Larval vision contributes to gregarious settlement in barnacles: adult red fluorescence as a possible visual signal

KAUST Repository

Matsumura, K.

2014-02-26

Gregarious settlement, an essential behavior for many barnacle species that can only reproduce by mating with a nearby barnacle, has long been thought to rely on larval ability to recognize chemical signals from conspecifics during settlement. However, the cyprid, the settlement stage larva in barnacles, has one pair of compound eyes that appear only at the late nauplius VI and cyprid stages, but the function(s) of these eyes remains unknown. Here we show that cyprids of the intertidal barnacle Balanus (=Amphibalanus) amphitrite can locate adult barnacles even in the absence of chemical cues, and prefer to settle around them probably via larval sense of vision. We also show that the cyprids can discriminate color and preferred to settle on red surfaces. Moreover, we found that shells of adult B. amphitrite emit red auto-fluorescence and the adult extracts with the fluorescence as a visual signal attracted cyprid larvae to settle around it. We propose that the perception of specific visual signals can be involved in behavior of zooplankton including marine invertebrate larvae, and that barnacle auto-fluorescence may be a specific signal involved in gregarious larval settlement.

Fluorescence In Situ Hybridization (FISH Signal Analysis Using Automated Generated Projection Images

Directory of Open Access Journals (Sweden)

Xingwei Wang

2012-01-01

Full Text Available Fluorescence in situ hybridization (FISH tests provide promising molecular imaging biomarkers to more accurately and reliably detect and diagnose cancers and genetic disorders. Since current manual FISH signal analysis is low-efficient and inconsistent, which limits its clinical utility, developing automated FISH image scanning systems and computer-aided detection (CAD schemes has been attracting research interests. To acquire high-resolution FISH images in a multi-spectral scanning mode, a huge amount of image data with the stack of the multiple three-dimensional (3-D image slices is generated from a single specimen. Automated preprocessing these scanned images to eliminate the non-useful and redundant data is important to make the automated FISH tests acceptable in clinical applications. In this study, a dual-detector fluorescence image scanning system was applied to scan four specimen slides with FISH-probed chromosome X. A CAD scheme was developed to detect analyzable interphase cells and map the multiple imaging slices recorded FISH-probed signals into the 2-D projection images. CAD scheme was then applied to each projection image to detect analyzable interphase cells using an adaptive multiple-threshold algorithm, identify FISH-probed signals using a top-hat transform, and compute the ratios between the normal and abnormal cells. To assess CAD performance, the FISH-probed signals were also independently visually detected by an observer. The Kappa coefficients for agreement between CAD and observer ranged from 0.69 to 1.0 in detecting/counting FISH signal spots in four testing samples. The study demonstrated the feasibility of automated FISH signal analysis that applying a CAD scheme to the automated generated 2-D projection images.

Detection and Interpretation of Fluorescence Signals Generated by Excitable Cells and Tissues

Science.gov (United States)

Costantino, Anthony J.

Part 1: High-Sensitivity Amplifiers for Detecting Fluorescence . Monitoring electrical activity and Cai 2+ transients in biological tissues and individual cells increasingly utilizes optical sensors based on voltage-dependent and Cai 2+-dependent fluorescent dyes. However, achieving satisfactory signal-to-noise ratios (SNR) often requires increased illumination intensities and/or dye concentrations, which results in photo-toxicity, photo-bleaching and other adverse effects limiting the utility of optical recordings. The most challenging are the recordings from individual cardiac myocytes and neurons. Here we demonstrate that by optimizing a conventional transimpedance topology one can achieve a 10-20 fold increase of sensitivity with photodiode-based recording systems (dependent on application). We provide a detailed comparative analysis of the dynamic and noise characteristics of different transimpedance amplifier topologies as well as the example(s) of their practical implementation. Part 2: Light-Scattering Models for Interpretation of Fluorescence Data. Current interest in understanding light transport in cardiac tissue has been motivated in part by increased use of voltage-sensitive and Ca i2+-sensitive fluorescent probes to map electrical impulse propagation and Cai2+-transients in the heart. The fluorescent signals are recorded using such probes represent contributions from different layers of myocardial tissue and are greatly affected by light scattering. The interpretation of these signals thus requires deconvolution which would not be possible without detailed models of light transport in the respective tissue. Which involves the experimental measurements of the absorption, scattering, and anisotropy coefficients, mua, mu s, and g respectively. The aim of the second part of our thesis was to derive a new method for deriving these parameters from high spatial resolution measurements of forward-directed flux (FDF). To this end, we carried out high spatial

