homogeneous binding assays.

  4. Facile preparation of Gd3+ doped carbon quantum dots: Photoluminescence materials with magnetic resonance response as magnetic resonance/fluorescence bimodal probes

    Science.gov (United States)

    Ren, X. Y.; Yuan, X. X.; Wang, Y. P.; Liu, C. L.; Qin, Y.; Guo, L. P.; Liu, L. H.

    2016-07-01

    There are a few bimodal molecular imaging probes constructed by gadolinium (3+) ions in combination with carbon quantum dots (CQDs), and the reported ones show such obvious drawbacks as low luminous efficiency and weak MRI contrast. In the paper, a kind of CQDs photoluminescence materials with magnetic resonance response was prepared by hydrothermal method and employing gadopentetate monomeglumine (GdPM) as a precusor. Here, the GdPM plays a role of not only carbon source, but also gadolinium (3+) sources. When the GdPM aqueous solution with a concentration of 4 mg mL-1 was pyrolyzed under 220 °C and 2.0 MPa for 8 h, an optimal CQDs was obtained which are doped with gadolinium (3+) ions in both chelates and Gd2O3 (named as Gd3+-CQDs). The average diameter of the Gd3+-CQDs is about 1.6 nm, which show a high photoluminescence quantum yield of 7.1%, as well as high longitudinal relaxivity (r1) of 9.87 mM-1 s-1. And owing to the unconspicuous cell toxicity, the Gd3+-CQDs show big possibility for clinical application in magnetic resonance/fluorescence bimodal molecular imaging.

  5. Special characteristics of fluorescence and resonance Rayleigh scattering for cadmium telluride nanocrystal aqueous solution and its interactions with aminoglycoside antibiotics

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    CdTe nanocrystals(CdTe NCs) were achieved by reaction of CdCl2 with KHTe solution and were capped with sodium mercaptoacetate.The product was detected by transmission electron microscopy(TEM),high-resolution transmission electron microscopy(HRTEM),energy dispersive spectroscopy(EDS),fluorescence spectra,ultraviolet-visible spectra and X-ray diffraction(XRD).The CdTe NCs are of cubic structure and the average size is about 5 nm.The fluorescence quantum yield of CdTe NCs aqueous solution increased from 37% to 97% after 20 d under room light.The maximum λem of fluorescence changed from 543 nm to 510 nm and the blue shift was 33 nm.CdTe NCs aqueous solution can be steady for at least 10 months at 4℃ in a refrigerator.The resonance Rayleigh scattering(RRS) of CdTe NCs in the aqueous solution was investigated.The maximum scattering peak was located at about 554 nm.The interactions of CdTe NCs with amikacin sulfate(AS) and micronomicin sulfate(MS) were investigated respectively.The effects of AS and MS on fluorescence and RRS of CdTe NCs were analyzed.It was found that AS and MS quenched the photoluminescence of CdTe NCs and enhanced RRS of CdTe NCs.Under optimum conditions,there are linear relationships between quenching intensity(F0-F),intensity of RRS(I-I0) and concentration of AS and MS.The detection limits(3б) of AS and MS are respectively 3.4 ng·mL-1 and 2.6 ng·mL-1 by the fluorescence quenching method,and 15.2 ng·mL-1 and 14.0 ng·mL-1 by the RRS method.The methods have high sensitivity,thus CdTe NCs may be used as fluorescence probes and RRS probes for the detection of aminoglycoside antibiotics.

  6. Handheld Fluorescence Resonance Energy Transfer (FRET)-Aptamer Sensor for Bone Markers

    Science.gov (United States)

    Bruno, John G.

    2015-01-01

    Astronauts lose significant bone mass during lengthy space flights. NASA wishes to monitor this bone loss in order to develop nutritional and exercise countermeasures. Operational Technologies Corporation (OpTech) has developed a handheld device that quantifies bone loss in a spacecraft environment. The innovation works by adding fluorescent dyes and quenchers to aptamers to enable pushbutton, one-step bind-and-detect FRET assays that can be freeze-dried, rehydrated with body fluids, and used to quantify bone loss.

  7. Monochromatic X-ray propagation in multi-Z media for imaging and diagnostics including Kα Resonance Fluorescence

    Science.gov (United States)

    Westphal, Maximillian; Lim, Sara; Nahar, Sultana; Pradhan, Anil

    2016-05-01

    Aimed at monochromatic X-ray imaging and therapy, broadband, monochromatic, and quasi-monochromatic X-ray sources and propagation through low and high-Z (HZ) media were studied with numerically and experimentally. Monte Carlo simulations were performed using the software package Geant4, and a new code Photx, to simulate X-ray image contrast, depth of penetration, and total attenuation. The data show that monochromatic and quasi-monochromatic X-rays achieve improved contrast at lower absorbed radiation doses compared to conventional broadband 120 kV or CT scans. Experimental quasi-monochromatic high-intensity laser-produced plasma sources and monochromatic synchrotron beam data are compared. Physical processes responsible for X-ray photoexcitation and absorption are numerically modelled, including a novel mechanism for accelerating Kα resonance fluorescence via twin monochromatic X-ray beam. Potential applications are medical diagnostics and high-Z material detection. Acknowledgement: Ohio Supercomputer Center, Columbus, OH.

  8. Depth profiles of pulmonary surfactant protein B in phosphatidylcholine bilayers, studied by fluorescence and electron spin resonance spectroscopy

    DEFF Research Database (Denmark)

    Cruz, A; Casals, C; Plasencia, I;

    1998-01-01

    Pulmonary surfactant-associated protein B (SP-B) has been isolated from porcine lungs and reconstituted in bilayers of dipalmitoylphosphatidylcholine (DPPC) or egg yolk phosphatidylcholine (PC) to characterize the extent of insertion of the protein into phospholipid bilayers. The parameters...... for the interaction of SP-B with DPPC or PC using different reconstitution protocols have been estimated from the changes induced in the fluorescence emission spectrum of the single protein tryptophan. All the different reconstituted SP-B-phospholipid preparations studied had similar Kd values for the binding....... These differences in the extent of insertion lead to qualitative and quantitative differences in the effect of the protein on the mobility of the phospholipid acyl chains, as studied by spin-label electron spin resonance (ESR) spectroscopy, and could represent different functional stages in the surfactant cycle...

  9. Addendum to "Determination of $\\gamma$-ray widths in $^{15}$N using nuclear resonance fluorescence''

    CERN Document Server

    Szücs, Tamás

    2015-01-01

    The determination of absolute widths of two observed levels above the proton threshold in $^{15}$N has been improved by a combined analysis of our recent $^{15}$N($\\gamma$,$\\gamma'$)$^{15}$N$^\\ast$ photon scattering data, resonance strengths $\\omega\\gamma$ of the $^{14}$C($p$,$\\gamma$)$^{15}$N reaction, and $\\gamma$-ray branchings $b_{\\gamma,i}$ in $^{15}$N. The revised data are compared to the adopted values, and some inconsistencies in the adopted values are illustrated.

  10. Förster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multi-scale molecular rulers

    Science.gov (United States)

    Ploetz, Evelyn; Lerner, Eitan; Husada, Florence; Roelfs, Martin; Chung, Sangyoon; Hohlbein, Johannes; Weiss, Shimon; Cordes, Thorben

    2016-09-01

    Advanced microscopy methods allow obtaining information on (dynamic) conformational changes in biomolecules via measuring a single molecular distance in the structure. It is, however, extremely challenging to capture the full depth of a three-dimensional biochemical state, binding-related structural changes or conformational cross-talk in multi-protein complexes using one-dimensional assays. In this paper we address this fundamental problem by extending the standard molecular ruler based on Förster resonance energy transfer (FRET) into a two-dimensional assay via its combination with protein-induced fluorescence enhancement (PIFE). We show that donor brightness (via PIFE) and energy transfer efficiency (via FRET) can simultaneously report on e.g., the conformational state of double stranded DNA (dsDNA) following its interaction with unlabelled proteins (BamHI, EcoRV, and T7 DNA polymerase gp5/trx). The PIFE-FRET assay uses established labelling protocols and single molecule fluorescence detection schemes (alternating-laser excitation, ALEX). Besides quantitative studies of PIFE and FRET ruler characteristics, we outline possible applications of ALEX-based PIFE-FRET for single-molecule studies with diffusing and immobilized molecules. Finally, we study transcription initiation and scrunching of E. coli RNA-polymerase with PIFE-FRET and provide direct evidence for the physical presence and vicinity of the polymerase that causes structural changes and scrunching of the transcriptional DNA bubble.

  11. A fluorescence detected magnetic resonance investigation of the carotenoid triplet states associated with Photosystem II of isolated spinach thylakoid membranes

    CERN Document Server

    Santabarbara, S; Carbonera, D; Heathcote, P

    2005-01-01

    The carotenoid triplet populations associated with the fluorescence emission chlorophyll forms of Photosystem II have been investigated in isolated spinach thylakoid membranes by means of fluorescence detected magnetic resonance in zero field (FDMR). The spectra collected in the 680-690 nm emission range, have been fitted by a global analysis procedure. At least five different carotenoid triplet states coupled to the terminal emitting chlorophyll forms of PS II, peaking at 682 nm, 687 nm and 692 nm, have been characterised. The triplets associated with the outer antenna emission forms, at 682 nm, have zero field splitting parameters D = 0.0385 cm/sup -1/, E = 0.00367 cm/sup -1/; D = 0.0404 cm/sup -1/, E = 0.00379 cm/sup -1/ and D = 0.0386 cm/sup -1/, E = 0.00406 cm/sup -1/ which are very similar to those previously reported for the xanthophylls of the isolated LHC II complex. Therefore the FDMR spectra recorded in this work provide insights into the organisation of the LHC II complex in the unperturbed enviro...

  12. Towards understanding the free and receptor bound conformation of neuropeptide Y by fluorescence resonance energy transfer studies.

    Science.gov (United States)

    Haack, Michael; Beck-Sickinger, Annette G

    2009-06-01

    Despite a considerable sequence identity of the three mammalian hormones of the neuropeptide Y family, namely neuropeptide Y, peptide YY and pancreatic polypeptide, their structure in solution is described to be different. A so-called pancreatic polypeptide-fold has been identified for pancreatic polypeptide, whereas the structure of the N-terminal segment of neuropeptide Y is unknown. This element is important for the binding of neuropeptide Y to two of its relevant receptors, Y(1) and Y(5), but not to the Y(2) receptor subtype. In this study now, three doubly fluorescent-labeled analogs of neuropeptide Y have been synthesized that still bind to the Y(5) receptor with high affinity to investigate the conformation in solution and, for the first time, to probe the conformational changes upon binding of the ligand to its receptor in cell membrane preparations. The results obtained from the fluorescence resonance energy transfer investigations clearly show considerable differences in transfer efficiency that depend both on the solvent as well as on the peptide concentration. However, the studies do not support a pancreatic polypeptide-like folding of neuropeptide Y in the presence of membranes that express the human Y(5) receptor subtype.

  13. Förster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multi-scale molecular rulers

    Science.gov (United States)

    Ploetz, Evelyn; Lerner, Eitan; Husada, Florence; Roelfs, Martin; Chung, SangYoon; Hohlbein, Johannes; Weiss, Shimon; Cordes, Thorben

    2016-01-01

    Advanced microscopy methods allow obtaining information on (dynamic) conformational changes in biomolecules via measuring a single molecular distance in the structure. It is, however, extremely challenging to capture the full depth of a three-dimensional biochemical state, binding-related structural changes or conformational cross-talk in multi-protein complexes using one-dimensional assays. In this paper we address this fundamental problem by extending the standard molecular ruler based on Förster resonance energy transfer (FRET) into a two-dimensional assay via its combination with protein-induced fluorescence enhancement (PIFE). We show that donor brightness (via PIFE) and energy transfer efficiency (via FRET) can simultaneously report on e.g., the conformational state of double stranded DNA (dsDNA) following its interaction with unlabelled proteins (BamHI, EcoRV, and T7 DNA polymerase gp5/trx). The PIFE-FRET assay uses established labelling protocols and single molecule fluorescence detection schemes (alternating-laser excitation, ALEX). Besides quantitative studies of PIFE and FRET ruler characteristics, we outline possible applications of ALEX-based PIFE-FRET for single-molecule studies with diffusing and immobilized molecules. Finally, we study transcription initiation and scrunching of E. coli RNA-polymerase with PIFE-FRET and provide direct evidence for the physical presence and vicinity of the polymerase that causes structural changes and scrunching of the transcriptional DNA bubble. PMID:27641327

  14. Bioimaging of geographically adjacent proteins in a single cell by quantum dot-based fluorescent resonance energy transfer.

    Science.gov (United States)

    Kang, Won Jun; Ko, Mee Hyang; Lee, Dong Soo; Kim, Soonhag

    2009-12-01

    Thousands of proteins are simultaneously involved in the maintenance of a single cancer cell. Fluorescent resonance energy transfer (FRET) is one of the most general techniques for imaging biologically interacting molecules in a cell. Here, we applied FRET to image the co-localization of two proteins that do not interact biologically (nucleolin and integrin α(v) β(3),) both of which are highly expressed in the plasma membrane of cancer cells. AS1411 aptamer, which targets nucleolin, was labeled by Cy3 (Cy3-AS1411) and arginine-glycine-aspartic acid (RGD) peptide, which targets integrin α(v) β(3) , was conjugated with quantum dot (525 nm, Qd) Qd arginine-glycine-aspartic acid (Qd-RGD). FRET activities between Cy3-AS1411 and Qd-RGD were measured in HeLa cells, a human cervical cancer cell line. FRET phenomena between Qd and Cy3 showed good compatibility according to proximity. The fluorescence signature using Qd-RGD and Cy3-AS1411 showed that nucleolin and integrin α(v) β(3) proteins were highly expressed in HeLa cells. Co-incubation of Qd-RGD and Cy3-AS1411 in a single HeLa cell demonstrated that the fluorescence overlay by FRET was quantitatively and geographically quite different from that of individual confocal images. These results suggest that Qd-based FRET analysis can provide information on geographical co-localization of proteins in naïve cells, which is very important for determining the molecular and cellular functions of genes involved in cancers and other clinical diseases.

  15. Computer simulations on resonant fluorescence spectra in atomic gases in two monochromatic laser fields of arbitrary intensity and magnetic field

    Science.gov (United States)

    Karagodova, Tamara Y.

    1996-03-01

    In the intense radiation fields with power density from 104W/cm2 to 109W/cm2 the essential modification of electronic states of atoms occurs displaying, in particular, in modifications of resonant fluorescence (rf) spectra. We use 'Fermi golden rule' for calculations of relative intensities and frequencies for rf multiplet for real multilevel initially unexcited atoms in two monochromatic laser fields of arbitrary intensity resonant to adjacent transitions of (Xi) or (Lambda) types and magnetic field, giving the level splittings of different values from Zeeman to Paschen-Back effect. The dependence of quasienergies on parameters obtained with the help of a sorting program permits us to define the values of parameters for which the states of the system are mixed and so to receive the correct probability amplitudes for instantaneous or adiabatic regimes of switching the perturbation. The analysis of the quasienergies and form of rf spectra permits us to get relations between the form of the spectra and modifications of electronic structure of the atom due to radiation fields and external magnetic field.

  16. Identification of weak autoionizing resonances observed through fluorescence from the satellite states of Ar{sup +}

    Energy Technology Data Exchange (ETDEWEB)

    McLaughlin, K.W.; Yenen, O.; Samson, J.A.R. [Univ. of Nebraska, Lincoln, NE (United States)] [and others

    1997-04-01

    Photoionization accompanied by excitation of the residual ionic state violates an independent electron model since, according to QED, photons interact only with individual electrons. By allowing measurements at a threshold event with high resolution, the observation of the fluorescence from the decay of these excited states (satellite states) is a sensitive method in the study of electron-electron interactions, providing complementary information to photoelectron spectroscopy. In the measurements reported here, an atomic beam of argon has been photoionized with 34 to 39 eV synchrotron radiation at beamline 9.0.1 of the Advanced Light Source. This energy range encompasses the 3p{sup 4} [{sup 3}P] 4p {sup 4}P, {sup 2}P, and {sup 2}D as well as the [{sup 1}D]4p {sup 2}F satellite states of Ar{sup +}. By observing the fine-structure resolved fluorescence from these satellite states, new Rydberg series and extensions of previously known series have been resolved with an energy resolution of 3 meV. With the high photon flux available from the high resolution monochromator of beamline 9.0.1, even the weakly excited [{sup 3}P] 4p ({sup 2}S) ns,d autoionizing structure has been observed for the first time.

  17. Characterization of the mode of incorporation of lipophilic compounds in solid dispersions at the nanoscale using fluorescence resonance energy transfer (FRET)

    NARCIS (Netherlands)

    van Drooge, Dirk Jan; Braeckmans, Kevin; Hinrichs, Wouter L. J.; Remaut, Katrien; De Smedt, Stefaan C.; Frijlink, Henderik W.

    2006-01-01

    Efficient engineering of solid dispersions stagnates by the current inability to establish the mode of drug distribution on a molecular level at a low drug load. This study describes the application of fluorescence resonance energy transfer (FRET) to characterize the mode of incorporation of dispers

  18. A new assay format for NF-kappaB based on a DNA triple helix and a fluorescence resonance energy transfer.

    Science.gov (United States)

    Altevogt, Dominik; Hrenn, Andrea; Kern, Claudia; Clima, Lilia; Bannwarth, Willi; Merfort, Irmgard

    2009-10-07

    Herein we report a feasibility study for a new concept to detect DNA binding protein NF-kappaB based on a DNA triple helix formation in combination with a fluorescence resonance energy transfer (FRET). The new principle avoids expensive antibodies and radioactivity and might have implications for assays of other DNA binding proteins.

  19. Rapid peptide based diagnosis: peptide-based Fluorescence Resonance Energy Transfer (FRET) protease substrates for the detection and diagnosis of bacillus spp

    NARCIS (Netherlands)

    Bikker, F.J.; Kaman, W.E.

    2014-01-01

    We describe the development of a highly specific protease-based Fluorescence Resonance Energy Transfer (FRET) assay for easy and rapid detection both in vitro and in vivo of Bacillus spp, including Bacillus anthracis. Synthetic substrates for B. anthracis proteases were designed and exposed to secre

  20. Resonances

    DEFF Research Database (Denmark)

    an impetus or drive to that account: change, innovation, rupture, or discontinuity. Resonances: Historical Essays on Continuity and Change explores the historiographical question of the modes of interrelation between these motifs in historical narratives. The essays in the collection attempt to realize...... theoretical consciousness through historical narrative ‘in practice’, by discussing selected historical topics from Western cultural history, within the disciplines of history, literature, visual arts, musicology, archaeology, philosophy, and theology. The title Resonances indicates the overall perspective...

  1. Interfacial transduction of nucleic acid hybridization using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Algar, W Russ; Krull, Ulrich J

    2009-01-06

    Fluorescence resonance energy transfer (FRET) using immobilized quantum dots (QDs) as energy donors was explored as a transduction method for the detection of nucleic acid hybridization at an interface. This research was motivated by the success of the QD-FRET-based transduction of nucleic acid hybridization in solution-phase assays. This new work represents a fundamental step toward the assembly of a biosensor, where immobilization of the selective chemistry on a surface is desired. After immobilizing QD-probe oligonucleotide conjugates on optical fibers, a demonstration of the retention of selectivity was achieved by the introduction of acceptor (Cy3)-labeled single-stranded target oligonucleotides. Hybridization generated the proximity required for FRET, and the resulting fluorescence spectra provided an analytical signal proportional to the amount of target. This research provides an important framework for the future development of nucleic acid biosensors based on QDs and FRET. The most important findings of this work are that (1) a QD-FRET solid-phase hybridization assay is viable and (2) a passivating layer of denatured bovine serum albumin alleviates nonspecific adsorption, ultimately resulting in (3) the potential for a reusable assay format and mismatch discrimination. In this, the first incarnation of a solid-phase QD-FRET hybridization assay, the limit of detection was found to be 5 nM, and the dynamic range was almost 2 orders of magnitude. Selective discrimination of the target was shown using a three-base-pairs mismatch from a fully complementary sequence. Despite a gradual loss of signal, reuse of the optical fibers over multiple cycles of hybridization and dehybridization was possible. Directions for further improvement of the analytical performance by optimizing the design of the QD-probe oligonucleotide interface are identified.

  2. In-capillary probing of quantum dots and fluorescent protein self-assembly and displacement using Förster resonance energy transfer.

    Science.gov (United States)

    Wang, Jianhao; Fan, Jie; Li, Jinchen; Liu, Li; Wang, Jianpeng; Jiang, Pengju; Liu, Xiaoqian; Qiu, Lin

    2017-02-01

    Herein, a Förster resonance energy transfer system was designed, which consisted of CdSe/ZnS quantum dots donor and mCherry fluorescent protein acceptor. The quantum dots and the mCherry proteins were conjugated to permit Förster resonance energy transfer. Capillary electrophoresis with fluorescence detection was used for the analyses for the described system. The quantum dots and mCherry were sequentially injected into the capillary, while the real-time fluorescence signal of donor and acceptor was simultaneously monitored by two channels with fixed wavelength detectors. An effective separation of complexes from free donor and acceptor was achieved. Results showed quantum dots and hexahistidine tagged mCherry had high affinity and the assembly was affected by His6 -mCherry/quantum dot molar ratio. The kinetics of the self-assembly was calculated using the Hill equation. The microscopic dissociation constant values for out of- and in-capillary assays were 10.49 and 23.39 μM, respectively. The capillary electrophoresis with fluorescence detection that monitored ligands competition assay further delineated the different binding capacities of histidine containing peptide ligands for binding sites on quantum dots. This work demonstrated a novel approach for the improvement of Förster resonance energy transfer for higher efficiency, increased sensitivity, intuitionistic observation, and low sample requirements of the in-capillary probing system.

  3. Fluorescent magnetic iron oxide nanoparticles for cardiac precursor cell selection from stromal vascular fraction and optimization for magnetic resonance imaging

    Directory of Open Access Journals (Sweden)

    Verma VK

    2015-01-01

    Full Text Available Vinod Kumar Verma,1 Suguna Ratnakar Kamaraju,1 Ravindranath Kancherla,1 Lakshmi K Kona,1 Syed Sultan Beevi,1 Tanya Debnath,1 Shalini P Usha,1 Rammohan Vadapalli,2 Ali Syed Arbab,3 Lakshmi Kiran Chelluri11Department of Transplant Biology, Immunology and Stem Cell Laboratory, Global Hospitals, 2Department of Imageology, Vijaya Radiology Centre, Hyderabad, India; 3Department of Biochemistry and Molecular Biology, Georgia Regents University, Augusta, GA, USAAbstract: Fluorescent magnetic iron oxide nanoparticles have been used to label cells for imaging as well as for therapeutic purposes. The purpose of this study was to modify the approach to develop a nanoprobe for cell selection and imaging with a direct therapeutic translational focus. The approach involves physical coincubation and adsorption of superparamagnetic iron oxide nanoparticle-polyethylene glycol (SPION-PEG complexes with a monoclonal antibody (mAb or a set of antibodies. Flow cytometry, confocal laser scanning microscopy, transmission electron microscopy, iron staining, and magnetic resonance imaging were used to assess cell viability, function, and labeling efficiency. This process has been validated by selecting adipose tissue-derived cardiac progenitor cells from the stromal vascular fraction using signal regulatory protein alpha (SIRPA/kinase domain receptor (KDR mAbs. These markers were chosen because of their sustained expression during cardiomyocyte differentiation. Sorting of cells positive for SIRPA and KDR allowed the enrichment of cardiac progenitors with 90% troponin-I positivity in differentiation cultures. SPION labeled cardiac progenitor cells (1×105 cells was mixed with gel and used for 3T magnetic resonance imaging at a concentration, as low as 12.5 µg of iron. The toxicity assays, at cellular and molecular levels, did not show any detrimental effects of SPION. Our study has the potential to achieve moderate to high specific cell selection for the dual purpose of

  4. Characterization of the AT180 epitope of phosphorylated Tau protein by a combined nuclear magnetic resonance and fluorescence spectroscopy approach

    Energy Technology Data Exchange (ETDEWEB)

    Amniai, Laziza [CNRS-UMR 8576 UGSF-IFR 147, Universite des Sciences et Technologies de Lille 1, 59655 Villeneuve d' Ascq Cedex (France); Lippens, Guy, E-mail: guy.lippens@univ-lille1.fr [CNRS-UMR 8576 UGSF-IFR 147, Universite des Sciences et Technologies de Lille 1, 59655 Villeneuve d' Ascq Cedex (France); Landrieu, Isabelle, E-mail: isabelle.landrieu@univ-lille1.fr [CNRS-UMR 8576 UGSF-IFR 147, Universite des Sciences et Technologies de Lille 1, 59655 Villeneuve d' Ascq Cedex (France)

    2011-09-09

    Highlights: {yields} pThr231 of the Tau protein is necessary for the binding of the AT180 antibody. {yields} pSer235 of the Tau protein does not interfere with the AT180 recognition of pThr231. {yields} Epitope mapping is efficiently achieved by combining NMR and FRET spectroscopy. -- Abstract: We present here the characterization of the epitope recognized by the AT180 monoclonal antibody currently used to define an Alzheimer's disease (AD)-related pathological form of the phosphorylated Tau protein. Some ambiguity remains as to the exact phospho-residue(s) recognized by this monoclonal: pThr231 or both pThr231 and pSer235. To answer this question, we have used a combination of nuclear magnetic resonance (NMR) and fluorescence spectroscopy to characterize in a qualitative and quantitative manner the phospho-residue(s) essential for the epitope recognition. Data from the first step of NMR experiments are used to map the residues bound by the antibodies, which were found to be limited to a few residues. A fluorophore is then chemically attached to a cystein residue introduced close-by the mapped epitope, at arginine 221, by mutagenesis of the recombinant protein. The second step of Foerster resonance energy transfer (FRET) between the AT180 antibody tryptophanes and the phospho-Tau protein fluorophore allows to calculate a dissociation constant Kd of 30 nM. We show that the sole pThr231 is necessary for the AT180 recognition of phospho-Tau and that phosphorylation of Ser235 does not interfere with the binding.

  5. Study of endothelial cell apoptosis using fluorescence resonance energy transfer (FRET) biosensor cell line with hemodynamic microfluidic chip system.

    Science.gov (United States)

    Yu, J Q; Liu, X F; Chin, L K; Liu, A Q; Luo, K Q

    2013-07-21

    To better understand how hyperglycemia induces endothelial cell dysfunction under the diabetic conditions, a hemodynamic microfluidic chip system was developed. The system combines a caspase-3-based fluorescence resonance energy transfer (FRET) biosensor cell line which can detect endothelial cell apoptosis in real-time, post-treatment effect and with a limited cell sample, by using a microfluidic chip which can mimic the physiological pulsatile flow profile in the blood vessel. The caspase-3-based FRET biosensor endothelial cell line (HUVEC-C3) can produce a FRET-based sensor protein capable of probing caspase-3 activation. When the endothelial cells undergo apoptosis, the color of the sensor cells changes from green to blue, thus sensing apoptosis. A double-labeling fluorescent technique (yo pro-1 and propidium iodide) was used to validate the findings revealed by the FRET-based caspase sensor. The results show high rates of apoptosis and necrosis of endothelial cells when high glucose concentration was applied in our hemodynamic microfluidic chip combined with an exhaustive pulsatile flow profile. The two apoptosis detection techniques (fluorescent method and FRET biosensor) are comparable; but FRET biosensor offers more advantages such as real-time observation and a convenient operating process to generate more accurate and reliable data. Furthermore, the activation of the FRET biosensor also confirms the endothelial cell apoptosis induced by the abnormal pulsatile shear stress and high glucose concentration is through caspase-3 pathway. A 12% apoptotic rate (nearly a 4-fold increase compared to the static condition) was observed when the endothelial cells were exposed to a high glucose concentration of 20 mM under 2 h exhaustive pulsatile shear stress of 30 dyne cm(-2) and followed with another 10 h normal pulsatile shear stress of 15 dyne cm(-2). Therefore, the most important finding of this study is to develop a novel endothelial cell apoptosis detection

  6. Nucleic Acid Sandwich Hybridization Assay with Quantum Dot-Induced Fluorescence Resonance Energy Transfer for Pathogen Detection

    Directory of Open Access Journals (Sweden)

    Cheng-Chung Chou

    2012-12-01

    Full Text Available This paper reports a nucleic acid sandwich hybridization assay with a quantum dot (QD-induced fluorescence resonance energy transfer (FRET reporter system. Two label-free hemagglutinin H5 sequences (60-mer DNA and 630-nt cDNA fragment of avian influenza viruses were used as the targets in this work. Two oligonucleotides (16 mers and 18 mers that specifically recognize two separate but neighboring regions of the H5 sequences were served as the capturing and reporter probes, respectively. The capturing probe was conjugated to QD655 (donor in a molar ratio of 10:1 (probe-to-QD, and the reporter probe was labeled with Alexa Fluor 660 dye (acceptor during synthesis. The sandwich hybridization assay was done in a 20 μL transparent, adhesive frame-confined microchamber on a disposable, temperature-adjustable indium tin oxide (ITO glass slide. The FRET signal in response to the sandwich hybridization was monitored by a homemade optical sensor comprising a single 400 nm UV light-emitting diode (LED, optical fibers, and a miniature 16-bit spectrophotometer. The target with a concentration ranging from 0.5 nM to 1 μM was successfully correlated with both QD emission decrease at 653 nm and dye emission increase at 690 nm. To sum up, this work is beneficial for developing a portable QD-based nucleic acid sensor for on-site pathogen detection.

  7. Enzymatic activity characterization of SARS coronavirus 3C-like protease by fluorescence resonance energy transfer technique

    Institute of Scientific and Technical Information of China (English)

    Shuai CHEN; Hua-liang JIANG; Li-li CHEN; Hai-bin LUO; Tao SUN; Jing CHEN; Fei YE; Jian-hua CAI; Jing-kang SHEN; Xu SHEN

    2005-01-01

    Aim: To characterize enzymatic activity of severe acute respiratory syndrome(SARS) coronavirus (CoV) 3C-like protease (3CLpro) and its four site-directed mutants. Methods: Based on the fluorescence resonance energy transfer (FRET)principle using 5-[(2'-aminoethyl)-amino] naphthelenesulfonic acid (EDANS) and 4-[[4-(dimethylamino) phenyl] azo] benzoic acid (Dabcyl) as the energy transfer pair, one fluorogenic substrate was designed for the evaluation of SARS-CoV 3CLpro proteolytic activity. Results: The kinetic parameters of the fluorogenic substrate have been determined as Km=404 μmol.L-1, kcat=1.08 min-1, and kcat/Km=2.7 gered activity switches, and site-directed mutagenesis analysis of SARS-CoV 3CLpro revealed that substitutions of His41, Cys145, and His163 resulted in complete loss of enzymatic activity, while replacement of Met162 with Ala caused strongly increased activity. Conclusion: This present work has provided valuable information for understanding the catalytic mechanism of SARS-CoV 3CLpro. This FRET-based assay might supply an ideal approach for the exploration SARSCoV 3CLpro putative inhibitors.

  8. Fluorescence resonance energy transfer imaging reveals that chemokine-binding modulates heterodimers of CXCR4 and CCR5 receptors.

    Directory of Open Access Journals (Sweden)

    Nilgun Isik

    Full Text Available BACKGROUND: Dimerization has emerged as an important feature of chemokine G-protein-coupled receptors. CXCR4 and CCR5 regulate leukocyte chemotaxis and also serve as a co-receptor for HIV entry. Both receptors are recruited to the immunological synapse during T-cell activation. However, it is not clear whether they form heterodimers and whether ligand binding modulates the dimer formation. METHODOLOGY/PRINCIPAL FINDINGS: Using a sensitive Fluorescence Resonance Energy Transfer (FRET imaging method, we investigated the formation of CCR5 and CXCR4 heterodimers on the plasma membrane of live cells. We found that CCR5 and CXCR4 exist as constitutive heterodimers and ligands of CCR5 and CXCR4 promote different conformational changes within these preexisting heterodimers. Ligands of CCR5, in contrast to a ligand of CXCR4, induced a clear increase in FRET efficiency, indicating that selective ligands promote and stabilize a distinct conformation of the heterodimers. We also found that mutations at C-terminus of CCR5 reduced its ability to form heterodimers with CXCR4. In addition, ligands induce different conformational transitions of heterodimers of CXCR4 and CCR5 or CCR5(STA and CCR5(Delta4. CONCLUSIONS/SIGNIFICANCE: Taken together, our data suggest a model in which CXCR4 and CCR5 spontaneously form heterodimers and ligand-binding to CXCR4 or CCR5 causes different conformational changes affecting heterodimerization, indicating the complexity of regulation of dimerization/function of these chemokine receptors by ligand binding.

  9. Laser-induced fluorescence resonance energy transfer for analysis of the quality of a DNA double helix

    Science.gov (United States)

    Bregadze, V. G.; Melikishvili, Z. G.; Giorgadze, T. G.; Khutsishvili, I. G.; Khuskivadze, T. B.; Jaliashvili, Z. V.; Sigua, K. I.

    2016-11-01

    The goal of this work is to use the method of the laser-induced fluorescence resonance energy transfer (FRET) of electronic excitation in a donor-acceptor pair of intercalators, (acridine orange (AO) as a donor and ethidium bromide (EB) as an acceptor), for the quantitative analysis of the quality of a DNA double helix. This approach obtains a visual picture of the defects of the genetic apparatus of tissue cells, particularly those of skin cells in real time and it can be used for the diagnosis of skin diseases and also in cosmetology. Transition metal (TM) ions such as Cu(II), Cu(I), Ag(I), silver nanoparticles (AgNPs), photo- and thermo effects were used to cause double helix defects in DNA. The concentration of DNA sites after exposure to Cu(II), Cu(I), Ag(I) ions, AgNPs impact, as well as laser irradiation (λ  =  457 nm) and temperature, which are applicable for intercalation, were estimated in relative units. The nanoscale FRET method enables the estimation of the concentration of double helix areas with high stability, applicable for intercalation in DNA after it was subjected to stress effect. It provides the opportunity to compare DNA-s of (1) different origin; (2) with various degrees of damage; (3) being in various functional states.

  10. Development of a Fluorescence Resonance Energy Transfer (FRET-Based DNA Biosensor for Detection of Synthetic Oligonucleotide of Ganoderma boninense

    Directory of Open Access Journals (Sweden)

    Noremylia Mohd Bakhori

    2013-12-01

    Full Text Available An optical DNA biosensor based on fluorescence resonance energy transfer (FRET utilizing synthesized quantum dot (QD has been developed for the detection of specific-sequence of DNA for Ganoderma boninense, an oil palm pathogen. Modified QD that contained carboxylic groups was conjugated with a single-stranded DNA probe (ssDNA via amide-linkage. Hybridization of the target DNA with conjugated QD-ssDNA and reporter probe labeled with Cy5 allows for the detection of related synthetic DNA sequence of Ganoderma boninense gene based on FRET signals. Detection of FRET emission before and after hybridization was confirmed through the capability of the system to produce FRET at 680 nm for hybridized sandwich with complementary target DNA. No FRET emission was observed for non-complementary system. Hybridization time, temperature and effect of different concentration of target DNA were studied in order to optimize the developed system. The developed biosensor has shown high sensitivity with detection limit of 3.55 × 10−9 M. TEM results show that the particle size of QD varies in the range between 5 to 8 nm after ligand modification and conjugation with ssDNA. This approach is capable of providing a simple, rapid and sensitive method for detection of related synthetic DNA sequence of Ganoderma boninense.

  11. Dark-resonance Doppler cooling and high fluorescence in trapped Ca-43 ions at intermediate magnetic field

    CERN Document Server

    Allcock, D T C; Sepiol, M A; Janacek, H A; Ballance, C J; Steane, A M; Lucas, D M; Stacey, D N

    2015-01-01

    We demonstrate simple and robust methods for Doppler cooling and obtaining high fluorescence from trapped 43Ca+ ions at a magnetic field of 146 Gauss. This field gives access to a magnetic-field-independent "atomic clock" qubit transition within the ground level hyperfine structure of the ion, but also causes the complex internal structure of the 64 states relevant to Doppler cooling to be spread over many times the atomic transition line-width. Using a time-dependent optical Bloch equation simulation of the system we develop a simple scheme to Doppler-cool the ion on a two-photon dark resonance, which is robust to typical experimental variations in laser intensities, detunings and polarizations. We experimentally demonstrate cooling to a temperature of 0.3 mK, slightly below the Doppler limit for the corresponding two-level system, and then use Raman sideband laser cooling to cool further to the ground states of the ion's radial motional modes. These methods will enable two-qubit entangling gates with this i...

  12. Monitoring of bystander effect of herpes simplex virus thymidine kinase/acyclovir system using fluorescence resonance energy transfer technique.

    Science.gov (United States)

    Xiong, Tao; Li, Yongjun; Ni, Fenge; Zhang, Feng

    2012-02-01

    Cytotoxic gene therapy mediated by gene transfer of the herpes simplex virus thymidine kinase (HSV-tk) gene followed by acyclovir (ACV) treatment has been reported to inhibit malignant tumor growth in a variety of studies. The magnitude of "bystander effect" is an essential factor for this anti-tumor approach in vivo. However, the mechanism by which HSV-tk/ACV brings "bystander effect" is poorly understood. In this report, the plasmid CD3 (ECFP-CRS-DsRed) and TK-GFP were transferred to the human adenoid cystic carcinoma line ACC-M cell line. The CD3-expressing cells apoptosis was monitored using fluorescence resonance energy transfer (FRET) technique. First, CD3 and TK-GFP co-expressing ACC-M cells apoptosis was monitored using FRET technique. The apoptosis was induced by ACV and initiated by caspase3. The FRET efficient was remarkably decreased and then disappeared during cellular apoptosis, which indicated that the TK-GFP expressing ACC-M cells apoptosis, induced by ACV, was via a caspase3-dependent pathway. Secondly, CD3 and TK-GFP mixed expressing ACC-M cells apoptosis, induced by ACV, were monitored using FRET technique. The apoptotic phenomena appeared in the CD3-expressing ACC-M cells. The results show that HSV-tk/ACV system killed ACC-M cells using its bystander effect. These results confirm that HSV-tk/ACV system is potential for cancer gene therapy.

  13. Fluorescence resonance energy transfer (FRET) in chemistry and biology: Non-Förster distance dependence of the FRET rate

    Indian Academy of Sciences (India)

    Sangeeta Saini; Harjinder Singh; Biman Bagchi

    2006-01-01

    Fluorescence resonance energy transfer (FRET) is a popular tool to study equilibrium and dynamical properties of polymers and biopolymers in condensed phases and is now widely used in conjunction with single molecule spectroscopy. In the data analysis, one usually employs the Förster expression which predicts (1/6) distance dependence of the energy transfer rate. However, critical analysis shows that this expression can be of rather limited validity in many cases. We demonstrate this by explicitly considering a donor-acceptor system, polyfluorene (PF6)-tetraphenylporphyrin (TPP), where the size of both donor and acceptor is comparable to the distance separating them. In such cases, one may expect much weaker distance (as 1/2 or even weaker) dependence. We have also considered the case of energy transfer from a dye to a nanoparticle. Here we find 1/4 distance dependence at large separations, completely different from Förster. We also discuss recent application of FRET to study polymer conformational dynamics.

  14. A Quantitative Theoretical Framework For Protein-Induced Fluorescence Enhancement-Förster-Type Resonance Energy Transfer (PIFE-FRET).

    Science.gov (United States)

    Lerner, Eitan; Ploetz, Evelyn; Hohlbein, Johannes; Cordes, Thorben; Weiss, Shimon

    2016-07-01

    Single-molecule, protein-induced fluorescence enhancement (PIFE) serves as a molecular ruler at molecular distances inaccessible to other spectroscopic rulers such as Förster-type resonance energy transfer (FRET) or photoinduced electron transfer. In order to provide two simultaneous measurements of two distances on different molecular length scales for the analysis of macromolecular complexes, we and others recently combined measurements of PIFE and FRET (PIFE-FRET) on the single molecule level. PIFE relies on steric hindrance of the fluorophore Cy3, which is covalently attached to a biomolecule of interest, to rotate out of an excited-state trans isomer to the cis isomer through a 90° intermediate. In this work, we provide a theoretical framework that accounts for relevant photophysical and kinetic parameters of PIFE-FRET, show how this framework allows the extraction of the fold-decrease in isomerization mobility from experimental data, and show how these results provide information on changes in the accessible volume of Cy3. The utility of this model is then demonstrated for experimental results on PIFE-FRET measurement of different protein-DNA interactions. The proposed model and extracted parameters could serve as a benchmark to allow quantitative comparison of PIFE effects in different biological systems.

  15. Interfacial Chemistry and the Design of Solid-Phase Nucleic Acid Hybridization Assays Using Immobilized Quantum Dots as Donors in Fluorescence Resonance Energy Transfer

    OpenAIRE

    Krull, Ulrich J.; W. Russ Algar

    2011-01-01

    The use of quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) offer several advantages for the development of multiplexed solid-phase QD-FRET nucleic acid hybridization assays. Designs for multiplexing have been demonstrated, but important challenges remain in the optimization of these systems. In this work, we identify several strategies based on the design of interfacial chemistry for improving sensitivity, obtaining lower limits of detection (LOD) and enabling th...

  16. Targeting human c-Myc promoter duplex DNA with actinomycin D by use of multi-way analysis of quantum-dot-mediated fluorescence resonance energy transfer

    DEFF Research Database (Denmark)

    Gholami, Somayeh; Kompany Zare, Mohsen

    2013-01-01

    pairs resulted in efficient energy transfer from drug to QD via fluorescence resonance energy transfer (FRET). Multi-way analysis of the three-way data array obtained from titration experiments was performed by use of restricted Tucker3 and hard trilinear decomposition (HTD). These techniques enable...... of multi-way chemometric methods to investigation of nano-biotechnological systems where several overlapped species coexist in solution....

  17. Photonic Crystal Surfaces as a General Purpose Platform for Label-Free and Fluorescent Assays

    OpenAIRE

    Cunningham, Brian T.

    2010-01-01

    Photonic crystal surfaces can be designed to provide a wide range of functions that are used to perform biochemical and cell-based assays. Detection of the optical resonant reflections from photonic crystal surfaces enables high sensitivity label-free biosensing, while the enhanced electromagnetic fields that occur at resonant wavelengths can be used to enhance the detection sensitivity of any surface-based fluorescence assay. Fabrication of photonic crystals from inexpensive plastic material...

  18. Determination of trace uranium by resonance fluorescence method coupled with photo-catalytic technology and dual cloud point extraction.

    Science.gov (United States)

    Li, Jiekang; Li, Guirong; Han, Qian

    2016-12-01

    In this paper, two kinds of salophens (Sal) with different solubilities, Sal1 and Sal2, have been respectively synthesized, and they all can combine with uranyl to form stable complexes: [UO2(2+)-Sal1] and [UO2(2+)-Sal2]. Among them, [UO2(2+)-Sal1] was used as ligand to extract uranium in complex samples by dual cloud point extraction (dCPE), and [UO2(2+)-Sal2] was used as catalyst for the determination of uranium by photocatalytic resonance fluorescence (RF) method. The photocatalytic characteristic of [UO2(2+)-Sal2] on the oxidized pyronine Y (PRY) by potassium bromate which leads to the decrease of RF intensity of PRY were studied. The reduced value of RF intensity of reaction system (ΔF) is in proportional to the concentration of uranium (c), and a novel photo-catalytic RF method was developed for the determination of trace uranium (VI) after dCPE. The combination of photo-catalytic RF techniques and dCPE procedure endows the presented methods with enhanced sensitivity and selectivity. Under optimal conditions, the linear calibration curves range for 0.067 to 6.57ngmL(-1), the linear regression equation was ΔF=438.0 c (ngmL(-1))+175.6 with the correlation coefficient r=0.9981. The limit of detection was 0.066ngmL(-1). The proposed method was successfully applied for the separation and determination of uranium in real samples with the recoveries of 95.0-103.5%. The mechanisms of the indicator reaction and dCPE are discussed.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-08-31

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

  20. Determination of trace uranium by resonance fluorescence method coupled with photo-catalytic technology and dual cloud point extraction

    Science.gov (United States)

    Li, Jiekang; Li, Guirong; Han, Qian

    2016-12-01

    In this paper, two kinds of salophens (Sal) with different solubilities, Sal1 and Sal2, have been respectively synthesized, and they all can combine with uranyl to form stable complexes: [UO22 +-Sal1] and [UO22 +-Sal2]. Among them, [UO22 +-Sal1] was used as ligand to extract uranium in complex samples by dual cloud point extraction (dCPE), and [UO22 +-Sal2] was used as catalyst for the determination of uranium by photocatalytic resonance fluorescence (RF) method. The photocatalytic characteristic of [UO22 +-Sal2] on the oxidized pyronine Y (PRY) by potassium bromate which leads to the decrease of RF intensity of PRY were studied. The reduced value of RF intensity of reaction system (ΔF) is in proportional to the concentration of uranium (c), and a novel photo-catalytic RF method was developed for the determination of trace uranium (VI) after dCPE. The combination of photo-catalytic RF techniques and dCPE procedure endows the presented methods with enhanced sensitivity and selectivity. Under optimal conditions, the linear calibration curves range for 0.067 to 6.57 ng mL- 1, the linear regression equation was ΔF = 438.0 c (ng mL- 1) + 175.6 with the correlation coefficient r = 0.9981. The limit of detection was 0.066 ng mL- 1. The proposed method was successfully applied for the separation and determination of uranium in real samples with the recoveries of 95.0-103.5%. The mechanisms of the indicator reaction and dCPE are discussed.