Mechanistic studies of the genetically encoded fluorescent protein voltage probe ArcLight.

Directory of Open Access Journals (Sweden)

Zhou Han

Full Text Available ArcLight, a genetically encoded fluorescent protein voltage probe with a large ΔF/ΔV, is a fusion between the voltage sensing domain of the Ciona instestinalis voltage sensitive phosphatase and super ecliptic pHluorin carrying a single mutation (A227D in the fluorescent protein. Without this mutation the probe produces only a very small change in fluorescence in response to voltage deflections (∼ 1%. The large signal afforded by this mutation allows optical detection of action potentials and sub-threshold electrical events in single-trials in vitro and in vivo. However, it is unclear how this single mutation produces a probe with such a large modulation of its fluorescence output with changes in membrane potential. In this study, we identified which residues in super ecliptic pHluorin (vs eGFP are critical for the ArcLight response, as a similarly constructed probe based on eGFP also exhibits large response amplitude if it carries these critical residues. We found that D147 is responsible for determining the pH sensitivity of the fluorescent protein used in these probes but by itself does not result in a voltage probe with a large signal. We also provide evidence that the voltage dependent signal of ArcLight is not simply sensing environmental pH changes. A two-photon polarization microscopy study showed that ArcLight's response to changes in membrane potential includes a reorientation of the super ecliptic pHluorin. We also explored different changes including modification of linker length, deletion of non-essential amino acids in the super ecliptic pHluorin, adding a farnesylation site, using tandem fluorescent proteins and other pH sensitive fluorescent proteins.

Fluorescent protein pair emit intracellular FRET signal suitable for FACS screening

International Nuclear Information System (INIS)

Johansson, Daniel X.; Brismar, Hjalmar; Persson, Mats A.A.

2007-01-01

The fluorescent proteins ECFP and HcRed were shown to give an easily resolved FRET-signal when expressed as a fusion inside mammalian cells. HeLa-tat cells expressing ECFP, pHcRed, or the fusion protein pHcRed-ECFP were analyzed by flow cytometry after excitation of ECFP. Cells expressing HcRed-ECFP, or ECFP and HcRed, were mixed and FACS-sorted for FRET positive cells: HcRed-ECFP cells were greatly enriched (72 times). Next, cloned human antibodies were fused with ECFP and expressed anchored to the ER membrane. Their cognate antigens (HIV-1 gp120 or gp41) were fused to HcRed and co-expressed in the ER. An increase of 13.5 ± 1.5% (mean ± SEM) and 8.0 ± 0.7% in ECFP fluorescence for the specific antibodies reacting with gp120 or gp41, respectively, was noted after photobleaching. A positive control (HcRed-ECFP) gave a 14.8 ± 2.6% increase. Surprisingly, the unspecific antibody (anti-TT) showed 12.1 ± 1.1% increase, possibly because overexpression in the limited ER compartment gave false FRET signals

Double-staining chromogenic in situ hybridization as a useful alternative to split-signal fluorescence in situ hybridization in lymphoma diagnostics

DEFF Research Database (Denmark)

van Rijk, A.; Svenstroup-Poulsen, T.; Jones, M.

2010-01-01

within the reach of every pathology laboratory. Design and Methods Our study was initiated to determine the consistency between chromogenic in situ hybridization and fluorescence in situ hybridization, both using split-signal probes developed for the detection of chromosomal breaks. Five hundred...... and actual signal were compared to the original fluorescence hybridization results. In addition, hematoxylin background staining intensity and signal intensity of the double-staining chromogenic in situ hybridization procedure were analyzed. Results With respect to the presence or absence of chromosomal...