  1. Rocket observation of atomic oxygen and night airglow: Measurement of concentration with an improved resonance fluorescence technique

    Directory of Open Access Journals (Sweden)

    K. Kita

    Full Text Available An improved resonant fluorescence instrument for measuring atomic oxygen concentration was developed to avoid the Doppler effect and the aerodynamic shock effect due to the supersonic motion of a rocket. The shock effect is reduced by adopting a sharp wedge-shaped housing and by scanning of the detector field of view to change the distance between the scattering volume and the surface of the housing. The scanning enables us to determine absolute values of atomic oxygen concentration from relative variation of the scattered light signal due to the self-absorption. The instrument was calibrated in the laboratory, and the numerical simulation reproduced the calibration result. Using the instrument, the altitude profile of atomic oxygen concentration was observed by a rocket experiment at Uchinoura (31°N on 28 January 1992. The data obtained from the rocket experiment were not perfectly free from the shock effect, but errors due to the effect were reduced by the data analysis procedure. The observed maximum concentration was 3.8× 1011 cm–3 at altitudes around 94 km. The systematic error is estimated to be less than ±0.7×1011 cm–3 and the relative random error is less than±0.07× 1011 cm–3at the same altitudes. The altitude profile of the OI 557.7-nm airglow was also observed in the same rocket experiment. The maximum volume emission rate was found to be 150 photons cm–3 s–1 at 94 km. The observed altitude profiles are compared with the MSIS model and other in situ observations.

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

    Science.gov (United States)

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

    2015-01-14

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

  3. A carbon dots-CdTe quantum dots fluorescence resonance energy transfer system for the analysis of ultra-trace chlortoluron in water.

    Science.gov (United States)

    Tao, Huilin; Liao, Xiufen; Sun, Chao; Xie, Xiangli; Zhong, Fuxin; Yi, Zhongsheng; Huang, Yipeng

    2015-02-05

    In this paper, a fluorescence resonance energy transfer (FRET) system between fluorescence carbon dots (CDs, donor) and CdTe quantum dots (CdTe, acceptor) was constructed, and a novel platform for sensitive and selective determination of chlortoluron was accordingly proposed. It was found that in Tris-HCl buffer solution at pH=8.7, energy transfer from CDs to CdTe occurred, which resulted in a great enhancement of the fluorescence intensity of CdTe. Upon the addition of chlortoluron, in terms of strong interaction between chlortoluron and CdTe QDs through the formation of chlortoluron-CdTe ground state complex, resulted in CdTe fluorescence quenching. Under optimal conditions, in range of 2.4×10(-10)molL(-1)-8.5×10(-8)molL(-1), the change of CdTe fluorescence intensity was in good linear relationship with the chlortoluron concentration, and the detection limit was 7.8×10(-11)molL(-1) (S/N=3). Most of common relevant substance, cations and anions did not interfere with the detection of chlortoluron. The proposed method was applied to determine chlortoluron in water samples with satisfactory results.

  4. Construction of fluorescence resonance energy transfer vectors and their application in study of structure and function of signal transducers and activators of transcription 1

    Institute of Scientific and Technical Information of China (English)

    Fujun Han; Yongfeng Luo; Nanhai Ge; Jun Xu

    2008-01-01

    Protein-protein interactions have been studied extensively by green fluorescent protein-based fluorescence resonance energy transfer (FRET). The fluorescent proteins (FP) can be fused either to the N- or C-terminus of a host protein, but it is difficult to predict which order will perturb the host protein the least and provide the largest FRET. Therefore, a researcher needs to fuse host proteins with FP at both the N- and C-termini and test every possible combination (N-N,N-C, or C-C) to promote the energy transfer efficiency.Consequently, researchers required to do many subelonings.Herein, we designed FRET vectors to make them more efficient. The expression vectors ofpCTP.YFP and pYFP-CFP were constructed with both cyan fluorescent protein (CFP)-yellow fluorescent protein (YFP) and YFP-CFP coding sequences flanked by two restriction enzyme sites, and with multiple cloning regions in the middle of both coding sequences. To select an optimal combination for FRET detection, we created plasmids encoding various fusion proteins of FP and signal transducers and activators of transcription 1 (STAT1). We found that the nuclear:cytoplasmic fluorescence intensity ratios of STAT1 -FP were significantly higher than those of FP-STAT1 at steady state,and fluorescence redistribution was only observed for STAT1-FP upon interferon gamma (IFNΥ) stimulation. In addition, positive FRET signals were only detected in the C-C interactions of STAT 1 homodimer. Taken together, these data indicate that fusing STATI at the N.terminus with Fpimpairs the interactions ofunphospborylated STAT1 homodimers and possibly diminishes its binding with DNA. In contrast, STATIFP was functional with respect to its activation. Moreover, the FRET vectors are able to facilitate FRET studies.

  5. In vivo magnetic resonance and fluorescence dual imaging of tumor sites by using dye-doped silica-coated iron oxide nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Haeyun; Lee, Chaedong [Seoul National University, Program in Nano Science and Technology, Graduate School of Convergence Science and Technology (Korea, Republic of); Nam, Gi-Eun [University of Ulsan College of Medicine, Department of Radiology, Asan Medical Center (Korea, Republic of); Quan, Bo [Seoul National University, Program in Nano Science and Technology, Graduate School of Convergence Science and Technology (Korea, Republic of); Choi, Hyuck Jae [University of Ulsan College of Medicine, Department of Radiology, Asan Medical Center (Korea, Republic of); Yoo, Jung Sun [Seoul National University, Department of Transdisciplinary Studies, Graduate School of Convergence Science and Technology, Smart Humanity Convergence Center (Korea, Republic of); Piao, Yuanzhe, E-mail: parkat9@snu.ac.kr [Seoul National University, Program in Nano Science and Technology, Graduate School of Convergence Science and Technology (Korea, Republic of)

    2016-02-15

    The difficulty in delineating tumor is a major obstacle for better outcomes in cancer treatment of patients. The use of single-imaging modality is often limited by inadequate sensitivity and resolution. Here, we present the synthesis and the use of monodisperse iron oxide nanoparticles coated with fluorescent silica nano-shells for fluorescence and magnetic resonance dual imaging of tumor. The as-synthesized core–shell nanoparticles were designed to improve the accuracy of diagnosis via simultaneous tumor imaging with dual imaging modalities by a single injection of contrast agent. The iron oxide nanocrystals (∼11 nm) were coated with Rhodamine B isothiocyanate-doped silica shells via reverse microemulsion method. Then, the core–shell nanoparticles (∼54 nm) were analyzed to confirm their size distribution by transmission electron microscopy and dynamic laser scattering. Photoluminescence spectroscopy was used to characterize the fluorescent property of the dye-doped silica shell-coated nanoparticles. The cellular compatibility of the as-prepared nanoparticles was confirmed by a trypan blue dye exclusion assay and the potential as a dual-imaging contrast agent was verified by in vivo fluorescence and magnetic resonance imaging. The experimental results show that the uniform-sized core–shell nanoparticles are highly water dispersible and the cellular toxicity of the nanoparticles is negligible. In vivo fluorescence imaging demonstrates the capability of the developed nanoparticles to selectively target tumors by the enhanced permeability and retention effects and ex vivo tissue analysis was corroborated this. Through in vitro phantom test, the core/shell nanoparticles showed a T2 relaxation time comparable to Feridex{sup ®} with smaller size, indicating that the as-made nanoparticles are suitable for imaging tumor. This new dual-modality-nanoparticle approach has promised for enabling more accurate tumor imaging.

  6. In vivo magnetic resonance and fluorescence dual imaging of tumor sites by using dye-doped silica-coated iron oxide nanoparticles

    Science.gov (United States)

    Jang, Haeyun; Lee, Chaedong; Nam, Gi-Eun; Quan, Bo; Choi, Hyuck Jae; Yoo, Jung Sun; Piao, Yuanzhe

    2016-02-01

    The difficulty in delineating tumor is a major obstacle for better outcomes in cancer treatment of patients. The use of single-imaging modality is often limited by inadequate sensitivity and resolution. Here, we present the synthesis and the use of monodisperse iron oxide nanoparticles coated with fluorescent silica nano-shells for fluorescence and magnetic resonance dual imaging of tumor. The as-synthesized core-shell nanoparticles were designed to improve the accuracy of diagnosis via simultaneous tumor imaging with dual imaging modalities by a single injection of contrast agent. The iron oxide nanocrystals ( 11 nm) were coated with Rhodamine B isothiocyanate-doped silica shells via reverse microemulsion method. Then, the core-shell nanoparticles ( 54 nm) were analyzed to confirm their size distribution by transmission electron microscopy and dynamic laser scattering. Photoluminescence spectroscopy was used to characterize the fluorescent property of the dye-doped silica shell-coated nanoparticles. The cellular compatibility of the as-prepared nanoparticles was confirmed by a trypan blue dye exclusion assay and the potential as a dual-imaging contrast agent was verified by in vivo fluorescence and magnetic resonance imaging. The experimental results show that the uniform-sized core-shell nanoparticles are highly water dispersible and the cellular toxicity of the nanoparticles is negligible. In vivo fluorescence imaging demonstrates the capability of the developed nanoparticles to selectively target tumors by the enhanced permeability and retention effects and ex vivo tissue analysis was corroborated this. Through in vitro phantom test, the core/shell nanoparticles showed a T2 relaxation time comparable to Feridex® with smaller size, indicating that the as-made nanoparticles are suitable for imaging tumor. This new dual-modality-nanoparticle approach has promised for enabling more accurate tumor imaging.

  7. In vitro observation of the molecular interaction between NodD and its inducer naringenin as monitored by fluorescence resonance energy transfer

    Institute of Scientific and Technical Information of China (English)

    Fengqing Li; Bihe Hou; Lei Chen; Zhujun Yao; Guofan Hong

    2008-01-01

    At initial stages in the Rhizobium legume symbiosis, most nodulation genes are controlled by NodD protein and plant inducers. Some genetic studies and other reports have suggested that NodD may be activated by its direct interaction with plant inducers. However, there has been no molecular evidence of such an inducing interaction. In this paper, we used fluorescence resonance energy transfer technique to see whether such an interaction exists between NodD and its activator, naringenin, in vitro. The tetracysteine motif (Cys-Cys-Pro-Gly-Cys-Cys) was genetically inserted into NodD to label NodD with 4′,5′-bis(1,3,2-dithioarsolan-2-yl) fluorescein (FlAsH). Naringenin was labeled with fluorescein by chemical linking. In the fluorescence resonance energy transfer experiments in vitro, the fluorescence intensity of one acceptor, NodD(90R6)-FlAsH, increased by 13%. This suggests that NodD may directly interact with inducer naringenin in vitro and that the reaction centre is likely near hinge region 1 of NodD.

  8. Gold Nanoparticle-Quantum Dot Fluorescent Nanohybrid: Application for Localized Surface Plasmon Resonance-induced Molecular Beacon Ultrasensitive DNA Detection

    Science.gov (United States)

    Adegoke, Oluwasesan; Park, Enoch Y.

    2016-11-01

    In biosensor design, localized surface plasmon resonance (LSPR)-induced signal from gold nanoparticle (AuNP)-conjugated reporter can produce highly sensitive nanohybrid systems. In order to retain the physicochemical properties of AuNPs upon conjugation, high colloidal stability in aqueous solution is needed. In this work, the colloidal stability with respect to the zeta potential (ZP) of four negatively charged thiol-functionalized AuNPs, thioglycolic (TGA)-AuNPs, 3-mercaptopropionic acid (MPA)-AuNPs, l-cysteine-AuNPs and l-glutathione (GSH)-AuNPs, and a cationic cyteamine-capped AuNPs was studied at various pHs, ionic strength, and NP concentration. A strong dependence of the ZP charge on the nanoparticle (NP) concentration was observed. High colloidal stability was exhibited between pH 3 and 9 for the negatively charged AuNPs and between pH 3 and 7 for the cationic AuNPs. With respect to the ionic strength, high colloidal stability was exhibited at ≤104 μM for TGA-AuNPs, l-cysteine-AuNPs, and GSH-AuNPs, whereas ≤103 μM is recommended for MPA-AuNPs. For the cationic AuNPs, very low ionic strength of ≤10 μM is recommended due to deprotonation at higher concentration. GSH-AuNPs were thereafter bonded to SiO2-functionalized alloyed CdZnSeS/ZnSe1.0S1.3 quantum dots (SiO2-Qdots) to form a plasmon-enhanced AuNP-SiO2-Qdots fluorescent nanohybrid. The AuNP-SiO2-Qdots conjugate was afterward conjugated to a molecular beacon (MB), thus forming an ultrasensitive LSPR-induced SiO2-Qdots-MB biosensor probe that detected a perfect nucleotide DNA sequence at a concentration as low as 10 fg/mL. The limit of detection was 11 fg/mL (1.4 fM) while the biosensor probe efficiently distinguished between single-base mismatch and noncomplementary sequence target.

  9. Intercomparison of peroxy radical measurements obtained at atmospheric conditions by laser-induced fluorescence and electron spin resonance spectroscopy

    Directory of Open Access Journals (Sweden)

    A. Hofzumahaus

    2009-03-01

    Full Text Available Measurements of hydroperoxy radical (HO2 and organic peroxy radical (RO2 concentrations were performed by two different techniques in the atmospheric simulation chamber SAPHIR in Jülich, Germany. The first technique was the well-established Matrix Isolation Electron Spin Resonance (MIESR, which provides absolute measurements with a time resolution of 30 min and high accuracy (10%, 2 σ. The other technique, ROxLIF, has been newly developed. It is based on the selective chemical conversion of ROx radicals (HO2 and RO2 to OH, which is detected with high sensitivity by laser-induced fluorescence (LIF. ROxLIF is calibrated by quantitative photolysis of water vapor at 185 nm and provides ambient measurements at a temporal resolution of 1 min and accuracy of 20% (2 σ. The measurements of HO2 and RO2 obtained by the two techniques were compared for two types of atmospheric simulation experiments. In one experiment, HO2 and CH3O2 radicals were produced by photooxidation of methane in air at tropospheric conditions. In the second experiment, HO2 and C2H5O2 were produced by ozonolysis of 1-butene in air at dark conditions. The radical concentrations were within the range of 16 to 100 pptv for HO2 and 12 to 45 pptv for RO2. Good agreement was found in the comparison of the ROxLIF and MIESR measurements within their combined experimental uncertainties. Linear regressions to the combined data set yield slopes of 1.02±0.13 (1 σ for RO2 and 0.98±0.08 (1 σ for HO2 without significant offsets. The results confirm the calibration of the ROxLIF instrument and demonstrate that it can be applied with good accuracy for measurements of atmospheric peroxy radical concentrations.

  10. A highly sensitive fluorescence resonance energy transfer aptasensor for staphylococcal enterotoxin B detection based on exonuclease-catalyzed target recycling strategy

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Shijia; Duan, Nuo; Ma, Xiaoyuan; Xia, Yu; Wang, Hongxin; Wang, Zhouping, E-mail: wangzp@jiangnan.edu.cn

    2013-06-11

    Graphical abstract: -- Highlights: •An ultrasensitive FRET aptasensor was developed for staphylococcal enterotoxin B determination. •SEB was recognized by SEB aptamer with high affinity and specificity. •The Mn{sup 2+} doped NaYF{sub 4}:Yb/Er UCNPs used as donor to quencher dye (BHQ{sub 3}) in new FRET. •The fluorescence intensity was prominently amplified using an exonuclease-catalyzed target recycling strategy. -- Abstract: An ultrasensitive fluorescence resonance energy transfer (FRET) bioassay was developed to detect staphylococcal enterotoxin B (SEB), a low molecular exotoxin, using an aptamer-affinity method coupled with upconversion nanoparticles (UCNPs)-sensing, and the fluorescence intensity was prominently enhanced using an exonuclease-catalyzed target recycling strategy. To construct this aptasensor, both fluorescence donor probes (complementary DNA{sub 1}–UCNPs) and fluorescence quencher probes (complementary DNA{sub 2}–Black Hole Quencher{sub 3} (BHQ{sub 3})) were hybridized to an SEB aptamer, and double-strand oligonucleotides were fabricated, which quenched the fluorescence of the UCNPs via FRET. The formation of an aptamer–SEB complex in the presence of the SEB analyte resulted in not only the dissociation of aptamer from the double-strand DNA but also both the disruption of the FRET system and the restoration of the UCNPs fluorescence. In addition, the SEB was liberated from the aptamer–SEB complex using exonuclease I, an exonuclease specific to single-stranded DNA, for analyte recycling by selectively digesting a particular DNA (SEB aptamer). Based on this exonuclease-catalyzed target recycling strategy, an amplified fluorescence intensity could be produced using different SEB concentrations. Using optimized experimental conditions produced an ultrasensitive aptasensor for the detection of SEB, with a wide linear range of 0.001–1 ng mL{sup −1} and a lower detection limit (LOD) of 0.3 pg mL{sup −1} SEB (at 3σ). The fabricated

  11. Absorption,fluorescence and resonance Rayleigh scattering spectra of hydrophobic hydrogen bonding of eosin Y/Triton X-100 nanoparticles and their analytical applications

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    In a weak acidic medium(pH 2.4-2.8),eosin Y molecules(H2L) could replace water molecules to associate with Triton X-100 to form hydrophobic hydrogen bonding complexes.These complexes could further aggregate to form nanoparticles through the squeezing action of the water phase and Van Der Waals force,resulting in changes in the absorption spectrum and fluorescence quenching of EY as well as the significant enhancement of resonance Rayleigh scattering.This enables the sensitive determination of Triton X-100 using the fading spectrophotometry,fluorescence quenching method and RRS method.Among them,the RRS method shows the highest sensitivity with a detection limit of 20.6 ng mL-1 for Triton X-100.The optimum experimental conditions and factors that affect the absorption,fluorescence and RRS spectra were tested.The effects of coexisting substances were investigated and the results showed good selectivity.Based on these results,new spectrophotometric methods,fluorescence quenching method and RRS method for the determination of Triton X-100,were established.The hydrogen bonding association of eosin Y with Triton X-100 and the formation of nanoparticles as well as their effects on related spectral characteristics were discussed utilizing infrared,transmission electron microscope technique and quantum chemical method.

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

    Science.gov (United States)

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

    2015-07-01

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

  13. State transition analysis of spontaneous branch migration of the Holliday junction by photon-based single-molecule fluorescence resonance energy transfer.

    Science.gov (United States)

    Okamoto, Kenji; Sako, Yasushi

    2016-02-01

    Branch migration of Holliday junction (HJ) DNA in solution is a spontaneous conformational change between multiple discrete states. We applied single-molecule fluorescence resonance energy transfer (smFRET) measurement to three-state branch migration. The photon-based variational Bayes-hidden Markov model (VB-HMM) method was applied to fluorescence signals to reproduce the state transition trajectories and evaluate the transition parameters, such as transition rate. The upper limit of time resolution suggested in simulation was nearly achieved for the state dynamics with relatively small FRET changes, and the distinctions in the populations of different states were successfully retrieved. We also discuss the suitability of the HJ as a standard sample for smFRET dynamics measurements and data analysis.

  14. Vapor cell based sodium laser guide star mechanism study lab-bench

    Science.gov (United States)

    Wang, Hongyan; Li, Lihang; Luo, Ruiyao; Li, Lei; Ning, Yu; Xi, Fengjie; Xu, Xiaojun

    2016-07-01

    Sodium laser guide star (LGS) is the key for the success of modern adaptive optics (AO) supported large ground based telescopes, however, for many field applications, Sodium LGS's brightness is still a limited factor. Large amounts of theoretical efforts have been paid to optimize Sodium LGS exciting parameters, that is, to fully discover potential of harsh environment surrounding mesospheric extreme thin sodium atoms under resonant excitation, whether quantum or Monte Carlo based. But till to now, only limited proposals are demonstrated with on-sky test due to the high cost and engineering complexities. To bridge the gap between theoretical modeling and on-sky test, we built a magnetic field controllable sodium cell based lab-bench, which includes a small scale sum-frequency single mode 589nm laser, with added amplitude, polarization, and phase modulators. We could perform quantitative resonant fluorescence study under single, multi-frequency, side-band optical re-pumping exciting with different polarization, also we could perform optical field modulation to study Larmor precession which is considered as one of devils of Sodium LGS, and we have the ability to generate beams contain orbital angular moment. Our preliminary sodium cell based optical re-pumping experiments have shown excellent consistence with Bloch equation predicted results, other experimental results will also be presented in the report, and these results will give a direct support that sodium cell based lab-bench study could help a Sodium LGS scientists a lot before their on-sky test.

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

    Science.gov (United States)

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

    2010-12-01

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

  16. Cell-based bioassays in microfluidic systems

    Science.gov (United States)

    Itle, Laura J.; Zguris, Jeanna C.; Pishko, Michael V.

    2004-12-01

    The development of cell-based bioassays for high throughput drug screening or the sensing of biotoxins is contingent on the development of whole cell sensors for specific changes in intracellular conditions and the integration of those systems into sample delivery devices. Here we show the feasibility of using a 5-(and-6)-carboxy SNARF-1, acetoxymethyl ester, acetate, a fluorescent dye capable of responding to changes in intracellular pH, as a detection method for the bacterial endotoxin, lipopolysaccharide. We used photolithography to entrap cells with this dye within poly(ethylene) glyocol diacrylate hydrogels in microfluidic channels. After 18 hours of exposure to lipopolysaccharide, we were able to see visible changes in the fluorescent pattern. This work shows the feasibility of using whole cell based biosensors within microfluidic networks to detect cellular changes in response to exogenous agents.

  17. Super-resolution Localization and Defocused Fluorescence Microscopy on Resonantly Coupled Single-Molecule, Single-Nanorod Hybrids.

    Science.gov (United States)

    Su, Liang; Yuan, Haifeng; Lu, Gang; Rocha, Susana; Orrit, Michel; Hofkens, Johan; Uji-i, Hiroshi

    2016-02-23

    Optical antennas made of metallic nanostructures dramatically enhance single-molecule fluorescence to boost the detection sensitivity. Moreover, emission properties detected at the optical far field are dictated by the antenna. Here we study the emission from molecule-antenna hybrids by means of super-resolution localization and defocused imaging. Whereas gold nanorods make single-crystal violet molecules in the tip's vicinity visible in fluorescence, super-resolution localization on the enhanced molecular fluorescence reveals geometrical centers of the nanorod antenna instead. Furthermore, emission angular distributions of dyes linked to the nanorod surface resemble that of nanorods in defocused imaging. The experimental observations are consistent with numerical calculations using the finite-difference time-domain method.

  18. Determination of the cationic amphiphilic drug-DNA binding mode and DNA-assisted fluorescence resonance energy transfer amplification.

    Science.gov (United States)

    Yaseen, Zahid; Banday, Abdul Rouf; Hussain, Mohammed Aamir; Tabish, Mohammad; Kabir-ud-Din

    2014-03-25

    Understanding the mechanism of drug-DNA binding is crucial for predicting the potential genotoxicity of drugs. Agarose gel electrophoresis, absorption, steady state fluorescence, and circular dichroism have been used in exploring the interaction of cationic amphiphilic drugs (CADs) such as amitriptyline hydrochloride (AMT), imipramine hydrochloride (IMP), and promethazine hydrochloride (PMT) with calf thymus or pUC19 DNA. Agarose gel electrophoresis assay, along with absorption and steady state fluorescence studies, reveal interaction between the CADs and DNA. A comparative study of the drugs with respect to the effect of urea, iodide induced quenching, and ethidium bromide (EB) exclusion assay reflects binding of CADs to the DNA primarily in an intercalative fashion. Circular dichroism data also support the intercalative mode of binding. Besides quenching, there is fluorescence exchange energy transfer (FRET) in between CADs and EB using DNA as a template.

  19. Low-energy d-d excitations in MnO studied by resonant x-ray fluorescence spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Butorin, S.M.; Guo, J.; Magnuson, M. [Uppsala Univ. (Sweden)] [and others

    1997-04-01

    Resonant soft X-ray emission spectroscopy has been demonstrated to possess interesting abilities for studies of electronic structure in various systems, such as symmetry probing, alignment and polarization dependence, sensitivity to channel interference, etc. In the present abstract the authors focus on the feasibility of resonant soft X-ray emission to probe low energy excitations by means of resonant electronic X-ray Raman scattering. Resonant X-ray emission can be regarded as an inelastic scattering process where a system in the ground state is transferred to a low excited state via a virtual core excitation. The energy closeness to a core excitation of the exciting radiation enhances the (generally) low probability for inelastic scattering at these wavelengths. Therefore soft X-ray emission spectroscopy (in resonant electronic Raman mode) can be used to study low energy d-d excitations in transition metal systems. The involvement of the intermediate core state allows one to use the selection rules of X-ray emission, and the appearance of the elastically scattered line in the spectra provides the reference to the ground state.

  20. Observation of resonance fluorescence and the Mollow triplet from a coherently driven site-controlled quantum dot

    DEFF Research Database (Denmark)

    Unsleber, Sebastian; Maier, Sebastian; McCutcheon, Dara;

    2015-01-01

    Resonant excitation of solid state quantum emitters has the potential to deterministically excite a localized exciton while ensuring a maximally coherent emission. In this work, we demonstrate the coherent coupling of an exciton localized in a lithographically positioned, site-controlled semicond...

  1. pH-Responsive Tumor-Targetable Theranostic Nanovectors Based on Core Crosslinked (CCL Micelles with Fluorescence and Magnetic Resonance (MR Dual Imaging Modalities and Drug Delivery Performance

    Directory of Open Access Journals (Sweden)

    Sidan Tian

    2016-06-01

    Full Text Available The development of novel theranostic nanovectors is of particular interest in treating formidable diseases (e.g., cancers. Herein, we report a new tumor-targetable theranostic agent based on core crosslinked (CCL micelles, possessing tumor targetable moieties and fluorescence and magnetic resonance (MR dual imaging modalities. An azide-terminated diblock copolymer, N3-POEGMA-b-P(DPA-co-GMA, was synthesized via consecutive atom transfer radical polymerization (ATRP, where OEGMA, DPA, and GMA are oligo(ethylene glycolmethyl ether methacrylate, 2-(diisopropylaminoethyl methacrylate, and glycidyl methacrylate, respectively. The resulting diblock copolymer was further functionalized with DOTA(Gd (DOTA is 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakisacetic acid or benzaldehyde moieties via copper(I-catalyzed alkyne-azide cycloaddition (CuAAC chemistry, resulting in the formation of DOTA(Gd-POEGMA-b-P(DPA-co-GMA and benzaldehyde-POEGMA-b-P(DPA-co-GMA copolymers. The resultant block copolymers co-assembled into mixed micelles at neutral pH in the presence of tetrakis[4-(2-mercaptoethoxyphenyl]ethylene (TPE-4SH, which underwent spontaneous crosslinking reactions with GMA residues embedded within the micellar cores, simultaneously switching on TPE fluorescence due to the restriction of intramolecular rotation. Moreover, camptothecin (CPT was encapsulated into the crosslinked cores at neutral pH, and tumor-targeting pH low insertion peptide (pHLIP, sequence: AEQNPIYWARYADWLFTTPLLLLDLALLVDADEGTCG moieties were attached to the coronas through the Schiff base chemistry, yielding a theranostic nanovector with fluorescence and MR dual imaging modalities and tumor-targeting capability. The nanovectors can be efficiently taken up by A549 cells, as monitored by TPE fluorescence. After internalization, intracellular acidic pH triggered the release of loaded CPT, killing cancer cells in a selective manner. On the other hand, the nanovectors labeled with DOTA

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

    Science.gov (United States)

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

    2016-09-15

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

  3. Synthesis, characterization, and photophysical properties of Bodipy-spirooxazine and -spiropyran conjugates: modulation of fluorescence resonance energy transfer behavior via acidochromic and photochromic switching.

    Science.gov (United States)

    Kong, Lingcan; Wong, Hok-Lai; Tam, Anthony Yiu-Yan; Lam, Wai Han; Wu, Lixin; Yam, Vivian Wing-Wah

    2014-02-12

    Two series of Bodipy-containing photochromic spirooxazine and spiropyran derivatives have been designed, synthesized and characterized by (1)H NMR, ESI mass spectrometry and elemental analysis. Their electrochemical and photochromic properties were investigated. The photophysical, ultrafast transient absorption, and fluorescence resonance energy transfer (FRET) properties from Bodipy (donor) to the ring-opened merocyanine (acceptor) were also studied. Upon photoexcitation, all the photochromic spirooxazine- and spiropyran-containing compounds exhibited reversible photochromism. Computational studies have been performed to provide further insights into the nature of the electronic transitions for the two classes of compounds. The rate constants and activation parameters for thermal bleaching reactions of compounds SO, SP-alkyne, 1-3, and 8-10 were determined through kinetic studies in acetonitrile. The thermal bleaching reaction rate of the spiropyran-containing compounds is found to be much slower than that of the spirooxazine-containing counterparts.

  4. 碳点的制备及其荧光共振能量转移%Preparation and Fluorescence Resonance Energy Transfer of Carbon Dots

    Institute of Scientific and Technical Information of China (English)

    张煌博; 曹学功; 孙向英

    2014-01-01

    Amino-modified carbon dots were synthesized by the low temperature carbonization of citric acid in the pres-ence of branched polyethylenimine in one step.The bright blue emission was observed under the excitation of ultraviolet rays.Fourier transform infrared spectroscopy and X-ray powder diffraction were used to characterize its structure.Mean-while,the fluorescence resonance energy transfer between amino-modified carbon dots and CdTe QDs in liquid or solid-liquid phase was studied.The results show that fluorescence resonance energy transfer efficiency in liquid is much larger than the solid-liquid phase and it also has a limit.%以枝状聚乙烯亚胺和柠檬酸为原料,低温熔融法一步合成水溶性的氨基化碳点,碳点在紫外光激发下发出明亮的蓝光。采用傅里叶变换红外光谱和 X 射线粉末衍射仪对其结构进行表征,并研究其与碲化镉量子点在液相和固液界面的荧光共振能量转移。实验结果表明:液相中的荧光共振能量转移效率远大于固液界面的荧光共振能量转移,且能量转移具有一定的限度。

  5. Fluorescence and Magnetic Resonance Dual-Modality Imaging-Guided Photothermal and Photodynamic Dual-Therapy with Magnetic Porphyrin-Metal Organic Framework Nanocomposites

    Science.gov (United States)

    Zhang, Hui; Li, Yu-Hao; Chen, Yang; Wang, Man-Man; Wang, Xue-Sheng; Yin, Xue-Bo

    2017-01-01

    Phototherapy shows some unique advantages in clinical application, such as remote controllability, improved selectivity, and low bio-toxicity, than chemotherapy. In order to improve the safety and therapeutic efficacy, imaging-guided therapy seems particularly important because it integrates visible information to speculate the distribution and metabolism of the probe. Here we prepare biocompatible core-shell nanocomposites for dual-modality imaging-guided photothermal and photodynamic dual-therapy by the in situ growth of porphyrin-metal organic framework (PMOF) on Fe3O4@C core. Fe3O4@C core was used as T2-weighted magnetic resonance (MR) imaging and photothermal therapy (PTT) agent. The optical properties of porphyrin were well remained in PMOF, and PMOF was therefore selected for photodynamic therapy (PDT) and fluorescence imaging. Fluorescence and MR dual-modality imaging-guided PTT and PDT dual-therapy was confirmed with tumour-bearing mice as model. The high tumour accumulation of Fe3O4@C@PMOF and controllable light excitation at the tumour site achieved efficient cancer therapy, but low toxicity was observed to the normal tissues. The results demonstrated that Fe3O4@C@PMOF was a promising dual-imaging guided PTT and PDT dual-therapy platform for tumour diagnosis and treatment with low cytotoxicity and negligible in vivo toxicity. PMID:28272454

  6. Fluorescence characterization of co-immobilization-induced multi-enzyme aggregation in a polymer matrix using Förster resonance energy transfer (FRET): toward the metabolon biomimic.

    Science.gov (United States)

    Wu, Fei; Minteer, Shelley D

    2013-08-12

    Sequential metabolic enzymes can form supramolecular complexes named metabolons in vivo through enzyme-enzyme association or aggregation to facilitate efficient substrate channeling. By separately labeling enzymes with lysine-targeting carboxylic acid succinimidyl ester fluorophores of distinct excitation wavelengths, this research presents a quantitative study of polymer-entrapment-induced in vitro multi-enzyme aggregation from three Krebs cycle enzymes using Förster resonance energy transfer (FRET) to find potential polymer materials for immobilizing enzyme cascades and inducing the metabolon biomimic formation on electrodes. The effect of hydrophobic modification of linear polyethylenimine, Nafion, and chitosan polymers on metabolon formation has been investigated through photobleaching FRET imaging in addition to traditional steady-state fluorescence spectroscopy. By partially destroying FRET acceptors of longer excitation wavelength, increased fluorescence from dequenched donors of shorter excitation wavelength was measured and enzyme interactions in terms of energy-transfer efficiencies were mapped point by point. Results show that trimethyloctadecylammonium-modified Nafion works best in inducing multi-enzyme aggregation and exhibits a promising future in immobilized metabolon biomimics with the most uniform enzyme organization, as indicated by the protein distance distribution.

  7. In Vivo Magnetic Resonance and Fluorescence Dual-Modality Imaging of Tumor Angiogenesis in Rats Using GEBP11 Peptide Targeted Magnetic Nanoparticles.

    Science.gov (United States)

    Su, Tao; Wang, Yabin; Wang, Jiinda; Han, Dong; Ma, Sai; Cao, Jianbo; Li, Xiujuan; Zhang, Ran; Qiao, Hongyu; Liang, Jimin; Liu, Gang; Yang, Bo; Liang, Shuhui; Nie, Yongzhan; Wu, Kaichun; Li, Jiayi; Cao, Feng

    2016-05-01

    Angiogenesis is an essential process for tumor progression. Tumor vasculature-targeting peptides have shown great potential for use in cancer imaging and therapy. Our previous studies have shown that GEBP11, a novel vasculature-specific binding peptide that exhibits high affinity and specificity to tumor angiogenesis, is a promising candidate for the diagnosis and targeted radiotherapy of gastric cancer. In the present study, we developed a novel magnetic resonance and fluorescence (MR/Fluo) dual-modality imaging probe by covalently coupling 2,3-dimercaptosuccinnic acid-coated paramagnetic nanoparticles (DMSA-MNPs) and Cy5.5 to the GEBP11 peptide. The probe Cy5.5-GEBP11-DMSA-MNPs (CGD-MNPs), with a hydrodynamic diameter of 82.8 ± 6.5 nm, exhibited good imaging properties, high stability and little cytotoxicity. In vivo MR/Fluo imaging revealed that CGD-MNPs were successfully applied to visualize tumor angiogenesis in SGC-7901 xenograft mouse models. Prussian blue and CD31 immunohistochemical staining confirmed that CGD-MNPs co-localized with tumor blood vessels. In conclusion, CGD-MNPs are promising candidates for use as MR and fluorescence imaging probes for visualizing gastric cancer angiogenesis in vivo.

  8. Study on the absorption and fluorescence and resonance Rayleigh scattering spectra of Cu (Ⅱ)-fluoroquinolone chelates with erythrosine and their applications

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    In pH 4.2-5.0 Britton-Robinson buffer solution medium, fluoroquinolone antibiotics (FLQs), such as ciprofloxacin (CIP), norfloxacin (NOR), ofloxacin (OF), levofloxacin (LEV), lomefloxacin (LOM), and sparfloxacin (SPA), react with Cu (Ⅱ) to form chelate cations, which further bind with erythrosine to form the ion association complexes. They can result in the changes of the absorption spectra. Simultaneously, erythrosine fades obviously and the maximum fading wavelength is located at 526 nm. The fading reactions have high sensitivities. Thus, new spectrophotometries of determination for these drugs are developed. The ion-association reactions result in the quenching of fluorescence, which also have high sensitivities. The detection limits for six antibiotics are in the range of 7.1-12.2 μg·L-1. Furthermore, the reactions can result in the enhancement of resonance Rayleigh scattering (RRS). The maximum scattering peaks of six ion-association complexes are located at 566 nm, and there are two small RRS peaks at 333 nm and 287 nm. The detection limits for fluoroquinolone antibiotics are in the range of 1.70-3.10 μg·L-1 for RRS method. Among the above three methods, the RRS method has the highest sensitivity. In this work, we investigated the spectral characteristics of the absorption, fluorescence and RRS, the optimum conditions of the reactions, and the properties of the analytical chemistry. In addition, the mechanism of reactions were discussed by density function theory (DFT) and AM1 methods.

  9. Intramolecular ex vivo Fluorescence Resonance Energy Transfer (FRET of Dihydropyridine Receptor (DHPR β1a Subunit Reveals Conformational Change Induced by RYR1 in Mouse Skeletal Myotubes.

    Directory of Open Access Journals (Sweden)

    Dipankar Bhattacharya

    Full Text Available The dihydropyridine receptor (DHPR β1a subunit is essential for skeletal muscle excitation-contraction coupling, but the structural organization of β1a as part of the macromolecular DHPR-ryanodine receptor type I (RyR1 complex is still debatable. We used fluorescence resonance energy transfer (FRET to probe proximity relationships within the β1a subunit in cultured skeletal myotubes lacking or expressing RyR1. The fluorescein biarsenical reagent FlAsH was used as the FRET acceptor, which exhibits fluorescence upon binding to specific tetracysteine motifs, and enhanced cyan fluorescent protein (CFP was used as the FRET donor. Ten β1a reporter constructs were generated by inserting the CCPGCC FlAsH binding motif into five positions probing the five domains of β1a with either carboxyl or amino terminal fused CFP. FRET efficiency was largest when CCPGCC was positioned next to CFP, and significant intramolecular FRET was observed for all constructs suggesting that in situ the β1a subunit has a relatively compact conformation in which the carboxyl and amino termini are not extended. Comparison of the FRET efficiency in wild type to that in dyspedic (lacking RyR1 myotubes revealed that in only one construct (H458 CCPGCC β1a -CFP FRET efficiency was specifically altered by the presence of RyR1. The present study reveals that the C-terminal of the β1a subunit changes conformation in the presence of RyR1 consistent with an interaction between the C-terminal of β1a and RyR1 in resting myotubes.

  10. A Geant4-based Simulation to Evaluate the Feasibility of Using Nuclear Resonance Fluorescence (NRF) in Determining Atomic Compositions of Body Tissue in Cancer Diagnostics and Irradiation

    Science.gov (United States)

    Gilbo, Yekaterina; Wijesooriya, Krishni; Liyanage, Nilanga

    2017-01-01

    Customarily applied in homeland security for identifying concealed explosives and chemical weapons, NRF (Nuclear Resonance Fluorescence) may have high potential in determining atomic compositions of body tissue. High energy photons incident on a target excite the target nuclei causing characteristic re-emission of resonance photons. As the nuclei of each isotope have well-defined excitation energies, NRF uniquely indicates the isotopic content of the target. NRF radiation corresponding to nuclear isotopes present in the human body is emitted during radiotherapy based on Bremsstrahlung photons generated in a linear electron accelerator. We have developed a Geant4 simulation in order to help assess NRF capabilities in detecting, mapping, and characterizing tumors. We have imported a digital phantom into the simulation using anatomical data linked to known chemical compositions of various tissues. Work is ongoing to implement the University of Virginia's cancer center treatment setup and patient geometry, and to collect and analyze the simulation's physics quantities to evaluate the potential of NRF for medical imaging applications. Preliminary results will be presented.

  11. Cyclodextrin-Based Metal-Organic Nanotube as Fluorescent Probe for Selective Turn-On Detection of Hydrogen Sulfide in Living Cells Based on H2S-Involved Coordination Mechanism.

    Science.gov (United States)

    Xin, Xuelian; Wang, Jingxin; Gong, Chuanfang; Xu, Hai; Wang, Rongming; Ji, Shijie; Dong, Hanxiao; Meng, Qingguo; Zhang, Liangliang; Dai, Fangna; Sun, Daofeng

    2016-02-25

    Hydrogen sulfide (H2S) has been considered as the third biologically gaseous messenger (gasotransmitter) after nitric oxide (NO) and carbon monoxide (CO). Fluorescent detection of H2S in living cells is very important to human health because it has been found that the abnormal levels of H2S in human body can cause Alzheimer's disease, cancers and diabetes. Herein, we develop a cyclodextrin-based metal-organic nanotube, CD-MONT-2, possessing a {Pb14} metallamacrocycle for efficient detection of H2S. CD-MONT-2' (the guest-free form of CD-MONT-2) exhibits turn-on detection of H2S with high selectivity and moderate sensitivity when the material was dissolved in DMSO solution. Significantly, CD-MONT-2' can act as a fluorescent turn-on probe for highly selective detection of H2S in living cells. The sensing mechanism in the present work is based on the coordination of H2S as the auxochromic group to the central Pb(II) ion to enhance the fluorescence intensity, which is studied for the first time.

  12. Graphene and graphene-like two-denominational materials based fluorescence resonance energy transfer (FRET) assays for biological applications.

    Science.gov (United States)

    Tian, Feng; Lyu, Jing; Shi, Jingyu; Yang, Mo

    2017-03-15

    In the past decades, Förster resonance energy transfer (FRET) has been applied in many biological applications to reveal the biological information at the nanoscale. Recently, graphene and graphene-like two-dimensional (2D) nanomaterials started to be used in FRET assays as donors or acceptors including graphene oxide (GO), graphene quantum dot (GQD), graphitic-carbon nitride nanosheets (g-C3N4) and transition metal dichalcogenides (e.g. MoS2, MnO2, and WS2). Due to the remarkable properties such as large surface to volume ratio, tunable energy band, photoluminescence and excellent biocompatibility, these 2D nanomaterials based FRET assays have shown great potential in various biological applications. This review summarizes the recent development of graphene and graphene-like 2D nanomaterials based FRET assays in applications of biosensing, bioimaging, and drug delivery monitoring.

  13. Fluorescence resonance energy transfer and molecular modeling studies on 4',6-diamidino-2-phenylindole (DAPI) complexes with tubulin.

    Science.gov (United States)

    Arbildua, José J; Brunet, Juan E; Jameson, David M; López, Maribel; Nova, Esteban; Lagos, Rosalba; Monasterio, Octavio

    2006-03-01

    The goal of this work was to determine the binding properties and location of 4',6-diamidino-2-phenylindole (DAPI) complexed with tubulin. Using fluorescence anisotropy, a dissociation constant of 5.2+/-0.4 microM for the DAPI-tubulin complex was determined, slightly lower than that for the tubulin S complex. The influence of the C-terminal region on the binding of DAPI to tubulin was also characterized. Using FRET experiments, and assuming a kappa2 value of 2/3, distances between Co2+ bound to its high-affinity binding site and the DAPI-binding site and 2',3'-O-(trinitrophenyl)guanosine 5'-triphosphate bound to the exchangeable nucleotide and the DAPI-binding site were found to be 20+/-2 A and 43+/-2 A, respectively. To locate potential DAPI-binding sites on tubulin, a molecular modeling study was carried out using the tubulin crystal structure and energy minimization calculations. The results from the FRET measurements were used to limit the possible location of DAPI in the tubulin structure. Several candidate binding sites were found and these are discussed in the context of the various properties of bound DAPI.

  14. Fluorescence resonance energy transfer and molecular modeling studies on 4′,6-diamidino-2-phenylindole (DAPI) complexes with tubulin

    Science.gov (United States)

    Arbildua, José J.; Brunet, Juan E.; Jameson, David M.; López, Maribel; Nova, Esteban; Lagos, Rosalba; Monasterio, Octavio

    2006-01-01

    The goal of this work was to determine the binding properties and location of 4′,6-diamidino-2-phenylindole (DAPI) complexed with tubulin. Using fluorescence anisotropy, a dissociation constant of 5.2 ± 0.4 μM for the DAPI–tubulin complex was determined, slightly lower than that for the tubulin S complex. The influence of the C-terminal region on the binding of DAPI to tubulin was also characterized. Using FRET experiments, and assuming a κ2 value of 2/3, distances between Co2+ bound to its high-affinity binding site and the DAPI-binding site and 2′,3′-O-(trinitrophenyl)guanosine 5′-triphosphate bound to the exchangeable nucleotide and the DAPI-binding site were found to be 20 ± 2 Å and 43 ± 2 Å, respectively. To locate potential DAPI-binding sites on tubulin, a molecular modeling study was carried out using the tubulin crystal structure and energy minimization calculations. The results from the FRET measurements were used to limit the possible location of DAPI in the tubulin structure. Several candidate binding sites were found and these are discussed in the context of the various properties of bound DAPI. PMID:16452620

  15. Study on the absorption and fluorescence and resonance Rayleigh scattering spectra of Cu (II)-fluoroquinolone chelates with erythrosine and their applications

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    In pH 4.2-5.0 Britton-Robinson buffer solution medium, fluoroquinolone antibiotics (FLQs), such as ciprofloxacin (CIP), norfloxacin (NOR), ofloxacin (OF), levofloxacin (LEV), lomefloxacin (LOM), and sparfloxacin (SPA), react with Cu (II) to form chelate cations, which further bind with erythrosine to form the ion association complexes. They can result in the changes of the absorption spectra. Simultane- ously, erythrosine fades obviously and the maximum fading wavelength is located at 526 nm. The fad- ing reactions have high sensitivities. Thus, new spectrophotometries of determination for these drugs are developed. The ion-association reactions result in the quenching of fluorescence, which also have high sensitivities. The detection limits for six antibiotics are in the range of 7.1-12.2 μg·L?1. Furthermore, the reactions can result in the enhancement of resonance Rayleigh scattering (RRS). The maximum scattering peaks of six ion-association complexes are located at 566 nm, and there are two small RRS peaks at 333 nm and 287 nm. The detection limits for fluoroquinolone antibiotics are in the range of 1.70 -3.10 μg·L?1 for RRS method. Among the above three methods, the RRS method has the highest sen- sitivity. In this work, we investigated the spectral characteristics of the absorption, fluorescence and RRS, the optimum conditions of the reactions, and the properties of the analytical chemistry. In addi- tion, the mechanism of reactions were discussed by density function theory (DFT) and AM1 methods.

  16. Oligomerization of the UDP-glucuronosyltransferase 1A proteins: homo- and heterodimerization analysis by fluorescence resonance energy transfer and co-immunoprecipitation.

    Science.gov (United States)

    Operaña, Theresa N; Tukey, Robert H

    2007-02-16

    UDP-glucuronosyltransferases (UGTs) are membrane-bound proteins localized to the endoplasmic reticulum and catalyze the formation of beta-d-glucopyranosiduronic acids (glucuronides) using UDP-glucuronic acid and acceptor substrates such as drugs, steroids, bile acids, xenobiotics, and dietary nutrients. Recent biochemical evidence indicates that the UGT proteins may oligomerize in the membrane, but conclusive evidence is still lacking. In the present study, we have used fluorescence resonance energy transfer (FRET) to study UGT1A oligomerization in live cells. This technique demonstrated that UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A7, UGT1A8, UGT1A9, and UGT1A10 self-oligomerize (homodimerize). Heterodimer interactions were also explored, and it was determined that UGT1A1 was capable of binding with UGT1A3, UGT1A4, UGT1A6, UGT1A7, UGT1A8, UGT1A9, and UGT1A10. In addition to the in vivo FRET analysis, UGT1A protein-protein interactions were demonstrated through co-immunoprecipitation experiments. Co-expression of hemagglutinin-tagged and cyan fluorescent protein-tagged UGT1A proteins, followed by immunoprecipitation with anti-hemagglutinin beads, illustrated the potential of each UGT1A protein to homodimerize. Co-immunoprecipitation results also confirmed that UGT1A1 was capable of forming heterodimer complexes with all of the UGT1A proteins, corroborating the FRET results in live cells. These preliminary studies suggest that the UGT1A family of proteins form oligomerized complexes in the membrane, a property that may influence function and substrate selectivity.

  17. Förster Resonance Energy Transfer Switchable Self-Assembled Micellar Nanoprobe: Ratiometric Fluorescent Trapping of Endogenous H2S Generation via Fluvastatin-Stimulated Upregulation.

    Science.gov (United States)

    Zhao, Chunchang; Zhang, Xiuli; Li, Kaibin; Zhu, Shaojia; Guo, Zhiqian; Zhang, Lili; Wang, Feiyi; Fei, Qiang; Luo, Sihang; Shi, Ping; Tian, He; Zhu, Wei-Hong

    2015-07-08

    H2S produced in small amounts by mammalian cells has been identified in mediating biological signaling functions. However, the in situ trapping of endogenous H2S generation is still handicapped by a lack of straightforward methods with high selectivity and fast response. Here, we encapsulate a semi-cyanine-BODIPY hybrid dye (BODInD-Cl) and its complementary energy donor (BODIPY1) into the hydrophobic interior of an amphiphilic copolymer (mPEG-DSPE), especially for building up a ratiometric fluorescent H2S nanoprobe with extraordinarily fast response. A remarkable red-shift in the absorption band with a gap of 200 nm in the H2S response can efficiently switch off the Förster resonance energy transfer (FRET) from BODIPY1 to BODInD-Cl, subsequently recovering the donor fluorescence. Impressively, both the interior hydrophobicity of supramolecular micelles and electron-withdrawing nature of indolium unit in BODInD-Cl can sharply increase aromatic nucleophilic substitution with H2S. The ratiometric strategy based on the unique self-assembled micellar aggregate NanoBODIPY achieves an extremely fast response, enabling in situ imaging of endogenous H2S production and mapping its physiological and pathological consequences. Moreover, the amphiphilic copolymer renders the micellar assembly biocompatible and soluble in aqueous solution. The established FRET-switchable macromolecular envelope around BODInD-Cl and BODIPY1 enables cellular uptake, and makes a breakthrough in the trapping of endogenous H2S generation within raw264.7 macrophages upon stimulation with fluvastatin. This study manifests that cystathione γ-lyase (CSE) upregulation contributes to endogenous H2S generation in fluvastatin-stimulated macrophages, along with a correlation between CSE/H2S and activating Akt signaling pathway.

  18. Imaging neuroinflammation in vivo in a neuropathic pain rat model with near-infrared fluorescence and ¹⁹F magnetic resonance.

    Directory of Open Access Journals (Sweden)

    Kiran Vasudeva

    Full Text Available Chronic neuropathic pain following surgery represents a serious worldwide health problem leading to life-long treatment and the possibility of significant disability. In this study, neuropathic pain was modeled using the chronic constriction injury (CCI. The CCI rats exhibit mechanical hypersensitivity (typical neuropathic pain symptom to mechanical stimulation of the affected paw 11 days post surgery, at a time when sham surgery animals do not exhibit hypersensitivity. Following a similar time course, TRPV1 gene expression appears to rise with the hypersensitivity to mechanical stimulation. Recent studies have shown that immune cells play a role in the development of neuropathic pain. To further explore the relationship between neuropathic pain and immune cells, we hypothesize that the infiltration of immune cells into the affected sciatic nerve can be monitored in vivo by molecular imaging. To test this hypothesis, an intravenous injection of a novel perfluorocarbon (PFC nanoemulsion, which is phagocytosed by inflammatory cells (e.g. monocytes and macrophages, was used in a rat CCI model. The nanoemulsion carries two distinct imaging agents, a near-infrared (NIR lipophilic fluorescence reporter (DiR and a ¹⁹F MRI (magnetic resonance imaging tracer, PFC. We demonstrate that in live rats, NIR fluorescence is concentrated in the area of the affected sciatic nerve. Furthermore, the ¹⁹FF MRI signal was observed on the sciatic nerve. Histological examination of the CCI sciatic nerve reveals significant infiltration of CD68 positive macrophages. These results demonstrate that the infiltration of immune cells into the sciatic nerve can be visualized in live animals using these methods.

  19. Differential detection of Trichinella papuae, T. spiralis and T. pseudospiralis by real-time fluorescence resonance energy transfer PCR and melting curve analysis.

    Science.gov (United States)

    Tantrawatpan, Chairat; Intapan, Pewpan M; Thanchomnang, Tongjit; Lulitanond, Viraphong; Boonmars, Thidarut; Wu, Zhiliang; Morakote, Nimit; Maleewong, Wanchai

    2012-04-30

    Trichinellosis caused by nematodes of Trichinella spp. is a zoonotic foodborne disease. Three Trichinella species of the parasite including Trichinella spiralis, Trichinella papuae and Trichinella pseudospiralis, have been etiologic agents of human trichinellosis in Thailand. Definite diagnosis of this helminthiasis is based on a finding of the Trichinella larva (e) in a muscle biopsy. The parasite species or genotype can be determined using molecular methods, e.g., polymerase chain reaction (PCR). This study has utilized real-time fluorescence resonance energy transfer PCR (real-time FRET PCR) and a melting curve analysis for the differential diagnosis of trichinellosis. Three common Trichinella species in Thailand were studied using one set of primers and fluorophore-labeled hybridization probes specific for the small subunit of the mitochondrial ribosomal RNA gene. Using fewer than 35 cycles as the cut-off for positivity and using different melting temperatures (T(m)), this assay detected T. spiralis, T. papuae and T. pseudospiralis in muscle tissue and found the mean T(m) ± SD values to be 51.79 ± 0.06, 66.09 ± 0.46 and 51.46 ± 0.09, respectively. The analytical sensitivity of the technique enabled the detection of a single Trichinella larva of each species, and the detection limit for the target DNA sequence was 16 copies of positive control plasmid. A test of the technique's analytical specificity showed no fluorescence signal for a panel of 19 non-Trichinella parasites or for human and mouse genomic DNA. Due to the sensitivity and specificity of the detection of these Trichinella species, as well as the fast and high-throughput nature of these tools, this method has application potential in differentiating non-encapsulated larvae of T. papuae from T. spiralis and T. pseudospiralis in tissues of infected humans and animals.

  20. Comparison of Superparamagnetic and Ultrasmall Superparamagnetic Iron Oxide Cell Labeling for Tracking Green Fluorescent Protein Gene Marker with Negative and Positive Contrast Magnetic Resonance Imaging

    Directory of Open Access Journals (Sweden)

    Zhuoli Zhang

    2009-05-01

    Full Text Available The objectives of this study were to investigate the feasibility of imaging green fluorescent protein (GFP-expressing cells labeled with iron oxide nanoparticles with the fast low-angle positive contrast steady-state free precession (FLAPS method and to compare them with the traditional negative contrast technique. The GFP-R3230Ac cell line (GFP cell was incubated for 24 hours using 20 μg Fe/mL concentration of superparamagnetic iron oxide (SPIO and ultrasmall superparamagnetic iron oxide (USPIO nanoparticles. Cell samples were prepared for iron content analysis and cell function evaluation. The labeled cells were imaged using positive contrast with FLAPS imaging, and FLAPS images were compared with negative contrast T2*-weighted images. The results demonstrated that SPIO and USPIO labeling of GFP cells had no effect on cell function or GFP expression. Labeled cells were successfully imaged with both positive and negative contrast magnetic resonance imaging (MRI. The labeled cells were observed as a narrow band of signal enhancement surrounding signal voids in FLAPS images and were visible as signal voids in T2*-weighted images. Positive contrast and negative contrast imaging were both valuable for visualizing labeled GFP cells. MRI of labeled cells with GFP expression holds potential promise for monitoring the temporal and spatial migration of gene markers and cells, thereby enhancing the understanding of cell- and gene-based therapeutic strategies.

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

    Science.gov (United States)

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

    2015-01-01

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

  2. Detection of Ehrlichia canis in canine blood samples by real-time fluorescence resonance energy transfer (FRET) PCR and melting curve analysis.

    Science.gov (United States)

    Kongklieng, Amornmas; Thanchomnang, Tongjit; Intapan, Pewpan M; Boonmars, Thidarut; Janwan, Penchom; Sanpool, Oranuch; Lulitanond, Viraphong; Taweethavonsawat, Piyanan; Chungpivat, Sudchit; Morakote, Nimit; Maleewong, Wanchai

    2014-09-01

    Ehrlichia canis is a small pleomorphic gram-negative, coccoid, obligatory intracellular bacterium and the cause of canine monocytic ehrlichiosis. A real-time fluorescence resonance energy transfer polymerase chain reaction (real-time FRET PCR) coupled with melting curve analysis was established for detection of E. canis infection in canine blood samples. The VirB9 gene was amplified using one pair of primers and the melting curve analysis was generated by heating the hybridizing probes and amplified products. Eight E. canis-infected dog blood samples were initially identified using the Giemsa staining/microscopic method followed by conventional PCR (cPCR)/Sanger sequencing for confirmation. The sensitivity and specificity of the real-time FRET PCR detection were 87.5% and 100%, respectively and the limit of detection was 6.6 x 10(3) copies of positive E. canis control plasmids. The real-time FRET PCR with melting curve analysis reported here is better than microscopic visualization or cPCR because the method is not affected by the false bias inherent in the microscopic method. Furthermore, many samples can be processed rapidly at the same time. This convenient tool is beneficial as an alternative assay for the epidemiologic study of canine ehrlichiosis as well as for eradication of these organisms in prevention and control programs in endemic areas.

  3. Determination of the electromagnetic dipole strength distribution in medium-heavy atomic nuclei by means of nuclear resonance fluorescence; Bestimmung der elektromagnetischen Dipolstaerkeverteilung in mittelschweren Atomkernen mittels Kernresonanzfluoreszenz

    Energy Technology Data Exchange (ETDEWEB)

    Massarczyk, Ralph Jens

    2011-01-17

    During the last hundred years several models were developed to describe the configuration of nuclei. These models have to make predictions, which should be comparable with experiments. As a standard type of experiment the nuclear resonance fluorescence was established. A nucleus is excited by irradiation with photons. By emitting one or more photons the nucleus decays back to the ground state. With this method it is possible to measure energy levels and to determine the strength of their excitation. A continuum of unresolved peaks gives additional strength. The existing setup at the linear electron accelerator ELBE of the Forschungszentrum Dresden-Rossendorf uses bremsstrahlung, produced as a secondary beam in a thin Niobium foil. During the years 2008/09 experiments on the nuclei of {sup 86}Kr and {sup 136}Ba took place there. In this work they will be analyzed. Photon flux and efficiency determination have been done as well as simulations on detector response and non-nuclear scattered background events. For this purpose the GEANT4 package was used. Finally the resulting cross sections were corrected for branching and feeding.

  4. Study on the interaction between albendazole and eosin Y by fluorescence, resonance Rayleigh scattering and frequency doubling scattering spectra and their analytical applications

    Science.gov (United States)

    Tian, Fengling; Huang, Wei; Yang, Jidong; Li, Qin

    In pH 3.25-3.35 Britton-Robinson (BR) buffer solution, albendazole (ABZ) could react with eosin Y (EY) to form a 1:1 ion-association complex, which not only results in the quenching of fluorescence, but also resulted in the great enhancement of resonance Rayleigh scattering (RRS) and frequency doubling scattering (FDS). Furthermore, a new RRS spectrum will appear, and the maximum RRS wavelength was located at about 356 nm. The detection limit for ABZ were 21.51 ng mL-1 for the fluorophotometry, 6.93 ng mL-1 for the RRS method and 12.89 ng mL-1 for the FDS method. Among them, the RRS method had the highest sensitivity. The experimental conditions were optimized and effects of coexisting substances were evaluated. Meanwhile, the influences of coexisting substances were tested. The methods have been successfully applied to the determination of ABZ in capsules and human urine samples. The composition and structure of the ion-association complex and the reaction mechanism were discussed.

  5. Interfacial chemistry and the design of solid-phase nucleic acid hybridization assays using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Algar, W Russ; Krull, Ulrich J

    2011-01-01

    The use of quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) offer several advantages for the development of multiplexed solid-phase QD-FRET nucleic acid hybridization assays. Designs for multiplexing have been demonstrated, but important challenges remain in the optimization of these systems. In this work, we identify several strategies based on the design of interfacial chemistry for improving sensitivity, obtaining lower limits of detection (LOD) and enabling the regeneration and reuse of solid-phase QD-FRET hybridization assays. FRET-sensitized emission from acceptor dyes associated with hybridization events at immobilized QD donors provides the analytical signal in these assays. The minimization of active sensing area reduces background from QD donor PL and allows the resolution of smaller amounts of acceptor emission, thus lowering the LOD. The association of multiple acceptor dyes with each hybridization event can enhance FRET efficiency, thereby improving sensitivity. Many previous studies have used interfacial protein layers to generate selectivity; however, transient destabilization of these layers is shown to prevent efficient regeneration. To this end, we report a protein-free interfacial chemistry and demonstrate the specific detection of as little as 2 pmol of target, as well as an improved capacity for regeneration.

  6. Rapid and accurate detection of the CFTR gene mutation 1811+1.6 kbA>G by real-time fluorescence resonance energy transfer PCR.

    Science.gov (United States)

    Reboul, Marie-Pierre; Higueret, Laurent; Biteau, Nicolas; Iron, Albert

    2005-10-01

    The CFTR gene mutation 1811+1.6 kbA>G has been reported as associated with a severe phenotype of cystic fibrosis with pancreatic insufficiency. This mutation has been identified as a rather common one in the South West of France and in the Iberian Peninsula. Because of the precise geographical origin of the subjects and its frequency, the mutation has to be investigated with accuracy. We have developed an original real-time Fluorescence Resonance Energy Transfer (FRET) PCR assay for genotyping the mutation 1811+1.6 kbA>G. It is based on the amplification of a region spanning the mutation with simultaneous detection of the amplicon by hybridization with a bi-probe followed by a melting curve analysis. The results obtained are identical with those resulting from either restriction fragment length polymorphism analysis or sequencing. The distinction between the wild type and the mutation 1811+1.6 kbA>G is easy because the corresponding melting points shows a difference of 6 or 9.5 degrees C depending on the associated SNP A/T located 16 bp downstream. We demonstrated that a FRET assay showed enough sensitivity to discriminate between two nucleotide polymorphisms (SNPs) in the sequence of the sensor. In conclusion, this method is specific, fast, easy to perform, reproducible, inexpensive as it uses only one bi-probe and well adapted to daily practice.

  7. Real-time PCR/MCA assay using fluorescence resonance energy transfer for the genotyping of resistance related DHPS-540 mutations in Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Dujardin Jean-Claude

    2008-03-01

    Full Text Available Abstract Background Sulphadoxine-pyrimethamine has been abandoned as first- or second-line treatment by most African malaria endemic countries in favour of artemisinin-based combination treatments, but the drug is still used as intermittent preventive treatment during pregnancy. However, resistance to sulphadoxine-pyrimethamine has been increasing in the past few years and, although the link between molecular markers and treatment failure has not been firmly established, at least for pregnant women, it is important to monitor such markers. Methods This paper reports a novel sensitive, semi-quantitative and specific real-time PCR and melting curve analysis (MCA assay using fluorescence resonance energy transfer (FRET for the detection of DHPS-540, an important predictor for SP resistance. FRET/MCA was evaluated using 78 clinical samples from malaria patients and compared to PCR-RFLP. Results Sixty-two samples were in perfect agreement between both assays. One sample showed a small wild type signal with FRET/MCA that indicates a polyclonal infection. Four samples were not able to generate enough material in both assays to distinguish mutant from wild-type infection, six samples gave no signal in PCR-RFLP and five samples gave no amplification in FRET/MCA. Conclusion FRET/MCA is an effective tool for the identification of SNPs in drug studies and epidemiological surveys on resistance markers in general and DHPS-540 mutation in particular.

  8. Mammalian Cell-Based Sensor System

    Science.gov (United States)

    Banerjee, Pratik; Franz, Briana; Bhunia, Arun K.

    Use of living cells or cellular components in biosensors is receiving increased attention and opens a whole new area of functional diagnostics. The term "mammalian cell-based biosensor" is designated to biosensors utilizing mammalian cells as the biorecognition element. Cell-based assays, such as high-throughput screening (HTS) or cytotoxicity testing, have already emerged as dependable and promising approaches to measure the functionality or toxicity of a compound (in case of HTS); or to probe the presence of pathogenic or toxigenic entities in clinical, environmental, or food samples. External stimuli or changes in cellular microenvironment sometimes perturb the "normal" physiological activities of mammalian cells, thus allowing CBBs to screen, monitor, and measure the analyte-induced changes. The advantage of CBBs is that they can report the presence or absence of active components, such as live pathogens or active toxins. In some cases, mammalian cells or plasma membranes are used as electrical capacitors and cell-cell and cell-substrate contact is measured via conductivity or electrical impedance. In addition, cytopathogenicity or cytotoxicity induced by pathogens or toxins resulting in apoptosis or necrosis could be measured via optical devices using fluorescence or luminescence. This chapter focuses mainly on the type and applications of different mammalian cell-based sensor systems.

  9. Application of the laser induced fluorescence to the investigation of highly magnetized plasmas, heated by ion cyclotron resonance; Fluorescence induite par laser sur des plasmas fortement magnetises, chauffes par resonnance cyclotron ionique

    Energy Technology Data Exchange (ETDEWEB)

    Pailloux, A. [CEA Centre d`Etudes de Saclay, 91 - Gif-sur-Yvette (France). Dept. des Procedes d`Enrichissement]|[Universite Louis Pasteur, 67 - Strasbourg (France)

    1997-12-31

    This work has been achieved in the frame of isotopic separation studies by in cyclotron resonance. For this purpose, in a highly magnetized (2 to 3 Tesla) and non-collisional (10{sup 12} ions/cm{sup 3}) plasma, composed of metallic ions, a wave near the ion cyclotron frequency is thrown in order to heat selectively a given species. A laser induced fluorescence (LIP) has been developed on barium and gadolinium plasmas. The Larmor gyration of ions greatly modifies the interaction, which has been modelled through the time-dependent Schroedinger equation. The obtained excitation probably has been integrated over all the ions excited in the measurement volume in order to check that the LIF still leads to the distribution function of ion velocities. The influence of the Larmor motion of ions on the spectral distribution of LIF has been derived both theoretically and experimentally. The LIF diagnostics has been achieved with a dye O`ring laser. The barium ion has been excited on the transition 6142 angstrom, using rhodamine 6G dye, and the gadolinium ion on the pseudo-triplet 3861 angstrom, using exalite dye. Data treatment has been developed taking into account the Zeeman effect and the different heating of isotopes. The ionic temperature (from 1 eV to some hundreds eV) has been measured as a function of radiofrequency heating. Our experimental results are in good agreement with the selective heating theory. Also, the ion velocity distribution function has been found locally Maxwellian. And the behaviour of the plasma has been studied as a function of control parameters of the plasma source. (author) 62 refs.

  10. Optical-optical double resonance, laser induced fluorescence, and revision of the signs of the spin-spin constants of the boron carbide (BC) free radical

    Energy Technology Data Exchange (ETDEWEB)

    Sunahori, Fumie X. [Department of Chemistry and Physics, Franklin College, Franklin, Indiana 46131 (United States); Nagarajan, Ramya; Clouthier, Dennis J., E-mail: dclaser@uky.edu [Department of Chemistry, University of Kentucky, Lexington, Kentucky 40506-0055 (United States)

    2015-12-14

    The cold boron carbide free radical (BC X {sup 4}Σ{sup −}) has been produced in a pulsed discharge free jet expansion using a precursor mixture of trimethylborane in high pressure argon. High resolution laser induced fluorescence spectra have been obtained for the B {sup 4}Σ{sup −}–X {sup 4}Σ{sup −} and E {sup 4}Π–X {sup 4}Σ{sup −} band systems of both {sup 11}BC and {sup 10}BC. An optical-optical double resonance (OODR) scheme was implemented to study the finer details of both band systems. This involved pumping a single rotational level of the B state with one laser and then recording the various allowed transitions from the intermediate B state to the final E state with a second laser by monitoring the subsequent E–X ultraviolet fluorescence. In this fashion, we were able to prove unambiguously that, contrary to previous studies, the spin-spin constant λ is negative in the ground state and positive in the B {sup 4}Σ{sup −} excited state. It has been shown that λ″ < 0 is in fact expected based on a semiempirical second order perturbation theory calculation of the magnitude of the spin-spin constant. The OODR spectra have also been used to validate our assignments of the complex and badly overlapped E {sup 4}Π–X {sup 4}Σ{sup −} 0-0 and 1-0 bands of {sup 11}BC. The E–X 0-0 band of {sup 10}BC was found to be severely perturbed. The ground state main electron configuration is …3σ{sup 2}4σ{sup 2}5σ{sup 1}1π{sup 2}2π{sup 0} and the derived bond lengths show that there is a 0.03 Å contraction in the B state, due to the promotion of an electron from the 4σ antibonding orbital to the 5σ bonding orbital. In contrast, the bond length elongates by 0.15 Å in the E state, a result of promoting an electron from the 5σ bonding orbital to the 2π antibonding orbitals.

  11. Optical-optical double resonance, laser induced fluorescence, and revision of the signs of the spin-spin constants of the boron carbide (BC) free radical

    Science.gov (United States)

    Sunahori, Fumie X.; Nagarajan, Ramya; Clouthier, Dennis J.

    2015-12-01

    The cold boron carbide free radical (BC X 4Σ-) has been produced in a pulsed discharge free jet expansion using a precursor mixture of trimethylborane in high pressure argon. High resolution laser induced fluorescence spectra have been obtained for the B 4Σ--X 4Σ- and E 4Π-X 4Σ- band systems of both 11BC and 10BC. An optical-optical double resonance (OODR) scheme was implemented to study the finer details of both band systems. This involved pumping a single rotational level of the B state with one laser and then recording the various allowed transitions from the intermediate B state to the final E state with a second laser by monitoring the subsequent E-X ultraviolet fluorescence. In this fashion, we were able to prove unambiguously that, contrary to previous studies, the spin-spin constant λ is negative in the ground state and positive in the B 4Σ- excited state. It has been shown that λ″ < 0 is in fact expected based on a semiempirical second order perturbation theory calculation of the magnitude of the spin-spin constant. The OODR spectra have also been used to validate our assignments of the complex and badly overlapped E 4Π-X 4Σ- 0-0 and 1-0 bands of 11BC. The E-X 0-0 band of 10BC was found to be severely perturbed. The ground state main electron configuration is …3σ24σ25σ11π22π0 and the derived bond lengths show that there is a 0.03 Å contraction in the B state, due to the promotion of an electron from the 4σ antibonding orbital to the 5σ bonding orbital. In contrast, the bond length elongates by 0.15 Å in the E state, a result of promoting an electron from the 5σ bonding orbital to the 2π antibonding orbitals.

  12. Optical-optical double resonance, laser induced fluorescence, and revision of the signs of the spin-spin constants of the boron carbide (BC) free radical.

    Science.gov (United States)

    Sunahori, Fumie X; Nagarajan, Ramya; Clouthier, Dennis J

    2015-12-14

    The cold boron carbide free radical (BC X (4)Σ(-)) has been produced in a pulsed discharge free jet expansion using a precursor mixture of trimethylborane in high pressure argon. High resolution laser induced fluorescence spectra have been obtained for the B (4)Σ(-)-X (4)Σ(-) and E (4)Π-X (4)Σ(-) band systems of both (11)BC and (10)BC. An optical-optical double resonance (OODR) scheme was implemented to study the finer details of both band systems. This involved pumping a single rotational level of the B state with one laser and then recording the various allowed transitions from the intermediate B state to the final E state with a second laser by monitoring the subsequent E-X ultraviolet fluorescence. In this fashion, we were able to prove unambiguously that, contrary to previous studies, the spin-spin constant λ is negative in the ground state and positive in the B (4)Σ(-) excited state. It has been shown that λ″ < 0 is in fact expected based on a semiempirical second order perturbation theory calculation of the magnitude of the spin-spin constant. The OODR spectra have also been used to validate our assignments of the complex and badly overlapped E (4)Π-X (4)Σ(-) 0-0 and 1-0 bands of (11)BC. The E-X 0-0 band of (10)BC was found to be severely perturbed. The ground state main electron configuration is …3σ(2)4σ(2)5σ(1)1π(2)2π(0) and the derived bond lengths show that there is a 0.03 Å contraction in the B state, due to the promotion of an electron from the 4σ antibonding orbital to the 5σ bonding orbital. In contrast, the bond length elongates by 0.15 Å in the E state, a result of promoting an electron from the 5σ bonding orbital to the 2π antibonding orbitals.

  13. Fluorescence resonance energy transfer between NaYF{sub 4}:Yb,Tm upconversion nanoparticles and gold nanorods: Near-infrared responsive biosensor for streptavidin

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Shuang; Wang, Jing; Xu, Wen; Chen, Boting; Yu, Wei; Xu, Lin; Song, Hongwei, E-mail: songhw@jlu.edu.cn

    2014-03-15

    We represent a fluorescence resonance energy transfer (FRET) system using upconversion nanoparticles (UCNPs) and the gold nanorods (GNRs) as the energy donor–acceptor pair for directly determining streptavidin in near-infrared (NIR) region. NaYF{sub 4}:Yb,Tm UCNPs, which had a strong emission at 800 nm under 980-nm excitation, were adopted as the energy donor. The GNRs, which demonstrated strong surface plasmon absorption around 800 nm, were chosen as acceptor to quench the 800 nm emissions of the UCNPs. There had the spectral overlap between the emission of the donor nanoparticles (UCNPs) and the absorption of the acceptor nanoparticles (GNRs). This UCNP-based FRET system was then used to determine the amount of streptavidin. In this system, NaYF{sub 4}:Yb,Tm UCNPs conjugated with biotin, while GNRs conjugated with streptavidin. When added GNRs into UCNPs, the streptavidin were preferred to bind with biotin and decreased spacing between the donor and acceptor NPs. Consequently, FRET occurred and a linear relationship between the luminescence quenching efficiency and the concentration of streptavidin was obtained. Owing to the aforementioned merits of UCNPs as an energy donor and the strong quenching ability of GNRs, satisfactory analytical performances have been acquired. -- Highlights: • NaYF4:Yb,Tm and GNRs are as NIR energy donor and quenching acceptor for FRET. • Linkage between biotin and streptavidin make the distance between the donors and the acceptors short enough for FRET. • The FRET system in this work was applicable for the detection of streptavidin. • The donor and acceptor NPs can be modified by proper molecules for other biological molecules detection.

  14. Paper-based solid-phase multiplexed nucleic acid hybridization assay with tunable dynamic range using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Noor, M Omair; Krull, Ulrich J

    2013-08-06

    A multiplexed solid-phase nucleic acid hybridization assay on a paper-based platform is presented using multicolor immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). The surface of paper was modified with imidazole groups to immobilize two types of QD-probe oligonucleotide conjugates that were assembled in solution. Green-emitting QDs (gQDs) and red-emitting QDs (rQDs) served as donors with Cy3 and Alexa Fluor 647 (A647) acceptors. The gQD/Cy3 FRET pair served as an internal standard, while the rQD/A647 FRET pair served as a detection channel, combining the control and analytical test zones in one physical location. Hybridization of dye-labeled oligonucleotide targets provided the proximity for FRET sensitized emission from the acceptor dyes, which served as an analytical signal. Hybridization assays in the multicolor format provided a limit of detection of 90 fmol and an upper limit of dynamic range of 3.5 pmol. The use of an array of detection zones was designed to provide improved analytical figures of merit compared to that which could be achieved on one type of array design in terms of relative concentration of multicolor QDs. The hybridization assays showed excellent resistance to nonspecific adsorption of oligonucleotides. Selectivity of the two-plex hybridization assay was demonstrated by single nucleotide polymorphism (SNP) detection at a contrast ratio of 50:1. Additionally, it is shown that the use of preformed QD-probe oligonucleotide conjugates and consideration of the relative number density of the two types of QD-probe conjugates in the two-color assay format is advantageous to maximize assay sensitivity and the upper limit of dynamic range.

  15. Paper-based solid-phase nucleic acid hybridization assay using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Noor, M Omair; Shahmuradyan, Anna; Krull, Ulrich J

    2013-02-05

    A paper-based solid-phase assay is presented for transduction of nucleic acid hybridization using immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). The surface of paper was modified with imidazole groups to immobilize QD-probe oligonucleotide conjugates that were assembled in solution. Green-emitting QDs (gQDs) were FRET-paired with Cy3 acceptor. Hybridization of Cy3-labeled oligonucleotide targets provided the proximity required for FRET-sensitized emission from Cy3, which served as an analytical signal. The assay exhibited rapid transduction of nucleic acid hybridization within minutes. Without any amplification steps, the limit of detection of the assay was found to be 300 fmol with the upper limit of the dynamic range at 5 pmol. The implementation of glutathione-coated QDs for the development of nucleic acid hybridization assay integrated on a paper-based platform exhibited excellent resistance to nonspecific adsorption of oligonucleotides and showed no reduction in the performance of the assay in the presence of large quantities of noncomplementary DNA. The selectivity of nucleic acid hybridization was demonstrated by single-nucleotide polymorphism (SNP) detection at a contrast ratio of 19 to 1. The reuse of paper over multiple cycles of hybridization and dehybridization was possible, with less than 20% reduction in the performance of the assay in five cycles. This work provides an important framework for the development of paper-based solid-phase QD-FRET nucleic acid hybridization assays that make use of a ratiometric approach for detection and analysis.

  16. Preliminary Study of the Efficacy of Using Nuclear Resonance Fluorescence with Quasi-Monoenergetic Gamma-Ray Sources for Nuclear Safeguards Assay

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, M S; McNabb, D P; Hall, J M; Gonzalez, J J

    2011-02-17

    We have studied the efficacy of using nuclear resonance fluorescence (NRF)-based techniques to assay spent nuclear fuel for Pu content using quasi-monoenergetic sources. We have developed two techniques to precisely determine the Pu content in a fuel rod/pin. One of our approaches is virtually free of systematic uncertainties. Using analytical models, we have determined the amount of time required to measure the Pu content in spent nuclear fuel rods and spent fuel assemblies to within 1% precision. We note that Pu content can be determined in a fuel assembly about as fast as in a single fuel pin. The performance of NRF-based assay techniques with improved photon sources, which are currently under development, will also estimated. For follow-on research we propose to: (1) Construct research prototype detection systems for both of the NRF-based assay systems proposed in this paper and measure their calibration curves; (2) Determine the systematic errors associated with both assay methods, explore ways to reduce the errors and fold the results into future performance calculations; (3) Develop an algorithm to assay a fuel assembly; (4) Perform validation measurements using a single pin and scaled assemblies; (5) Research and develop current-mode detection and/or threshold detection techniques to improve assay times; (6) Characterize the flux of newly constructed sources and fold the results into the calculations presented here to determine the feasibility of a variety of proposed sources; and (7) Collaborate with others in the safeguards community to build a prototype system and perform an NRF-based assay demonstration on spent fuel.

  17. Dual fluorescence resonance energy transfer assay between tunable upconversion nanoparticles and controlled gold nanoparticles for the simultaneous detection of Pb²⁺ and Hg²⁺.

    Science.gov (United States)

    Wu, Shijia; Duan, Nuo; Shi, Zhao; Fang, Congcong; Wang, Zhouping

    2014-10-01

    In this work, we presented a novel dual fluorescence resonance energy transfer (FRET) system for the simultaneous detection of Pb(2+) and Hg(2+). This system employed two color upconversion nanoparticles (UCNPs) as the donors, and controlled gold nanoparticles (AuNPs) as the acceptors. The two donor-acceptor pairs were fabricated by hybridizing the aptamers and their corresponding complementary DNA. Thus, the green and red upconversion fluorescence could be quenched because of a good overlap between the UCNPs fluorescence emission and the AuNPs absorption spectrum. In the presence of Pb(2+) and Hg(2+), the aptamers preferred to bind to their corresponding analytes and formed a G-quadruplexes structure for Pb(2+) and the hairpin-like structure for Hg(2+). As a result, the dual FRET was disrupted, and the green and red upconversion fluorescence was restored. Under optimized experimental conditions, the relative fluorescence intensity increased as the metal ion concentrations were increased, allowing for the quantification of Pb(2+) and Hg(2+). The relationships between the fluorescence intensity and plotting logarithms of ion concentrations were linear in the range from 0.1 to 100 nM for Pb(2+) and 0.5 to 500 nM for Hg(2+), and the detection limits of Pb(2+) and Hg(2+) were 50 pM and 150 pM, respectively. As a practical application, the aptasensor was used to monitor Pb(2+) and Hg(2+) levels in naturally contaminated samples and human serum samples. Ultimately, this type of dual FRET could be used to detect other metal ions or contaminants in food safety analysis and environment monitoring.

  18. 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

    Science.gov (United States)

    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.

  19. [Cell based therapy for COPD].

    Science.gov (United States)

    Kubo, Hiroshi

    2007-04-01

    To develop a new cell based therapy for chronic obstructive pulmonary disease (COPD), we need to understand 1) the role of tissue-specific and bone marrow-derived stem cells, 2) extracellular matrix, and 3) growth factors. Recently, bronchioalveolar stem cells were identified in murine distal lungs. Impairment of these stem cells may cause improper lung repair after inflammation, resulting in pulmonary emphysema. Bone marrow-derived cells are necessary to repair injured lungs. However, the long term role of these cells is not understood yet. Although we need more careful analysis and additional experiments, growth factors, such as hepatocyte growth factor, are good candidates for the new cell based therapy for COPD. Lung was believed as a non-regenerative organ. Based on these recent reports about lung regeneration and stem cells, however, new strategies to treat COPD and a new point of view to understand the pathophysiology of COPD are rising.

  20. Surface plasmon resonance-enhanced fluorescence implementation of a single-step competition assay: demonstration of fatty acid measurement using an anti-fatty acid monoclonal antibody and a Cy5-labeled fatty acid.

    Science.gov (United States)

    Vareiro, Margarida M L M; Tranchant, Isabelle; Maplin, Sandra; Zak, Kris; Gani, M M; Slevin, Christopher J; Hailes, Helen C; Tabor, Alethea B; Cameron, Petra J; Jenkins, A Toby A; Williams, David E

    2008-06-15

    The development of a single-step, separation-free method for measurement of low concentrations of fatty acid using a surface plasmon resonance-enhanced fluorescence competition assay with a surface-bound antibody is described. The assay behavior was unexpectedly complex. A nonlinear coverage-dependent self-quenching of emission from surface-bound fluorescent label was deduced from the response kinetics and attributed to a surface plasmon-mediated energy transfer between adsorbed fluorophores, modified by the effects of plasmon interference. Principles of assay design to avoid complications from such effects are discussed. An anti-fatty acid mouse monoclonal antibody reacting to the alkyl chain was prepared and supported on a gold chip at a spacing appropriate for surface-plasmon field-enhanced fluorescence spectroscopy (SPEFS), by applying successively a self-assembled biotinylated monolayer, then streptavidin, then biotinylated protein A, and then the antibody, which was crosslinked to the protein A. Synthesis of a fluorescently (Cy5) tagged C-11 fatty acid is reported. SPEFS was used to follow the kinetics of the binding of the labeled fatty acid to the antibody, and to implement a competition assay with free fatty acid (undecanoic acid), sensitive at the 1 microM scale, a sensitivity limit caused by the low affinity of antibodies for free fatty acids, rather than the SPEFS technique itself. Free fatty acid concentration in human serum is in the range 0.1-1mM, suggesting that this measurement approach could be applied in a clinical diagnostic context. Finally, a predictive, theoretical model of fatty acid binding was developed that accounted for the observed "overshoot" kinetics.

  1. A fluorescence resonance energy transfer (FRET) based "Turn-On" nanofluorescence sensor using a nitrogen-doped carbon dot-hexagonal cobalt oxyhydroxide nanosheet architecture and application to α-glucosidase inhibitor screening.

    Science.gov (United States)

    Li, Guoliang; Kong, Weiheng; Zhao, Mei; Lu, Shuaimin; Gong, Peiwei; Chen, Guang; Xia, Lian; Wang, Hua; You, Jinmao; Wu, Yongning

    2016-05-15

    The medicines targeted at α-glucosidase played an important role in anti-diabetes and anti-HIV therapy. Unfortunately, the method based on fluorescent assay strategy for α-glucosidase inhibitor screening remains poorly investigated. In this study, a novel "Turn On" fluorescence sensor platform has been developed for trace α-glucosidase inhibitor screening from natural medicines. Firstly, carbon dots were prepared by one-pot synthesis and used as the signal output. Combining with the carbon dots, cobalt oxyhydroxide (CoOOH) nanoflakes were employed to build the fluorescence resonance energy transfer (FRET) based sensor platform. Secondly, L-ascorbic acid-2-O-α-D-glucopyranosyl (AAG) was innovatively introduced as α-glucosidase substrate. With hydrolysis of AAG by α-glucosidase, ascorbic acids (AA) were released that can rapidly reduce CoOOH nanoflakes to Co(2+), and then FRET was stopped accompanying with the fluorescence recovery of CDs. The sensor platform was ultrasensitive to AA with a detection limit of 5 nM, ensuring the sensitive monitoring of enzyme activity. Acarbose was used as the inhibitor model and its inhibition rate is proportional to the logarithm of concentration in range of 10(-9)-10(-3)M with the correlation coefficient of R(2)=0.996, and an ultralow limit of detection of ~1×10(-9)M was obtained. The inhibiting ability of seven compounds isolated from natural medicines was also evaluated. The constructed sensor platform was proven to be sensitive and selective as well as cost-effective, facile and reliable, making it promising as a candidate for trace α-glucosidase inhibitor screening.

  2. Analysis of the near-resonant fluorescence spectra of a single rubidium atom localized in a three-dimensional optical lattice

    CERN Document Server

    Kim, Wookrae; Kim, Jung-Ryul; Lee, Yea-Lee; Ihm, Jisoon; An, Kyungwon

    2010-01-01

    Supplementary information is presented on the recent work by W. Kim et al. on the matter-wave-tunneling-induced broadening in the near-resonant spectra of a single rubidium atom localized in a three-dimensional optical lattice in a strong Lamb-Dicke regime.

  3. A novel approach to identify non-palpable breast lesions combining fluorescent liposomes and magnetic resonance-guided high intensity focused ultrasound-triggered release

    NARCIS (Netherlands)

    Oerlemans, Chris; Nijsen, Frank; van Amersfoort, Miranda; van Bloois, Louis; Heijman, Edwin; Luijten, Peter; Mali, Willem; Storm, Gert

    2011-01-01

    The combination of fluorescein-containing liposomes (FCL) and magnetic resonance-guided high intensity focused ultrasound (MR-HIFU)-triggered release is a promising approach for lesion demarcation and more efficient removal of non-palpable breast lesions. Exposure of FCL to ablation temperatures (60

  4. Fluorescence Resonance Energy Transfer of the Tb(III)-Nd(III) Binary System in Molten LiCl-KCl Eutectic Salt

    Energy Technology Data Exchange (ETDEWEB)

    Kim, B. Y. [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of); Yun, J. I. [KAIST, Daejeon (Korea, Republic of)

    2015-05-15

    The lanthanides act as a neutron poison in nuclear reactor with large neutron absorption cross section. For that reason, very low amount of lanthanides is required in the recovered U/TRU ingot product from pyrochemical process. In view of that, the investigation of thermodynamic properties and chemical behaviors of lanthanides in molten chloride salt are necessary to estimate the performance efficiency of pyrochemical process. However, there are uncertainties about knowledge and understanding of basic mechanisms in pyrochemical process, such as chemical speciation and redox behaviors due to the lack of in-situ monitoring methods for high temperature molten salt. The spectroscopic analysis is one of the probable techniques for in-situ qualitative and quantitative analysis. Recently, a few fluorescence spectroscopic measurements on single lanthanide element in molten LiCl-KCl eutectic have been investigated. The fluorescence intensity and the fluorescence lifetime of Tb(III) were decreased as increasing the concentration of Nd(III), demonstrating collisional quenching between donor ions and acceptor ions. The Forster distance (..0) of Tb(III)-Nd(III) binary system in molten LiCl-KCl eutectic was determined in the specific range of .... (0.1-1.0) and .. (1.387-1.496)

  5. Biomolecule-to-fluorescent-color encoder: modulation of fluorescence emission via DNA structural changes.

    Science.gov (United States)

    Nishimura, Takahiro; Ogura, Yusuke; Yamada, Kenji; Ohno, Yuko; Tanida, Jun

    2014-07-01

    A biomolecule-to-fluorescent-color (B/F) encoder for optical readout of biomolecular information is proposed. In the B/F encoder, a set of fluorescence wavelengths and their intensity levels are used for coding of a biomolecular signal. A hybridization chain reaction of hairpin DNAs labeled with fluorescent reporters was performed to generate the fluorescence color codes. The fluorescence is modulated via fluorescence resonance energy transfer, which is controlled by DNA structural changes. The results demonstrate that fluorescent color codes can be configured based on two wavelengths and five intensities using the B/F encoder, and the assigned codes can be retrieved via fluorescence measurements.

  6. Cell-based microfluidic platform for mimicking human olfactory system.

    Science.gov (United States)

    Lee, Seung Hwan; Oh, Eun Hae; Park, Tai Hyun

    2015-12-15

    Various attempts have been made to mimic the human olfactory system using human olfactory receptors (hORs). In particular, OR-expressed cell-based odorant detection systems mimic the smell sensing mechanism of humans, as they exploit endogenous cellular signaling pathways. However, the majority of such cell-based studies have been performed in the liquid phase to maintain cell viability, and liquid odorants were used as detection targets. Here, we present a microfluidic device for the detection of gaseous odorants which more closely mimics the human olfactory system. Cells expressing hOR were cultured on a porous membrane. The membrane was then flipped over and placed between two compartments. The upper compartment is the gaseous part where gaseous odorants are supplied, while the lower compartment is the aqueous part where viable cells are maintained in the liquid medium. Using this simple microfluidic device, we were able to detect gaseous odorant molecules by a fluorescence signal. The fluorescence signal was generated by calcium influx resulting from the interaction between odorant molecules and the hOR. The system allowed detection of gaseous odorant molecules in real-time, and the findings showed that the fluorescence responses increased dose-dependently in the range of 0-2 ppm odorant. In addition, the system can discriminate among gaseous odorant molecules. This microfluidic system closely mimics the human olfactory system in the sense that the submerged cells detect gaseous odorants.

  7. Fluorescence resonance energy transfer biosensors that detect Ran conformational changes and a Ran x GDP-importin-beta -RanBP1 complex in vitro and in intact cells.