Synthesis and characterization of photoswitchable fluorescent silica nanoparticles.

Science.gov (United States)

Fölling, Jonas; Polyakova, Svetlana; Belov, Vladimir; van Blaaderen, Alfons; Bossi, Mariano L; Hell, Stefan W

2008-01-01

We have designed and synthesized a new functional (amino reactive) highly efficient fluorescent molecular switch (FMS) with a photochromic diarylethene and a rhodamine fluorescent dye. The reactive group in this FMS -N-hydroxysuccinimide ester- allows selective labeling of amino containing molecules or other materials. In ethanolic solutions, the compound displays a large fluorescent quantum yield of 52 % and a large fluorescence modulation ratio (94 %) between two states that may be interconverted with red and near-UV light. Silica nanoparticles incorporating the new FMS were prepared and characterized, and their spectroscopic and switching properties were also studied. The dye retained its properties after the incorporation into the silica, thereby allowing light-induced reversible high modulation of the fluorescence signal of a single particle for up to 60 cycles, before undergoing irreversible photobleaching. Some applications of these particles in fluorescence microscopy are also demonstrated. In particular, subdiffraction images of nanoparticles were obtained, in the focal plane of a confocal microscope.

Hyperspectral small animal fluorescence imaging: spectral selection imaging

Science.gov (United States)

Leavesley, Silas; Jiang, Yanan; Patsekin, Valery; Hall, Heidi; Vizard, Douglas; Robinson, J. Paul

2008-02-01

Molecular imaging is a rapidly growing area of research, fueled by needs in pharmaceutical drug-development for methods for high-throughput screening, pre-clinical and clinical screening for visualizing tumor growth and drug targeting, and a growing number of applications in the molecular biology fields. Small animal fluorescence imaging employs fluorescent probes to target molecular events in vivo, with a large number of molecular targeting probes readily available. The ease at which new targeting compounds can be developed, the short acquisition times, and the low cost (compared to microCT, MRI, or PET) makes fluorescence imaging attractive. However, small animal fluorescence imaging suffers from high optical scattering, absorption, and autofluorescence. Much of these problems can be overcome through multispectral imaging techniques, which collect images at different fluorescence emission wavelengths, followed by analysis, classification, and spectral deconvolution methods to isolate signals from fluorescence emission. We present an alternative to the current method, using hyperspectral excitation scanning (spectral selection imaging), a technique that allows excitation at any wavelength in the visible and near-infrared wavelength range. In many cases, excitation imaging may be more effective at identifying specific fluorescence signals because of the higher complexity of the fluorophore excitation spectrum. Because the excitation is filtered and not the emission, the resolution limit and image shift imposed by acousto-optic tunable filters have no effect on imager performance. We will discuss design of the imager, optimizing the imager for use in small animal fluorescence imaging, and application of spectral analysis and classification methods for identifying specific fluorescence signals.

Heme Gazing: Illuminating Eukaryotic Heme Trafficking, Dynamics, and Signaling with Fluorescent Heme Sensors.

Science.gov (United States)

Hanna, David A; Martinez-Guzman, Osiris; Reddi, Amit R

2017-04-04

Heme (iron protoporphyrin IX) is an essential protein prosthetic group and signaling molecule required for most life on Earth. All heme-dependent processes require the dynamic and rapid mobilization of heme from sites of synthesis or uptake to hemoproteins present in virtually every subcellular compartment. The cytotoxicity and hydrophobicity of heme necessitate that heme mobilization be carefully controlled to mitigate the deleterious effects of this essential toxin. Indeed, a number of disorders, including certain cancers, cardiovascular diseases, and aging and age-related neurodegenerative diseases, are tied to defects in heme homeostasis. However, the molecules and mechanisms that mediate heme transport and trafficking, and the dynamics of these processes, are poorly understood. This is in large part due to the lack of physical tools for probing cellular heme. Herein, we discuss the recent development of fluorescent probes that can monitor and image kinetically labile heme with respect to its mobilization and role in signaling. In particular, we will highlight how heme gazing with these tools can uncover new heme trafficking factors upon being integrated with genetic screens and illuminate the concentration, subcellular distribution, and dynamics of labile heme in various physiological contexts. Altogether, the monitoring of labile heme, along with recent biochemical and cell biological studies demonstrating the reversible regulation of certain cellular processes by heme, is challenging us to reconceptualize heme from being a static cofactor buried in protein active sites to a dynamic and mobile signaling molecule.