    Science.gov (United States)

    Plafker, Kendra; Macara, Ian G

    2002-08-16

    The Ran GTPase plays a central role in nucleocytoplasmic transport. Association of Ran x GTP with transport carriers (karyopherins) triggers the loading/unloading of export or import cargo, respectively. The C-terminal tail of Ran x GTP is deployed in an extended conformation when associated with a Ran binding domain or importins. To monitor tail orientation, a Ran-GFP fusion was labeled with the fluorophore Alexa546. Fluorescence resonance energy transfer (FRET) occurs efficiently between the green fluorescent protein (GFP) and Alexa546 for Ran x GDP and Ran x GTP, suggesting that the tail is tethered in both states. However, Ran x GTP complexes with importin-beta, RanBP1, and Crm1 all show reduced FRET consistent with tail extension. Displacement of the C-terminal tail of Ran by karyopherins may be a general mechanism to facilitate RanBP1 binding. A Ran x GDP-RanBP1-importin-beta complex also displayed a low FRET signal. To detect this complex in vivo, a bipartite biosensor consisting of Ran-Alexa546 plus GST-GFP-RanBP1, was co-injected into the cytoplasm of cells. The Ran redistributed predominantly to the nucleus, and RanBP1 remained cytoplasmic. Nonetheless, a robust cytoplasmic FRET signal was detectable, which suggests that a significant fraction of cytoplasmic Ran.GDP may exist in a ternary complex with RanBP1 and importins.

  8. Zn1‑x Gd x S (x = 0.1, 0.2 and 0.3) nanoparticles for magnetic resonance imaging and optical fluorescence imaging

    Science.gov (United States)

    Venkatesha, N.; Poojar, Pavan; Qurishi, Yasrib; Geethanath, Sairam; Srivastava, Chandan

    2017-03-01

    This work proposes chemically synthesized Gd doped ZnS nanoparticles based system for potential use as contrast enhancing agent for both optical fluorescence imaging and magnetic resonance imaging. Two different Gd doped ZnS nanoparticle systems were synthesized. These systems were (i) graphene oxide–Zn1‑x Gd x S (x = 0.1, 0.2 and 0.3) nanoparticle composites and (ii) chitosan coated Zn1‑x Gd x S (x = 0.1, 0.2 and 0.3) nanoparticles. Gd formed solid solution with ZnS in all the six as-synthesized samples. Gd doped ZnS nanoparticles in both cases exhibited both longitudinal and transverse relaxivity values. A clear dependence the relaxivity values on the composition of the nanoparticles and the nanoparticle environment (presence and absence of graphene oxide) was observed. Between the two cases, values of the longitudinal and transverse relaxivity were higher for the graphene oxide–Zn1‑x Gd x S composites. It is also shown that Gd doped ZnS nanoparticle can be used for fluorescence imaging also. Gd doped ZnS nanoparticle exhibited biocompatibility towards the MCF-7 cell line.

  9. Application of a real-time fluorescence resonance energy transfer polymerase chain reaction assay with melting curve analysis for the detection of Paragonimus heterotremus eggs in the feces of experimentally infected cats.

    Science.gov (United States)

    Tantrawatpan, Chairat; Intapan, Pewpan M; Thanchomnang, Tongjit; Sanpool, Oranuch; Janwan, Penchom; Lulitanond, Viraphong; Anamnart, Witthaya; Maleewong, Wanchai

    2013-09-01

    Paragonimus heterotremus is a medically important lung fluke that causes human and animal paragonimiasis in Southeast Asia, including Thailand. In the current study, a real-time fluorescence resonance energy transfer polymerase chain reaction (real-time FRET PCR) with melting curve analysis was developed and evaluated to detect P. heterotremus eggs in the feces of experimentally infected cats. The detection limit of this method for the P. heterotremus DNA sequence was 3 × 10(2) copies of the positive control plasmid and 10(-3) ng of P. heterotremus genomic DNA. The assay system could detect 10 eggs of P. heterotremus per gram of cat feces. No fluorescence signal was observed when DNA purified from 16 other organisms or genomic DNA from cats and human beings were tested. Real-time FRET PCR yielded positive results for all fecal samples from 17 P. heterotremus-infected cats and showed a negative relationship (r = -0.852, P analysis. This assay can be useful for the detection of, and epidemiological studies on, P. heterotremus infection in endemic areas.

  10. Detection of Babesia canis vogeli and Hepatozoon canis in canine blood by a single-tube real-time fluorescence resonance energy transfer polymerase chain reaction assay and melting curve analysis.

    Science.gov (United States)

    Kongklieng, Amornmas; Intapan, Pewpan M; Boonmars, Thidarut; Thanchomnang, Tongjit; Janwan, Penchom; Sanpool, Oranuch; Lulitanond, Viraphong; Taweethavonsawat, Piyanan; Chungpivat, Sudchit; Maleewong, Wanchai

    2015-03-01

    A real-time fluorescence resonance energy transfer polymerase chain reaction (qFRET PCR) coupled with melting curve analysis was developed for detection of Babesia canis vogeli and Hepatozoon canis infections in canine blood samples in a single tube assay. The target of the assay was a region within the 18S ribosomal RNA gene amplified in either species by a single pair of primers. Following amplification from the DNA of infected dog blood, a fluorescence melting curve analysis was done. The 2 species, B. canis vogeli and H. canis, could be detected and differentiated in infected dog blood samples (n = 37) with high sensitivity (100%). The detection limit for B. canis vogeli was 15 copies of a positive control plasmid, and for H. canis, it was 150 copies of a positive control plasmid. The assay could simultaneously distinguish the DNA of both parasites from the DNA of controls. Blood samples from 5 noninfected dogs were negative, indicating high specificity. Several samples can be run at the same time. The assay can reduce misdiagnosis and the time associated with microscopic examination, and is not prone to the carryover contamination associated with the agarose gel electrophoresis step of conventional PCR. In addition, this qFRET PCR method would be useful to accurately determine the range of endemic areas or to discover those areas where the 2 parasites co-circulate.

  11. Rapid detection of the factor XIII Val34Leu (163 G-->T) polymorphism by real-time PCR using fluorescence resonance energy transfer detection and melting curve analysis.

    Science.gov (United States)

    Shemirani, Amir H; Muszbek, László

    2004-01-01

    The Val34Leu polymorphism in the A subunit of blood coagulation factor XIII (FXIII-A) is located in the activation peptide, just three amino acids upstream of the thrombin cleavage site. The Val-->Leu replacement accelerates the rate of the proteolytic activation of FXIII and it seems to provide protection against myocardial infarction. Methods available for the assessment of the FXIII-A Val34Leu polymorphism are rather time-consuming, laborious and not easily applicable for large-scale studies. In this study a new method based on real-time PCR with fluorescence resonance energy transfer (FRET) detection and melting curve analysis was developed. The rapid, simple method was adapted to the widely used real-time PCR instrument, LightCycler (Roche Diagnostics). The results showed 100% coincidence with those obtained by the traditional PCR-restriction fragment length polymorphism (RFLP) assay and fluorescent DNA sequencing. Using this method, an allele frequency of 24.2% was obtained (n=113), which well agrees with the allele frequency obtained by PCR-RFLP on a different group of the same ethnic Hungarian population (25.9%).

  12. Electron spin resonance, thermoluminescence and fluorescence studies on BaSO{sub 4}:Eu and BaSO{sub 4}:Eu, P thermoluminescent phosphors

    Energy Technology Data Exchange (ETDEWEB)

    Gundo Rao, T.K.; Shinde, S.S.; Bhatt, B.C.; Srivastava, J.K.; Nambi, K.S.V. [Regional Sophisticated Instrumentation Centre, Indian Inst. of Technol., Bombay (India)

    1995-08-07

    The paramagnetic radicals formed in the TL phosphors BaSO{sub 4}:Eu and BaSO{sub 4}:Eu, P have been studied using the technique of ESR. Two radicals assigned to SO{sub 2}{sup -}, were observed at room temperature. One of them exhibits a hyperfine interaction with a nearby cation. Step annealing measurements suggest a connection between these radicals and the TL glow peaks at 170 and 215 degrees C. Co-doping BaSO{sub 4}:Eu with phosphorus enhanced its TL efficiency. ESR and fluorescence emission measurements indicate that the enhancement results from an increased intake of divalent europium into the lattice. (author)

  13. Rapid detection and differentiation of Clonorchis sinensis and Opisthorchis viverrini eggs in human fecal samples using a duplex real-time fluorescence resonance energy transfer PCR and melting curve analysis.

    Science.gov (United States)

    Sanpool, Oranuch; Intapan, Pewpan M; Thanchomnang, Tongjit; Janwan, Penchom; Lulitanond, Viraphong; Doanh, Pham Ngoc; Van Hien, Hoang; Dung, Do Trung; Maleewong, Wanchai; Nawa, Yukifumi

    2012-07-01

    We developed a single step duplex real-time fluorescence resonance energy transfer (FRET) PCR merged with melting curve analysis for the fast detection and differentiation of Clonorchis sinensis and Opisthorchis viverrini eggs in human fecal samples. Two species of mitochondrial NADH dehydrogenase subunit 2 (nad2) DNA elements, the 165-bp nad2 product of C. sinensis and the 209-bp nad2 product of O. viverrini, were amplified by species-specific primers, and the fluorescence melting curve analyses were generated from hybrid of amplicons and two pairs of species-specific fluorophore-labeled probes. By their different fluorescence channels and melting temperatures, both C. sinensis and O. viverrini eggs in infected human fecal samples were detected and differentiated with high (100%) sensitivity and specificity. Detection limit was as little as a single C. sinensis egg and two O. viverrini eggs in 100 mg of fecal sample. The assay could distinguish the DNA of both parasites from the DNA of negative fecal samples and fecal samples with other parasitosis, as well as from the well-defined genomic DNA of human leukocytes and other parasites. It can reduce labor time of microscopic examination and is not prone to carry over contamination of agarose electrophoresis. Our duplex real-time FRET PCR method would be useful to determine the accurate range of endemic areas and/or to discover the co-endemic areas of two liver flukes, C. sinensis and O. viverrini, in Asia. This method also would be helpful for the differential diagnosis of the suspected cases of liver fluke infections among travelers who had visited the endemic countries of those parasites.

  14. A dual-modal magnetic nanoparticle probe for preoperative and intraoperative mapping of sentinel lymph nodes by magnetic resonance and near infrared fluorescence imaging.

    Science.gov (United States)

    Zhou, Zhengyang; Chen, Hongwei; Lipowska, Malgorzata; Wang, Liya; Yu, Qiqi; Yang, Xiaofeng; Tiwari, Diana; Yang, Lily; Mao, Hui

    2013-07-01

    The ability to reliably detect sentinel lymph nodes for sentinel lymph node biopsy and lymphadenectomy is important in clinical management of patients with metastatic cancers. However, the traditional sentinel lymph node mapping with visible dyes is limited by the penetration depth of light and fast clearance of the dyes. On the other hand, sentinel lymph node mapping with radionucleotide technique has intrinsically low spatial resolution and does not provide anatomic details in the sentinel lymph node mapping procedure. This work reports the development of a dual modality imaging probe with magnetic resonance and near infrared imaging capabilities for sentinel lymph node mapping using magnetic iron oxide nanoparticles (10 nm core size) conjugated with a near infrared molecule with emission at 830 nm. Accumulation of magnetic iron oxide nanoparticles in sentinel lymph nodes leads to strong T2 weighted magnetic resonance imaging contrast that can be potentially used for preoperative localization of sentinel lymph nodes, while conjugated near infrared molecules provide optical imaging tracking of lymph nodes with a high signal to background ratio. The new magnetic nanoparticle based dual imaging probe exhibits a significant longer lymph node retention time. Near infrared signals from nanoparticle conjugated near infrared dyes last up to 60 min in sentinel lymph node compared to that of 25 min for the free near infrared dyes in a mouse model. Furthermore, axillary lymph nodes, in addition to sentinel lymph nodes, can be also visualized with this probe, given its slow clearance and sufficient sensitivity. Therefore, this new dual modality imaging probe with the tissue penetration and sensitive detection of sentinel lymph nodes can be applied for preoperative survey of lymph nodes with magnetic resonance imaging and allows intraoperative sentinel lymph node mapping using near infrared optical devices.

  15. Photonic Crystal Surfaces as a General Purpose Platform for Label-Free and Fluorescent Assays.

    Science.gov (United States)

    Cunningham, Brian T

    2010-04-01

    Photonic crystal surfaces can be designed to provide a wide range of functions that are used to perform biochemical and cell-based assays. Detection of the optical resonant reflections from photonic crystal surfaces enables high sensitivity label-free biosensing, while the enhanced electromagnetic fields that occur at resonant wavelengths can be used to enhance the detection sensitivity of any surface-based fluorescence assay. Fabrication of photonic crystals from inexpensive plastic materials over large surface areas enables them to be incorporated into standard formats that include microplates, microarrays, and microfluidic channels. This report reviews the design of photonic crystal biosensors, their associated detection instrumentation, and biological applications. Applications including small molecule high throughput screening, cell membrane integrin activation, gene expression analysis, and protein biomarker detection are highlighted. Recent results in which photonic crystal surfaces are used for enhancing the detection of Surface-Enhanced Raman Spectroscopy, and the development of high resolution photonic crystal-based laser biosensors are also described.

  16. Fluorescence spectroscopy

    DEFF Research Database (Denmark)

    Bagatolli, Luis

    2016-01-01

    Fluorescence spectroscopy is a powerful experimental tool used by scientists from many disciplines. During the last decades there have been important developments on distinct fluorescence methods, particularly those related to the study of biological phenomena. This chapter discusses the foundati......Fluorescence spectroscopy is a powerful experimental tool used by scientists from many disciplines. During the last decades there have been important developments on distinct fluorescence methods, particularly those related to the study of biological phenomena. This chapter discusses...

  17. Fluorescence resonance energy transfer measured by spatial photon migration in CdSe-ZnS quantum dots colloidal systems as a function of concentration

    Energy Technology Data Exchange (ETDEWEB)

    Azevedo, G.; Monte, A. F. G.; Reis, A. F.; Messias, D. N. [Laboratório de Espectroscopia Óptica, Instituto de Física, Universidade Federal de Uberlândia, Uberlândia, MG 38400-902 (Brazil)

    2014-11-17

    The study of the spatial photon migration as a function of the concentration brings into attention the problem of the energy transfer in quantum dot embedded systems. By measuring the photon propagation and its spatial dependence, it is possible to understand the whole dynamics in a quantum dot system, and also improve their concentration dependence to maximize energy propagation due to radiative and non-radiative processes. In this work, a confocal microscope was adapted to scan the spatial distribution of photoluminescence from CdSe-ZnS core-shell quantum dots in colloidal solutions. The energy migration between the quantum dots was monitored by the direct measurement of the photon diffusion length, according to the diffusion theory. We observed that the photon migration length decreases by increasing the quantum dot concentration, this kind of behavior has been regarded as a signature of Förster resonance energy transfer in the system.

  18. Cell-Based Biosensors Principles and Applications

    CERN Document Server

    Wang, Ping

    2009-01-01

    Written by recognized experts the field, this leading-edge resource is the first book to systematically introduce the concept, technology, and development of cell-based biosensors. You find details on the latest cell-based biosensor models and novel micro-structure biosensor techniques. Taking an interdisciplinary approach, this unique volume presents the latest innovative applications of cell-based biosensors in a variety of biomedical fields. The book also explores future trends of cell-based biosensors, including integrated chips, nanotechnology and microfluidics. Over 140 illustrations hel

  19. Near Infrared Microspectroscopy, Fluorescence Microspectroscopy, Infrared Chemical Imaging and High Resolution Nuclear Magnetic Resonance Analysis of Soybean Seeds, Somatic Embryos and Single Cells

    CERN Document Server

    Baianu, I C; Hofmann, N E; Korban, S S; Lozano, P; You, T; AOCS 94th Meeting, Kansas

    2002-01-01

    Novel methodologies are currently being developed and established for the chemical analysis of soybean seeds, embryos and single cells by Fourier Transform Infrared (FT-IR), Fourier Transform Near Infrared (FT-NIR) Microspectroscopy, Fluorescence and High-Resolution NMR (HR-NMR). The first FT-NIR chemical images of biological systems approaching one micron resolution are presented here. Chemical images obtained by FT-NIR and FT-IR Microspectroscopy are presented for oil in soybean seeds and somatic embryos under physiological conditions. FT-NIR spectra of oil and proteins were obtained for volumes as small as two cubic microns. Related, HR-NMR analyses of oil contents in somatic embryos are also presented here with nanoliter precision. Such 400 MHz 1H NMR analyses allowed the selection of mutagenized embryos with higher oil content (e.g. ~20%) compared to non-mutagenized control embryos. Moreover, developmental changes in single soybean seeds and/or somatic embryos may be monitored by FT-NIR with a precision ...

  20. Fourier Transform Near Infrared Microspectroscopy, Infrared Chemical Imaging, High-Resolution Nuclear Magnetic Resonance and Fluorescence Microspectroscopy Detection of Single Cancer Cells and Single Viral Particles

    CERN Document Server

    Baianu,I C; Hofmann, N E; Korban, S S; Lozano, P; You, T

    2004-01-01

    Single Cancer Cells from Human tumors are being detected and imaged by Fourier Transform Infrared (FT-IR), Fourier Transform Near Infrared (FT-NIR)Hyperspectral Imaging and Fluorescence Correlation Microspectroscopy. The first FT-NIR chemical, microscopic images of biological systems approaching one micron resolution are here reported. Chemical images obtained by FT-NIR and FT-IR Microspectroscopy are also presented for oil in soybean seeds and somatic embryos under physiological conditions. FT-NIR spectra of oil and proteins were obtained for volumes as small as two cubic microns. Related, HR-NMR analyses of oil contents in somatic embryos as well as 99% accurate calibrations are also presented here with nanoliter precision. Such high-resolution, 400 MHz H-1 NMR analyses allowed the selection of mutagenized embryos with higher oil content (e.g. >~20%) compared to the average levels in non-mutagenized control embryos. Moreover, developmental changes in single soybean seeds and/or somatic embryos may be monito...

  1. Comparative Fluorescence Resonance Energy-Transfer Study in Pluronic Triblock Copolymer Micelle and Niosome Composed of Biological Component Cholesterol: An Investigation of Effect of Cholesterol and Sucrose on the FRET Parameters.

    Science.gov (United States)

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

    2016-01-14

    The formation of pluronic triblock copolymer (F127)-cholesterol-based niosome and its interaction with sugar (sucrose) molecules have been investigated. The morphology of F127-cholesterol -based niosome in the presence of sucrose has been successfully demonstrated using dynamic light scattering (DLS) and transmission electron microscopic (TEM) techniques. The DLS profiles and TEM images clearly suggest that the size of the niosome aggregates increases significantly in the presence of sucrose. In addition to structural characterization, a detailed comparative fluorescence resonance energy transfer (FRET) study has been carried out in these F127-containing aggregates, involving coumarin 153 (C153) as donor (D) and rhodamine 6G (R6G) as an acceptor (A) to monitor the dynamic heterogeneity of the systems. Besides, time-resolved anisotropy and fluorescence correlation spectroscopy measurements have been carried out to monitor the rotational and lateral diffusion motion in these F127-cholesterol-based aggregates using C153 and R6G, respectively. During the course of FRET study, we have observed multiple time constants of FRET inside the F127-cholesterol-based niosomes in contrast with the F127 micelle. This corresponds to the presence of more than one preferential donor-acceptor (D-A) distance in niosomes than in F127 micelle. FRET has also been successfully used to probe the effect of sucrose on the morphology of F127-cholesterol-based niosome. In the presence of sucrose, the time constant of FRET further increases as the D-A distances increase in sucrose-decorated niosome. Finally, the excitation-wavelength-dependent FRET studies have indicated that as the excitation of donor molecules varies from 408 to 440 nm the contribution of the faster rise component of the acceptor enhances considerably, which clearly establishes the dynamics heterogeneity of both systems. Our findings also indicate that FRET is completely intravesicular in nature in these block copolymer

  2. An ion-insensitive cAMP biosensor for long term quantitative ratiometric fluorescence resonance energy transfer (FRET) measurements under variable physiological conditions.

    Science.gov (United States)

    Salonikidis, Petrus S; Niebert, Marcus; Ullrich, Tim; Bao, Guobin; Zeug, Andre; Richter, Diethelm W

    2011-07-01

    Ratiometric measurements with FRET-based biosensors in living cells using a single fluorescence excitation wavelength are often affected by a significant ion sensitivity and the aggregation behavior of the FRET pair. This is an important problem for quantitative approaches. Here we report on the influence of physiological ion concentration changes on quantitative ratiometric measurements by comparing different FRET pairs for a cAMP-detecting biosensor. We exchanged the enhanced CFP/enhanced YFP FRET pair of an established Epac1-based biosensor by the fluorophores mCerulean/mCitrine. In the case of enhanced CFP/enhanced YFP, we showed that changes in proton, and (to a lesser extent) chloride ion concentrations result in incorrect ratiometric FRET signals, which may exceed the dynamic range of the biosensor. Calcium ions have no direct, but an indirect pH-driven effect by mobilizing protons. These ion dependences were greatly eliminated when mCerulean/mCitrine fluorophores were used. For such advanced FRET pairs the biosensor is less sensitive to changes in ion concentration and allows consistent cAMP concentration measurements under different physiological conditions, as occur in metabolically active cells. In addition, we verified that the described FRET pair exchange increased the dynamic range of the FRET efficiency response. The time window for stable experimental conditions was also prolonged by a faster biosensor expression rate in transfected cells and a greatly reduced tendency to aggregate, which reduces cytotoxicity. These properties were verified in functional tests in single cells co-expressing the biosensor and the 5-HT(1A) receptor.

  3. Label-free characterization of carbonic anhydrase-novel inhibitor interactions using surface plasmon resonance, isothermal titration calorimetry and fluorescence-based thermal shift assays.

    Science.gov (United States)

    Rogez-Florent, Tiphaine; Duhamel, Laetitia; Goossens, Laurence; Six, Perrine; Drucbert, Anne-Sophie; Depreux, Patrick; Danzé, Pierre-Marie; Landy, David; Goossens, Jean-François; Foulon, Catherine

    2014-01-01

    This work describes the development of biophysical unbiased methods to study the interactions between new designed compounds and carbonic anhydrase II (CAII) enzyme. These methods have to permit both a screening of a series of sulfonamide derivatives and the identification of a lead compound after a thorough study of the most promising molecules. Interactions data were collected using surface plasmon resonance (SPR) and thermal shift assay (TSA). In the first step, experiments were performed with bovine CAII isoform and were extended to human CAII. Isothermal titration calorimetry (ITC) experiments were also conducted to obtain thermodynamics parameters necessary for the processing of the TSA data. Results obtained with this reference methodology demonstrate the effectiveness of SPR and TSA. KD values obtained from SPR data were in perfect accordance with ITC. For TSA, despite the fact that the absolute values of KD were quite different, the same affinity scale was obtained for all compounds. The binding affinities of the analytes studied vary by more than 50 orders of magnitude; for example, the KD value determined by SPR were 6 ± 4 and 299 ± 25 nM for compounds 1 and 3, respectively. This paper discusses some of the theoretical and experimental aspects of the affinity-based methods and evaluates the protein consumption to develop methods for the screening of further new compounds. The double interest of SPR, that is, for screening and for the quick thorough study of the interactions parameters (ka , kd , and KD ), leads us to choose this methodology for the study of new potential inhibitors.

  4. Comparison of two DNA targets for the diagnosis of Toxoplasmosis by real-time PCR using fluorescence resonance energy transfer hybridization probes

    Directory of Open Access Journals (Sweden)

    Ernault Pauline

    2003-05-01

    Full Text Available Abstract Background Toxoplasmosis is an infectious disease caused by the parasitic protozoan Toxoplasma gondii. It is endemic worldwide and, depending on the geographic location, 15 to 85% of the human population are asymptomatically infected. Routine diagnosis is based on serology. The parasite has emerged as a major opportunistic pathogen for immunocompromised patients, in whom it can cause life-threatening disease. Moreover, when a pregnant woman develops a primary Toxoplasma gondii infection, the parasite may be transmitted to the fetus and cause serious damnage. For these two subpopulations, a rapid and accurate diagnosis is required to initiate treatment. Serological diagnosis of active infection is unreliable because reactivation is not always accompanied by changes in antibody levels, and the presence of IgM does not necessarily indicate recent infection. Application of quantitative PCR has evolved as a sensitive, specific, and rapid method for the detection of Toxoplasma gondii DNA in amniotic fluid, blood, tissue samples, and cerebrospinal fluid. Methods Two separate, real-time fluorescence PCR assays were designed and evaluated with clinical samples. The first, targeting the 35-fold repeated B1 gene, and a second, targeting a newly described multicopy genomic fragment of Toxoplasma gondii. Amplicons of different intragenic copies were analyzed for sequence heterogeneity. Results Comparative LightCycler experiments were conducted with a dilution series of Toxoplasma gondii genomic DNA, 5 reference strains, and 51 Toxoplasma gondii-positive amniotic fluid samples revealing a 10 to 100-fold higher sensitivity for the PCR assay targeting the newly described 529-bp repeat element of Toxoplasma gondii. Conclusion We have developed a quantitative LightCycler PCR protocol which offer rapid cycling with real-time, sequence-specific detection of amplicons. Results of quantitative PCR demonstrate that the 529-bp repeat element is repeated more

  5. Imaging protein complex formation in the autophagy pathway: analysis of the interaction of LC3 and Atg4BC74A in live cells using Förster resonance energy transfer and fluorescence recovery after photobleaching

    Science.gov (United States)

    Kraft, Lewis J.; Kenworthy, Anne K.

    2012-01-01

    The protein microtubule-associated protein 1, light chain 3 (LC3) functions in autophagosome formation and plays a central role in the autophagy pathway. Previously, we found LC3 diffuses more slowly in cells than is expected for a freely diffusing monomer, suggesting it may constitutively associate with a macromolecular complex containing other protein components of the pathway. In the current study, we used Förster resonance energy transfer (FRET) microscopy and fluorescence recovery after photobleaching (FRAP) to investigate the interactions of LC3 with Atg4BC74A, a catalytically inactive mutant of the cysteine protease involved in lipidation and de-lipidation of LC3, as a model system to probe protein complex formation in the autophagy pathway. We show Atg4BC74A is in FRET proximity with LC3 in both the cytoplasm and nucleus of living cells, consistent with previous biochemical evidence that suggests these proteins directly interact. In addition, overexpressed Atg4BC74A diffuses significantly more slowly than predicted based on its molecular weight, and its translational diffusion coefficient is significantly slowed upon coexpression with LC3 to match that of LC3 itself. Taken together, these results suggest Atg4BC74A and LC3 are contained within the same multiprotein complex and that this complex exists in both the cytoplasm and nucleoplasm of living cells.

  6. Constitutive oligomerization of human D2 dopamine receptors expressed in Spodoptera frugiperda 9 (Sf9) and in HEK293 cells. Analysis using co-immunoprecipitation and time-resolved fluorescence resonance energy transfer.

    Science.gov (United States)

    Gazi, Lucien; López-Giménez, Juan F; Rüdiger, Martin P; Strange, Philip G

    2003-10-01

    Human D2Long (D2L) and D2Short (D2S) dopamine receptor isoforms were modified at their N-terminus by the addition of a human immunodeficiency virus (HIV) or a FLAG epitope tag. The receptors were then expressed in Spodoptera frugiperda 9 (Sf9) cells using the baculovirus system, and their oligomerization was investigated by means of co-immunoprecipitation and time-resolved fluorescence resonance energy transfer (FRET). [3H]Spiperone labelled D2 receptors in membranes prepared from Sf9 cells expressing epitope-tagged D2L or D2S receptors, with a pKd value of approximately 10. Co-immunoprecipitation using antibodies specific for the tags showed constitutive homo-oligomerization of D2L and D2S receptors in Sf9 cells. When the FLAG-tagged D2S and HIV-tagged D2L receptors were co-expressed, co-immunoprecipitation showed that the two isoforms can also form hetero-oligomers in Sf9 cells. Time-resolved FRET with europium and XL665-labelled antibodies was applied to whole Sf9 cells and to membranes from Sf9 cells expressing epitope-tagged D2 receptors. In both cases, constitutive homo-oligomers were revealed for D2L and D2S isoforms. Time-resolved FRET also revealed constitutive homo-oligomers in HEK293 cells expressing FLAG-tagged D2S receptors. The D2 receptor ligands dopamine, R-(-)propylnorapomorphine, and raclopride did not affect oligomerization of D2L and D2S in Sf9 and HEK293 cells. Human D2 dopamine receptors can therefore form constitutive oligomers in Sf9 cells and in HEK293 cells that can be detected by different approaches, and D2 oligomerization in these cells is not regulated by ligands.

  7. Fluorogenic Cell-Based Biosensors for Monitoring Microbes

    Science.gov (United States)

    Curtis, Theresa; Salazar, Noe; Tabb, Joel; Chase, Chris

    2010-01-01

    Fluorogenic cell-based sensor systems for detecting microbes (especially pathogenic ones) and some toxins and allergens are undergoing development. These systems harness the natural signaltransduction and amplification cascades that occur in mast cells upon activation with antigens. These systems include (1) fluidic biochips for automated containment of samples, reagents, and wastes and (2) sensitive, compact fluorometers for monitoring the fluorescent responses of mast cells engineered to contain fluorescent dyes. It should be possible to observe responses within minutes of adding immune complexes. The systems have been shown to work when utilizing either immunoglobulin E (IgE) antibodies or traditionally generated rat antibodies - a promising result in that it indicates that the systems could be developed to detect many target microbes. Chimeric IgE antibodies and rat immunoglobulin G (IgG) antibodies could be genetically engineered for recognizing biological and chemical warfare agents and airborne and food-borne allergens. Genetic engineering efforts thus far have yielded (1) CD14 chimeric antibodies that recognize both Grampositive and Gram-negative bacteria and bind to the surfaces of mast cells, eliciting a degranulation response and (2) rat IgG2a antibodies that act similarly in response to low levels of canine parvovirus.

  8. Cell-Based Therapies for Diabetic Complications

    Science.gov (United States)

    Bernardi, Stella; Severini, Giovanni Maria; Zauli, Giorgio; Secchiero, Paola

    2012-01-01

    In recent years, accumulating experimental evidence supports the notion that diabetic patients may greatly benefit from cell-based therapies, which include the use of adult stem and/or progenitor cells. In particular, mesenchymal stem cells and the circulating pool of endothelial progenitor cells have so far been the most studied populations of cells proposed for the treatment of vascular complications affecting diabetic patients. We review the evidence supporting their use in this setting, the therapeutic benefits that these cells have shown so far as well as the challenges that cell-based therapies in diabetic complications put out. PMID:21822425

  9. Cell-Based Therapies for Diabetic Complications

    Directory of Open Access Journals (Sweden)

    Stella Bernardi

    2012-01-01

    Full Text Available In recent years, accumulating experimental evidence supports the notion that diabetic patients may greatly benefit from cell-based therapies, which include the use of adult stem and/or progenitor cells. In particular, mesenchymal stem cells and the circulating pool of endothelial progenitor cells have so far been the most studied populations of cells proposed for the treatment of vascular complications affecting diabetic patients. We review the evidence supporting their use in this setting, the therapeutic benefits that these cells have shown so far as well as the challenges that cell-based therapies in diabetic complications put out.

  10. A cell-based screen reveals that the albendazole metabolite, albendazole sulfone, targets Wolbachia.

    OpenAIRE

    Serbus, Laura R.; Frederic Landmann; Bray, Walter M.; Pamela M White; Jordan Ruybal; R Scott Lokey; Alain Debec; William Sullivan

    2012-01-01

    Wolbachia endosymbionts carried by filarial nematodes give rise to the neglected diseases African river blindness and lymphatic filariasis afflicting millions worldwide. Here we identify new Wolbachia-disrupting compounds by conducting high-throughput cell-based chemical screens using a Wolbachia-infected, fluorescently labeled Drosophila cell line. This screen yielded several Wolbachia-disrupting compounds including three that resembled Albendazole, a widely used anthelmintic drug that targe...

  11. CdTe量子点-铜酞菁复合体系荧光共振能量转移的研究∗%Fluorescence resonance energy transfer b etween CdTe quantum dots and copp er phthalo cyanine

    Institute of Scientific and Technical Information of China (English)

    何志聪; 李芳; 李牧野; 魏来

    2015-01-01

    The fluorescence resonance energy transfer in CdTe quantum dots (QDs)-copper phthalocyanine (CuPc) is inves-tigated by ultrafast time-resolved spectroscopy technique equipped with femtosecond laser (780 nm, 76 MHz, 130 fs). The results show that the fluorescence lifetime of CdTe QDs decreases with the increase of CuPc concentration, and the energy transfer efficiency is found to increase with the increase of CuPc concentration. Moreover, the influence of the laser excitation power on the energy transfer efficiency is also studied. It is found that transfer efficiency decreases as excitation laser power increases, the physical mechanism is the thermal activation in the high power and the excited state transitions of high order induced by two-photon. The energy transfer efficiency can reach 43.8%, when the laser power is 200 mW, via two-photon excitation. This study indicates that the CdTe QDs-CuPc composite system has high potential as the third generation of photosensitizers.%以波长为780 nm、重复频率为76 MHz、脉宽为130 fs的飞秒激光作为激发光源,采用超快时间分辨光谱技术研究了CdTe量子点-铜酞菁复合体系的荧光共振能量转移.实验结果表明,在780 nm的双光子激发条件下,复合体系中CdTe量子点的荧光寿命随着铜酞菁溶液浓度的增加而减少,荧光共振能量转移效率增加.同时也研究了激发功率对荧光共振能量转移效率的影响.结果表明,随着激发光功率的增加,复合体系溶液中CdTe量子点的荧光寿命增加,荧光共振能量转移效率减小,其物理机理是因为高激发功率下的热效应和由双光子诱导的高阶激发态的跃迁.当激发光功率为200 mW时,双光子荧光共振能量转移效率为43.8%.研究表明CdTe量子点-铜酞菁复合体系是非常有潜力的第三代光敏剂.

  12. Fluorescent microspheres

    Science.gov (United States)

    Rembaum, A.

    1978-01-01

    Latex particles with attached antibodies have potential biochemical and environmental applications. Human red blood cells and lymphocytes have been labeled with fluorescent microspheres by either direct or indirect immunological technique. Immunolatex spheres can also be used for detecting and localizing specific cell surface receptors. Hormones and toxins may also be bondable.

  13. A magnetic cell-based sensor.

    Science.gov (United States)

    Wang, Hua; Mahdavi, Alborz; Tirrell, David A; Hajimiri, Ali

    2012-11-07

    Cell-based sensing represents a new paradigm for performing direct and accurate detection of cell- or tissue-specific responses by incorporating living cells or tissues as an integral part of a sensor. Here we report a new magnetic cell-based sensing platform by combining magnetic sensors implemented in the complementary metal-oxide-semiconductor (CMOS) integrated microelectronics process with cardiac progenitor cells that are differentiated directly on-chip. We show that the pulsatile movements of on-chip cardiac progenitor cells can be monitored in a real-time manner. Our work provides a new low-cost approach to enable high-throughput screening systems as used in drug development and hand-held devices for point-of-care (PoC) biomedical diagnostic applications.

  14. Stem cell-based approaches in dentistry

    OpenAIRE

    Mitsiadis, T A; Orsini, G.; Jimenez-Rojo, L

    2015-01-01

    Repair of dental pulp and periodontal lesions remains a major clinical challenge. Classical dental treatments require the use of specialised tissue-adapted materials with still questionable efficacy and durability. Stem cell-based therapeutic approaches could offer an attractive alternative in dentistry since they can promise physiologically improved structural and functional outcomes. These therapies necessitate a sufficient number of specific stem cell populations for implantation. Dental m...

  15. Cell based assay for hypoglycemic drugs screening

    Institute of Scientific and Technical Information of China (English)

    LiZHANG; Juan-juanHU; Guan-huaDU

    2004-01-01

    OBJECTIVE: To establish a cell based assay for hypoglyc emicdrugs. METHODS: The five cell lines, BALB/c3T3, HepG2, NIH3T3, Be17402, and L929 were incubated with insulin (0-125n mol/L) for 48 h. Their sensitivities to insulin were studied by detecting glucose consumption. The dose-response and time-response relationship between the sensitive cell line (BALB/c 3T3)

  16. Ontology for cell-based geographic information

    Science.gov (United States)

    Zheng, Bin; Huang, Lina; Lu, Xinhai

    2009-10-01

    Inter-operability is a key notion in geographic information science (GIS) for the sharing of geographic information (GI). That requires a seamless translation among different information sources. Ontology is enrolled in GI discovery to settle the semantic conflicts for its natural language appearance and logical hierarchy structure, which are considered to be able to provide better context for both human understanding and machine cognition in describing the location and relationships in the geographic world. However, for the current, most studies on field ontology are deduced from philosophical theme and not applicable for the raster expression in GIS-which is a kind of field-like phenomenon but does not physically coincide to the general concept of philosophical field (mostly comes from the physics concepts). That's why we specifically discuss the cell-based GI ontology in this paper. The discussion starts at the investigation of the physical characteristics of cell-based raster GI. Then, a unified cell-based GI ontology framework for the recognition of the raster objects is introduced, from which a conceptual interface for the connection of the human epistemology and the computer world so called "endurant-occurrant window" is developed for the better raster GI discovery and sharing.

  17. Fluorescence applications in molecular neurobiology.

    Science.gov (United States)

    Taraska, Justin W; Zagotta, William N

    2010-04-29

    Macromolecules drive the complex behavior of neurons. For example, channels and transporters control the movements of ions across membranes, SNAREs direct the fusion of vesicles at the synapse, and motors move cargo throughout the cell. Understanding the structure, assembly, and conformational movements of these and other neuronal proteins is essential to understanding the brain. Developments in fluorescence have allowed the architecture and dynamics of proteins to be studied in real time and in a cellular context with great accuracy. In this review, we cover classic and recent methods for studying protein structure, assembly, and dynamics with fluorescence. These methods include fluorescence and luminescence resonance energy transfer, single-molecule bleaching analysis, intensity measurements, colocalization microscopy, electron transfer, and bimolecular complementation analysis. We present the principles of these methods, highlight recent work that uses the methods, and discuss a framework for interpreting results as they apply to molecular neurobiology.

  18. Sensitive-cell-based fish chromatophore biosensor

    Science.gov (United States)

    Plant, Thomas K.; Chaplen, Frank W.; Jovanovic, Goran; Kolodziej, Wojtek; Trempy, Janine E.; Willard, Corwin; Liburdy, James A.; Pence, Deborah V.; Paul, Brian K.

    2004-07-01

    A sensitive biosensor (cytosensor) has been developed based on color changes in the toxin-sensitive colored living cells of fish. These chromatophores are highly sensitive to the presence of many known and unknown toxins produced by microbial pathogens and undergo visible color changes in a dose-dependent manner. The chromatophores are immobilized and maintained in a viable state while potential pathogens multiply and fish cell-microbe interactions are monitored. Low power LED lighting is used to illuminate the chromatophores which are magnified using standard optical lenses and imaged onto a CCD array. Reaction to toxins is detected by observing changes is the total area of color in the cells. These fish chromatophores are quite sensitive to cholera toxin, Staphococcus alpha toxin, and Bordatella pertussis toxin. Numerous other toxic chemical and biological agents besides bacterial toxins also cause readily detectable color effects in chromatophores. The ability of the chromatophore cell-based biosensor to distinguish between different bacterial pathogens was examined. Toxin producing strains of Salmonella enteritis, Vibrio parahaemolyticus, and Bacillus cereus induced movement of pigmented organelles in the chromatophore cells and this movement was measured by changes in the optical density over time. Each bacterial pathogen elicited this measurable response in a distinctive and signature fashion. These results suggest a chromatophore cell-based biosensor assay may be applicable for the detection and identification of virulence activities associated with certain air-, food-, and water-borne bacterial pathogens.

  19. Photothermal resonance

    DEFF Research Database (Denmark)

    2014-01-01

    The present invention relates to a method for detecting photo-thermal absorbance of a material utilising a mechanically temperature sensitive resonator (20) and a sample being arrange in thermal communication with the temperature sensitive resonator. The present invention further relates to an ap......The present invention relates to a method for detecting photo-thermal absorbance of a material utilising a mechanically temperature sensitive resonator (20) and a sample being arrange in thermal communication with the temperature sensitive resonator. The present invention further relates...... to an apparatus for detecting photo-thermal absorbance of a sample....

  20. A dual mode targeting probe for distinguishing HER2-positive breast cancer cells using silica-coated fluorescent magnetic nanoparticles

    Science.gov (United States)

    Li, Jia; An, Yan-Li; Zang, Feng-Chao; Zong, Shen-Fei; Cui, Yi-Ping; Teng, Gao-Jun

    2013-10-01

    We report a composite nanoprobe based on silica-coated magnetic nanoparticles (NPs) for distinguishing breast cancers at different HER2 statuses. The nanoprobe has a core-shell structure, with Fe3O4 NPs as the magnetic core and dye-embedded silica as the fluorescent shell, whose average size is about 150 nm. Besides, the outmost surfaces of the probes were modified with specific antibodies to endow the probe with a targeting ability. With such a structure, the nanoprobe can accomplish dual mode targeting of human breast cancer cells based on fluorescence and magnetic resonance imaging (MRI). In the experiments, three human breast cancer cell lines were used to test the targeting ability of the nanoprobe. Specifically, SKBR3 cells with a high HER2 expression level were used as the model target cells, while MCF7 cells with a lower HER2 expression levels and HER2-negative MDA-MB-231 cells were used as the controls. Both the fluorescence and MRI imaging results confirmed that the nanoprobe can distinguish three cancer cell lines with different HER2 expression levels. With the dual mode imaging and specific targeting properties, we anticipate that the presented nanoprobe may have a great potential in the diagnosis and treatment of cancerous diseases.

  1. Cell-based semiquantitative assay for sulfated glycosaminoglycans facilitating the identification of chondrogenesis.

    Science.gov (United States)

    Yen, Ching-Yu; Wu, Yu-Wei; Hsiung, Chao-Nan; Yeh, Min-I; Lin, Yi-Ming; Lee, Sheng-Yang

    2015-10-01

    Glycosaminoglycans (GAGs), in particular chondroitin sulfate, are an accepted marker of chondrogenic cells. In this study, a cell-based sulfated GAG assay for identifying the chondrogenesis of mesenchymal stem cells was developed. Based on fluorescent staining using safranin O and 4',6-diamidino-2-phenylindole (DAPI), this method was highly sensitive. The results were both qualitative and quantitative. The method is suitable for identifying the chondrogenic process and also for screening compounds. The method may be helpful for discovering novel bioactive compounds for cartilage regeneration.

  2. Nanoantenna array-induced fluorescence enhancement and reduced lifetimes

    DEFF Research Database (Denmark)

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

    2008-01-01

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

  3. Fluorescence of Alexa Fluor dye tracks protein folding

    NARCIS (Netherlands)

    Lindhoud, S.; Westphal, A.H.; Borst, J.W.; Visser, A.J.W.G.; Mierlo, van C.P.M.

    2012-01-01

    Fluorescence spectroscopy is an important tool for the characterization of protein folding. Often, a protein is labeled with appropriate fluorescent donor and acceptor probes and folding-induced changes in Förster Resonance Energy Transfer (FRET) are monitored. However, conformational changes of the

  4. Binding of several benzodiazepines to bovine serum albumin: Fluorescence study

    Science.gov (United States)

    Machicote, Roberta G.; Pacheco, María E.; Bruzzone, Liliana

    2010-10-01

    The interactions of lorazepam, oxazepam and bromazepam with bovine serum albumin (BSA) were studied by fluorescence spectrometry. The Stern-Volmer quenching constants and corresponding thermodynamic parameters Δ H, Δ G and Δ S were calculated. The binding constants and the number of binding sites were also investigated. The distances between the donor (BSA) and the acceptors (benzodiazepines) were obtained according to fluorescence resonance energy transfer and conformational changes of BSA were observed from synchronous fluorescence spectra.

  5. Nonlinear resonances

    CERN Document Server

    Rajasekar, Shanmuganathan

    2016-01-01

    This introductory text presents the basic aspects and most important features of various types of resonances and anti-resonances in dynamical systems. In particular, for each resonance, it covers the theoretical concepts, illustrates them with case studies, and reviews the available information on mechanisms, characterization, numerical simulations, experimental realizations, possible quantum analogues, applications and significant advances made over the years. Resonances are one of the most fundamental phenomena exhibited by nonlinear systems and refer to specific realizations of maximum response of a system due to the ability of that system to store and transfer energy received from an external forcing source. Resonances are of particular importance in physical, engineering and biological systems - they can prove to be advantageous in many applications, while leading to instability and even disasters in others. The book is self-contained, providing the details of mathematical derivations and techniques invo...

  6. Coupled external cavity photonic crystal enhanced fluorescence.

    Science.gov (United States)

    Pokhriyal, Anusha; Lu, Meng; Ge, Chun; Cunningham, Brian T

    2014-05-01

    We report a fundamentally new approach to enhance fluorescence in which surface adsorbed fluorophore-tagged biomolecules are excited on a photonic crystal surface that functions as a narrow bandwidth and tunable mirror of an external cavity laser. This scheme leads to ∼10× increase in the electromagnetic enhancement factor compared to ordinary photonic crystal enhanced fluorescence. In our experiments, the cavity automatically tunes its lasing wavelength to the resonance wavelength of the photonic crystal, ensuring optimal on-resonance coupling even in the presence of variable device parameters and variations in the density of surface-adsorbed capture molecules. We achieve ∼10(5) × improvement in the limit of detection of a fluorophore-tagged protein compared to its detection on an unpatterned glass substrate. The enhanced fluorescence signal and easy optical alignment make cavity-coupled photonic crystals a viable approach for further reducing detection limits of optically-excited light emitters that are used in biological assays.

  7. Cell-based technologies for Huntington's disease

    Directory of Open Access Journals (Sweden)

    Mônica Santoro Haddad

    Full Text Available ABSTRACT Huntington's disease (HD is a fatal genetic disorder, which causes the progressive breakdown of neurons in the human brain. HD deteriorates human physical and mental abilities over time and has no cure. Stem cell-based technologies are promising novel treatments, and in HD, they aim to replace lost neurons and/or to prevent neural cell death. Herein we discuss the use of human fetal tissue (hFT, neural stem cells (NSCs of hFT origin or embryonic stem cells (ESCs and induced pluripotent stem cells (IPSCs, in clinical and pre-clinical studies. The in vivo use of mesenchymal stem cells (MSCs, which are derived from non-neural tissues, will also be discussed. All these studies prove the potential of stem cells for transplantation therapy in HD, demonstrating cell grafting and the ability to differentiate into mature neurons, resulting in behavioral improvements. We claim that there are still many problems to overcome before these technologies become available for HD patient treatment, such as: a safety regarding the use of NSCs and pluripotent stem cells, which are potentially teratogenic; b safety regarding the transplantation procedure itself, which represents a risk and needs to be better studied; and finally c technical and ethical issues regarding cells of fetal and embryonic origin.

  8. Stem Cell-Based Gene Therapy.

    Science.gov (United States)

    Bagnis; Mannoni

    1997-01-01

    Many researchers and clinicians wonder if gene therapy remains a way to treat genetic or acquired life-threatening diseases. For the last few years, many experimental, pre-clinical, and clinical data have been published showing that it is possible to transfer with relatively high efficiency new genetic information (transgene) in many cells or tissues including both hematopoietic progenitor cells and differentiated cells. Based on experimental works, addition of the normal gene to cells with deletions, mutations, or alterations of the corresponding endogenous one has been shown to reverse the phenotype and to restore (in some case) the functional defect. In spite of very attractive preliminary results, however, suggesting the feasibility and safety of this process, therapeutically efficient gene transfer and expression in targeted cells or tissues must be proven. In this review, we will focus primarily on the attempts to use gene transfer in hematopoietic stem cells as a model for more general genetic manipulations of stem cells. Hematopoietic stem cells are included in a subset of bone marrow, cord blood, or peripheral blood cells identified by the expression of the CD34 antigen on their membrane.

  9. Plasmonic Molecular Nanohybrids—Spectral Dependence of Fluorescence Quenching

    Directory of Open Access Journals (Sweden)

    Maria Olejnik

    2012-01-01

    Full Text Available We demonstrate strong spectral dependence of the efficiency of fluorescence quenching in molecular systems composed of organic dyes and gold nanoparticles. In order to probe the coupling with metallic nanoparticles we use dyes with varied spectral overlap between the plasmon resonance and their absorption. Hybrid molecular structures were obtained via conjugation of metallic nanoparticles with the dyes using biotin-streptavidin linkage. For dyes featuring absorption above the plasmon excitation in gold nanoparticles, laser excitation induces minute changes in the fluorescence intensity and its lifetime for both conjugated and non-conjugated mixtures, which are the reference. In contrast, when the absorption of the dye overlaps with the plasmon resonance, the effect is quite dramatic, reaching 85% and 95% fluorescence quenching for non-conjugated and conjugated mixtures, respectively. The degree of fluorescence quenching strongly depends upon the concentration of metallic nanoparticles. Importantly, the origin of the fluorescence quenching is different in the case of the conjugated mixture, as evidenced by time-resolved fluorescence. For conjugated mixtures of dyes resonant with plasmon, excitation features two-exponential decay. This is in contrast to the single exponential decay measured for the off-resonant configuration. The results provide valuable insight into spectral dependence of the fluorescence quenching in molecular assemblies involving organic dyes and metallic nanoparticles.

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

    Directory of Open Access Journals (Sweden)

    Bobin George Abraham

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

  11. Refractometric Sensing of Heavy Oils in Fluorescent Core Microcapillaries

    Directory of Open Access Journals (Sweden)

    Zamora V.

    2015-03-01

    Full Text Available The refractometric sensing of calibrated heavy oils (density > 1 000 kg/m3 is demonstrated using fluorescent-core microcapillaries. A 25-micron capillary channel was first coated with a high-index layer of fluorescent silicon Quantum Dots (QD. This QD film supports the development of cylindrical Whispering Gallery Mode (WGM resonances inside the capillary. Heavy oils spanning a wide range of refractive index were pumped into the capillary channel, causing large shifts in the fluorescence WGM resonant wavelengths. The sensitivity for heavy oils approached 250 nm per Refractive Index Unit (nm/RIU at the higher oil indices, which is the highest sensitivity so far observed for a refractometric sensor operating in the fluorescence mode. This suggests that fluorescent core microcapillaries may be a viable microfluidic alternative for refractometric or chemical sensing in various stages of oil and gas processing, monitoring and usage.

  12. Fundamentals of fluorescence and fluorescence microscopy.

    Science.gov (United States)

    Wolf, David E

    2013-01-01

    This chapter discusses the fundamental physics of fluorescence. The application of fluorescence to microscopy represents an important transition in the development of microscopy, particularly as it applies to biology. It enables quantitating the amounts of specific molecules within a cell, determining whether molecules are complexing on a molecular level, measuring changes in ionic concentrations within cells and organelles, and measuring molecular dynamics. This chapter also discusses the issues important to quantitative measurement of fluorescence and focuses on four of quantitative measurements of fluorescence--boxcar-gated detection, streak cameras, photon correlation, and phase modulation. Although quantitative measurement presents many pitfalls to the beginner, it also presents significant opportunities to one skilled in the art. This chapter also examines how fluorescence is measured in the steady state and time domain and how fluorescence is applied in the modern epifluorescence microscope.

  13. Refractometric sensing with fluorescent-core microcapillaries.

    Science.gov (United States)

    Manchee, C P K; Zamora, V; Silverstone, J W; Veinot, J G C; Meldrum, A

    2011-10-24

    Capillaries present a promising structure for microfluidic refractive index sensors. We demonstrate a capillary-type fluorescent core microcavity sensor based on whispering gallery mode (WGM) resonances. The device consists of a microcapillary having a layer of fluorescent silicon quantum dots (QDs) coated on the channel surface. The high effective index of the QD layer confines the electric field near the capillary channel and causes the development of WGM resonances in the fluorescence spectrum. Solutions consisting of sucrose dissolved in water were pumped through the capillary while the fluorescence WGMs were measured with a spectrometer. The device showed a refractometric sensitivity of 9.8 nm/RIU (up to 13.8 nm/RIU for higher solution refractive index) and a maximum detection limit of ~7.2 x 10(-3) RIU. Modeling the field inside the capillary structure, which is analogous to a layered hollow ring resonator, shows that sensitivities as high as 100 nm/RIU and detection limits as low as ~10(-5) RIU may be achievable by optimizing the QD film thickness.

  14. Multiplexed magnetic nanoparticle-antibody conjugates (MNPs-ABS) based prognostic detection of ovarian cancer biomarkers, CA-125, β-2M and ApoA1 using fluorescence spectroscopy with comparison of surface plasmon resonance (SPR) analysis.

    Science.gov (United States)

    Pal, Manoj K; Rashid, Mohammad; Bisht, Manisha

    2015-11-15

    A multiplexed MNPs-Abs based fluorescence spectroscopic system in analysis of serum biomarkers; CA-125, β2-M and ApoA1 for the early detection of ovarian cancer was first time proposed. The lowest detection limits measured in multiplexed setup were 0.26 U/mL, 0.55 ng/mL and 7.7 ng/mL respectively for CA-125, β2-M and ApoA1. A comparative real sample analysis of healthy normal (Control), benign and ovarian cancer patients with SPR has also been done to validate the process. Moreover CA-125 detection only confirms 50-60% of early stage disease. This multiplexed system achieved sensitivity and specificity up to 94% and 98% respectively to distinguish early stage ovarian cancer patients from healthy individuals.

  15. Multiquark resonances

    Science.gov (United States)

    Esposito, A.; Pilloni, A.; Polosa, A. D.

    2017-01-01

    Multiquark resonances are undoubtedly experimentally observed. The number of states and the amount of details on their properties have been growing over the years. It is very recent the discovery of two pentaquarks and the confirmation of four tetraquarks, two of which had not been observed before. We mainly review the theoretical understanding of this sector of particle physics phenomenology and present some considerations attempting a coherent description of the so called X and Z resonances. The prominent problems plaguing theoretical models, like the absence of selection rules limiting the number of states predicted, motivate new directions in model building. Data are reviewed going through all of the observed resonances with particular attention to their common features and the purpose of providing a starting point to further research.

  16. Multiquark Resonances

    CERN Document Server

    Esposito, A.; Polosa, A.D.

    2016-01-01

    Multiquark resonances are undoubtedly experimentally observed. The number of states and the amount of details on their properties has been growing over the years. It is very recent the discovery of two pentaquarks and the confirmation of four tetraquarks, two of which had not been observed before. We mainly review the theoretical understanding of this sector of particle physics phenomenology and present some considerations attempting a coherent description of the so called X and Z resonances. The prominent problems plaguing theoretical models, like the absence of selection rules limiting the number of states predicted, motivate new directions in model building. Data are reviewed going through all of the observed resonances with particular attention to their common features and the purpose of providing a starting point to further research.

  17. Resonating Statements

    DEFF Research Database (Denmark)

    Hjelholt, Morten; Jensen, Tina Blegind

    2015-01-01

    IT projects are often complex arrangements of technological components, social actions, and organizational transformation that are difficult to manage in practice. This paper takes an analytical discourse perspective to explore the process of legitimizing IT projects. We introduce the concept...... of resonating statements to highlight how central actors navigate in various discourses over time. Particularly, the statements and actions of an IT project manager are portrayed to show how individuals can legitimize actions by connecting statements to historically produced discourses. The case study...... of an IT project in a Danish local government spans a two-year time period and demonstrates a double-loop legitimization process. First, resonating statements are produced to localize a national IT initiative to support the specificity of a local government discourse. Second, the resonating statements are used...

  18. Principles of fluorescence techniques

    CERN Document Server

    2016-01-01

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

  19. Reviews in fluorescence 2010

    CERN Document Server

    Geddes, Chris D

    2011-01-01

    ""Reviews in Fluorescence 2010"", the seventh volume of the book serial from Springer, serves as a comprehensive collection of current trends and emerging hot topics in the field of fluorescence and closely related disciplines. It summarizes the year's progress in fluorescence and its applications, with authoritative analytical reviews specialized enough to be attractive to professional researchers, yet also appealing to the wider audience of scientists in related disciplines of fluorescence. ""Reviews in Fluorescence"" offers an essential reference material for any lab working in the fluoresc

  20. A Guide to Fluorescent Protein FRET Pairs

    Science.gov (United States)

    Bajar, Bryce T.; Wang, Emily S.; Zhang, Shu; Lin, Michael Z.; Chu, Jun

    2016-01-01

    Förster or fluorescence resonance energy transfer (FRET) technology and genetically encoded FRET biosensors provide a powerful tool for visualizing signaling molecules in live cells with high spatiotemporal resolution. Fluorescent proteins (FPs) are most commonly used as both donor and acceptor fluorophores in FRET biosensors, especially since FPs are genetically encodable and live-cell compatible. In this review, we will provide an overview of methods to measure FRET changes in biological contexts, discuss the palette of FP FRET pairs developed and their relative strengths and weaknesses, and note important factors to consider when using FPs for FRET studies. PMID:27649177

  1. Baryon Resonances

    CERN Document Server

    Oset, E; Sun, Bao Xi; Vacas, M J Vicente; Ramos, A; Gonzalez, P; Vijande, J; Torres, A Martinez; Khemchandani, K

    2009-01-01

    In this talk I show recent results on how many excited baryon resonances appear as systems of one meson and one baryon, or two mesons and one baryon, with the mesons being either pseudoscalar or vectors. Connection with experiment is made including a discussion on old predictions and recent results for the photoproduction of the $\\Lambda(1405)$ resonance, as well as the prediction of one $1/2^+$ baryon state around 1920 MeV which might have been seen in the $\\gamma p \\to K^+ \\Lambda$ reaction.

  2. Baryon Resonances

    Energy Technology Data Exchange (ETDEWEB)

    Oset, E. [Departamento de Fisica Teorica and IFIC, Centro Mixto Universidad de Valencia-CSIC, Institutos de Investigacion de Paterna, Aptdo. 22085, 46071 Valencia (Spain); Sarkar, S. [Variable Energy Cyclotron Centre, 1/AF, Bidhannagar, Kolkata 700064 (India); Sun Baoxi [Institute of Theoretical Physics, College of Applied Sciences, Beijing University of Technology, Beijing 100124 (China); Vicente Vacas, M.J. [Departamento de Fisica Teorica and IFIC, Centro Mixto Universidad de Valencia-CSIC, Institutos de Investigacion de Paterna, Aptdo. 22085, 46071 Valencia (Spain); Ramos, A. [Departament d' Estructura i Constituents de la Materia and Institut de Ciencies del Cosmos, Universitat de Barcelona, 08028 Barcelona (Spain); Gonzalez, P. [Departamento de Fisica Teorica and IFIC, Centro Mixto Universidad de Valencia-CSIC, Institutos de Investigacion de Paterna, Aptdo. 22085, 46071 Valencia (Spain); Vijande, J. [Departamento de Fisica Atomica Molecular y Nuclear and IFIC, Centro Mixto Universidad de Valencia-CSIC, Institutos de Investigacion de Paterna, Aptdo. 22085, 46071 Valencia (Spain); Martinez Torres, A. [Departamento de Fisica Teorica and IFIC, Centro Mixto Universidad de Valencia-CSIC, Institutos de Investigacion de Paterna, Aptdo. 22085, 46071 Valencia (Spain); Khemchandani, K. [Centro de Fisica Computacional, Departamento de Fisica, Universidade de Coimbra, P-3004-516 Coimbra (Portugal)

    2010-04-01

    In this talk I show recent results on how many excited baryon resonances appear as systems of one meson and one baryon, or two mesons and one baryon, with the mesons being either pseudoscalar or vectors. Connection with experiment is made including a discussion on old predictions and recent results for the photoproduction of the {lambda}(1405) resonance, as well as the prediction of one 1/2{sup +} baryon state around 1920 MeV which might have been seen in the {gamma}p{yields}K{sup +}{lambda} reaction.

  3. Neuroaesthetic Resonance

    DEFF Research Database (Denmark)

    Brooks, Anthony Lewis

    2013-01-01

    sessions are achieved via adaptive action-analyzed activities. These interactive virtual environments are designed to empower patients’ creative and/or playful expressions via digital feedback stimuli. Unconscious self- pushing of limits result from innate distractive mechanisms offered by the alternative...... the unencumbered motion-to-computer-generated activities - ‘Music Making’, ‘Painting’, ‘Robotic’ and ‘Video Game’ control. A focus of this position paper is to highlight how Aesthetic Resonance, in this context, relates to the growing body of research on Neuroaesthetics to evolve Neuroaesthetic Resonance....

  4. A Caco-2 cell-based quantitative antioxidant activity assay for antioxidants.

    Science.gov (United States)

    Wan, Hongxia; Liu, Dong; Yu, Xiangying; Sun, Haiyan; Li, Yan

    2015-05-15

    A Caco-2 cell-based antioxidant activity (CAA) assay for quantitative evaluation of antioxidants was developed by optimizing seeding density and culture time of Caco-2 cells, incubation time and concentration of fluorescent probe (2',7'-dichlorofluorescin diacetate, DCFH-DA), incubation way and incubation time of antioxidants (pure phytochemicals) and DCFH-DA with cells, and detection time of fluorescence. Results showed that the CAA assay was of good reproducibility and could be used to evaluate the antioxidant activity of antioxidants at the following conditions: seeding density of 5 × 10(4)/well, cell culture time of 24h, co-incubation of 60 μM DCFH-DA and pure phytochemicals with Caco-2 cells for 20 min and fluorescence recorded for 90 min. Additionally, a significant correlation was observed between CAA values and rat plasma ORAC values following the intake of antioxidants for selected pure phytochemicals (R(2) = 0.815, p < 0.01), demonstrating the good biological relevance of CAA assay.

  5. Autostereogram resonators

    Science.gov (United States)

    Leavey, Sean; Rae, Katherine; Murray, Adam; Courtial, Johannes

    2012-09-01

    Autostereograms, or "Magic Eye" pictures, are repeating patterns designed to give the illusion of depth. Here we discuss optical resonators that create light patterns which, when viewed from a suitable position by a monocular observer, are autostereograms of the three-dimensional shape of one of the mirror surfaces.

  6. Rb optical resonance inside a random porous medium

    CERN Document Server

    Villalba, S; Laliotis, A; Lenci, L; Barreiro, S; Lezama, A

    2012-01-01

    We studied absorption and fluorescence of Rb atoms confined to the interstitial cavities of a random porous glass. Due to the diffusive light propagation in the porous sample, resonant light absorption is almost entirely compensated by atomic fluorescence at low atomic densities. For higher densities, radiation trapping increases the probability of non-radiative decay via atom-wall collisions. A simple connection of the fluorescence/absorption yield to the sample porosity is given.

  7. Fluorescence correlation spectroscopy

    NARCIS (Netherlands)

    Hink, M.A.; Verveer, P.J.

    2015-01-01

    Fluorescence fluctuation spectroscopy techniques allow the quantification of fluorescent molecules present at the nanomolar concentration level. After a brief introduction to the technique, this chapter presents a protocol including background information in order to measure and quantify the molecul

  8. Safe biodegradable fluorescent particles

    Science.gov (United States)

    Martin, Sue I.; Fergenson, David P.; Srivastava, Abneesh; Bogan, Michael J.; Riot, Vincent J.; Frank, Matthias

    2010-08-24

    A human-safe fluorescence particle that can be used for fluorescence detection instruments or act as a safe simulant for mimicking the fluorescence properties of microorganisms. The particle comprises a non-biological carrier and natural fluorophores encapsulated in the non-biological carrier. By doping biodegradable-polymer drug delivery microspheres with natural or synthetic fluorophores, the desired fluorescence can be attained or biological organisms can be simulated without the associated risks and logistical difficulties of live microorganisms.

  9. Fluorescence imaging of soybean flavonol isolines

    Science.gov (United States)

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

    1998-07-01

    region of the spectrum when excited with radiation in the blue region of the spectrum. Thus, green fluorescence emission due to kaempferol glycosides excited by the blue fluorescent compounds with UV excitation (resonance energy excitation) could become a factor in the fluorescence studies of in vivo plants.

  10. Time Resolved Resonance Raman Conference Royal Institution, London United Kingdom,

    Science.gov (United States)

    1983-01-01

    polar molecules. The rejection of fluorescence which is practically inherent _s most if not all resonance Raman experiments was discussed is great ...presented may be devided into two sections. 1) chemical processes presented in the rorninq and 2) biological in the afternoon. The resonance Raman spectra

  11. Cell-based Assays to Identify Inhibitors of Viral Disease

    Science.gov (United States)

    Green, Neil; Ott, Robert D.; Isaacs, Richard J.; Fang, Hong

    2009-01-01

    Background Antagonizing the production of infectious virus inside cells requires drugs that can cross the cell membrane without harming host cells. Objective It is therefore advantageous to establish intracellular potency of anti-viral drug candidates early in the drug-discovery pipeline. Methods To this end, cell-based assays are being developed and employed in high-throughput drug screening, ranging from assays that monitor replication of intact viruses to those that monitor activity of specific viral proteins. While numerous cell-based assays have been developed and investigated, rapid counter screens are also needed to define the specific viral targets of identified inhibitors and to eliminate nonspecific screening hits. Results/Conclusions Here, we describe the types of cell-based assays being used in antiviral drug screens and evaluate the equally important counter screens that are being employed to reach the full potential of cell-based high-throughput screening. PMID:19750206

  12. Fluorescence energy transfer enhancement in aluminum nanoapertures

    CERN Document Server

    de Torres, Juan; Moparthi, Satish Babu; Grigoriev, Victor; Wenger, Jérome

    2015-01-01

    Zero-mode waveguides (ZMWs) are confining light into attoliter volumes, enabling single molecule fluorescence experiments at physiological micromolar concentrations. Among the fluorescence spectroscopy techniques that can be enhanced by ZMWs, F\\"{o}rster resonance energy transfer (FRET) is one of the most widely used in life sciences. Combining zero-mode waveguides with FRET provides new opportunities to investigate biochemical structures or follow interaction dynamics at micromolar concentration with single molecule resolution. However, prior to any quantitative FRET analysis on biological samples, it is crucial to establish first the influence of the ZMW on the FRET process. Here, we quantify the FRET rates and efficiencies between individual donor-acceptor fluorophore pairs diffusing in aluminum zero-mode waveguides. Aluminum ZMWs are important structures thanks to their commercial availability and the large literature describing their use for single molecule fluorescence spectroscopy. We also compare the ...

  13. Novel patient cell-based HTS assay for identification of small molecules for a lysosomal storage disease.

    Directory of Open Access Journals (Sweden)

    Haifeng Geng

    Full Text Available Small molecules have been identified as potential therapeutic agents for lysosomal storage diseases (LSDs, inherited metabolic disorders caused by defects in proteins that result in lysosome dysfunctional. Some small molecules function assisting the folding of mutant misfolded lysosomal enzymes that are otherwise degraded in ER-associated degradation. The ultimate result is the enhancement of the residual enzymatic activity of the deficient enzyme. Most of the high throughput screening (HTS assays developed to identify these molecules are single-target biochemical assays. Here we describe a cell-based assay using patient cell lines to identify small molecules that enhance the residual arylsulfatase A (ASA activity found in patients with metachromatic leukodystrophy (MLD, a progressive neurodegenerative LSD. In order to generate sufficient cell lines for a large scale HTS, primary cultured fibroblasts from MLD patients were transformed using SV40 large T antigen. These SV40 transformed (SV40t cells showed to conserve biochemical characteristics of the primary cells. Using a specific colorimetric substrate para-nitrocatechol sulfate (pNCS, detectable ASA residual activity were observed in primary and SV40t fibroblasts from a MLD patient (ASA-I179S cultured in multi-well plates. A robust fluorescence ASA assay was developed in high-density 1,536-well plates using the traditional colorimetric pNCS substrate, whose product (pNC acts as "plate fluorescence quencher" in white solid-bottom plates. The quantitative cell-based HTS assay for ASA generated strong statistical parameters when tested against a diverse small molecule collection. This cell-based assay approach can be used for several other LSDs and genetic disorders, especially those that rely on colorimetric substrates which traditionally present low sensitivity for assay-miniaturization. In addition, the quantitative cell-based HTS assay here developed using patient cells creates an

  14. Resonance Energy Transfer Molecular Imaging Application in Biomedicine

    Directory of Open Access Journals (Sweden)

    NIE Da-hong1,2;TANG Gang-hua1,3

    2016-11-01

    Full Text Available Resonance energy transfer molecular imaging (RETI can markedly improve signal intensity and tissue penetrating capacity of optical imaging, and have huge potential application in the deep-tissue optical imaging in vivo. Resonance energy transfer (RET is an energy transition from the donor to an acceptor that is in close proximity, including non-radiative resonance energy transfer and radiative resonance energy transfer. RETI is an optical imaging technology that is based on RET. RETI mainly contains fluorescence resonance energy transfer imaging (FRETI, bioluminescence resonance energy transfer imaging (BRETI, chemiluminescence resonance energy transfer imaging (CRETI, and radiative resonance energy transfer imaging (RRETI. RETI is the hot field of molecular imaging research and has been widely used in the fields of biology and medicine. This review mainly focuses on RETI principle and application in biomedicine.

  15. Synthesis and high content cell-based profiling of simplified analogues of the microtubule stabilizer (+)-discodermolide.

    Science.gov (United States)

    Minguez, Jose M; Giuliano, Kenneth A; Balachandran, Raghavan; Madiraju, Charitha; Curran, Dennis P; Day, Billy W

    2002-12-01

    (+)-Discodermolide, a C24:4, trihydroxylated, octamethyl, carbamate-bearing fatty acid lactone originally isolated from a Caribbean sponge, has proven to be the most potent of the microtubule-stabilizing agents. Recent studies suggest that it or its analogues may have advantages over other classes of microtubule-stabilizing agents. (+)-Discodermolide's complex molecular architecture has made structure-activity relationship analysis in this class of compounds a formidable task. The goal of this study was to prepare simplified analogues of (+)-discodermolide and to analyze their biological activities to expand structure-activity relationships. A small library of analogues was prepared wherein the (+)-discodermolide methyl groups at C-14 and C-16 and the C-7 hydroxyl were removed, and the lactone was replaced by simple esters. The library components were analyzed for microtubule-stabilizing actions in vitro, antiproliferative activity against a small panel of human carcinoma cells, and cell signaling, microtubule architecture and mitotic spindle alterations by a multiparameter fluorescence cell-based screening technique. The results show that even drastic structural simplification can lead to analogues with actions related to microtubule targeting and signal transduction, but that these subtle effects were illuminated only through the high information content cell-based screen.

  16. Non-blinking quantum dot with a plasmonic nanoshell resonator

    Science.gov (United States)

    Ji, Botao; Giovanelli, Emerson; Habert, Benjamin; Spinicelli, Piernicola; Nasilowski, Michel; Xu, Xiangzhen; Lequeux, Nicolas; Hugonin, Jean-Paul; Marquier, Francois; Greffet, Jean-Jacques; Dubertret, Benoit

    2015-02-01

    Colloidal semiconductor quantum dots are fluorescent nanocrystals exhibiting exceptional optical properties, but their emission intensity strongly depends on their charging state and local environment. This leads to blinking at the single-particle level or even complete fluorescence quenching, and limits the applications of quantum dots as fluorescent particles. Here, we show that a single quantum dot encapsulated in a silica shell coated with a continuous gold nanoshell provides a system with a stable and Poissonian emission at room temperature that is preserved regardless of drastic changes in the local environment. This novel hybrid quantum dot/silica/gold structure behaves as a plasmonic resonator with a strong Purcell factor, in very good agreement with simulations. The gold nanoshell also acts as a shield that protects the quantum dot fluorescence and enhances its resistance to high-power photoexcitation or high-energy electron beams. This plasmonic fluorescent resonator opens the way to a new family of plasmonic nanoemitters with robust optical properties.

  17. A multifunctional core-shell nanoparticle for dendritic cell-based cancer immunotherapy

    Science.gov (United States)

    Cho, Nam-Hyuk; Cheong, Taek-Chin; Min, Ji Hyun; Wu, Jun Hua; Lee, Sang Jin; Kim, Daehong; Yang, Jae-Seong; Kim, Sanguk; Kim, Young Keun; Seong, Seung-Yong

    2011-10-01

    Dendritic cell-based cancer immunotherapy requires tumour antigens to be delivered efficiently into dendritic cells and their migration to be monitored in vivo. Nanoparticles have been explored as carriers for antigen delivery, but applications have been limited by the toxicity of the solvents used to make nanoparticles, and by the need to use transfection agents to deliver nanoparticles into cells. Here we show that an iron oxide-zinc oxide core-shell nanoparticle can deliver carcinoembryonic antigen into dendritic cells while simultaneously acting as an imaging agent. The nanoparticle-antigen complex is efficiently taken up by dendritic cells within one hour and can be detected in vitro by confocal microscopy and in vivo by magnetic resonance imaging. Mice immunized with dendritic cells containing the nanoparticle-antigen complex showed enhanced tumour antigen specific T-cell responses, delayed tumour growth and better survival than controls.

  18. Preclinical fluorescent mouse models of pancreatic cancer

    Science.gov (United States)

    Bouvet, Michael; Hoffman, Robert M.

    2007-02-01

    Here we describe our cumulative experience with the development and preclinical application of several highly fluorescent, clinically-relevant, metastatic orthotopic mouse models of pancreatic cancer. These models utilize the human pancreatic cancer cell lines which have been genetically engineered to selectively express high levels of the bioluminescent green fluorescent (GFP) or red fluorescent protein (RFP). Fluorescent tumors are established subcutaneously in nude mice, and tumor fragments are then surgically transplanted onto the pancreas. Locoregional tumor growth and distant metastasis of these orthotopic implants occurs spontaneously and rapidly throughout the abdomen in a manner consistent with clinical human disease. Highly specific, high-resolution, real-time visualization of tumor growth and metastasis may be achieved in vivo without the need for contrast agents, invasive techniques, or expensive imaging equipment. We have shown a high correlation between florescent optical imaging and magnetic resonance imaging in these models. Alternatively, transplantation of RFP-expressing tumor fragments onto the pancreas of GFP-expressing transgenic mice may be used to facilitate visualization of tumor-host interaction between the pancreatic tumor fragments and host-derived stroma and vasculature. Such in vivo models have enabled us to serially visualize and acquire images of the progression of pancreatic cancer in the live animal, and to demonstrate the real-time antitumor and antimetastatic effects of several novel therapeutic strategies on pancreatic malignancy. These fluorescent models are therefore powerful and reliable tools with which to investigate human pancreatic cancer and therapeutic strategies directed against it.

  19. Recent Advances in Development of Genetically Encoded Fluorescent Sensors.

    Science.gov (United States)

    Sanford, Lynn; Palmer, Amy

    2017-01-01

    Genetically encoded fluorescent sensors are essential tools in modern biological research, and recent advances in fluorescent proteins (FPs) have expanded the scope of sensor design and implementation. In this review we compare different sensor platforms, including Förster resonance energy transfer (FRET) sensors, fluorescence-modulated single FP-based sensors, translocation sensors, complementation sensors, and dimerization-based sensors. We discuss elements of sensor design and engineering for each platform, including the incorporation of new types of FPs and sensor screening techniques. Finally, we summarize the wide range of sensors in the literature, exploring creative new sensor architectures suitable for different applications.

  20. Demonstration of a visual cell-based assay for screening glucose transporter 4 translocation modulators in real time

    Indian Academy of Sciences (India)

    Maleppillil Vavachan Vijayakumar; Amrendra Kumar Ajay; Manoj Kumar Bhat

    2010-12-01

    Insulin-stimulated translocation of glucose transporter 4 (GLUT4) to cell membrane leading to glucose uptake is the rate-limiting step in diabetes. It is also a defined target of antidiabetic drug research. Existing GLUT4 translocation assays are based on time-consuming immunoassays and are hampered by assay variability and low sensitivity. We describe a real-time, visual, cell-based qualitative GLUT4 translocation assay using CHO-HIRc-myc-GLUT4eGFP cells that stably express myc- and eGFP-tagged GLUT4 in addition to human insulin receptor (HIRc). GLUT4 translocation is visualized by live cell imaging based on GFP fluorescence by employing a cooled charge-coupled device camera attached to a fluorescent microscope. This video imaging method and further quantitative analysis of GLUT4 on the cell membrane provide rapid and foolproof visual evidence that this method is suitable for screening GLUT4 translocation modulators.

  1. Recent Progress on Plasmon-Enhanced Fluorescence

    Directory of Open Access Journals (Sweden)

    Dong Jun

    2015-12-01

    Full Text Available The optically generated collective electron density waves on metal–dielectric boundaries known as surface plasmons have been of great scientific interest since their discovery. Being electromagnetic waves on gold or silver nanoparticle’s surface, localised surface plasmons (LSP can strongly enhance the electromagnetic field. These strong electromagnetic fields near the metal surfaces have been used in various applications like surface enhanced spectroscopy (SES, plasmonic lithography, plasmonic trapping of particles, and plasmonic catalysis. Resonant coupling of LSPs to fluorophore can strongly enhance the emission intensity, the angular distribution, and the polarisation of the emitted radiation and even the speed of radiative decay, which is so-called plasmon enhanced fluorescence (PEF. As a result, more and more reports on surface-enhanced fluorescence have appeared, such as SPASER-s, plasmon assisted lasing, single molecule fluorescence measurements, surface plasmoncoupled emission (SPCE in biological sensing, optical orbit designs etc. In this review, we focus on recent advanced reports on plasmon-enhanced fluorescence (PEF. First, the mechanism of PEF and early results of enhanced fluorescence observed by metal nanostructure will be introduced. Then, the enhanced substrates, including periodical and nonperiodical nanostructure, will be discussed and the most important factor of the spacer between molecule and surface and wavelength dependence on PEF is demonstrated. Finally, the recent progress of tipenhanced fluorescence and PEF from the rare-earth doped up-conversion (UC and down-conversion (DC nanoparticles (NPs are also commented upon. This review provides an introduction to fundamentals of PEF, illustrates the current progress in the design of metallic nanostructures for efficient fluorescence signal amplification that utilises propagating and localised surface plasmons.

  2. Recent Progress on Plasmon-Enhanced Fluorescence

    Science.gov (United States)

    Dong, Jun; Zhang, Zhenglong; Zheng, Hairong; Sun, Mentao

    2015-12-01

    The optically generated collective electron density waves on metal-dielectric boundaries known as surface plasmons have been of great scientific interest since their discovery. Being electromagnetic waves on gold or silver nanoparticle's surface, localised surface plasmons (LSP) can strongly enhance the electromagnetic field. These strong electromagnetic fields near the metal surfaces have been used in various applications like surface enhanced spectroscopy (SES), plasmonic lithography, plasmonic trapping of particles, and plasmonic catalysis. Resonant coupling of LSPs to fluorophore can strongly enhance the emission intensity, the angular distribution, and the polarisation of the emitted radiation and even the speed of radiative decay, which is so-called plasmon enhanced fluorescence (PEF). As a result, more and more reports on surface-enhanced fluorescence have appeared, such as SPASER-s, plasmon assisted lasing, single molecule fluorescence measurements, surface plasmoncoupled emission (SPCE) in biological sensing, optical orbit designs etc. In this review, we focus on recent advanced reports on plasmon-enhanced fluorescence (PEF). First, the mechanism of PEF and early results of enhanced fluorescence observed by metal nanostructure will be introduced. Then, the enhanced substrates, including periodical and nonperiodical nanostructure, will be discussed and the most important factor of the spacer between molecule and surface and wavelength dependence on PEF is demonstrated. Finally, the recent progress of tipenhanced fluorescence and PEF from the rare-earth doped up-conversion (UC) and down-conversion (DC) nanoparticles (NPs) are also commented upon. This review provides an introduction to fundamentals of PEF, illustrates the current progress in the design of metallic nanostructures for efficient fluorescence signal amplification that utilises propagating and localised surface plasmons.

  3. Fluorescent Nanoparticle Uptake for Brain Tumor Visualization

    Directory of Open Access Journals (Sweden)

    Rachel Tréhin

    2006-04-01

    Full Text Available Accurate delineation of tumor margins is vital to the successful surgical resection of brain tumors. We have previously developed a multimodal nanoparticle CLIO-Cy5.5, which is detectable by both magnetic resonance imaging and fluorescence, to assist in intraoperatively visualizing tumor boundaries. Here we examined the accuracy of tumor margin determination of orthotopic tumors implanted in hosts with differing immune responses to the tumor. Using a nonuser-based signal intensity method applied to fluorescent micrographs of 9L gliosarcoma green fluorescent protein (GFP tumors, mean overestimations of 2 and 24 µm were obtained using Cy5.5 fluorescence, compared to the true tumor margin determined by GFP fluorescence, in nude mice and rats, respectively. To resolve which cells internalized the nanoparticle and to quantitate degree of uptake, tumors were disaggregated and cells were analyzed by flow cytometry and fluorescence microscopy. Nanoparticle uptake was seen in both CD11b+ cells (representing activated microglia and macrophages and tumor cells in both animal models by both methods. CD11b+ cells were predominantly found at the tumor margin in both hosts, but were more pronounced at the margin in the rat model. Additional metastatic (CT26 colon and primary (Gli36 glioma brain tumor models likewise demonstrated that the nanoparticle was internalized both by tumor cells and by host cells. Together, these observations suggest that fluorescent nanoparticles provide an accurate method of tumor margin estimation based on a combination of tumor cell and host cell uptake for primary and metastatic tumors in animal model systems and offer potential for clinical translation.

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

    Science.gov (United States)

    Wang, Qing; Zhang, Yongkui; Li, Hui

    2017-03-15

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

  5. Tailoring cyanine dark states for improved optically modulated fluorescence recovery.

    Science.gov (United States)

    Mahoney, Daniel P; Owens, Eric A; Fan, Chaoyang; Hsiang, Jung-Cheng; Henary, Maged M; Dickson, Robert M

    2015-04-02

    Cyanine dyes are well-known for their bright fluorescence and utility in biological imaging. However, cyanines also readily photoisomerize to produce nonemissive dark states. Co-illumination with a secondary, red-shifted light source on-resonance with the longer wavelength absorbing dark state reverses the photoisomerization and returns the cyanine dye to the fluorescent manifold, increasing steady-state fluorescence intensity. Modulation of this secondary light source dynamically alters emission intensity, drastically improving detection sensitivity and facilitating fluorescence signals to be recovered from an otherwise overwhelming background. Red and near-IR emitting cyanine derivatives have been synthesized with varying alkyl chain lengths and halogen substituents to alter dual-laser fluorescence enhancement. Photophysical properties and enhancement with dual laser modulation were coupled with density functional calculations to characterize substituent effects on dark state photophysics, potentially improving detection in high background biological environments.

  6. Stem Cell-Based Therapies in Chagasic Cardiomyopathy

    Directory of Open Access Journals (Sweden)

    Antonio Carlos Campos de Carvalho

    2015-01-01

    Full Text Available Chagas disease is caused by Trypanosoma cruzi and can lead to a dilated cardiomyopathy decades after the prime infection by the parasite. As with other dilated cardiomyopathies, conventional pharmacologic therapies are not always effective and as heart failure progresses patients need heart transplantation. Therefore alternative therapies are highly desirable and cell-based therapies have been investigated in preclinical and clinical studies. In this paper we review the main findings of such studies and discuss future directions for stem cell-based therapies in chronic chagasic cardiomyopathy.

  7. Cell-based biosensors: Towards the development of cellular monitoring

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Cell-based biosensors (CBBs), a research hotspot of biosensors, which treat living cells as sensing elements, can detect the functional information of biologically active analytes. They characterize with high sensitivity, excellent selectivity and rapid response, and have been applied in many fields, such as biomedicine, environmental monitoring and pharmaceutical screening. Recently cell-cultured technology, silicon microfabrication technology and genetic technology have promoted exploration of CBBs dramatically. To elucidate the novel research findings and applications of cell- based biosensors, this paper summarizes various research approaches, presents some challenges and proposes the research trends.

  8. Recent developments in fluorescence-based microscopy applied in biomedical sciences

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The present short review aims to give an overview of the most recent de velopments in fluorescence microscopy and its applications in biomedical science s. Apart from improvements in well-established methods based on conventional fl u orescence microscopy and confocal microscopy (fluorescence in situ hybridisa tion (FISH), tyramide signal amplification (TSA) in immunocytochemistry, new fluorop hores), more recently introduced techniques like fluorescence resonance energy t ransfer (FRET), fluorescence recovery after photobleaching (FRAP), multiphoton m icroscopy and fluorescence correlation spectroscopy (FCS) will be discussed.

  9. Fluorescence antibunching microscopy

    CERN Document Server

    Schwartz, Osip

    2011-01-01

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

  10. LEDs for fluorescence microscopy

    NARCIS (Netherlands)

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

    2004-01-01

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

  11. Fluorescence of atopic allergens

    NARCIS (Netherlands)

    Berrens, L.

    1967-01-01

    Purified atopic allergens have been found to emit flue fluorescence upon irradiation with ultraviolet light of 365 mμ wavelength. The maximum of fluorescence is in the region 445–490 mμ and the intensity is of the same order of magnitude for different atopic allergens. Synthetic model compounds, inc

  12. Global Distribution of Businesses Marketing Stem Cell-Based Interventions.

    Science.gov (United States)

    Berger, Israel; Ahmad, Amina; Bansal, Akhil; Kapoor, Tanvir; Sipp, Douglas; Rasko, John E J

    2016-08-04

    A structured search reveals that online marketing of stem-cell-based interventions is skewed toward developed economies including the United States, Ireland, Australia, and Germany. Websites made broad, imprecise therapeutic claims and frequently failed to detail procedures. Widespread marketing poses challenges to regulators, bioethicists, and those seeking realistic hope from therapies.

  13. Metal-enhanced fluorescence of carbon nanotubes.

    Science.gov (United States)

    Hong, Guosong; Tabakman, Scott M; Welsher, Kevin; Wang, Hailiang; Wang, Xinran; Dai, Hongjie

    2010-11-17

    The photoluminescence (PL) quantum yield of single-walled carbon nanotubes (SWNTs) is relatively low, with various quenching effects by metallic species reported in the literature. Here, we report the first case of metal enhanced fluorescence (MEF) of surfactant-coated carbon nanotubes on nanostructured gold substrates. The photoluminescence quantum yield of SWNTs is observed to be enhanced more than 10-fold. The dependence of fluorescence enhancement on metal-nanotube distance and on the surface plasmon resonance (SPR) of the gold substrate for various SWNT chiralities is measured to reveal the mechanism of enhancement. Surfactant-coated SWNTs in direct contact with metal exhibit strong MEF without quenching, suggesting a small quenching distance for SWNTs on the order of the van der Waals distance, beyond which the intrinsically fast nonradiative decay rate in nanotubes is little enhanced by metal. The metal enhanced fluorescence of SWNTs is attributed to radiative lifetime shortening through resonance coupling of SWNT emission to the reradiating dipolar plasmonic modes in the metal.

  14. Theory of analytical curves in atomic fluorescence flame spectrometry

    NARCIS (Netherlands)

    Hooymayers, H.P.

    1968-01-01

    An explicit expression for the intensity of atomic resonance fluorescence as a function of atomic concentration in a flame is derived under certain idealized conditions. The expression is generally valid for a pure Doppler absorption line profile as well as for a combined Doppler and collisional bro

  15. Experiments with Helmholtz Resonators.

    Science.gov (United States)

    Greenslade, Thomas B., Jr.

    1996-01-01

    Presents experiments that use Helmholtz resonators and have been designed for a sophomore-level course in oscillations and waves. Discusses the theory of the Helmholtz resonator and resonance curves. (JRH)

  16. MRI (Magnetic Resonance Imaging)

    Science.gov (United States)

    ... and Procedures Medical Imaging MRI (Magnetic Resonance Imaging) MRI (Magnetic Resonance Imaging) Share Tweet Linkedin Pin it More sharing options ... usually given through an IV in the arm. MRI Research Programs at FDA Magnetic Resonance Imaging (MRI) ...

  17. Development of a Cell-Based Functional Assay for the Detection of Clostridium botulinum Neurotoxin Types A and E

    Directory of Open Access Journals (Sweden)

    Uma Basavanna

    2013-01-01

    Full Text Available The standard procedure for definitive detection of BoNT-producing Clostridia is a culture method combined with neurotoxin detection using a standard mouse bioassay (MBA. The mouse bioassay is highly sensitive and specific, but it is expensive and time-consuming, and there are ethical concerns due to use of laboratory animals. Cell-based assays provide an alternative to the MBA in screening for BoNT-producing Clostridia. Here, we describe a cell-based assay utilizing a fluorescence reporter construct expressed in a neuronal cell model to study toxin activity in situ. Our data indicates that the assay can detect as little as 100 pM BoNT/A activity within living cells, and the assay is currently being evaluated for the analysis of BoNT in food matrices. Among available in vitro assays, we believe that cell-based assays are widely applicable in high-throughput screenings and have the potential to at least reduce and refine animal assays if not replace it.

  18. Fluorescence and Spectral Imaging

    Directory of Open Access Journals (Sweden)

    Ralph S. DaCosta

    2007-01-01

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

  19. Regenerative feedback resonant circuit

    Science.gov (United States)

    Jones, A. Mark; Kelly, James F.; McCloy, John S.; McMakin, Douglas L.

    2014-09-02

    A regenerative feedback resonant circuit for measuring a transient response in a loop is disclosed. The circuit includes an amplifier for generating a signal in the loop. The circuit further includes a resonator having a resonant cavity and a material located within the cavity. The signal sent into the resonator produces a resonant frequency. A variation of the resonant frequency due to perturbations in electromagnetic properties of the material is measured.

  20. Fluorescent carbon nanowires made by pyrolysis of DNA nanofibers and plasmon-assisted emission enhancement of their fluorescence.

    Science.gov (United States)

    Nakao, Hidenobu; Tokonami, Shiho; Yamamoto, Yojiro; Shiigi, Hiroshi; Takeda, Yoshihiko

    2014-10-14

    We report on a facile method for preparing fluorescent carbon nanowires (CNWs) with pyrolysis of highly aligned DNA nanofibers as carbon sources. Silver nanoparticle (AgNP)-doped CNWs were also produced using pyrolysis of DNA nanofibers with well-attached AgNPs, indicating emission enhancement assisted by localized plasmon resonances.

  1. Highly thermostable fluorescent proteins

    Science.gov (United States)

    Bradbury, Andrew M.; Waldo, Geoffrey S.; Kiss, Csaba

    2011-03-22

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  2. [Development of Fluorescence Sensing Mechanism for Cell Functional Analysis].