Double-labeled donor probe can enhance the signal of fluorescence resonance energy transfer (FRET) in detection of nucleic acid hybridization

Science.gov (United States)

Okamura, Yukio; Kondo, Satoshi; Sase, Ichiro; Suga, Takayuki; Mise, Kazuyuki; Furusawa, Iwao; Kawakami, Shigeki; Watanabe, Yuichiro

2000-01-01

A set of fluorescently-labeled DNA probes that hybridize with the target RNA and produce fluorescence resonance energy transfer (FRET) signals can be utilized for the detection of specific RNA. We have developed probe sets to detect and discriminate single-strand RNA molecules of plant viral genome, and sought a method to improve the FRET signals to handle in vivo applications. Consequently, we found that a double-labeled donor probe labeled with Bodipy dye yielded a remarkable increase in fluorescence intensity compared to a single-labeled donor probe used in an ordinary FRET. This double-labeled donor system can be easily applied to improve various FRET probes since the dependence upon sequence and label position in enhancement is not as strict. Furthermore this method could be applied to other nucleic acid substances, such as oligo RNA and phosphorothioate oligonucleotides (S-oligos) to enhance FRET signal. Although the double-labeled donor probes labeled with a variety of fluorophores had unexpected properties (strange UV-visible absorption spectra, decrease of intensity and decay of donor fluorescence) compared with single-labeled ones, they had no relation to FRET enhancement. This signal amplification mechanism cannot be explained simply based on our current results and knowledge of FRET. Yet it is possible to utilize this double-labeled donor system in various applications of FRET as a simple signal-enhancement method. PMID:11121494

Effects of Depilation-Induced Skin Pigmentation and Diet-Induced Fluorescence on In Vivo Fluorescence Imaging

OpenAIRE

Kwon, Sunkuk; Sevick-Muraca, Eva M.

2017-01-01

Near-infrared fluorescence imaging (NIRFI) and far-red fluorescence imaging (FRFI) were used to investigate effects of depilation-induced skin pigmentation and diet-induced background fluorescence on fluorescent signal amplitude and lymphatic contraction frequency in C57BL6 mice. Far-red fluorescent signal amplitude, but not frequency, was affected by diet-induced fluorescence, which was removed by feeding the mice an alfalfa-free diet, and skin pigmentation further impacted the amplitude mea...

Simultaneous recording of fluorescence and electrical signals by photometric patch electrode in deep brain regions in vivo.

Science.gov (United States)

Hirai, Yasuharu; Nishino, Eri; Ohmori, Harunori

2015-06-01

Despite its widespread use, high-resolution imaging with multiphoton microscopy to record neuronal signals in vivo is limited to the surface of brain tissue because of limited light penetration. Moreover, most imaging studies do not simultaneously record electrical neural activity, which is, however, crucial to understanding brain function. Accordingly, we developed a photometric patch electrode (PME) to overcome the depth limitation of optical measurements and also enable the simultaneous recording of neural electrical responses in deep brain regions. The PME recoding system uses a patch electrode to excite a fluorescent dye and to measure the fluorescence signal as a light guide, to record electrical signal, and to apply chemicals to the recorded cells locally. The optical signal was analyzed by either a spectrometer of high light sensitivity or a photomultiplier tube depending on the kinetics of the responses. We used the PME in Oregon Green BAPTA-1 AM-loaded avian auditory nuclei in vivo to monitor calcium signals and electrical responses. We demonstrated distinct response patterns in three different nuclei of the ascending auditory pathway. On acoustic stimulation, a robust calcium fluorescence response occurred in auditory cortex (field L) neurons that outlasted the electrical response. In the auditory midbrain (inferior colliculus), both responses were transient. In the brain-stem cochlear nucleus magnocellularis, calcium response seemed to be effectively suppressed by the activity of metabotropic glutamate receptors. In conclusion, the PME provides a powerful tool to study brain function in vivo at a tissue depth inaccessible to conventional imaging devices. Copyright © 2015 the American Physiological Society.