    Science.gov (United States)

    Ojida, Akio; Takashima, Ippei

    2016-01-01

      Fluorescence probes are now widely used as indispensable tools in many cell functional analyses. At present, the design of fluorescent probes largely depends on the limited numbers of established sensing mechanisms such as photo-induced electron transfer (PET), photo-induced charge transfer (PCT), and fluorescence resonance energy transfer (FRET). Although these mechanisms are versatile in metal ion sensing, introduction of a new sensing mechanism is highly desirable not only to design a more sophisticated probe with high selectivity and sensitivity but also to expand the diversity of the sensing targets to unveil biological phenomena. In this article, we report the design of dual emission fluorescent probes for metal ions based on a unique fluorescence-sensing mechanism. The fluorescent probes incorporating this sensing mechanism displayed a large emission red-shift on complexation with metal ions such as Cd(II) and Ag(I). X-ray crystallography and theoretical computational calculations of the Cd(II) and Ag(I) complexes revealed that the emission shift was induced by non-coordination contact between the aromatic ring of fluorophore and the metal ion (arene-metal ion contact; AM-contact), which modulates the energy levels of molecular orbitals. The fluorescent probe was successfully applied to in cell ratiometric bioimaging of bioactive hydrogen sulfide (H2S). These successful applications highlight the usefulness of this sensing mechanism in biological fluorescence analysis.

  3. UV fluorescence enhancement from nanostructured aluminum materials

    Science.gov (United States)

    Montanari, Danielle E.; Dean, Nathan; Poston, Pete E.; Blair, Steve; Harris, Joel M.

    2016-09-01

    Interest in label-free detection of biomolecules has given rise to the need for UV plasmonic materials. DNA bases and amino acid residues have electronic resonances in the UV which allow for sensitive detection of these species by surface-enhanced UV fluorescence spectroscopy. Electrochemical roughening has been used extensively to generate plasmonically-active metal surfaces that produce localized enhancement of excitation and emission of electromagnetic radiation from surface-bound molecules. Electrochemically roughened gold and silver surfaces produce enhancement in the visible and near-IR regions, but to the best of our knowledge, application of this technique for producing UV-enhancing substrates has not been reported. Using electropolishing of aluminum, we are able to generate nanostructured surfaces that produce enhanced spectroscopic detection of molecules in the UV. Aluminum is a natural choice for substrate composition as it exhibits a relatively large quality factor in the UV. We have fabricated electropolished aluminum films with nanometer scale roughness and have studied UV-excited fluorescence enhancement from submonolayer coverage of tryptophan on these substrates using a UV-laser based spectrometer. Quantitative dosing by dip-coating was used to deposit known surface concentrations of the aromatic amino acid tryptophan, so that fluorescence enhancement could be evaluated. Compared to a dielectric substrate (surface-oxidized silicon), we observe a 180-fold enhancement in the total fluorescence emitted by tryptophan on electropolished aluminum under photobleaching conditions, allowing detection of sub-monolayer coverages of molecules essential for development of biosensor technologies.

  4. Plasmonic antennas and zero mode waveguides to enhance single molecule fluorescence detection and fluorescence correlation spectroscopy towards physiological concentrations

    CERN Document Server

    Punj, Deep; Moparthi, Satish Babu; de Torres, Juan; Grigoriev, Victor; Rigneault, Hervé; Wenger, Jérôme

    2014-01-01

    Single-molecule approaches to biology offer a powerful new vision to elucidate the mechanisms that underpin the functioning of living cells. However, conventional optical single molecule spectroscopy techniques such as F\\"orster fluorescence resonance energy transfer (FRET) or fluorescence correlation spectroscopy (FCS) are limited by diffraction to the nanomolar concentration range, far below the physiological micromolar concentration range where most biological reaction occur. To breach the diffraction limit, zero mode waveguides and plasmonic antennas exploit the surface plasmon resonances to confine and enhance light down to the nanometre scale. The ability of plasmonics to achieve extreme light concentration unlocks an enormous potential to enhance fluorescence detection, FRET and FCS. Single molecule spectroscopy techniques greatly benefit from zero mode waveguides and plasmonic antennas to enter a new dimension of molecular concentration reaching physiological conditions. The application of nano-optics...

  5. Fluorescent silver nanoparticles via exploding wire technique

    Indian Academy of Sciences (India)

    Alqudami Abdullah; S Annapoorni

    2005-11-01

    Aqueous solution containing spherical silver nanoparticles of 20–80 nm size have been generated using a newly developed novel electro-exploding wire (EEW) technique where thin silver wires have been exploded in double distilled water. Structural properties of the resulted nanoparticles have been studied by means of X-ray diffractometer (XRD) and transmission electron microscopy (TEM). The absorption spectrum of the aqueous solution of silver nanoparticles showed the appearance of a broad surface plasmon resonance (SPR) peak centered at a wavelength of 390 nm. The theoretically generated SPR peak seems to be in good agreement with the experimental one. Strong green fluorescence emission was observed from the water-suspended silver nanoparticles excited with light of wavelengths 340, 360 and 390 nm. The fluorescence of silver nanoparticles could be due to the excitation of the surface plasmon coherent electronic motion with the small size effect and the surface effect considerations.

  6. Potential National Security Applications of Nuclear Resonance Fluorescence Methods

    Energy Technology Data Exchange (ETDEWEB)

    Warren, Glen A.; Peplowski, Patrick N.; Caggiano, Joseph A.

    2009-06-09

    The objective of this report is to document the initial investigation into the possible research issues related to the development of NRF-based national security applications. The report discusses several potential applications ranging from measuring uranium enrichment in UF6 canisters to characterization of gas samples. While these applications are varied, there are only a few research issues that need to be addressed to understand the limitation of NRF in solving these problems. These research issues range from source and detector development to measuring small samples. The next effort is to determine how best to answer the research issues, followed by a prioritization of those questions to ensure that the most important are addressed. These issues will be addressed through either analytical calculations, computer simulations, analysis of previous data or collection of new measurements. It will also be beneficial to conduct a thorough examination of a couple of the more promising applications in order to develop concrete examples of how NRF may be applied in specific situations. The goals are to develop an understanding of whether the application of NRF is limited by technology or physics in addressing national security applications, to gain a motivation to explore those possible applications, and to develop a research roadmap so that those possibilities may be made reality.

  7. Resonance Fluorescence from Semiconductor Quantum Dots: Beyond the Mollow Triplet

    DEFF Research Database (Denmark)

    Lund, Anders Mølbjerg; Nielsen, Per Kær; Lorke, Michael;

    2012-01-01

    analytical theory has been derived, which quantitatively accounts for the appearance and position of the peaks. This theory explains the physics responsible for the multiple peaks. By considering the time-dependent spectrum we demonstrate a time ordering of the side peaks, which is further evidence...

  8. Fluorescent resonant excitation energy transfer in linear polyenes

    Science.gov (United States)

    Das, Mousumi; Ramasesha, S.

    2010-03-01

    We have studied the dynamics of excitation transfer between two conjugated polyene molecules whose intermolecular separation is comparable to the molecular dimensions. We have employed a correlated electron model that includes both the charge-charge, charge-bond, and bond-bond intermolecular electron repulsion integrals. We have shown that the excitation transfer rate varies as inverse square of donor-acceptor separation R-2 rather than as R-6, suggested by the Förster type of dipolar approximation. Our time-evolution study also shows that the orientational dependence on excitation transfer at a fixed short donor-acceptor separation cannot be explained by Förster type of dipolar approximation beyond a certain orientational angle of rotation of an acceptor polyene with respect to the donor polyene. The actual excitation transfer rate beyond a certain orientational angle is faster than the Förster type of dipolar approximation rate. We have also studied the excitation transfer process in a pair of push-pull polyenes for different push-pull strengths. We have seen that, depending on the push-pull strength, excitation transfer could occur to other dipole coupled states. Our study also allows for the excitation energy transfer to optically dark states which are excluded by Förster theory since the one-photon transition intensity to these states (from the ground state) is zero.

  9. A sandwiched microarray platform for benchtop cell-based high throughput screening

    Science.gov (United States)

    Wu, Jinhui; Wheeldon, Ian; Guo, Yuqi; Lu, Tingli; Du, Yanan; Wang, Ben; He, Jiankang; Hu, Yiqiao; Khademhosseini, Ali

    2010-01-01

    The emergence of combinatorial chemistries and the increased discovery of natural compounds have led to the production of expansive libraries of drug candidates and vast numbers of compounds with potentially interesting biological activities. Despite broad interest in high throughput screening (HTS) across varied fields of biological research, there has not been an increase in accessible HTS technologies. Here, we present a simple microarray sandwich system suitable for screening chemical libraries in cell-based assays at the benchtop. The microarray platform delivers chemical compounds to isolated cell cultures by ‘sandwiching’ chemical-laden arrayed posts with cell-seeded microwells. In this way, an array of sealed cell-based assays was generated without cross-contamination between neighboring assays. After chemical exposure, cell viability was analyzed by fluorescence detection of cell viability indicator assays on a per microwell basis in a standard microarray scanner. We demonstrate the efficacy of the system by generating four hits from toxicology screens towards MCF-7 human breast cancer cells. Three of the hits were identified in a combinatorial screen of a library of natural compounds in combination with verapamil, a P-glycoprotein inhibitor. A fourth hit, 9-methoxy-camptothecin, was identified by screening the natural compound library in the absence of verapamil. The method developed here miniaturizes existing HTS systems and enables the screening of a wide array of individual or combinatorial libraries in a reproducible and scalable manner. We anticipate broad application of such a system as it is amenable to combinatorial drug screening in a simple, robust and portable platform. PMID:20965560

  10. Enhancement of cell-based therapeutic angiogenesis using a novel type of injectable scaffolds of hydroxyapatite-polymer nanocomposite microspheres.

    Directory of Open Access Journals (Sweden)

    Yohei Mima

    Full Text Available BACKGROUND: Clinical trials demonstrate the effectiveness of cell-based therapeutic angiogenesis in patients with severe ischemic diseases; however, their success remains limited. Maintaining transplanted cells in place are expected to augment the cell-based therapeutic angiogenesis. We have reported that nano-hydroxyapatite (HAp coating on medical devices shows marked cell adhesiveness. Using this nanotechnology, HAp-coated poly(l-lactic acid (PLLA microspheres, named nano-scaffold (NS, were generated as a non-biological, biodegradable and injectable cell scaffold. We investigate the effectiveness of NS on cell-based therapeutic angiogenesis. METHODS AND RESULTS: Bone marrow mononuclear cells (BMNC and NS or control PLLA microspheres (LA were intramuscularly co-implanted into mice ischemic hindlimbs. When BMNC derived from enhanced green fluorescent protein (EGFP-transgenic mice were injected into ischemic muscle, the muscle GFP level in NS+BMNC group was approximate fivefold higher than that in BMNC or LA+BMNC groups seven days after operation. Kaplan-Meier analysis demonstrated that NS+BMNC markedly prevented hindlimb necrosis (P<0.05 vs. BMNC or LA+BMNC. NS+BMNC revealed much higher induction of angiogenesis in ischemic tissues and collateral blood flow confirmed by three-dimensional computed tomography angiography than those of BMNC or LA+BMNC groups. NS-enhanced therapeutic angiogenesis and arteriogenesis showed good correlations with increased intramuscular levels of vascular endothelial growth factor and fibroblast growth factor-2. NS co-implantation also prevented apoptotic cell death of transplanted cells, resulting in prolonged cell retention. CONCLUSION: A novel and feasible injectable cell scaffold potentiates cell-based therapeutic angiogenesis, which could be extremely useful for the treatment of severe ischemic disorders.

  11. Stem Cell-Based Therapies for Ischemic Stroke

    Directory of Open Access Journals (Sweden)

    Lei Hao

    2014-01-01

    Full Text Available In recent years, stem cell-based approaches have attracted more attention from scientists and clinicians due to their possible therapeutical effect on stroke. Animal studies have demonstrated that the beneficial effects of stem cells including embryonic stem cells (ESCs, inducible pluripotent stem cells (iPSCs, neural stem cells (NSCs, and mesenchymal stem cell (MSCs might be due to cell replacement, neuroprotection, endogenous neurogenesis, angiogenesis, and modulation on inflammation and immune response. Although several clinical studies have shown the high efficiency and safety of stem cell in stroke management, mainly MSCs, some issues regarding to cell homing, survival, tracking, safety, and optimal cell transplantation protocol, such as cell dose and time window, should be addressed. Undoubtably, stem cell-based gene therapy represents a novel potential therapeutic strategy for stroke in future.

  12. Mesenchymal Stem Cell-Based Therapy for Prostate Cancer

    Science.gov (United States)

    2014-09-01

    Mesenchymal Stem Cell-Based Therapy for Prostate Cancer PRINCIPAL INVESTIGATOR: John Isaacs; Jeffrey Karp ...clinical trials for CRPC. The team is composed of Drs. Jeffrey Karp Co-Director of Regenerative Therapeutics at the Brigham & Women’s Hospital...encapsulating a PSA-activated thapsigargin-based prodrug (G115, Fig. 5) were generated by the Karp lab with the properties outlined in Table 7. These

  13. Mesenchymal Stem Cell Based Therapy for Prostate Cancer

    Science.gov (United States)

    2015-11-01

    Prostate: From Birth to Death and Potential Applications in Between. Prostate Cancer Foundation Tumor Microenvironment/ Immunology Working Group...Award Number: W81XWH-13-1-0304 TITLE: Mesenchymal Stem Cell-Based Therapy for Prostate Cancer PRINCIPAL INVESTIGATOR: John Isaacs CONTRACTING...Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions

  14. Glial progenitor cell-based treatment of the childhood leukodystrophies

    DEFF Research Database (Denmark)

    Osorio, M Joana; Goldman, Steven A

    2016-01-01

    The childhood leukodystrophies comprise a group of hereditary disorders characterized by the absence, malformation or destruction of myelin. These disorders share common clinical, radiological and pathological features, despite their diverse molecular and genetic etiologies. Oligodendrocytes...... genetic editing of pluripotent stem cells. Yet these challenges notwithstanding, the promise of glial progenitor cell-based treatment of the childhood myelin disorders offers hope to the many victims of this otherwise largely untreatable class of disease....

  15. Cell-based detection of microbial biomaterial contaminations.

    Science.gov (United States)

    Roch, Toralf; Ma, Nan; Kratz, Karl; Lendlein, Andreas

    2015-01-01

    A major challenge in biomaterial synthesis and functionalization is the prevention of microbial contaminations such as endotoxins (lipopolysaccharides (LPS)). In addition to LPS, which are exclusively expressed by Gram negative bacteria, also other microbial products derived from fungi or Gram positive bacteria can be found as contaminations in research laboratories. Typically, the Limulus amebocyte lysate (LAL)-test is used to determine the endotoxin levels of medical devices. However, this test fails to detect material-bound LPS and other microbial contaminations and, as demonstrated in this study, detects LPS from various bacterial species with different sensitivities.In this work, a cell-based assay using genetically engineered RAW macrophages, which detect not only soluble but also material-bound microbial contaminations is introduced.The sensitivity of this cell-line towards different LPS species and different heat-inactivated microbes was investigated. As proof of principle a soft hydrophobic poly(n-butyl acrylate) network (cPnBA), which may due to adhesive properties strongly bind microbes, was deliberately contaminated with heat-inactivated bacteria. While the LAL-test failed to detect the microbial contamination, the cell-based assay clearly detected material-bound microbial contaminations. Our data demonstrate that a cell-based detection system should routinely be used as supplement to the LAL-test to determine microbial contaminations of biomaterials.

  16. Enhancing molecule fluorescence with asymmetrical plasmonic antennas.

    Science.gov (United States)

    Lu, Guowei; Liu, Jie; Zhang, Tianyue; Shen, Hongming; Perriat, Pascal; Martini, Matteo; Tillement, Olivier; Gu, Ying; He, Yingbo; Wang, Yuwei; Gong, Qihuang

    2013-07-21

    We propose and justify by the finite-difference time-domain method an efficient strategy to enhance the spontaneous emission of a fluorophore with a multi-resonance plasmonic antenna. The custom-designed asymmetrical antenna consists of two plasmonic nanoparticles with different sizes and is able to couple efficiently to free space light through multiple localized surface plasmon resonances. This design simultaneously permits a large near-field excitation near the antenna as well as a high quantum efficiency, which results in an unusual and significant enhancement of the fluorescence of a single emitter. Such an asymmetrical antenna presents intrinsic advantages over single particle or dimer based antennas made using two identical nanostructures. This promising concept can be exploited in the large domain of light-matter interaction processes involving multiple frequencies.

  17. Enhanced image reconstruction of three-dimensional fluorescent assays by subtractive structured-light illumination microscopy.

    Science.gov (United States)

    Choi, Jong-ryul; Kim, Donghyun

    2012-10-01

    We investigate improved image reconstruction of structured light illumination for high-resolution imaging of three-dimensional (3D) cell-based assays. For proof of concept, an in situ fluorescence optical detection system was built with a digital micromirror device as a spatial light modulator, for which phase and tilting angle in a grid pattern were varied to implement specific image reconstruction schemes. Subtractive reconstruction algorithms based on structured light illumination were used to acquire images of fluorescent microbeads deposited as a two-dimensional monolayer or in 3D alginate matrix. We have confirmed that an optical subtraction algorithm improves axial and lateral resolution by effectively removing out-of-focus fluorescence. The results suggest that subtractive image reconstruction can be useful for structured illumination microscopy of broad types of cell-based assays with high image resolution.

  18. Plasmonic-enhanced two-photon fluorescence with single gold nanoshell

    Science.gov (United States)

    Zhang, TianYue; Lu, GuoWei; Shen, HongMing; Perriat, P.; Martini, M.; Tillement, O.; Gong, QiHuang

    2014-06-01

    Single gold nanoshell with mutilpolar plasmon resonances is proposed to enhance two-photon fluorescence efficiently. The single emitter single nanoshell configuration is studied systematically by employing the finite-difference time-domain method. The emitter located inside or outside the nanoshell at various positions leads to a significantly different enhancement effect. The fluorescent emitter placed outside the nanoshell can achieve large fluorescence intensity given that both the position and orientation of the emission dipole are optimally controlled. In contrast, for the case of the emitter placed inside the nanoshell, it can experience substantial two-photon fluorescence enhancement without strict requirements upon the position and dipole orientations. Metallic nanoshell encapsulating many fluorescent emitters should be a promising nanocomposite configuration for bright two-photon fluorescence label. The results provide a comprehensive understanding about the plasmonic-enhanced two-photon fluorescence behaviors, and the nanocomposite configuration has great potential for optical detecting, imaging and sensing in biological applications.

  19. Cell-Based Odorant Sensor Array for Odor Discrimination Based on Insect Odorant Receptors.

    Science.gov (United States)

    Termtanasombat, Maneerat; Mitsuno, Hidefumi; Misawa, Nobuo; Yamahira, Shinya; Sakurai, Takeshi; Yamaguchi, Satoshi; Nagamune, Teruyuki; Kanzaki, Ryohei

    2016-07-01

    The olfactory system of living organisms can accurately discriminate numerous odors by recognizing the pattern of activation of several odorant receptors (ORs). Thus, development of an odorant sensor array based on multiple ORs presents the possibility of mimicking biological odor discrimination mechanisms. Recently, we developed novel odorant sensor elements with high sensitivity and selectivity based on insect OR-expressing Sf21 cells that respond to target odorants by displaying increased fluorescence intensity. Here we introduce the development of an odorant sensor array composed of several Sf21 cell lines expressing different ORs. In this study, an array pattern of four cell lines expressing Or13a, Or56a, BmOR1, and BmOR3 was successfully created using a patterned polydimethylsiloxane film template and cell-immobilizing reagents, termed biocompatible anchor for membrane (BAM). We demonstrated that BAM could create a clear pattern of Sf21 sensor cells without impacting their odorant-sensing performance. Our sensor array showed odorant-specific response patterns toward both odorant mixtures and single odorant stimuli, allowing us to visualize the presence of 1-octen-3-ol, geosmin, bombykol, and bombykal as an increased fluorescence intensity in the region of Or13a, Or56a, BmOR1, and BmOR3 cell lines, respectively. Therefore, we successfully developed a new methodology for creating a cell-based odorant sensor array that enables us to discriminate multiple target odorants. Our method might be expanded into the development of an odorant sensor capable of detecting a large range of environmental odorants that might become a promising tool used in various applications including the study of insect semiochemicals and food contamination.

  20. Nine New Fluorescent Probes

    Science.gov (United States)

    Lin, Tsung-I.; Jovanovic, Misa V.; Dowben, Robert M.

    1989-06-01

    Absorption and fluorescence spectroscopic studies are reported here for nine new fluorescent probes recently synthesized in our laboratories: four pyrene derivatives with substituents of (i) 1,3-diacetoxy-6,8-dichlorosulfonyl, (ii) 1,3-dihydroxy-6,8-disodiumsulfonate, (iii) 1,3-disodiumsulfonate, and (iv) l-ethoxy-3,6,8-trisodiumsulfonate groups, and five [7-julolidino] coumarin derivatives with substituents of (v) 3-carboxylate-4-methyl, (vi) 3- methylcarboxylate, (vii) 3-acetate-4-methyl, (viii) 3-propionate-4-methyl, and (ix) 3-sulfonate-4-methyl groups. Pyrene compounds i and ii and coumarin compounds v and vi exhibit interesting absorbance and fluorescence properties: their absorption maxima are red shifted compared to the parent compound to the blue-green region, and the band width broadens considerably. All four blue-absorbing dyes fluoresce intensely in the green region, and the two pyrene compounds emit at such long wavelengths without formation of excimers. The fluorescence properties of these compounds are quite environment-sensitive: considerable spectral shifts and fluorescence intensity changes have been observed in the pH range from 3 to 10 and in a wide variety of polar and hydrophobic solvents with vastly different dielectric constants. The high extinction and fluorescence quantum yield of these probes make them ideal fluorescent labeling reagents for proteins, antibodies, nucleic acids, and cellular organelles. The pH and hydrophobicity-dependent fluorescence changes can be utilized as optical pH and/or hydrophobicity indicators for mapping environmental difference in various cellular components in a single cell. Since all nine probes absorb in the UV, but emit at different wavelengths in the visible, these two groups of compounds offer an advantage of utilizing a single monochromatic light source (e.g., a nitrogen laser) to achieve multi-wavelength detection for flow cytometry application. As a first step to explore potential application in

  1. Cell-free measurements of brightness of fluorescently labeled antibodies

    Science.gov (United States)

    Zhou, Haiying; Tourkakis, George; Shi, Dennis; Kim, David M.; Zhang, Hairong; Du, Tommy; Eades, William C.; Berezin, Mikhail Y.

    2017-02-01

    Validation of imaging contrast agents, such as fluorescently labeled imaging antibodies, has been recognized as a critical challenge in clinical and preclinical studies. As the number of applications for imaging antibodies grows, these materials are increasingly being subjected to careful scrutiny. Antibody fluorescent brightness is one of the key parameters that is of critical importance. Direct measurements of the brightness with common spectroscopy methods are challenging, because the fluorescent properties of the imaging antibodies are highly sensitive to the methods of conjugation, degree of labeling, and contamination with free dyes. Traditional methods rely on cell-based assays that lack reproducibility and accuracy. In this manuscript, we present a novel and general approach for measuring the brightness using antibody-avid polystyrene beads and flow cytometry. As compared to a cell-based method, the described technique is rapid, quantitative, and highly reproducible. The proposed method requires less than ten microgram of sample and is applicable for optimizing synthetic conjugation procedures, testing commercial imaging antibodies, and performing high-throughput validation of conjugation procedures.

  2. Random search for a dark resonance

    Science.gov (United States)

    Kiilerich, Alexander Holm; Mølmer, Klaus

    2017-02-01

    A pair of resonant laser fields can drive a three-level system into a dark state where it ceases to absorb and emit radiation due to destructive interference. We propose a scheme to search for this resonance by randomly changing the frequency of one of the fields each time a fluorescence photon is detected. The longer the system is probed, the more likely the frequency is close to resonance and the system populates the dark state. Due to the correspondingly long waiting times between detection events, the evolution is nonergodic and the precision of the frequency estimate does not follow from the conventional Cramér-Rao bound of parameter estimation. Instead, a Lévy statistical analysis yields the scaling of the estimation error with time for precision probing of this kind.

  3. Fluorescence of Alexa fluor dye tracks protein folding.

    Science.gov (United States)

    Lindhoud, Simon; Westphal, Adrie H; Visser, Antonie J W G; Borst, Jan Willem; van Mierlo, Carlo P M

    2012-01-01

    Fluorescence spectroscopy is an important tool for the characterization of protein folding. Often, a protein is labeled with appropriate fluorescent donor and acceptor probes and folding-induced changes in Förster Resonance Energy Transfer (FRET) are monitored. However, conformational changes of the protein potentially affect fluorescence properties of both probes, thereby profoundly complicating interpretation of FRET data. In this study, we assess the effects protein folding has on fluorescence properties of Alexa Fluor 488 (A488), which is commonly used as FRET donor. Here, A488 is covalently attached to Cys69 of apoflavodoxin from Azotobacter vinelandii. Although coupling of A488 slightly destabilizes apoflavodoxin, the three-state folding of this protein, which involves a molten globule intermediate, is unaffected. Upon folding of apoflavodoxin, fluorescence emission intensity of A488 changes significantly. To illuminate the molecular sources of this alteration, we applied steady state and time-resolved fluorescence techniques. The results obtained show that tryptophans cause folding-induced changes in quenching of Alexa dye. Compared to unfolded protein, static quenching of A488 is increased in the molten globule. Upon populating the native state both static and dynamic quenching of A488 decrease considerably. We show that fluorescence quenching of Alexa Fluor dyes is a sensitive reporter of conformational changes during protein folding.

  4. Fluorescence of Alexa fluor dye tracks protein folding.

    Directory of Open Access Journals (Sweden)

    Simon Lindhoud

    Full Text Available Fluorescence spectroscopy is an important tool for the characterization of protein folding. Often, a protein is labeled with appropriate fluorescent donor and acceptor probes and folding-induced changes in Förster Resonance Energy Transfer (FRET are monitored. However, conformational changes of the protein potentially affect fluorescence properties of both probes, thereby profoundly complicating interpretation of FRET data. In this study, we assess the effects protein folding has on fluorescence properties of Alexa Fluor 488 (A488, which is commonly used as FRET donor. Here, A488 is covalently attached to Cys69 of apoflavodoxin from Azotobacter vinelandii. Although coupling of A488 slightly destabilizes apoflavodoxin, the three-state folding of this protein, which involves a molten globule intermediate, is unaffected. Upon folding of apoflavodoxin, fluorescence emission intensity of A488 changes significantly. To illuminate the molecular sources of this alteration, we applied steady state and time-resolved fluorescence techniques. The results obtained show that tryptophans cause folding-induced changes in quenching of Alexa dye. Compared to unfolded protein, static quenching of A488 is increased in the molten globule. Upon populating the native state both static and dynamic quenching of A488 decrease considerably. We show that fluorescence quenching of Alexa Fluor dyes is a sensitive reporter of conformational changes during protein folding.

  5. Stroboscopic fluorescence lifetime imaging.

    Science.gov (United States)

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

    2009-03-30

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

  6. Resonance and Neck Length for a Spherical Resonator

    Directory of Open Access Journals (Sweden)

    Emily Corning

    2011-06-01

    Full Text Available The relationship between the neck length of a spherical resonator and its period of fundamental resonance was investigated. This was done by measuring the frequency of fundamental resonance of the resonator at 6 different neck lengths. It was found that its resonance resembled Helmholtz resonance but was not that of ideal Helmholtz resonance.

  7. Enhancement of single-molecule fluorescence signals by colloidal silver nanoparticles in studies of protein translation.

    Science.gov (United States)

    Bharill, Shashank; Chen, Chunlai; Stevens, Benjamin; Kaur, Jaskiran; Smilansky, Zeev; Mandecki, Wlodek; Gryczynski, Ignacy; Gryczynski, Zygmunt; Cooperman, Barry S; Goldman, Yale E

    2011-01-25

    Metal-enhanced fluorescence (MEF) increased total photon emission of Cy3- and Cy5-labeled ribosomal initiation complexes near 50 nm silver particles 4- and 5.5-fold, respectively. Fluorescence intensity fluctuations above shot noise, at 0.1-5 Hz, were greater on silver particles. Overall signal-to-noise ratio was similar or slightly improved near the particles. Proximity to silver particles did not compromise ribosome function, as measured by codon-dependent binding of fluorescent tRNA, dynamics of fluorescence resonance energy transfer between adjacent tRNAs in the ribosome, and tRNA translocation induced by elongation factor G.

  8. Dendritic cell-based cancer immunotherapy for colorectal cancer.

    Science.gov (United States)

    Kajihara, Mikio; Takakura, Kazuki; Kanai, Tomoya; Ito, Zensho; Saito, Keisuke; Takami, Shinichiro; Shimodaira, Shigetaka; Okamoto, Masato; Ohkusa, Toshifumi; Koido, Shigeo

    2016-05-01

    Colorectal cancer (CRC) is one of the most common cancers and a leading cause of cancer-related mortality worldwide. Although systemic therapy is the standard care for patients with recurrent or metastatic CRC, the prognosis is extremely poor. The optimal sequence of therapy remains unknown. Therefore, alternative strategies, such as immunotherapy, are needed for patients with advanced CRC. This review summarizes evidence from dendritic cell-based cancer immunotherapy strategies that are currently in clinical trials. In addition, we discuss the possibility of antitumor immune responses through immunoinhibitory PD-1/PD-L1 pathway blockade in CRC patients.

  9. Cell-based reparative therapies for multiple sclerosis.

    Science.gov (United States)

    Ben-Hur, Tamir; Fainstein, Nina; Nishri, Yossi

    2013-11-01

    The strong rationale for cell-based therapy in multiple sclerosis is based on the ability of stem and precursor cells of neural and mesenchymal origin to attenuate neuroinflammation, to facilitate endogenous repair processes, and to participate directly in remyelination, if directed towards a myelin-forming fate. However, there are still major gaps in knowledge regarding induction of repair in chronic multiple sclerosis lesions, and whether transplanted cells can overcome the multiple environmental inhibitory factors which underlie the failure of endogenous repair. Major challenges in clinical translation include the determination of the optimal cellular platform, the route of cell delivery, and candidate patients for treatment.

  10. Sorting live stem cells based on Sox2 mRNA expression.

    Directory of Open Access Journals (Sweden)

    Hans M Larsson

    Full Text Available While cell sorting usually relies on cell-surface protein markers, molecular beacons (MBs offer the potential to sort cells based on the presence of any expressed mRNA and in principle could be extremely useful to sort rare cell populations from primary isolates. We show here how stem cells can be purified from mixed cell populations by sorting based on MBs. Specifically, we designed molecular beacons targeting Sox2, a well-known stem cell marker for murine embryonic (mES and neural stem cells (NSC. One of our designed molecular beacons displayed an increase in fluorescence compared to a nonspecific molecular beacon both in vitro and in vivo when tested in mES and NSCs. We sorted Sox2-MB(+SSEA1(+ cells from a mixed population of 4-day retinoic acid-treated mES cells and effectively isolated live undifferentiated stem cells. Additionally, Sox2-MB(+ cells isolated from primary mouse brains were sorted and generated neurospheres with higher efficiency than Sox2-MB(- cells. These results demonstrate the utility of MBs for stem cell sorting in an mRNA-specific manner.

  11. A cell-based screen reveals that the albendazole metabolite, albendazole sulfone, targets Wolbachia.

    Science.gov (United States)

    Serbus, Laura R; Landmann, Frederic; Bray, Walter M; White, Pamela M; Ruybal, Jordan; Lokey, R Scott; Debec, Alain; Sullivan, William

    2012-09-01

    Wolbachia endosymbionts carried by filarial nematodes give rise to the neglected diseases African river blindness and lymphatic filariasis afflicting millions worldwide. Here we identify new Wolbachia-disrupting compounds by conducting high-throughput cell-based chemical screens using a Wolbachia-infected, fluorescently labeled Drosophila cell line. This screen yielded several Wolbachia-disrupting compounds including three that resembled Albendazole, a widely used anthelmintic drug that targets nematode microtubules. Follow-up studies demonstrate that a common Albendazole metabolite, Albendazole sulfone, reduces intracellular Wolbachia titer both in Drosophila melanogaster and Brugia malayi, the nematode responsible for lymphatic filariasis. Significantly, Albendazole sulfone does not disrupt Drosophila microtubule organization, suggesting that this compound reduces titer through direct targeting of Wolbachia. Accordingly, both DNA staining and FtsZ immunofluorescence demonstrates that Albendazole sulfone treatment induces Wolbachia elongation, a phenotype indicative of binary fission defects. This suggests that the efficacy of Albendazole in treating filarial nematode-based diseases is attributable to dual targeting of nematode microtubules and their Wolbachia endosymbionts.

  12. Improvement of Bioactive Compound Classification through Integration of Orthogonal Cell-Based Biosensing Methods

    Directory of Open Access Journals (Sweden)

    Goran N. Jovanovic

    2007-01-01

    Full Text Available Lack of specificity for different classes of chemical and biological agents, and false positives and negatives, can limit the range of applications for cell-based biosensors. This study suggests that the integration of results from algal cells (Mesotaenium caldariorum and fish chromatophores (Betta splendens improves classification efficiency and detection reliability. Cells were challenged with paraquat, mercuric chloride, sodium arsenite and clonidine. The two detection systems were independently investigated for classification of the toxin set by performing discriminant analysis. The algal system correctly classified 72% of the bioactive compounds, whereas the fish chromatophore system correctly classified 68%. The combined classification efficiency was 95%. The algal sensor readout is based on fluorescence measurements of changes in the energy producing pathways of photosynthetic cells, whereas the response from fish chromatophores was quantified using optical density. Change in optical density reflects interference with the functioning of cellular signal transduction networks. Thus, algal cells and fish chromatophores respond to the challenge agents through sufficiently different mechanisms of action to be considered orthogonal.

  13. A liposomal system for contrast-enhanced magnetic resonance imaging of molecular targets

    NARCIS (Netherlands)

    Mulder, WJM; Strijkers, GJ; Griffioen, AW; van Bloois, L; Molema, G; Storm, G; Koning, GA; Nicolay, K

    2004-01-01

    Pegylated paramagnetic and fluorescent immunoliposomes were designed to enable the parallel detection of the induced expression of molecular markers on endothelial cells with magnetic resonance imaging (MRI) and fluorescence microscopy. MRI is capable of three-dimensional noninvasive imaging of opaq

  14. Fluorescent polymer coated capillaries as optofluidic refractometric sensors.

    Science.gov (United States)

    Rowland, Kristopher J; François, Alexandre; Hoffmann, Peter; Monro, Tanya M

    2013-05-06

    A capillary microresonator platform for refractometric sensing is demonstrated by coating the interior of thick-walled silica capillaries with a sub-wavelength layer of high refractive index, dye-doped polymer. No intermediate processing, such as etching or tapering, of the capillary is required. Side illumination and detection of the polymer layer reveals a fluorescence spectrum that is periodically modulated by whispering gallery mode resonances within the layer. Using a Fourier technique to calculate the spectral resonance shifts, the fabricated capillary resonators exhibited refractometric sensitivities up to approximately 30 nm/RIU upon flowing aqueous glucose through them. These sensors could be readily integrated with existing biological and chemical separation platforms such as capillary electrophoresis and gas chromatography where such thick walled capillaries are routinely used with polymer coatings. A review of the modelling required to calculate whispering gallery eigenmodes of such inverted cylindrical resonators is also presented.

  15. Integral resonator gyroscope

    Science.gov (United States)

    Shcheglov, Kirill V. (Inventor); Challoner, A. Dorian (Inventor); Hayworth, Ken J. (Inventor); Wiberg, Dean V. (Inventor); Yee, Karl Y. (Inventor)

    2008-01-01

    The present invention discloses an inertial sensor having an integral resonator. A typical sensor comprises a planar mechanical resonator for sensing motion of the inertial sensor and a case for housing the resonator. The resonator and a wall of the case are defined through an etching process. A typical method of producing the resonator includes etching a baseplate, bonding a wafer to the etched baseplate, through etching the wafer to form a planar mechanical resonator and the wall of the case and bonding an end cap wafer to the wall to complete the case.

  16. Cooperative fluorescence from a strongly driven dilute cloud of atoms

    DEFF Research Database (Denmark)

    Ott, Johan Raunkjær; Wubs, Martijn; Lodahl, Peter;

    2013-01-01

    We investigate cooperative fluorescence in a dilute cloud of strongly driven two-level emitters. Starting from the Heisenberg equations of motion, we compute the first-order scattering corrections to the saturation of the excited-state population and to the resonance-fluorescence spectrum, which...... both require going beyond the state-of-the-art linear-optics approach to describe collective phenomena. A dipole blockade is observed due to long-range dipole-dipole coupling that vanishes at stronger driving fields. Furthermore, we compute the inelastic component of the light scattered by a cloud...

  17. Cell-Based Strategies for Meniscus Tissue Engineering

    Science.gov (United States)

    Niu, Wei; Guo, Weimin; Han, Shufeng; Zhu, Yun; Liu, Shuyun; Guo, Quanyi

    2016-01-01

    Meniscus injuries remain a significant challenge due to the poor healing potential of the inner avascular zone. Following a series of studies and clinical trials, tissue engineering is considered a promising prospect for meniscus repair and regeneration. As one of the key factors in tissue engineering, cells are believed to be highly beneficial in generating bionic meniscus structures to replace injured ones in patients. Therefore, cell-based strategies for meniscus tissue engineering play a fundamental role in meniscal regeneration. According to current studies, the main cell-based strategies for meniscus tissue engineering are single cell type strategies; cell coculture strategies also were applied to meniscus tissue engineering. Likewise, on the one side, the zonal recapitulation strategies based on mimicking meniscal differing cells and internal architectures have received wide attentions. On the other side, cell self-assembling strategies without any scaffolds may be a better way to build a bionic meniscus. In this review, we primarily discuss cell seeds for meniscus tissue engineering and their application strategies. We also discuss recent advances and achievements in meniscus repair experiments that further improve our understanding of meniscus tissue engineering. PMID:27274735

  18. Stem Cell-Based Cell Therapy for Glomerulonephritis

    Directory of Open Access Journals (Sweden)

    Meiling Jin

    2014-01-01

    Full Text Available Glomerulonephritis (GN, characterized by immune-mediated inflammatory changes in the glomerular, is a common cause of end stage renal disease. Therapeutic options for glomerulonephritis applicable to all cases mainly include symptomatic treatment and strategies to delay progression. In the attempt to yield innovative interventions fostering the limited capability of regeneration of renal tissue after injury and the uncontrolled pathological process by current treatments, stem cell-based therapy has emerged as novel therapy for its ability to inhibit inflammation and promote regeneration. Many basic and clinical studies have been performed that support the ability of various stem cell populations to ameliorate glomerular injury and improve renal function. However, there is a long way before putting stem cell-based therapy into clinical practice. In the present article, we aim to review works performed with respect to the use of stem cell of different origins in GN, and to discuss the potential mechanism of therapeutic effect and the challenges for clinical application of stem cells.

  19. Cell-Based Strategies for Meniscus Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Wei Niu

    2016-01-01

    Full Text Available Meniscus injuries remain a significant challenge due to the poor healing potential of the inner avascular zone. Following a series of studies and clinical trials, tissue engineering is considered a promising prospect for meniscus repair and regeneration. As one of the key factors in tissue engineering, cells are believed to be highly beneficial in generating bionic meniscus structures to replace injured ones in patients. Therefore, cell-based strategies for meniscus tissue engineering play a fundamental role in meniscal regeneration. According to current studies, the main cell-based strategies for meniscus tissue engineering are single cell type strategies; cell coculture strategies also were applied to meniscus tissue engineering. Likewise, on the one side, the zonal recapitulation strategies based on mimicking meniscal differing cells and internal architectures have received wide attentions. On the other side, cell self-assembling strategies without any scaffolds may be a better way to build a bionic meniscus. In this review, we primarily discuss cell seeds for meniscus tissue engineering and their application strategies. We also discuss recent advances and achievements in meniscus repair experiments that further improve our understanding of meniscus tissue engineering.

  20. FLEX: fluorescence explorer

    NARCIS (Netherlands)

    Stoll, M.Ph.; Court, A.J.; Smorenburg, C.; Visser, H.; Crocco, L.; Heilimo, J.; Honig, A.

    1999-01-01

    FLEX is a scientifically driven space mission to provide demonstration/validation of the instrumentation and technique for measuring the natural fluorescence of vegetation in the Fraunhofer lines. The payload consists of high spectral resolution (0.1-0.3 nm) CCD imaging grating spectrometer with two

  1. Fluorescence Experiments with Quinine

    Science.gov (United States)

    O'Reilly, James E.

    1975-01-01

    Describes a series of experiments which illustrate the analytical capabilities of fluorescence, and outlines two straightforward analyses involving real analyses. These experiments are suitable for an undergraduate instrumental analysis course and require approximately six to seven hours of laboratory time. (MLH)

  2. Surface-Enhanced X-Ray Fluorescence

    Science.gov (United States)

    Anderson, Mark

    2010-01-01

    Surface-enhanced x-ray fluorescence (SEn-XRF) spectroscopy is a form of surface- enhanced spectroscopy that was conceived as a means of obtaining greater sensitivity in x-ray fluorescence (XRF) spectroscopy. As such, SEn-XRF spectroscopy joins the ranks of such other, longer-wavelength surface-enhanced spectroscopies as those based on surface-enhanced Raman scattering (SERS), surface-enhanced resonance Raman scattering (SERRS), and surfaceenhanced infrared Raman absorption (SEIRA), which have been described in previous NASA Tech Briefs articles. XRF spectroscopy has been used in analytical chemistry for determining the elemental compositions of small samples. XRF spectroscopy is rapid and quantitative and has been applied to a variety of metal and mineralogical samples. The main drawback of XRF spectroscopy as practiced heretofore is that sensitivity has not been as high as required for some applications. In SEn-XRF as in the other surface-enhanced spectroscopies, one exploits several interacting near-field phenomena, occurring on nanotextured surfaces, that give rise to local concentrations of incident far-field illumination. In this case, the far-field illumination comes from an x-ray source. Depending on the chemical composition and the geometry of a given nanotextured surface, these phenomena could include the lightning-rod effect (concentration of electric fields at the sharpest points on needlelike surface features), surface plasmon resonances, and grazing incidence geometric effects. In the far field, the observable effect of these phenomena is an increase in the intensity of the spectrum of interest - in this case, the x-ray fluorescence spectrum of chemical elements of interest that may be present within a surface layer at distances no more than a few nanometers from the surface.

  3. Spin coupling and resonance

    NARCIS (Netherlands)

    Zielinski, M.L.; van Lenthe, J.H.

    2008-01-01

    The resonating block localize wave function (RBLW) method is introduced, a resonating modification of the block localized wave functions introduced by Mo et al. [Mo, Y.; Peyerimhoff, S. D. J. Chem. Phys. 1998, 109, 1687].This approach allows the evaluation of resonance energies following Pauling’s r

  4. Controlling Parametric Resonance

    DEFF Research Database (Denmark)

    Galeazzi, Roberto; Pettersen, Kristin Ytterstad

    2012-01-01

    Parametric resonance is a resonant phenomenon which takes place in systems characterized by periodic variations of some parameters. While seen as a threatening condition, whose onset can drive a system into instability, this chapter advocates that parametric resonance may become an advantage if t...

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

    Science.gov (United States)

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

    2012-01-01

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

  6. Study on binding and fluorescence energy transfer efficiency of Rhodamine B with Pluronic F127-gold nanohybrid using optical spectroscopy methods

    Science.gov (United States)

    Antonisamy, Jenif Dsouza; Swain, Jitendriya; Dash, Sasmita

    2017-02-01

    This work focuses on the binding efficiency and fluorescence resonance energy transfer (FRET) of fluorescent dye Rhodamine B (Rh B) to Pluronic F127-gold nanohybrid. The formation of gold nanoparticles inside Rh B doped Pluronic F127 copolymer have been characterized using dynamic light scattering study, HR-TEM images, UV-visible spectra and fluorescence studies. Fluorescence quenching and the constant fluorescence lifetime of the Rhodamine B present in the cavity of Pluronic F127-gold nanohybrid suggested a strong binding ability (3.5 × 103 L mol- 1), static nature of quenching and better energy transfer efficiency of fluorescent dye towards Pluronic F127-gold (Au) nanohybrids.

  7. Synthesis and Fluorescence Spectra of Triazolylcoumarin Fluorescent Dyes

    Institute of Scientific and Technical Information of China (English)

    PENG Xian-fu; LI Hong-qi

    2009-01-01

    Much attention is devoted to fluorescent dyes especially those with potential in versatile applications. Reactions under "click" conditions between nonfluorescent 3 - azidocoumarins and terminal alkynes produced 3 -(1, 2, 3- triazol- 1 - yl)cournarins, a novel type of fluorescent dyes with intense fluorescence. The structures of the new coumarins were characterized by 1H NMR, MS, and IR spectra. Fluorescence spectra measurement demonstrated excellent fluorescence performance of the triazolylcoumarins and this click reaction is a promising candidate for bioconjugation and bioimaging applications since both azide and alkynes are quite inert to biological systems.

  8. Fluorescence nanoscopy. Methods and applications

    OpenAIRE

    Requejo-Isidro, Jose

    2013-01-01

    Fluorescence nanoscopy refers to the experimental techniques and analytical methods used for fluorescence imaging at a resolution higher than conventional, diffraction-limited, microscopy. This review explains the concepts behind fluorescence nanoscopy and focuses on the latest and promising developments in acquisition techniques, labelling strategies to obtain highly detailed super-resolved images and in the quantitative methods to extract meaningful information from them.

  9. Fabrication et caracterisation de cristaux photoniques pour exaltation de fluorescence

    Science.gov (United States)

    Gascon, Annabelle

    2011-12-01

    In today's world, there is a pressing need for point-of-care molecular analysis that is fast, inexpensive and transportable. Lab-on-a- chips are designed to fulfill that need. They are micro-electromechanical systems (MEMS), fabricated with microelectronic techniques, that use the analytes physical properties to detect their presence in liquid samples. This detection can be performed by attaching the analyte to quantum dots. These quantum dots are semiconducting nanoparticles with narrow fluorescence band. In our project, we use a tuneable system with a two-slab photonic crystal that serves as a tuneable optical filter, detecting the presence and wavelength of these quantum dots. Photonic crystals are dielectrics with a variable refractive index, with a period near the visible light wavelength. They are called photonic crystals because they have a photonic band gap just as atomic crystals, periodic structure of atoms, have an electronic band gap. They are photonic because photons instead of electrons propagate through them. They can also enhance fluorescence from quantum dots at the photonic crystals guided resonance wavelength. My project objectives are to: (1) Fabricate two-slab photonic crystal, (2) Characterize photonic crystals, (3) Place quantum dots on photonic crystals, (4) Measure fluorescence enhancement. The device made during this project consists of a silicon wafer on which were deposited a 200 nm silicon nitride layer, then a 200 nm silicon dioxide layer and finally another 200 nm silicon nitride layer. An electron-beam lithography defines the photonic crystals and the MEMS. The photonic crystals are square lattices of holes 180 nm in diameter, at a period of 460 nm, etched through the two silicon nitride slabs. The two slabs are etched in a single step of Reactive Ion Etching (RIE). Then, the silicon under the photonic crystal is etched from the backside up to the nitride by deep-RIE. Finally, the oxide layer is removed in order to completely suspend

  10. Ovenized microelectromechanical system (MEMS) resonator

    Science.gov (United States)

    Olsson, Roy H; Wojciechowski, Kenneth; Kim, Bongsang

    2014-03-11

    An ovenized micro-electro-mechanical system (MEMS) resonator including: a substantially thermally isolated mechanical resonator cavity; a mechanical oscillator coupled to the mechanical resonator cavity; and a heating element formed on the mechanical resonator cavity.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2011-09-07

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

  12. Cell-Based Microarrays for In Vitro Toxicology

    Science.gov (United States)

    Wegener, Joachim

    2015-07-01

    DNA/RNA and protein microarrays have proven their outstanding bioanalytical performance throughout the past decades, given the unprecedented level of parallelization by which molecular recognition assays can be performed and analyzed. Cell microarrays (CMAs) make use of similar construction principles. They are applied to profile a given cell population with respect to the expression of specific molecular markers and also to measure functional cell responses to drugs and chemicals. This review focuses on the use of cell-based microarrays for assessing the cytotoxicity of drugs, toxins, or chemicals in general. It also summarizes CMA construction principles with respect to the cell types that are used for such microarrays, the readout parameters to assess toxicity, and the various formats that have been established and applied. The review ends with a critical comparison of CMAs and well-established microtiter plate (MTP) approaches.

  13. Improved Computational Model of Grid Cells Based on Column Structure

    Institute of Scientific and Technical Information of China (English)

    Yang Zhou; Dewei Wu; Weilong Li; Jia Du

    2016-01-01

    To simulate the firing pattern of biological grid cells, this paper presents an improved computational model of grid cells based on column structure. In this model, the displacement along different directions is processed by modulus operation, and the obtained remainder is associated with firing rate of grid cell. Compared with the original model, the improved parts include that: the base of modulus operation is changed, and the firing rate in firing field is encoded by Gaussian⁃like function. Simulation validates that the firing pattern generated by the improved computational model is more consistent with biological characteristic than original model. Besides, the firing pattern is badly influenced by the cumulative positioning error, but the computational model can also generate the regularly hexagonal firing pattern when the real⁃time positioning results are modified.

  14. Microencapsulating and Banking Living Cells for Cell-Based Medicine

    Directory of Open Access Journals (Sweden)

    Wujie Zhang

    2011-01-01

    Full Text Available A major challenge to the eventual success of the emerging cell-based medicine such as tissue engineering, regenerative medicine, and cell transplantation is the limited availability of the desired cell sources. This challenge can be addressed by cell microencapsulation to overcome the undesired immune response (i.e., to achieve immunoisolation so that non-autologous cells can be used to treat human diseases, and by cell/tissue preservation to bank living cells for wide distribution to end users so that they are readily available when needed in the future. This review summarizes the status quo of research in both cell microencapsulation and banking the microencapsulated cells. It is concluded with a brief outlook of future research directions in this important field.

  15. Printing technologies for biomolecule and cell-based applications.

    Science.gov (United States)

    Ihalainen, Petri; Määttänen, Anni; Sandler, Niklas

    2015-10-30

    Biomolecules, such as enzymes, proteins and other biomacromolecules (polynucleotides, polypeptides, polysaccharides and DNA) that are immobilized on solid surfaces are relevant to many areas of science and technology. These functionalized surfaces have applications in biosensors, chromatography, diagnostic immunoassays, cell culturing, DNA microarrays and other analytical techniques. Printing technologies offer opportunities in this context. The main interests in printing biomolecules are in immobilizing them on surfaces for sensors and catalysts or for controlled delivery of protein-based drugs. Recently, there have been significant developments in the use of inkjet printing for dispensing of proteins, biomacromolecules and cells. This review discusses the use of roll-to-roll and inkjet printing technologies in manufacturing of biomolecule and cell-based applications.

  16. Cell-Based Microarrays for In Vitro Toxicology.

    Science.gov (United States)

    Wegener, Joachim

    2015-01-01

    DNA/RNA and protein microarrays have proven their outstanding bioanalytical performance throughout the past decades, given the unprecedented level of parallelization by which molecular recognition assays can be performed and analyzed. Cell microarrays (CMAs) make use of similar construction principles. They are applied to profile a given cell population with respect to the expression of specific molecular markers and also to measure functional cell responses to drugs and chemicals. This review focuses on the use of cell-based microarrays for assessing the cytotoxicity of drugs, toxins, or chemicals in general. It also summarizes CMA construction principles with respect to the cell types that are used for such microarrays, the readout parameters to assess toxicity, and the various formats that have been established and applied. The review ends with a critical comparison of CMAs and well-established microtiter plate (MTP) approaches.

  17. Cell based assays for anti-Plasmodium activity evaluation.

    Science.gov (United States)

    Mokgethi-Morule, Thabang; N'Da, David D

    2016-03-10

    Malaria remains one of the most common and deadly infectious diseases worldwide. The severity of this global public health challenge is reflected by the approximately 198 million people, who were reportedly infected in 2013 and by the more than 584,000 related deaths in that same year. The rising emergence of drug resistance towards the once effective artemisinin combination therapies (ACTs) has become a serious concern and warrants more robust drug development strategies, with the objective of eradicating malaria infections. The intricate biology and life cycle of Plasmodium parasites complicate the understanding of the disease in such a way that would enhance the development of more effective chemotherapies that would achieve radical clinical cure and that would prevent disease relapse. Phenotypic cell based assays have for long been a valuable approach and involve the screening and analysis of diverse compounds with regards to their activities towards whole Plasmodium parasites in vitro. To achieve the Millennium Development Goal (MDG) of malaria eradication by 2020, new generation drugs that are active against all parasite stages (erythrocytic (blood), exo-erythrocytic (liver stages and gametocytes)) are needed. Significant advances are being made in assay development to overcome some of the practical challenges of assessing drug efficacy, particularly in the liver and transmission stage Plasmodium models. This review discusses primary screening models and the fundamental progress being made in whole cell based efficacy screens of anti-malarial activity. Ongoing challenges and some opportunities for improvements in assay development that would assist in the discovery of effective, safe and affordable drugs for malaria treatments are also discussed.

  18. Electrochemical As(III) whole-cell based biochip sensor.

    Science.gov (United States)

    Cortés-Salazar, Fernando; Beggah, Siham; van der Meer, Jan Roelof; Girault, Hubert H

    2013-09-15

    The development of a whole-cell based sensor for arsenite detection coupling biological engineering and electrochemical techniques is presented. This strategy takes advantage of the natural Escherichia coli resistance mechanism against toxic arsenic species, such as arsenite, which consists of the selective intracellular recognition of arsenite and its pumping out from the cell. A whole-cell based biosensor can be produced by coupling the intracellular recognition of arsenite to the generation of an electrochemical signal. Hereto, E. coli was equipped with a genetic circuit in which synthesis of beta-galactosidase is under control of the arsenite-derepressable arsR-promoter. The E. coli reporter strain was filled in a microchip containing 16 independent electrochemical cells (i.e. two-electrode cell), which was then employed for analysis of tap and groundwater samples. The developed arsenic-sensitive electrochemical biochip is easy to use and outperforms state-of-the-art bacterial bioreporters assays specifically in its simplicity and response time, while keeping a very good limit of detection in tap water, i.e. 0.8ppb. Additionally, a very good linear response in the ranges of concentration tested (0.94ppb to 3.75ppb, R(2)=0.9975 and 3.75 ppb to 30ppb, R(2)=0.9991) was obtained, complying perfectly with the acceptable arsenic concentration limits defined by the World Health Organization for drinking water samples (i.e. 10ppb). Therefore, the proposed assay provides a very good alternative for the portable quantification of As (III) in water as corroborated by the analysis of natural groundwater samples from Swiss mountains, which showed a very good agreement with the results obtained by atomic absorption spectroscopy.

  19. Determination of binding specificity between receptors and probes covering HIV-1 gp41 NHR in fluorescence resonance energy transfer assay%HIV-1gp41NHR序列靶点专一性的荧光共振能转移测定

    Institute of Scientific and Technical Information of China (English)

    王昆; 郑保华; 蔡利锋; 刘克良; 李艳妮

    2013-01-01

    目的 确定并利用荧光共振能转移法(FRET)对以HIV-1 gp41 N端七联重复序列(NHR)为靶点的融合抑制剂进行筛选和作用机制研究.方法 FRET采用金属络合物多肽技术设计针对不同结合位点、结合强度可调、涵盖全部NHR序列的靶点和探针,对HIV融合抑制剂进行高通量筛选.由于HIV在进行膜融合时其gp41的N端NHR和C端CHR可形成稳定的六螺旋结构,因此,利用圆二色谱仪对FRET所使用的靶点/探针对的结合强度进行验证,确定对应的靶点/探针对可形成稳定的六螺旋结构;同时,借助细胞活性测试测定抑制剂的活性,验证FRET是否可用于筛选以HIV-1 gp41 NHR为靶点的抑制剂.结果与结论 FRET中使用的靶点/探针均可形成螺旋度较高的六螺旋结构,其中Fe(Env2.0)3/CP2及Fe(Env5.0)3/CP5形成的α螺旋度分别高达89.6%和84.7%.FRET所使用的靶点/探针对专一性强、结合作用强,可用于进行HIV-1融合抑制剂的筛选和机制研究.%Objective To identify and optimize a fluorescence resonance energy transfer ( FRET )-based assay to screen fusion inhibitors which target HIV-1 gp41 N-terminal heptad repeats( NHR ) and to study their mechanisms. Methods The FRET assay using metallopeptide technology was applied to detect HIV fusion inhibitors in a high-throughput mode, by which receptors and probes were designed to cover all the possible NHR binding sites with flexible binding affinities. The assay biochemically represented a stable fusion-active conformation of the envelope protein gp41 of HIV-1, which was a six-helix bundle formed by NHR and C-terminal heptad repeat ( CHR ) in the process of virus-cell membrane fusion. The six-helix bundle formed by an NHR-derived receptor and CHR-derived probe was characterized by circular dichroism to verify the effective receptor/probe pair, and the FRET assay was verified by comparing the result of the assay with a well-established cell-cell fusion assay using known

  20. Magnetic resonance energy and topological resonance energy.

    Science.gov (United States)

    Aihara, Jun-Ichi

    2016-04-28

    Ring-current diamagnetism of a polycyclic π-system is closely associated with thermodynamic stability due to the individual circuits. Magnetic resonance energy (MRE), derived from the ring-current diamagnetic susceptibility, was explored in conjunction with graph-theoretically defined topological resonance energy (TRE). For many aromatic molecules, MRE is highly correlative with TRE with a correlation coefficient of 0.996. For all π-systems studied, MRE has the same sign as TRE. The only trouble with MRE may be that some antiaromatic and non-alternant species exhibit unusually large MRE-to-TRE ratios. This kind of difficulty can in principle be overcome by prior geometry-optimisation or by changing spin multiplicity. Apart from the semi-empirical resonance-theory resonance energy, MRE is considered as the first aromatic stabilisation energy (ASE) defined without referring to any hypothetical polyene reference.

  1. Dual channel fluorescence engineering on the nano-patterened metamaterial (Conference Presentation)

    Science.gov (United States)

    Wu, Huijun; Tseng, Ming Lun; Tsai, Wei Yi; Chen, Ting-Yu; Chien, Yihsin; Chang, Yun-Chorng; Tsai, Din Ping

    2016-09-01

    Upconversion fluorescence from Lanthanide-doped nanocrystals has attracted widespread interests because of its greatly potential applications in various fields, such as photonic crystal lasers, material science, biological therapy, and so on. However, the relatively low quantum yield (typically chemical methods, plasmonic structures have been adopted as another strategy to improve the radiation efficiency and control the relaxation process of the upcovnersion nanocrystals. We designed the anti-symmetric split ring resonators with various periods and the fishnet structures. The surface plasmon resonance peaks of the structure shift as the periods varies. For example, in a multi-layered plamsonic metasurface with the period of 250nm, both the electric and magnetic modes could be generated simultaneously when excited by the incident light with proper polarization. This plasmonic structure provides two different channels for the enhancement of upconversion fluorescence. The resonance peak of 650nm is magnetic resonance mode, while the peak of 980nm is electric resonance mode. The resonance peak of 980nm coincides with the absorption band of the Lanthanide-dopoed nanocrystal, and the peak of 650nm matches with its emission band. We found that the upconversion fluorescence intensity could be enhanced more than 10 times when the electric resonance frequency of the metasurface matches with the absorption band of the upconversion nanocrystals, while the magnetic mode overlaps with its emission band. This is due to the local density of optical states was significantly enhanced by the plasmonic metasurface. The detailed results and mechanism will be discussed.

  2. Fluorescent biosensors for high throughput screening of protein kinase inhibitors.

    Science.gov (United States)

    Prével, Camille; Pellerano, Morgan; Van, Thi Nhu Ngoc; Morris, May C

    2014-02-01

    High throughput screening assays aim to identify small molecules that interfere with protein function, activity, or conformation, which can serve as effective tools for chemical biology studies of targets involved in physiological processes or pathways of interest or disease models, as well as templates for development of therapeutics in medicinal chemistry. Fluorescent biosensors constitute attractive and powerful tools for drug discovery programs, from high throughput screening assays, to postscreen characterization of hits, optimization of lead compounds, and preclinical evaluation of candidate drugs. They provide a means of screening for inhibitors that selectively target enzymatic activity, conformation, and/or function in vitro. Moreover, fluorescent biosensors constitute useful tools for cell- and image-based, multiplex and multiparametric, high-content screening. Application of fluorescence-based sensors to screen large and complex libraries of compounds in vitro, in cell-based formats or whole organisms requires several levels of optimization to establish robust and reproducible assays. In this review, we describe the different fluorescent biosensor technologies which have been applied to high throughput screens, and discuss the prerequisite criteria underlying their successful application. Special emphasis is placed on protein kinase biosensors, since these enzymes constitute one of the most important classes of therapeutic targets in drug discovery.

  3. Multifunctional fluorescent magnetic nanoparticles for lung cancer stem cells research.

    Science.gov (United States)

    Zhou, Xuan; Chen, Lisha; Wang, Anxin; Ma, Yufei; Zhang, Hailu; Zhu, Yimin

    2015-10-01

    In this paper, a multifunctional peptide-fluorescent-magnetic nanocomposites (Fe₃O₄@PEI@Cy5.5@PEG@HCBP-1 NPs) was synthesized via a layer-by-layer approach for potential application to cancer diagnoses. The multifunctional nanocomposites have great dispersibility and homogeneous particle sizes in aqueous solution. Meanwhile, it has perfect hemocompatibility and satisfying cytocompatibility in a relatively high concentration. Data from in vitro cytotoxicity assay indicated that the nanocomposites could recognize the lung cancer stem cells (CSCs) specifically and enrich the HCBP-1 positive CSCs from H460 tumor xenografts effectively. Additionally, the results of in vivo live fluorescent imaging and magnetic resonance imaging (MRI) showed that the nanocomposites could identify lung CSCs in tumor xenografts. These results suggested that the nanocomposites could be used as a potential cancer diagnostic agent through modifying diverse fluorescence dyes and targeting ligands on its surface.

  4. Coumarin amide derivatives as fluorescence chemosensors for cyanide anions

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Qianqian [School of Material Science and Engineering, Shandong Provincial Key Laboratory of Preparation and Measurement of Building Materials, University of Jinan, Jinan 250022, Shandong (China); Liu, Zhiqiang [State Key Laboratory of Crystal Materials, Shandong University, Jinan 250100, Shandong (China); Cao, Duxia, E-mail: duxiacao@ujn.edu.cn [School of Material Science and Engineering, Shandong Provincial Key Laboratory of Preparation and Measurement of Building Materials, University of Jinan, Jinan 250022, Shandong (China); Guan, Ruifang, E-mail: mse_guanrf@ujn.edu.cn [School of Material Science and Engineering, Shandong Provincial Key Laboratory of Preparation and Measurement of Building Materials, University of Jinan, Jinan 250022, Shandong (China); Wang, Kangnan; Shan, Yanyan; Xu, Yongxiao; Ma, Lin [School of Material Science and Engineering, Shandong Provincial Key Laboratory of Preparation and Measurement of Building Materials, University of Jinan, Jinan 250022, Shandong (China)

    2015-07-01

    Four coumarin amide derivatives with 4-methyl coumarin or pyrene as terminal group have been synthesized. Their photophysical properties and recognition properties for cyanide anions have been examined. The results indicate that the compounds can recognize cyanide anions with obvious absorption and fluorescence spectra change, at the same time, obvious color and fluorescence change can be observed by naked eye. The in situ hydrogen nuclear magnetic resonance spectra and photophysical properties change confirm that Michael additions between the chemosensors and cyanide anions take place at the 4-position of coumarin. - Highlights: • Four coumarin amide derivatives with 4-methyl coumarin or pyrene as terminal group were synthesized. • The compounds can recognize cyanide anions with obvious absorption and fluorescence spectra change. • Michael additions between the chemosensors and cyanide anions take place at the 4-position of coumarin.

  5. Nanostructured surfaces and detection instrumentation for photonic crystal enhanced fluorescence.

    Science.gov (United States)

    Chaudhery, Vikram; George, Sherine; Lu, Meng; Pokhriyal, Anusha; Cunningham, Brian T

    2013-04-26

    Photonic crystal (PC) surfaces have been demonstrated as a compelling platform for improving the sensitivity of surface-based fluorescent assays used in disease diagnostics and life science research. PCs can be engineered to support optical resonances at specific wavelengths at which strong electromagnetic fields are utilized to enhance the intensity of surface-bound fluorophore excitation. Meanwhile, the leaky resonant modes of PCs can be used to direct emitted photons within a narrow range of angles for more efficient collection by a fluorescence detection system. The multiplicative effects of enhanced excitation combined with enhanced photon extraction combine to provide improved signal-to-noise ratios for detection of fluorescent emitters, which in turn can be used to reduce the limits of detection of low concentration analytes, such as disease biomarker proteins. Fabrication of PCs using inexpensive manufacturing methods and materials that include replica molding on plastic, nano-imprint lithography on quartz substrates result in devices that are practical for single-use disposable applications. In this review, we will describe the motivation for implementing high-sensitivity fluorescence detection in the context of molecular diagnosis and gene expression analysis though the use of PC surfaces. Recent efforts to improve the design and fabrication of PCs and their associated detection instrumentation are summarized, including the use of PCs coupled with Fabry-Perot cavities and external cavity lasers.

  6. Nanostructured Surfaces and Detection Instrumentation for Photonic Crystal Enhanced Fluorescence

    Directory of Open Access Journals (Sweden)

    Brian T. Cunningham

    2013-04-01

    Full Text Available Photonic crystal (PC surfaces have been demonstrated as a compelling platform for improving the sensitivity of surface-based fluorescent assays used in disease diagnostics and life science research. PCs can be engineered to support optical resonances at specific wavelengths at which strong electromagnetic fields are utilized to enhance the intensity of surface-bound fluorophore excitation. Meanwhile, the leaky resonant modes of PCs can be used to direct emitted photons within a narrow range of angles for more efficient collection by a fluorescence detection system. The multiplicative effects of enhanced excitation combined with enhanced photon extraction combine to provide improved signal-to-noise ratios for detection of fluorescent emitters, which in turn can be used to reduce the limits of detection of low concentration analytes, such as disease biomarker proteins. Fabrication of PCs using inexpensive manufacturing methods and materials that include replica molding on plastic, nano-imprint lithography on quartz substrates result in devices that are practical for single-use disposable applications. In this review, we will describe the motivation for implementing high-sensitivity fluorescence detection in the context of molecular diagnosis and gene expression analysis though the use of PC surfaces. Recent efforts to improve the design and fabrication of PCs and their associated detection instrumentation are summarized, including the use of PCs coupled with Fabry-Perot cavities and external cavity lasers.

  7. Aspect ratio dependent fluorescence quenching of eosin Y by gold nanorods.

    Science.gov (United States)

    Weng, Guojun; Li, Jianjun; Zhang, Li; Zhao, Junwu

    2014-06-01

    Gold nanorods of different aspect ratios had been synthesized using seed mediated growth method. The formed gold nanorods had been characterized by the absorption and transmission electron microscopy (TEM) measurements. The obtained gold nanorods were used to study the quenched effect on fluorescence of Eosin Y. Experimental results revealed that Eosin Y molecules adsorbed on the metallic surfaces, suffering strong quenching of their fluorescence and the quenching efficiency was different for different aspect ratio. Using dielectric coated gold nanorods model, the probable mechanism of aspect ratio dependent quenching efficiency was obtained by numerical calculation based on fluorescence resonance energy transfer and quasi-static theory. The calculation results showed that the non-monotonic changing of fluorescence quenching was attributed to competing effects of aspect ratio and the dielectric constant of coated shell on surface plasmon resonance.

  8. A detection instrument for enhanced-fluorescence and label-free imaging on photonic crystal surfaces.

    Science.gov (United States)

    Block, Ian D; Mathias, Patrick C; Ganesh, Nikhil; Jones, Sarah I; Dorvel, Brian R; Chaudhery, Vikram; Vodkin, Lila O; Bashir, Rashid; Cunningham, Brian T

    2009-07-20

    We report on the design and demonstration of an optical imaging system capable of exciting surface-bound fluorophores within the resonant evanescent electric field of a photonic crystal surface and gathering fluorescence emission that is directed toward the imaging objective by the photonic crystal. The system also has the ability to quantify shifts in the local resonance angle induced by the adsorption of biomolecules on the photonic crystal surface for label-free biomolecular imaging. With these two capabilities combined within a single detection system, we demonstrate label-free images self-registered to enhanced fluorescence images with 328x more sensitive fluorescence detection relative to a glass surface. This technique is applied to a DNA microarray where label-free quantification of immobilized capture DNA enables improved quality control and subsequent enhanced fluorescence detection of dye-tagged hybridized DNA yields 3x more genes to be detected versus commercially available microarray substrates.

  9. FRET-Based Localization of Fluorescent Protein Insertions Within the Ryanodine Receptor Type 1

    OpenAIRE

    Raina, Shweta A.; Jeffrey Tsai; Montserrat Samsó; Fessenden, James D.

    2012-01-01

    Fluorescent protein (FP) insertions have often been used to localize primary structure elements in mid-resolution 3D cryo electron microscopic (EM) maps of large protein complexes. However, little is known as to the precise spatial relationship between the location of the fused FP and its insertion site within a larger protein. To gain insights into these structural considerations, Förster resonance energy transfer (FRET) measurements were used to localize green fluorescent protein (GFP) inse...

  10. Fluorescence energy transfer in quantum dot/azo dye complexes in polymer track membranes

    Science.gov (United States)

    Gromova, Yulia A.; Orlova, Anna O.; Maslov, Vladimir G.; Fedorov, Anatoly V.; Baranov, Alexander V.

    2013-10-01

    Fluorescence resonance energy transfer in complexes of semiconductor CdSe/ZnS quantum dots with molecules of heterocyclic azo dyes, 1-(2-pyridylazo)-2-naphthol and 4-(2-pyridylazo) resorcinol, formed at high quantum dot concentration in the polymer pore track membranes were studied by steady-state and transient PL spectroscopy. The effect of interaction between the complexes and free quantum dots on the efficiency of the fluorescence energy transfer and quantum dot luminescence quenching was found and discussed.

  11. Driving the atom by atomic fluorescence: Analytic results for the power and noise spectra

    OpenAIRE

    2000-01-01

    We study how the spectral properties of resonance fluorescence propagate through a two-atom system. Within the weak-driving-field approximation we find that, as we go from one atom to the next, the power spectrum exhibits both subnatural linewidth narrowing and large asymmetries while the noise spectrum of the squeezed quadrature narrows but remains otherwise unchanged. Analytical results for the observed spectral features of the fluorescence are provided and their origin is thoroughly discus...

  12. The resonant body transistor.

    Science.gov (United States)

    Weinstein, Dana; Bhave, Sunil A

    2010-04-14

    This paper introduces the resonant body transistor (RBT), a silicon-based dielectrically transduced nanoelectromechanical (NEM) resonator embedding a sense transistor directly into the resonator body. Combining the benefits of FET sensing with the frequency scaling capabilities and high quality factors (Q) of internal dielectrically transduced bar resonators, the resonant body transistor achieves >10 GHz frequencies and can be integrated into a standard CMOS process for on-chip clock generation, high-Q microwave circuits, fundamental quantum-state preparation and observation, and high-sensitivity measurements. An 11.7 GHz bulk-mode RBT is demonstrated with a quality factor Q of 1830, marking the highest frequency acoustic resonance measured to date on a silicon wafer.

  13. Resonance energy transfer: Dye to metal nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Wari, M. N.; Pujar, G. H.; Inamdar, S. R., E-mail: him-lax3@yahoo.com [Laser Spectroscopy Programme, Department of Physics, Karnatak University, Dharwad-580003 (India)

    2015-06-24

    In the present study, surface energy transfer (SET) from Coumarin 540A (C540 A) to Gold nanoparticle (Au) is demonstrated. The observed results show pronounced effect on the photoluminescence intensity and shortening of the lifetime of Coumarin 540A upon interaction with the spherical gold nanoparticle, also there are measured effects on radiative rate of the dye. Experimental results are analyzed with fluorescence resonance energy transfer (FRET) and SET theories. The results obtained from distance-dependent quenching provide experimental evidence that the efficiency curve slope and distance of quenching is best modeled by surface energy transfer process.

  14. Dynamically generated resonances

    CERN Document Server

    Oset, E; Sarkar, S; Sun, Bao Xi; Vacas, M J Vicente; González, P; Vijande, J; Jido, D; Sekihara, T; Torres, A Martinez; Khemchandani, K

    2009-01-01

    In this talk I report on recent work related to the dynamical generation of baryonic resonances, some made up from pseudoscalar meson-baryon, others from vector meson-baryon and a third type from two meson-one baryon systems. We can establish a correspondence with known baryonic resonances, reinforcing conclusions previously drawn and bringing new light on the nature of some baryonic resonances of higher mass.

  15. Raman Scattering at Resonant or Near-Resonant Conditions: A Generalized Short-Time Approximation

    Institute of Scientific and Technical Information of China (English)

    Abdelsalam Mohammed; Yu-Ping Sun; Quan Miao; Hans (A)gren; Faris Gel'mukhanov

    2012-01-01

    We investigate the dynamics of resonant Raman scattering in the course of the frequency detuning.The dephasing in the time domain makes the scattering fast when the photon energy is tuned from the absorption resonance.This makes frequency detuning to act as a camera shutter with a regulated scattering duration and provides a practical tool of controlling the scattering time in ordinary stationary measurements.The theory is applied to resonant Raman spectra of a couple of few-mode model systems and to trans-1,3,5-hexatriene and guanine-cytosine (G-C) Watson-Crick base pairs (DNA) molecules.Besides some particular physical effects,the regime of fast scattering leads to a simplification of the spectrum as well as to the scattering theory itself.Strong overtones appear in the Raman spectra when the photon frequency is tuned in the resonant region,while in the mode of fast scattering,the overtones are gradually quenched when the photon frequency is tuned more than one vibrational quantum below the first absorption resonance.The detuning from the resonant region thus leads to a strong purification of the Raman spectrum from the contamination by higher overtones and soft modes and purifies the spectrum also in terms of avoidance of dissociation and interfering fluorescence decay of the resonant state.This makes frequency detuning a very useful practical tool in the analysis of the resonant Raman spectra of complex systems and considerably improves the prospects for using the Raman effect for detection of foreign substances at ultra-low concentrations.

  16. Electron paramagnetic resonance

    CERN Document Server

    Al'tshuler, S A

    2013-01-01

    Electron Paramagnetic Resonance is a comprehensive text on the field of electron paramagnetic resonance, covering both the theoretical background and the results of experiment. This book is composed of eight chapters that cover theoretical materials and experimental data on ionic crystals, since these are the materials that have been most extensively studied by the methods of paramagnetic resonance. The opening chapters provide an introduction to the basic principles of electron paramagnetic resonance and the methods of its measurement. The next chapters are devoted to the theory of spectra an

  17. Combined enhanced fluorescence and label-free biomolecular detection with a photonic crystal surface.

    Science.gov (United States)

    Mathias, Patrick C; Ganesh, Nikhil; Chan, Leo L; Cunningham, Brian T

    2007-04-20

    A 2D photonic crystal surface with a different period in each lateral direction is demonstrated to detect biomolecules using two distinct sensing modalities. The sensing mechanisms both rely on the generation of a resonant reflection peak at one of two specific wavelengths, depending on the polarization of light that is incident on the photonic crystal. One polarization results in a resonant reflection peak in the visible spectrum to coincide with the excitation wavelength of a fluorophore, while the orthogonal polarization results in a resonant reflection peak at an infrared wavelength which is used for label-free detection of adsorbed biomolecules. The photonic crystal resonance for fluorescence excitation causes enhanced near fields at the structure surface, resulting in increased signal from fluorophores within 100 nm of the device surface. Label-free detection is performed by illuminating the photonic crystal with white light and monitoring shifts in the peak reflected wavelength of the infrared resonance with a high-resolution imaging detection instrument. Rigorous coupled-wave analysis was used to determine optimal dimensions for the photonic crystal structure, and devices were fabricated using a polymer-based nanoreplica molding approach. Fluorescence-based and label-free detection were demonstrated using arrays of spots of dye-conjugated streptavidin. Quantification of the fluorescent signal showed that the fluorescence output from protein spots on the photonic crystal was increased by up to a factor of 35, and deposited spots were also imaged in the label-free detection mode.

  18. Highly confined, enhanced surface fluorescence imaging with two-dimensional silver nanoparticle sheets

    Energy Technology Data Exchange (ETDEWEB)

    Usukura, Eiji; Shinohara, Shuhei; Okamoto, Koichi; Tamada, Kaoru, E-mail: tamada@ms.ifoc.kyushu-u.ac.jp [Institute for Materials Chemistry and Engineering, Kyushu University, Fukuoka 812-8581 (Japan); Lim, Jaehoon; Char, Kookheon [The National Creative Research Center for Intelligent Hybrid, School of Chemical and Biological Engineering, Seoul National University, Seoul 151-744 (Korea, Republic of)

    2014-03-24

    A method of obtaining highly confined, enhanced surface fluorescence imaging is proposed using two-dimensional (2D) silver nanoparticle (AgMy) sheets. This technique is based on the localized surface plasmon resonance excited homogeneously on a 2D silver nanoparticle sheet. The AgMy sheets are fabricated at the air–water interface by self-assembly and transferred onto hydrophobic glass substrates. These sheets can enhance the fluorescence only when the excitation wavelength overlaps with the plasmon resonance wavelength. To confirm the validity of this technique, two separate test experiments are performed. One is the epifluorescence microscope imaging of a quantum dot 2D sheet on the AgMy 2D sheet with a SiO{sub 2} spacer layer, where the fluorescence is maximized with the 20 nm SiO{sub 2} layer, determined by the Förster resonance energy transfer distances. The second experiment is the imaging of a single fluorescence bead with a total internal reflection fluorescent microscope. We confirmed that the AgMy sheet provides a 4-fold increase in fluorescence with a 160-nm spatial resolution at 30 ms/frame snapshot. The AgMy sheet will be a powerful tool for high sensitivity and high-resolution real time bioimaging at nanointerfaces.

  19. Fibroblast Cell-Based Therapy for Experimental Autoimmune Diabetes.

    Directory of Open Access Journals (Sweden)

    Reza B Jalili

    Full Text Available Type 1 diabetes (T1D results from autoimmune destruction of insulin producing β cells of the pancreatic islets. Curbing autoimmunity at the initiation of T1D can result in recovery of residual β cells and consequently remission of diabetes. Here we report a cell-based therapy for autoimmune diabetes in non-obese diabetic (NOD mice using dermal fibroblasts. This was achieved by a single injection of fibroblasts, expressing the immunoregulatory molecule indoleamine 2,3 dioxygenase (IDO, into peritoneal cavity of NOD mice shortly after the onset of overt hyperglycemia. Mice were then monitored for reversal of hyperglycemia and changes in inflammatory/regulatory T cell profiles. Blood glucose levels dropped into the normal range in 82% of NOD mice after receiving IDO-expressing fibroblasts while all control mice remained diabetic. We found significantly reduced islet inflammation, increased regulatory T cells, and decreased T helper 17 cells and β cell specific autoreactive CD8+ T cells following IDO cell therapy. We further showed that some of intraperitoneal injected fibroblasts migrated to local lymph nodes and expressed co-inhibitory molecules. These findings suggest that IDO fibroblasts therapy can reinstate self-tolerance and alleviate β cell autoreactivity in NOD mice, resulting in remission of autoimmune diabetes.

  20. Fibroblast Cell-Based Therapy for Experimental Autoimmune Diabetes.

    Science.gov (United States)

    Jalili, Reza B; Zhang, Yun; Hosseini-Tabatabaei, Azadeh; Kilani, Ruhangiz T; Khosravi Maharlooei, Mohsen; Li, Yunyuan; Salimi Elizei, Sanam; Warnock, Garth L; Ghahary, Aziz

    2016-01-01

    Type 1 diabetes (T1D) results from autoimmune destruction of insulin producing β cells of the pancreatic islets. Curbing autoimmunity at the initiation of T1D can result in recovery of residual β cells and consequently remission of diabetes. Here we report a cell-based therapy for autoimmune diabetes in non-obese diabetic (NOD) mice using dermal fibroblasts. This was achieved by a single injection of fibroblasts, expressing the immunoregulatory molecule indoleamine 2,3 dioxygenase (IDO), into peritoneal cavity of NOD mice shortly after the onset of overt hyperglycemia. Mice were then monitored for reversal of hyperglycemia and changes in inflammatory/regulatory T cell profiles. Blood glucose levels dropped into the normal range in 82% of NOD mice after receiving IDO-expressing fibroblasts while all control mice remained diabetic. We found significantly reduced islet inflammation, increased regulatory T cells, and decreased T helper 17 cells and β cell specific autoreactive CD8+ T cells following IDO cell therapy. We further showed that some of intraperitoneal injected fibroblasts migrated to local lymph nodes and expressed co-inhibitory molecules. These findings suggest that IDO fibroblasts therapy can reinstate self-tolerance and alleviate β cell autoreactivity in NOD mice, resulting in remission of autoimmune diabetes.

  1. Photonic crystal enhanced silicon cell based thermophotovoltaic systems.

    Science.gov (United States)

    Yeng, Yi Xiang; Chan, Walker R; Rinnerbauer, Veronika; Stelmakh, Veronika; Senkevich, Jay J; Joannopoulos, John D; Soljacic, Marin; Čelanović, Ivan

    2015-02-09

    We report the design, optimization, and experimental results of large area commercial silicon solar cell based thermophotovoltaic (TPV) energy conversion systems. Using global non-linear optimization tools, we demonstrate theoretically a maximum radiative heat-to-electricity efficiency of 6.4% and a corresponding output electrical power density of 0.39 W cm(-2) at temperature T = 1660 K when implementing both the optimized two-dimensional (2D) tantalum photonic crystal (PhC) selective emitter, and the optimized 1D tantalum pentoxide - silicon dioxide PhC cold-side selective filter. In addition, we have developed an experimental large area TPV test setup that enables accurate measurement of radiative heat-to-electricity efficiency for any emitter-filter-TPV cell combination of interest. In fact, the experimental results match extremely well with predictions of our numerical models. Our experimental setup achieved a maximum output electrical power density of 0.10W cm(-2) and radiative heat-to-electricity efficiency of 1.18% at T = 1380 K using commercial wafer size back-contacted silicon solar cells.

  2. [Safety monitoring of cell-based medicinal products (CBMPs)].

    Science.gov (United States)

    Funk, Markus B; Frech, Marion; Spranger, Robert; Keller-Stanislawski, Brigitte

    2015-11-01

    Cell-based medicinal products (CBMPs), a category of advanced-therapy medicinal products (ATMPs), are authorised for the European market by the European Commission by means of the centralized marketing authorisation. By conforming to the German Medicinal Products Act (Sec. 4b AMG), national authorisation can be granted by the Paul-Ehrlich-Institut in Germany exclusively for ATMPs not based on a routine manufacturing procedure. In both procedures, quality, efficacy, and safety are evaluated and the risk-benefit balance is assessed. For the centralised procedure, mainly controlled clinical trial data must be submitted, whereas the requirements for national procedures could be modified corresponding to the stage of development of the ATMP. After marketing authorization, the marketing authorization/license holder is obligated to report all serious adverse reactions to the competent authority and to provide periodic safety update reports. If necessary, post-authorization safety studies could be imposed. On the basis of these regulatory measures, the safety of advanced therapies can be monitored and improved.

  3. A MULTISCALE, CELL-BASED FRAMEWORK FOR MODELING CANCER DEVELOPMENT

    Energy Technology Data Exchange (ETDEWEB)

    JIANG, YI [Los Alamos National Laboratory

    2007-01-16

    Cancer remains to be one of the leading causes of death due to diseases. We use a systems approach that combines mathematical modeling, numerical simulation, in vivo and in vitro experiments, to develop a predictive model that medical researchers can use to study and treat cancerous tumors. The multiscale, cell-based model includes intracellular regulations, cellular level dynamics and intercellular interactions, and extracellular level chemical dynamics. The intracellular level protein regulations and signaling pathways are described by Boolean networks. The cellular level growth and division dynamics, cellular adhesion and interaction with the extracellular matrix is described by a lattice Monte Carlo model (the Cellular Potts Model). The extracellular dynamics of the signaling molecules and metabolites are described by a system of reaction-diffusion equations. All three levels of the model are integrated through a hybrid parallel scheme into a high-performance simulation tool. The simulation results reproduce experimental data in both avasular tumors and tumor angiogenesis. By combining the model with experimental data to construct biologically accurate simulations of tumors and their vascular systems, this model will enable medical researchers to gain a deeper understanding of the cellular and molecular interactions associated with cancer progression and treatment.

  4. A novel multi-functional cell-based microphysiometer

    Institute of Scientific and Technical Information of China (English)

    XU Ying; XU Gaixia; LIU Qingjun; CAI Hua; LI Yan; LI Rong; WANG Ping

    2006-01-01

    This paper presents a novel multi-functional microphysiometer for simultaneous measurements of several extracellular ion concentrations and action potential measurement in living cells based on MLAPS (multi-light addressable potentiometric sensor). In the microphysiometer, sorts of sensitive membranes are illuminated in parallel with n light sources at working frequencies, and the response amplitudes of each frequency component can be measured on-line by parallel processing algorithm. In the experiments, the relations of the extracellular environmental H+, Na +, K +, Ca2 + under the effects of western medicines (dilantin, phenobarbital sodium) and Chinese drugs (scutellaria, medlar, hemlock parsley) were analyzed, and the effects of several drugs were evaluated. Moreover, the action potential signals of different cell types (cardiac myocytes and neurons) could be measured and analyzed by LAPS. By detecting these parameters, the system can monitor the real-time process of the cell metabolism and action potential, observe the functional responses of different kinds of membrane-bound receptors, and evaluate the activities of drugs.

  5. Neural stem cell-based treatment for neurodegenerative diseases.

    Science.gov (United States)

    Kim, Seung U; Lee, Hong J; Kim, Yun B

    2013-10-01

    Human neurodegenerative diseases such as Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD) are caused by a loss of neurons and glia in the brain or spinal cord. Neurons and glial cells have successfully been generated from stem cells such as embryonic stem cells (ESCs), mesenchymal stem cells (MSCs) and neural stem cells (NSCs), and stem cell-based cell therapies for neurodegenerative diseases have been developed. A recent advance in generation of a new class of pluripotent stem cells, induced pluripotent stem cells (iPSCs), derived from patients' own skin fibroblasts, opens doors for a totally new field of personalized medicine. Transplantation of NSCs, neurons or glia generated from stem cells in animal models of neurodegenerative diseases, including PD, HD, ALS and AD, demonstrates clinical improvement and also life extension of these animals. Additional therapeutic benefits in these animals can be provided by stem cell-mediated gene transfer of therapeutic genes such as neurotrophic factors and enzymes. Although further research is still needed, cell and gene therapy based on stem cells, particularly using neurons and glia derived from iPSCs, ESCs or NSCs, will become a routine treatment for patients suffering from neurodegenerative diseases and also stroke and spinal cord injury.

  6. Highly efficient vacuum processed BHJ solar cell based on merocyanines

    Energy Technology Data Exchange (ETDEWEB)

    Steinmann, Vera; Kronenberg, Nils M.; Lenze, Martin R.; Hertel, Dirk; Meerholz, Klaus [Department fuer Chemie, Universitaet Koeln (Germany); Buerckstuemmer, Hannah; Wuerthner, Frank [Institut fuer Organische Chemie, Roentgen Research Center for Complex Material Systems, Universitaet Wuerzburg (Germany)

    2011-07-01

    Bulk heterojunction (BHJ) organic solar cells have attracted considerable interest due to their potential for large-scale, cost-effective and environmentally friendly power generation. Small molecules have been successfully introduced in solution- (SOL) as well as vacuum- (VAC) processed devices, reporting efficiencies (PCE) up to 4.4% and 5.7% respectively. For simple layer stack devices (2-3 layers) based on CuPc as electron donor and C{sub 60} as electron acceptor PCEs up to 5.0% have been achieved. Recently, we presented a direct comparison of highly efficient SOL and VAC BHJ cells based on merocyanine dyes (MC) with a similarly simple layer stack as reported in the literature. Our most efficient devices exhibited PCEs up to 4.9%. Further optimizations on the VAC processed cells led to high PCEs exceeding 6% while keeping the same simple layer stack. In addition, these cells have demonstrated exceptional performance even at lower light intensities. Due to the simple chemical variability of MC dyes, they are ideally suited for tandem solar cells. We present first attempts in this direction.

  7. A fluorescence scanning electron microscope

    Directory of Open Access Journals (Sweden)

    Takaaki Kanemaru

    2010-01-01

    Full Text Available Fluorescence techniques are widely used in biological research to examine molecular localization, while electron microscopy can provide unique ultrastructural information. To date, correlative images from both fluorescence and electron microscopy have been obtained separately using two different instruments, i.e. a fluorescence microscope (FM and an electron microscope (EM. In the current study, a scanning electron microscope (SEM (JEOL JXA8600 M was combined with a fluorescence digital camera microscope unit and this hybrid instrument was named a fluorescence SEM (FL-SEM. In the labeling of FL-SEM samples, both Fluolid, which is an organic EL dye, and Alexa Fluor, were employed. We successfully demonstrated that the FL-SEM is a simple and practical tool for correlative fluorescence and electron microscopy.

  8. Fluorescence-Based Sensors

    Science.gov (United States)

    Orellana, Guillermo

    The natural luminescent phenomena (from the Latin words "lumen" and "essentia", i.e., "made of light") such as northern lights (aurora borealis), marine brightness, glow-worms, shining putrid fish scales, "bluish"- appearing water when contained in certain wooden cups (quinine fluorescence), some stones heated at high temperatures with reducing agents (BaS phosphorescence), or light emitted while crushing sugar (triboluminescence) already fascinated our ancestors. Nowadays we understand that ultraviolet and visible emission of light originates from a competitive deactivation pathway of the lowest electronic excited state of atoms and molecules that produces the so called luminescence (the sub-terms fluorescence and phosphorescence just designate whether the return of the excited to the ground state is an "allowed" or "forbidden" process, namely it is fast or slow, the loosely-defined border between them being a 1-μs-1 rate constant). Actually, luminescence is the only method to generate light in the known Universe regardless it is powered by the nuclear reactions in the stars, the ohmical heating in bulbs, an electric discharge, the absorption of light or a (bio)chemical reaction (chemiluminescence).

  9. The TALE Fluorescence Detectors

    Science.gov (United States)

    Jui, Charles

    2009-05-01

    The TALE fluorescence detectors are designed to extend the threshold for fluorescence observation by TA down to 3x10^16 eV. It will comprise two main components. The first is a set of 24 telescopes working in stereo, with an existing TA FD station at ˜6 km separation. These will cover between 3-31 degrees in elevation and have azimuthal coverage maximizing the stereo aperture in the 10^18-10^19 eV energy range. The second component consists of 15 telescopes equipped with 4m diameter mirrors and covering the sky between 31 and 73 degrees in elevation. The larger mirror size pushes the physics threshold down to 3x10^16 eV, and provides view of the shower maximum for the lower energy events. The Tower detector will cover one quadrant in azimuth and operate in hybrid mode with the TALE infill array to provide redundant composition measurements from both shower maximum information and muon-to-electron ratio.

  10. Stochastic Resonance in Protein Folding Dynamics.

    Science.gov (United States)

    Davtyan, Aram; Platkov, Max; Gruebele, Martin; Papoian, Garegin A

    2016-05-04

    Although protein folding reactions are usually studied under static external conditions, it is likely that proteins fold in a locally fluctuating cellular environment in vivo. To mimic such behavior in in vitro experiments, the local temperature of the solvent can be modulated either harmonically or using correlated noise. In this study, coarse-grained molecular simulations are used to investigate these possibilities, and it is found that both periodic and correlated random fluctuations of the environment can indeed accelerate folding kinetics if the characteristic frequencies of the applied fluctuations are commensurate with the internal timescale of the folding reaction; this is consistent with the phenomenon of stochastic resonance observed in many other condensed-matter processes. To test this theoretical prediction, the folding dynamics of phosphoglycerate kinase under harmonic temperature fluctuations are experimentally probed using Förster resonance energy transfer fluorescence measurements. To analyze these experiments, a combination of theoretical approaches is developed, including stochastic simulations of folding kinetics and an analytical mean-field kinetic theory. The experimental observations are consistent with the theoretical predictions of stochastic resonance in phosphoglycerate kinase folding. When combined with an alternative experiment on the protein VlsE using a power spectrum analysis, elaborated in Dave et al., ChemPhysChem 2016, 10.1002/cphc.201501041, the overall data overwhelmingly point to the experimental confirmation of stochastic resonance in protein folding dynamics.

  11. Split ring resonator resonance assisted terahertz antennas

    CERN Document Server

    Galal, Hossam; Vitiello, Miriam S

    2016-01-01

    We report on the computational development of novel architectures of low impedance broadband antennas, for efficient detection of Terahertz (THz) frequency beams. The conceived Split Ring Resonator Resonance Assisted (SRR RA) antennas are based on both a capacitive and inductive scheme, exploiting a 200 Ohm and 400 Ohm impedance, respectively. Moreover, the impedance is tunable by varying the coupling parameters in the exploited geometry, allowing for better matching with the detector circuit for maximum power extraction. Our simulation results have been obtained by assuming a 1.5 THz operation frequency.

  12. Fluorescence of ceramic color standards.

    Science.gov (United States)

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

    2010-04-20

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

  13. Resonance and Fractal Geometry

    NARCIS (Netherlands)

    Broer, Henk W.

    2012-01-01

    The phenomenon of resonance will be dealt with from the viewpoint of dynamical systems depending on parameters and their bifurcations. Resonance phenomena are associated to open subsets in the parameter space, while their complement corresponds to quasi-periodicity and chaos. The latter phenomena oc

  14. Modeling and control of fuel cell based distributed generation systems

    Science.gov (United States)

    Jung, Jin Woo

    This dissertation presents circuit models and control algorithms of fuel cell based distributed generation systems (DGS) for two DGS topologies. In the first topology, each DGS unit utilizes a battery in parallel to the fuel cell in a standalone AC power plant and a grid-interconnection. In the second topology, a Z-source converter, which employs both the L and C passive components and shoot-through zero vectors instead of the conventional DC/DC boost power converter in order to step up the DC-link voltage, is adopted for a standalone AC power supply. In Topology 1, two applications are studied: a standalone power generation (Single DGS Unit and Two DGS Units) and a grid-interconnection. First, dynamic model of the fuel cell is given based on electrochemical process. Second, two full-bridge DC to DC converters are adopted and their controllers are designed: an unidirectional full-bridge DC to DC boost converter for the fuel cell and a bidirectional full-bridge DC to DC buck/boost converter for the battery. Third, for a three-phase DC to AC inverter without or with a Delta/Y transformer, a discrete-time state space circuit model is given and two discrete-time feedback controllers are designed: voltage controller in the outer loop and current controller in the inner loop. And last, for load sharing of two DGS units and power flow control of two DGS units or the DGS connected to the grid, real and reactive power controllers are proposed. Particularly, for the grid-connected DGS application, a synchronization issue between an islanding mode and a paralleling mode to the grid is investigated, and two case studies are performed. To demonstrate the proposed circuit models and control strategies, simulation test-beds using Matlab/Simulink are constructed for each configuration of the fuel cell based DGS with a three-phase AC 120 V (L-N)/60 Hz/50 kVA and various simulation results are presented. In Topology 2, this dissertation presents system modeling, modified space

  15. Fluorescence dynamics of green fluorescent protein in AOT reversed micelles

    NARCIS (Netherlands)

    Uskova, M.A.; Borst, J.W.; Hink, M.A.; Hoek, van A.; Schots, A.; Klyachko, N.L.; Visser, A.J.W.G.

    2000-01-01

    We have used the enhanced green fluorescent protein (EGFP) to investigate the properties of surfactant-entrapped water pools in organic solvents (reversed micelles) with steady-state and time-resolved fluorescence methods. The surfactant used was sodium bis(2-ethylhexyl)sulfosuccinate (AOT) and the

  16. The promise of fuel cell-based automobiles

    Indian Academy of Sciences (India)

    A K Shukla; C L Jackson; K Scott

    2003-02-01

    Fuel cell-based automobiles have gained attention in the last few years due to growing public concern about urban air pollution and consequent environmental problems. From an analysis of the power and energy requirements of a modern car, it is estimated that a base sustainable power of $ca$. 50 kW supplemented with short bursts up to 80 kW will suffice in most driving requirements. The energy demand depends greatly on driving characteristics but under normal usage is expected to be 200 Wh/km. The advantages and disadvantages of candidate fuel-cell systems and various fuels are considered together with the issue of whether the fuel should be converted directly in the fuel cell or should be reformed to hydrogen onboard the vehicle. For fuel cell vehicles to compete successfully with conventional internal-combustion engine vehicles, it appears that direct conversion fuel cells using probably hydrogen, but possibly methanol, are the only realistic contenders for road transportation applications. Among the available fuel cell technologies, polymer–electrolyte fuel cells directly fueled with hydrogen appear to be the best option for powering fuel cell vehicles as there is every prospect that these will exceed the performance of the internal-combustion engine vehicles but for their first cost. A target cost of $ 50/kW would be mandatory to make polymer–electrolyte fuel cells competitive with the internal combustion engines and can only be achieved with design changes that would substantially reduce the quantity of materials used. At present, prominent car manufacturers are deploying important research and development efforts to develop fuel cell vehicles and are projecting to start production by 2005.

  17. Development of a Drosophila cell-based error correction assay

    Directory of Open Access Journals (Sweden)

    Jeffrey D. Salemi

    2013-07-01

    Full Text Available Accurate transmission of the genome through cell division requires microtubules from opposing spindle poles to interact with protein super-structures called kinetochores that assemble on each sister chromatid. Most kinetochores establish erroneous attachments that are destabilized through a process called error correction. Failure to correct improper kinetochore-microtubule (kt-MT interactions before anaphase onset results in chromosomal instability (CIN, which has been implicated in tumorigenesis and tumor adaptation. Thus, it is important to characterize the molecular basis of error correction to better comprehend how CIN occurs and how it can be modulated. An error correction assay has been previously developed in cultured mammalian cells in which incorrect kt-MT attachments are created through the induction of monopolar spindle assembly via chemical inhibition of kinesin-5. Error correction is then monitored following inhibitor wash out. Implementing the error correction assay in Drosophila melanogaster S2 cells would be valuable because kt-MT attachments are easily visualized and the cells are highly amenable to RNAi and high-throughput screening. However, Drosophila kinesin-5 (Klp61F is unaffected by available small molecule inhibitors. To overcome this limitation, we have rendered S2 cells susceptible to kinesin-5 inhibitors by functionally replacing Klp61F with human kinesin-5 (Eg5. Eg5 expression rescued the assembly of monopolar spindles typically caused by Klp61F depletion. Eg5-mediated bipoles collapsed into monopoles due to the activity of kinesin-14 (Ncd when treated with the kinesin-5 inhibitor S-trityl-L-cysteine (STLC. Furthermore, bipolar spindles reassembled and error correction was observed after STLC wash out. Importantly, error correction in Eg5-expressing S2 cells was dependent on the well-established error correction kinase Aurora B. This system provides a powerful new cell-based platform for studying error correction and

  18. High content cell-based assay for the inflammatory pathway

    Science.gov (United States)

    Mukherjee, Abhishek; Song, Joon Myong

    2015-07-01

    Cellular inflammation is a non-specific immune response to tissue injury that takes place via cytokine network orchestration to maintain normal tissue homeostasis. However chronic inflammation that lasts for a longer period, plays the key role in human diseases like neurodegenerative disorders and cancer development. Understanding the cellular and molecular mechanisms underlying the inflammatory pathways may be effective in targeting and modulating their outcome. Tumor necrosis factor alpha (TNF-α) is a pro-inflammatory cytokine that effectively combines the pro-inflammatory features with the pro-apoptotic potential. Increased levels of TNF-α observed during acute and chronic inflammatory conditions are believed to induce adverse phenotypes like glucose intolerance and abnormal lipid profile. Natural products e. g., amygdalin, cinnamic acid, jasmonic acid and aspirin have proven efficacy in minimizing the TNF-α induced inflammation in vitro and in vivo. Cell lysis-free quantum dot (QDot) imaging is an emerging technique to identify the cellular mediators of a signaling cascade with a single assay in one run. In comparison to organic fluorophores, the inorganic QDots are bright, resistant to photobleaching and possess tunable optical properties that make them suitable for long term and multicolor imaging of various components in a cellular crosstalk. Hence we tested some components of the mitogen activated protein kinase (MAPK) pathway during TNF-α induced inflammation and the effects of aspirin in HepG2 cells by QDot multicolor imaging technique. Results demonstrated that aspirin showed significant protective effects against TNF-α induced cellular inflammation. The developed cell based assay paves the platform for the analysis of cellular components in a smooth and reliable way.

  19. Development of a Drosophila cell-based error correction assay.

    Science.gov (United States)

    Salemi, Jeffrey D; McGilvray, Philip T; Maresca, Thomas J

    2013-01-01

    Accurate transmission of the genome through cell division requires microtubules from opposing spindle poles to interact with protein super-structures called kinetochores that assemble on each sister chromatid. Most kinetochores establish erroneous attachments that are destabilized through a process called error correction. Failure to correct improper kinetochore-microtubule (kt-MT) interactions before anaphase onset results in chromosomal instability (CIN), which has been implicated in tumorigenesis and tumor adaptation. Thus, it is important to characterize the molecular basis of error correction to better comprehend how CIN occurs and how it can be modulated. An error correction assay has been previously developed in cultured mammalian cells in which incorrect kt-MT attachments are created through the induction of monopolar spindle assembly via chemical inhibition of kinesin-5. Error correction is then monitored following inhibitor wash out. Implementing the error correction assay in Drosophila melanogaster S2 cells would be valuable because kt-MT attachments are easily visualized and the cells are highly amenable to RNAi and high-throughput screening. However, Drosophila kinesin-5 (Klp61F) is unaffected by available small molecule inhibitors. To overcome this limitation, we have rendered S2 cells susceptible to kinesin-5 inhibitors by functionally replacing Klp61F with human kinesin-5 (Eg5). Eg5 expression rescued the assembly of monopolar spindles typically caused by Klp61F depletion. Eg5-mediated bipoles collapsed into monopoles due, in part, to kinesin-14 (Ncd) activity when treated with the kinesin-5 inhibitor S-trityl-L-cysteine (STLC). Furthermore, bipolar spindles reassembled and error correction was observed after STLC wash out. Importantly, error correction in Eg5-expressing S2 cells was dependent on the well-established error correction kinase Aurora B. This system provides a powerful new cell-based platform for studying error correction and CIN.

  20. Fundamentals of nanomechanical resonators

    CERN Document Server

    Schmid, Silvan; Roukes, Michael Lee

    2016-01-01

    This authoritative book introduces and summarizes the latest models and skills required to design and optimize nanomechanical resonators, taking a top-down approach that uses macroscopic formulas to model the devices. The authors cover the electrical and mechanical aspects of nano electromechanical system (NEMS) devices. The introduced mechanical models are also key to the understanding and optimization of nanomechanical resonators used e.g. in optomechanics. Five comprehensive chapters address: The eigenmodes derived for the most common continuum mechanical structures used as nanomechanical resonators; The main sources of energy loss in nanomechanical resonators; The responsiveness of micro and nanomechanical resonators to mass, forces, and temperature; The most common underlying physical transduction mechanisms; The measurement basics, including amplitude and frequency noise. The applied approach found in this book is appropriate for engineering students and researchers working with micro and nanomechanical...

  1. Monolithic MACS micro resonators

    Science.gov (United States)

    Lehmann-Horn, J. A.; Jacquinot, J.-F.; Ginefri, J. C.; Bonhomme, C.; Sakellariou, D.

    2016-10-01

    Magic Angle Coil Spinning (MACS) aids improving the intrinsically low NMR sensitivity of heterogeneous microscopic samples. We report on the design and testing of a new type of monolithic 2D MACS resonators to overcome known limitations of conventional micro coils. The resonators' conductors were printed on dielectric substrate and tuned without utilizing lumped element capacitors. Self-resonance conditions have been computed by a hybrid FEM-MoM technique. Preliminary results reported here indicate robust mechanical stability, reduced eddy currents heating and negligible susceptibility effects. The gain in B1 /√{ P } is in agreement with the NMR sensitivity enhancement according to the principle of reciprocity. A sensitivity enhancement larger than 3 has been achieved in a monolithic micro resonator inside a standard 4 mm rotor at 500 MHz. These 2D resonators could offer higher performance micro-detection and ease of use of heterogeneous microscopic substances such as biomedical samples, microscopic specimens and thin film materials.

  2. Neutrino Production of Resonances

    CERN Document Server

    Paschos, E A; Yu, J Y; Paschos, Emmanuel A.; Sakuda, Makoto; Yu, Ji--Young

    2004-01-01

    We take a fresh look at the analysis of resonance production by neutrinos. We consider three resonances $P_{33}, P_{11}$ and $S_{11}$ with a detailed discussion of their form factors. The article presents results for free proton and neutron targets and discusses the corrections which appear on nuclear targets. The Pauli suppression factor is derived in the Fermi gas model and shown to apply to resonance production. The importance of the various resonances is demonstrated with numerical calculations. The $\\Delta$-resonance is described by two formfactors and its differential cross sections are compared with experimental data. The article is self-contained and could be helpful to readers who wish to reproduce and use these cross sections.

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

    Directory of Open Access Journals (Sweden)

    Sean C Warren

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

  4. Silica nanocapsules of fluorescent conjugated polymers and superparamagnetic nanocrystals for dual-mode cellular imaging.

    Science.gov (United States)

    Tan, Happy; Wang, Miao; Yang, Chang-Tong; Pant, Shilpa; Bhakoo, Kishore Kumar; Wong, Siew Yee; Chen, Zhi-Kuan; Li, Xu; Wang, John

    2011-06-01

    We describe here a facile and benign synthetic strategy to integrate the fluorescent behavior of conjugated polymers and superparamagnetic properties of iron oxide nanocrystals into silica nanocapsules, forming a new type of bifunctional magnetic fluorescent silica nanocapsule (BMFSN). The resultant BMFSNs are uniform, colloidally stable in aqueous medium, and exhibit the desired dual functionality of fluorescence and superparamagnetism in a single entity. Four conjugated polymers with different emissions were used to demonstrate the versatility of employing this class of fluorescent materials for the preparation of BMFSNs. The applicability of BMFSNs in cellular imaging was studied by incubating them with human liver cancer cells, the result of which demonstrated that the cells could be visualized by dual-mode fluorescence and magnetic resonance imaging. Furthermore, the superparamagnetic behavior of the BMFSNs was exploited for in vitro magnetic-guided delivery of the nanocapsules into the cancer cells, thereby highlighting their potential for targeting biomedical applications.

  5. A General Strategy for the Semisynthesis of Ratiometric Fluorescent Sensor Proteins with Increased Dynamic Range.

    Science.gov (United States)

    Xue, Lin; Prifti, Efthymia; Johnsson, Kai

    2016-04-27

    We demonstrate how a combination of self-labeling protein tags and unnatural amino acid technology permits the semisynthesis of ratiometric fluorescent sensor proteins with unprecedented dynamic range in vitro and on live cells. To generate such a sensor, a binding protein is labeled with a fluorescent competitor of the analyte using SNAP-tag in conjugation with a second fluorophore that is positioned in vicinity of the binding site of the binding protein using unnatural amino acid technology. Binding of the analyte by the sensor displaces the tethered fluorescent competitor from the binding protein and disrupts fluorescence resonance energy transfer between the two fluorophores. Using this design principle, we generate a ratiometric fluorescent sensor protein for methotrexate that exhibits large dynamic ranges both in vitro (ratio changes up to 32) and on cell surfaces (ratio change of 13). The performance of these semisynthetic sensor proteins makes them attractive for applications in basic research and diagnostics.

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

    Science.gov (United States)

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

    2012-12-01

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

  7. Fluorescent glutathione probe based on MnO2-phenol formaldehyde resin nanocomposite.

    Science.gov (United States)

    Wang, Xudong; Wang, Dan; Guo, Yali; Yang, Chengduan; Liu, Xiaoyu; Iqbal, Anam; Liu, Weisheng; Qin, Wenwu; Yan, Dan; Guo, Huichen

    2016-03-15

    MnO2-phenol formaldehyde resin (MnO2-PFR) nanocomposite is successfully prepared by a simple chemical reduction process. The resultant MnO2-PFR nanocomposite is well characterized. The absorption band of non-fluorescent MnO2 nanosheets overlaps well with the fluorescence emission of PFR nanoparticles. The green fluorescence of PFR in this nanocomposite can be effectively quenched by fluorescence resonance energy transfer from PFR to MnO2. In the presence of glutathione (GSH), the fluorescence of PFR could be recovered due to MnO2 was reduced to Mn(2+) by GSH. The nanocomposite can be use for detecting glutathione in blood serum.

  8. Green fluorescence of terbium ions in lithium fluoroborate glasses for fibre lasers and display devices

    Indian Academy of Sciences (India)

    G R DILLIP; C MADHUKAR REDDY; M RAJESH; SHIVANAND CHAURASIA; B DEVA PRASAD RAJU; S W JOO

    2016-06-01

    In this paper, for the first time, the visible fluorescence properties, resonance energy transfer mechanism responsible for non-radiative decay rates of ${}^5$D$_4$ $\\to$ ${}^7$F$_5$ transition and also quenching of fluorescence intensity of the ${}^5$D$_3$ $\\to$ ${}^7$F$_5$ transition of various concentrations of Tb$^{3+}$ ions in LBZLFB glasses are reported. Optical absorption, fluorescence spectra and quantum efficiencies are measured and analysed. Green fluorescence related to ${}^5$D$_4$ $\\to$ ${}^7$F$_5$ (548 nm) transition is registered under excitation of 378 nm of Tb$^{3+}$ ions. Based on excitation and fluorescence measurements, several spectroscopic parameters for Tb$^{3+}$ ions are examined as a function of concentration by Judd–Ofelt theory to judge the suitability of studied glasses for display devices and fibre lasers.

  9. Fluorescence energy transfer in the bi-fluorescent S-layer tandem fusion protein ECFP-SgsE-YFP.

    Science.gov (United States)

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

    2010-12-01

    This work reports for the first time on the fabrication of a bi-functional S-layer tandem fusion protein which is able to self-assemble on solid supports without losing its functionality. Two variants of the green fluorescent protein (GFP) were genetically combined with a self-assembly system having the remarkable opportunity to interact with each other and act as functional nanopatterning biocoating. The S-layer protein SgsE of Geobacillus stearothermophilus NRS 2004/3a was fused with the cyan ECFP donor protein at the SgsE N-terminus and with the yellow YFP acceptor protein at the C-terminus. The fluorescence energy transfer was studied with spectrofluorimetry, confocal microscopy and flow cytometry, whilst protein self-assembly (on silicon dioxide particles) and structural investigations were carried out with atomic force microscopy (AFM). The fluorescence resonance energy transfer efficiency of reassembled SgsE tandem protein was 20.0 ± 6.1% which is almost the same transfer efficiency shown in solution (19.6 ± 0.1%). This work shows that bi-fluorescent S-layer fusion proteins self-assemble on silica particles retaining their fluorescent properties.

  10. Twisting Fluorescence through Extrinsic Chiral Antennas.

    Science.gov (United States)

    Yan, Chen; Wang, Xiaolong; Raziman, T V; Martin, Olivier J F

    2017-03-22

    Plasmonic antennas and planar structures have been undergoing intensive developments in order to control the scattering and absorption of light. One specific class, extrinsic chiral surfaces, that does not possess 2-fold rotational symmetry exhibits strong asymmetric transmission for different circular polarizations under obliquely incident illumination. In this work, we show that the design of those surfaces can be optimized with complex multipolar resonances in order to twist the fluorescence emission from nearby molecules. While this emission is usually dipolar and linearly polarized, the interaction with these resonances twists it into a multipolar radiation pattern with opposite helicity in different directions. The proposed structure maximizes this effect and provides control over the polarization of light. Splitting of left- and right-handed circularly polarized light is experimentally obtained in the backward direction. These results highlight the intricate interplay between the near-field absorption and the far-field scattering of a plasmonic nanostructure and are further used for modifying the emission of incoherent quantum sources. Our finding can potentially lead to the development of polarization- and angle-resolved ultracompact optical devices.

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

    Science.gov (United States)

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

    2014-11-21

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

  12. Reliability of acridine orange fluorescence microscopy in oral cytodiagnosis

    Directory of Open Access Journals (Sweden)

    Nilima Prakash

    2011-01-01

    Full Text Available Context and Aims: The oral cavity is the most predominant location in the head and neck region for primary malignant epithelial tumors. Oral cancer is estimated to be the sixth most common malignancy. Early recognition is imperative for successful treatment and good prognosis. Exfoliative cytology is a simple and reasonably effective technique for rapid initial evaluation of a suspicious oral lesion. The present study was conducted to determine the reliability of acridine orange fluorescence microscopy for cytodiagnosis as a more rapid and easier method for the final evaluation of the cytological specimen. Materials and Methods: Smears were collected from 20 individuals with oral lesions suspicious of malignancy, oral lesions not suggestive of malignancy and normal buccal mucosa. One smear was stained with Papanicolaou stain and another one with acridine orange stain. The differences in the study group and control group were compared by means of the χ2 (Chi-square test. The results were considered statistically significant whenever P was <0.05. Results: The acridine orange fluorescence stain reliably demonstrated malignant cells based on the differential fluorescence - a cytochemical criterion. The efficacy of the stain was higher than the conventional Papanicolaou stain in screening of oral lesions suspicious of malignancy. However, the acridine orange fluorescence stain did not differentiate effectively between malignant cells and rapidly proliferating cells, as the technique is based on the nucleic acid content. Conclusion: The fluorescent acridine orange method can be used reliably for the screening of carcinomas and it is especially helpful in the follow-up detection of recurrent carcinoma in previously treated cases.

  13. Resonance Radiation and Excited Atoms

    Science.gov (United States)

    Mitchell, Allan C. G.; Zemansky, Mark W.

    2009-06-01

    1. Introduction; 2. Physical and chemical effects connected with resonance radiation; 3. Absorption lines and measurements of the lifetime of the resonance state; 4. Collision processes involving excited atoms; 5. The polarization of resonance radiation; Appendix; Index.

  14. X-ray Fluorescence Sectioning

    CERN Document Server

    Cong, Wenxiang

    2012-01-01

    In this paper, we propose an x-ray fluorescence imaging system for elemental analysis. The key idea is what we call "x-ray fluorescence sectioning". Specifically, a slit collimator in front of an x-ray tube is used to shape x-rays into a fan-beam to illuminate a planar section of an object. Then, relevant elements such as gold nanoparticles on the fan-beam plane are excited to generate x-ray fluorescence signals. One or more 2D spectral detectors are placed to face the fan-beam plane and directly measure x-ray fluorescence data. Detector elements are so collimated that each element only sees a unique area element on the fan-beam plane and records the x-ray fluorescence signal accordingly. The measured 2D x-ray fluorescence data can be refined in reference to the attenuation characteristics of the object and the divergence of the beam for accurate elemental mapping. This x-ray fluorescence sectioning system promises fast fluorescence tomographic imaging without a complex inverse procedure. The design can be ad...

  15. Assessing Photosynthesis by Fluorescence Imaging

    Science.gov (United States)

    Saura, Pedro; Quiles, Maria Jose

    2011-01-01

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

  16. Optical Properties of Fluorescent Dyes

    Institute of Scientific and Technical Information of China (English)

    李戎; 陈东辉

    2001-01-01

    Fluorescent dyes have been widely used these years.Because of the special optical performance, conventional CCM systems seem to be unable to predict the recipes of fabrics dyed with fluorescent dyes. In order to enhance the functions of CCM systems, the optical properties of fluorescent dyes in their absorption region were investigated. It has been found that there was a fixed maximum absorption wavelength for each fluorescent dyes whatever its concentration is. Both absorption region and maximum absorption wavelength of the dyes in solution are the same to those in fabric, and that the absorption is directly proportional to the concentration of the dye. So the optical properties obtained in solutions cna be applied for describing the optics performance of fluorescent dyes in fabrics.

  17. Advances in magnetic resonance 10

    CERN Document Server

    Waugh, John S

    2013-01-01

    Advances in Magnetic Resonance, Volume 10, presents a variety of contributions to the theory and practice of magnetic resonance. The book contains three chapters that examine superoperators in magnetic resonance; ultrasonically modulated paramagnetic resonance; and the utility of electron paramagnetic resonance (EPR) and electron-nuclear double-resonance (ENDOR) techniques for studying low-frequency modes of atomic fluctuations and their significance for understanding the mechanism of structural phase transitions in solids.

  18. Tunable multiwalled nanotube resonator

    Science.gov (United States)

    Zettl, Alex K.; Jensen, Kenneth J.; Girit, Caglar; Mickelson, William E.; Grossman, Jeffrey C.

    2011-03-29

    A tunable nanoscale resonator has potential applications in precise mass, force, position, and frequency measurement. One embodiment of this device consists of a specially prepared multiwalled carbon nanotube (MWNT) suspended between a metal electrode and a mobile, piezoelectrically controlled contact. By harnessing a unique telescoping ability of MWNTs, one may controllably slide an inner nanotube core from its outer nanotube casing, effectively changing its length and thereby changing the tuning of its resonance frequency. Resonant energy transfer may be used with a nanoresonator to detect molecules at a specific target oscillation frequency, without the use of a chemical label, to provide label-free chemical species detection.

  19. Spin Resonance Strength Calculations

    Science.gov (United States)

    Courant, E. D.

    2009-08-01

    In calculating the strengths of depolarizing resonances it may be convenient to reformulate the equations of spin motion in a coordinate system based on the actual trajectory of the particle, as introduced by Kondratenko, rather than the conventional one based on a reference orbit. It is shown that resonance strengths calculated by the conventional and the revised formalisms are identical. Resonances induced by radiofrequency dipoles or solenoids are also treated; with rf dipoles it is essential to consider not only the direct effect of the dipole but also the contribution from oscillations induced by it.

  20. Spin resonance strength calculations

    Energy Technology Data Exchange (ETDEWEB)

    Courant,E.D.

    2008-10-06

    In calculating the strengths of depolarizing resonances it may be convenient to reformulate the equations of spin motion in a coordinate system based on the actual trajectory of the particle, as introduced by Kondratenko, rather than the conventional one based on a reference orbit. It is shown that resonance strengths calculated by the conventional and the revised formalisms are identical. Resonances induced by radiofrequency dipoles or solenoids are also treated; with rf dipoles it is essential to consider not only the direct effect of the dipole but also the contribution from oscillations induced by it.

  1. Resonant power converters

    CERN Document Server

    Kazimierczuk, Marian K

    2012-01-01

    This book is devoted to resonant energy conversion in power electronics. It is a practical, systematic guide to the analysis and design of various dc-dc resonant inverters, high-frequency rectifiers, and dc-dc resonant converters that are building blocks of many of today's high-frequency energy processors. Designed to function as both a superior senior-to-graduate level textbook for electrical engineering courses and a valuable professional reference for practicing engineers, it provides students and engineers with a solid grasp of existing high-frequency technology, while acquainting them wit

  2. Magnetic Resonance Force Microscopy System

    Data.gov (United States)

    Federal Laboratory Consortium — The Magnetic Resonance Force Microscopy (MRFM) system, developed by ARL, is the world's most sensitive nuclear magnetic resonance (NMR) spectroscopic analysis tool,...

  3. The Tracking Resonance Frequency Method for Photoacoustic Measurements Based on the Phase Response

    Science.gov (United States)

    Suchenek, Mariusz

    2017-04-01

    One of the major issues in the use of the resonant photoacoustic cell is the resonance frequency of the cell. The frequency is not stable, and its changes depend mostly on temperature and gas mixture. This paper presents a new method for tracking resonance frequency, where both the amplitude and phase are calculated from the input samples. The stimulating frequency can be adjusted to the resonance frequency of the cell based on the phase. This method was implemented using a digital measurement system with an analog to digital converter, field programmable gate array (FPGA) and a microcontroller. The resonance frequency was changed by the injection of carbon dioxide into the cell. A theoretical description and experimental results are also presented.

  4. Two-dimensional fluorescence spectroscopy of laser-produced plasmas

    Energy Technology Data Exchange (ETDEWEB)

    Harilal, Sivanandan S.; LaHaye, Nicole L.; Phillips, Mark C.

    2016-08-01

    We use a two-dimensional laser-induced fluorescence spectroscopy technique to measure the coupled absorption and emission properties of atomic species in plasmas produced via laser ablation of solid aluminum targets at atmospheric pressure. Emission spectra from the Al I 394.4 nm and Al I 396.15 nm transitions are measured while a frequency-doubled, continuous-wave, Ti:Sapphire laser is tuned across the Al I 396.15 nm transition. The resulting two-dimensional spectra show the energy coupling between the two transitions via increased emission intensity for both transitions during resonant absorption of the continuous-wave laser at one transition. Time-delayed and gated detection of the emission spectrum is used to isolate the resonantly-excited fluorescence emission from the thermally-excited emission from the plasma. In addition, the tunable continuous-wave laser measures the absorption spectrum of the Al transition with ultra-high resolution after the plasma has cooled, resulting in narrower spectral linewidths than observed in emission spectra. Our results highlight that fluorescence spectroscopy employing continuous-wave laser re-excitation after pulsed laser ablation combines benefits of both traditional emission and absorption spectroscopic methods.

  5. Improved "optical highlighter" probes derived from discosoma red fluorescent protein.

    Science.gov (United States)

    Robinson, Lisbeth C; Marchant, Jonathan S

    2005-02-01

    The tetrameric red fluorescent protein, DsRed, undergoes a rapid red to green color change evoked by short wavelength (lambda highlighter" probe for tracking live cells, organelles, and fusion proteins. This color change results from selective bleaching of the "mature" red-emitting species of DsRed and an enhancement of emission from the "immature" green species, likely caused by dequenching of fluorescence resonance energy transfer occurring within the protein tetramer. Here, we have examined the role of residues known to influence the rate and completeness of chromophore maturation on the cellular and biophysical properties of DsRed mutants. Surprisingly, a single amino acid mutation (N42Q) with increased basal green emission yet rapid chromophore maturation displayed a multiphoton-evoked color change that was brighter, more consistent, more vivid, and easier to evoke than DsRed, despite the larger proportion of green chromophores. Rapidly maturing mutants with more complete chromophore maturation, exhibited little color change and increased resistance to multiphoton bleaching. We describe improved optical and cell biological properties for two DsRed-derived variants which we showcase in photolabeling studies, and discuss these data in terms of implications for fluorescence resonance energy transfer-based probes.

  6. Oligomerization of epidermal growth factor receptors (EGFR) on A431 cells studied by time-resolved fluorescence imaging microscopy: a stereochemical model for tyrosine kinase receptor activation

    NARCIS (Netherlands)

    Th.W.J. Gadella; T.M. Jovin

    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 en- ergy transfer: donor photobleaching fluorescence reso- nance energy transfer (pbFRET) microscopy and

  7. Rise-time of FRET-acceptor fluorescence tracks protein folding

    NARCIS (Netherlands)

    Lindhoud, S.; Westphal, A.H.; Van Mierlo, C.P.M.; Visser, A.J.W.G.; Borst, J.W.

    2014-01-01

    Uniform labeling of proteins with fluorescent donor and acceptor dyes with an equimolar ratio is paramount for accurate determination of Förster resonance energy transfer (FRET) efficiencies. In practice, however, the labeled protein population contains donor-labeled molecules that have no correspon

  8. Ultimate statistical physics: fluorescence of a single atom

    Science.gov (United States)

    Pomeau, Yves; Le Berre, Martine; Ginibre, Jean

    2016-10-01

    We discuss the statistics of emission of photons by a single atom or ion illuminated by a laser beam at the frequency of quasi-resonance between two energy levels, a situation that corresponds to real experiments. We extend this to the case of two laser beams resonant with the energy differences between two excited levels and the ground state (three level atom in V-configuration). We use a novel approach to this type of problem by considering a Kolmogorov equation for the probability distribution of the atomic state, which takes into account first the deterministic evolution of this state under the effect of the incoming laser beam and second the random emission of photons during the spontaneous decay of the excited state(s) to the ground state. This approach yields solvable equations in the two level atom case. For the three level atom case we set the problem and clearly define its frame. The results obtained are valid in both opposite limits of rare and frequent spontaneous decay, compared to the period of the optical Rabi oscillations due to the interaction between resonant excitation and atomic levels. Our analysis gives access to various statistical properties of the fluorescence light, including one showing that its fluctuations in time are not invariants under time reversal. This result makes evident the fundamentally irreversible character of quantum measurements, represented here by the emission of photons of fluorescence.

  9. Resonant Diphoton Phenomenology Simplified

    CERN Document Server

    Panico, Giuliano; Wulzer, Andrea

    2016-01-01

    A framework is proposed to describe resonant diphoton phenomenology at hadron colliders in full generality. It can be employed for a comprehensive model-independent interpretation of the experimental data. Within the general framework, few benchmark scenarios are defined as representative of the various phenomenological options and/or of motivated new physics scenarios. Their usage is illustrated by performing a characterization of the 750 GeV excess, based on a recast of available experimental results. We also perform an assessment of which properties of the resonance could be inferred, after discovery, by a careful experimental study of the diphoton distributions. These include the spin J of the new particle and its dominant production mode. Partial information on its CP-parity can also be obtained, but only for J >= 2. The complete determination of the resonance CP properties requires studying the pattern of the initial state radiation that accompanies the resonance production.

  10. Resonances in QCD

    Energy Technology Data Exchange (ETDEWEB)

    Lutz, Matthias F. M.; Lange, Jens Sören; Pennington, Michael; Bettoni, Diego; Brambilla, Nora; Crede, Volker; Eidelman, Simon; Gillitzer, Albrecht; Gradl, Wolfgang; Lang, Christian B.; Metag, Volker; Nakano, Takashi; Nieves, Juan; Neubert, Sebastian; Oka, Makoto; Olsen, Stephen L.; Pappagallo, Marco; Paul, Stephan; Pelizäus, Marc; Pilloni, Alessandro; Prencipe, Elisabetta; Ritman, Jim; Ryan, Sinead; Thoma, Ulrike; Uwer, Ulrich; Weise, Wolfram

    2016-04-01

    We report on the EMMI Rapid Reaction Task Force meeting 'Resonances in QCD', which took place at GSI October 12-14, 2015 (Fig.~1). A group of 26 people met to discuss the physics of resonances in QCD. The aim of the meeting was defined by the following three key questions; what is needed to understand the physics of resonances in QCD?; where does QCD lead us to expect resonances with exotic quantum numbers?; and what experimental efforts are required to arrive at a coherent picture? For light mesons and baryons only those with up, down and strange quark content were considered. For heavy-light and heavy-heavy meson systems, those with charm quarks were the focus.This document summarizes the discussions by the participants, which in turn led to the coherent conclusions we present here.

  11. Resonant Thermoelectric Nanophotonics

    CERN Document Server

    Mauser, Kelly W; Kim, Seyoon; Fleischman, Dagny; Atwater, Harry A

    2016-01-01

    Photodetectors are typically based on photocurrent generation from electron-hole pairs in semiconductor structures and on bolometry for wavelengths that are below bandgap absorption. In both cases, resonant plasmonic and nanophotonic structures have been successfully used to enhance performance. In this work, we demonstrate subwavelength thermoelectric nanostructures designed for resonant spectrally selective absorption, which creates large enough localized temperature gradients to generate easily measureable thermoelectric voltages. We show that such structures are tunable and are capable of highly wavelength specific detection, with an input power responsivity of up to 119 V/W (referenced to incident illumination), and response times of nearly 3 kHz, by combining resonant absorption and thermoelectric junctions within a single structure, yielding a bandgap-independent photodetection mechanism. We report results for both resonant nanophotonic bismuth telluride-antimony telluride structures and chromel-alumel...

  12. Resonance Production in Jet

    CERN Document Server

    Markert, Christina

    2007-01-01

    Hadronic resonances with short life times and strong coupling to the dense medium may exhibit mass shifts and width broadening as signatures of chiral symmetry restoration at the phase transition between hadronic and partonic matter. Resonances with different lifetimes are also used to extract information about the time evolution and temperature of the expanding hadronic medium. In order to collect information about the early stage (at the phase transition) of a heavy-ion collision, resonances and decay particles which are unaffected by the hadronic medium have to be used. We explore a possible new technique to extract signals from the early stage through the selection of resonances from jets. A first attempt of this analysis, using the reconstructed $\\phi$(1020) from 200 GeV Au+Au collisions in STAR, is presented.

  13. Triple-resonant transducers.

    Science.gov (United States)

    Butler, Stephen C

    2012-06-01

    A detailed analysis is presented of two novel multiple-resonant transducers which produce a wider transmit response than that of a conventional Tonpilz-type transducer. These multi-resonant transducers are Tonpilz-type longitudinal vibrators that produce three coupled resonances and are referred to as triple-resonant transducers (TRTs). One of these designs is a mechanical series arrangement of a tail mass, piezoelectric ceramic stack, central mass, compliant spring, second central mass, second compliant spring, and a piston-radiating head mass. The other TRT design is a mechanical series arrangement of a tail mass, piezoelectric ceramic stack, central mass, compliant spring, and head mass with a quarter-wave matching layer of poly(methyl methacrylate) on the head mass. Several prototype transducer element designs were fabricated that demonstrated proof-of-concept.

  14. Magnetic Resonance Sensors

    Directory of Open Access Journals (Sweden)

    Robert H. Morris

    2014-11-01

    Full Text Available Magnetic Resonance finds countless applications, from spectroscopy to imaging, routinely in almost all research and medical institutions across the globe. It is also becoming more frequently used for specific applications in which the whole instrument and system is designed for a dedicated application. With beginnings in borehole logging for the petro-chemical industry Magnetic Resonance sensors have been applied to fields as varied as online process monitoring for food manufacture and medical point of care diagnostics. This great diversity is seeing exciting developments in magnetic resonance sensing technology published in application specific journals where they are often not seen by the wider sensor community. It is clear that there is enormous interest in magnetic resonance sensors which represents a significant growth area. The aim of this special edition of Sensors was to address the wide distribution of relevant articles by providing a forum to disseminate cutting edge research in this field in a single open source publication.[...

  15. Resonances in QCD

    Science.gov (United States)

    Lutz, Matthias F. M.; Lange, Jens Sören; Pennington, Michael; Bettoni, Diego; Brambilla, Nora; Crede, Volker; Eidelman, Simon; Gillitzer, Albrecht; Gradl, Wolfgang; Lang, Christian B.; Metag, Volker; Nakano, Takashi; Nieves, Juan; Neubert, Sebastian; Oka, Makoto; Olsen, Stephen L.; Pappagallo, Marco; Paul, Stephan; Pelizäus, Marc; Pilloni, Alessandro; Prencipe, Elisabetta; Ritman, Jim; Ryan, Sinead; Thoma, Ulrike; Uwer, Ulrich; Weise, Wolfram

    2016-04-01

    We report on the EMMI Rapid Reaction Task Force meeting 'Resonances in QCD', which took place at GSI October 12-14, 2015. A group of 26 people met to discuss the physics of resonances in QCD. The aim of the meeting was defined by the following three key questions: What is needed to understand the physics of resonances in QCD? Where does QCD lead us to expect resonances with exotic quantum numbers? What experimental efforts are required to arrive at a coherent picture? For light mesons and baryons only those with up, down and strange quark content were considered. For heavy-light and heavy-heavy meson systems, those with charm quarks were the focus. This document summarizes the discussions by the participants, which in turn led to the coherent conclusions we present here.

  16. Resonances in QCD

    CERN Document Server

    Lutz, Matthias F M; Pennington, Michael; Bettoni, Diego; Brambilla, Nora; Crede, Volker; Eidelman, Simon; Gillitzer, Albrecht; Gradl, Wolfgang; Lang, Christian B; Metag, Volker; Nieves, Juan; Neubert, Sebastian; Oka, Makoto; Olsen, Steve L; Pappagallo, Marco; Paul, Stephan; Pelizäus, Marc; Pilloni, Alessandro; Prencipe, Elisabetta; Ritman, Jim; Ryan, Sinead; Thoma, Ulrike; Uwer, Ulrich; Weise, Wolfram

    2015-01-01

    We report on the EMMI Rapid Reaction Task Force meeting 'Resonances in QCD', which took place at GSI October 12-14, 2015. A group of 26 people met to discuss the physics of resonances in QCD. The aim of the meeting was defined by the following three key questions: What is needed to understand the physics of resonances in QCD? Where does QCD lead us to expect resonances with exotic quantum numbers? What experimental efforts are required to arrive at a coherent picture? For light mesons and baryons only those with ${\\it up}$, ${\\it down}$ and ${\\it strange}$ quark content were considered. For heavy-light and heavy-heavy meson systems, those with ${\\it charm}$ quarks were the focus. This document summarizes the discussions by the participants, which in turn led to the coherent conclusions we present here.

  17. Lattices of dielectric resonators

    CERN Document Server

    Trubin, Alexander

    2016-01-01

    This book provides the analytical theory of complex systems composed of a large number of high-Q dielectric resonators. Spherical and cylindrical dielectric resonators with inferior and also whispering gallery oscillations allocated in various lattices are considered. A new approach to S-matrix parameter calculations based on perturbation theory of Maxwell equations, developed for a number of high-Q dielectric bodies, is introduced. All physical relationships are obtained in analytical form and are suitable for further computations. Essential attention is given to a new unified formalism of the description of scattering processes. The general scattering task for coupled eigen oscillations of the whole system of dielectric resonators is described. The equations for the  expansion coefficients are explained in an applicable way. The temporal Green functions for the dielectric resonator are presented. The scattering process of short pulses in dielectric filter structures, dielectric antennas  and lattices of d...

  18. Piezoelectric MEMS resonators

    CERN Document Server

    Piazza, Gianluca

    2017-01-01

    This book introduces piezoelectric microelectromechanical (pMEMS) resonators to a broad audience by reviewing design techniques including use of finite element modeling, testing and qualification of resonators, and fabrication and large scale manufacturing techniques to help inspire future research and entrepreneurial activities in pMEMS. The authors discuss the most exciting developments in the area of materials and devices for the making of piezoelectric MEMS resonators, and offer direct examples of the technical challenges that need to be overcome in order to commercialize these types of devices. Some of the topics covered include: Widely-used piezoelectric materials, as well as materials in which there is emerging interest Principle of operation and design approaches for the making of flexural, contour-mode, thickness-mode, and shear-mode piezoelectric resonators, and examples of practical implementation of these devices Large scale manufacturing approaches, with a focus on the practical aspects associate...

  19. Perspective on resonances of metamaterials.

    Science.gov (United States)

    Min, Li; Huang, Lirong

    2015-07-27

    Electromagnetic resonance as the most important characteristic of metamaterials enables lots of exotic phenomena, such as invisible, negative refraction, man-made magnetism, etc. Conventional LC-resonance circuit model as the most authoritative and classic model is good at explaining and predicting the fundamental resonance wavelength of a metamaterial, while feels hard for high-order resonances, especially for resonance intensity (strength of resonance, determining on the performance and efficiency of metamaterial-based devices). In present work, via an easy-to-understand mass-spring model, we present a different and comprehensive insight for the resonance mechanism of metamaterials, through which both the resonance wavelengths (including the fundamental and high-order resonance wavelengths) and resonance intensities of metamaterials can be better understood. This developed theory has been well verified by different-material and different-structure resonators. This perspective will provide a broader space for exploring novel optical devices based on metamaterials (or metasurfaces).

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

    Science.gov (United States)

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

    2012-05-25

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