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

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Bryan Sands

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

Signals from fluorescent materials on the surface of silicon micro-strip sensors

CERN Document Server

Sperlich, Dennis; The ATLAS collaboration

2017-01-01

For the High-Luminosity Upgrade of the Large Hadron Collider at CERN, the ATLAS Inner Detector will be replaced with a new, all-silicon tracker. In order to minimise the amount of material in the detector, circuit boards with readout electronics will be glued on to the active area of the sensor. Several adhesives investigated to be used for the construction of detector modules were found to become fluorescent when exposed to UV light. These adhesives could become a light source in the high-radiation environment of the ATLAS detector. The effect of fluorescent material covering the sensor surface in a high- radiation environment has been studied for a silicon micro-strip sensor using a micro-focused X-ray beam. By pointing the beam both inside the sensor and parallel to the sensor surface, the sensor responses from direct hits and fluorescence can be compared with high precision. This contribution presents a setup to study the susceptibility of silicon strip sensors to light contamination from fluorescent mate...

Fluorescence enhancement in large-scale self-assembled gold nanoparticle double arrays

International Nuclear Information System (INIS)

Chekini, M.; Bierwagen, J.; Cunningham, A.; Bürgi, T.; Filter, R.; Rockstuhl, C.

2015-01-01

Localized surface plasmon resonances excited in metallic nanoparticles confine and enhance electromagnetic fields at the nanoscale. This is particularly pronounced in dimers made from two closely spaced nanoparticles. When quantum emitters, such as dyes, are placed in the gap of those dimers, their absorption and emission characteristics can be modified. Both processes have to be considered when aiming to enhance the fluorescence from the quantum emitters. This is particularly challenging for dimers, since the electromagnetic properties and the enhanced fluorescence sensitively depend on the distance between the nanoparticles. Here, we use a layer-by-layer method to precisely control the distances in such systems. We consider a dye layer deposited on top of an array of gold nanoparticles or integrated into a central position of a double array of gold nanoparticles. We study the effect of the spatial arrangement and the average distance on the plasmon-enhanced fluorescence. We found a maximum of a 99-fold increase in the fluorescence intensity of the dye layer sandwiched between two gold nanoparticle arrays. The interaction of the dye layer with the plasmonic system also causes a spectral shift in the emission wavelengths and a shortening of the fluorescence life times. Our work paves the way for large-scale, high throughput, and low-cost self-assembled functionalized plasmonic systems that can be used as efficient light sources

Detecting crop population growth using chlorophyll fluorescence imaging.

Science.gov (United States)

Wang, Heng; Qian, Xiangjie; Zhang, Lan; Xu, Sailong; Li, Haifeng; Xia, Xiaojian; Dai, Liankui; Xu, Liang; Yu, Jingquan; Liu, Xu

2017-12-10

For both field and greenhouse crops, it is challenging to evaluate their growth information on a large area over a long time. In this work, we developed a chlorophyll fluorescence imaging-based system for crop population growth information detection. Modular design was used to make the system provide high-intensity uniform illumination. This system can perform modulated chlorophyll fluorescence induction kinetics measurement and chlorophyll fluorescence parameter imaging over a large area of up to 45  cm×34  cm. The system can provide different lighting intensity by modulating the duty cycle of its control signal. Results of continuous monitoring of cucumbers in nitrogen deficiency show the system can reduce the judge error of crop physiological status and improve monitoring efficiency. Meanwhile, the system is promising in high throughput application scenarios.

MATtrack: A MATLAB-Based Quantitative Image Analysis Platform for Investigating Real-Time Photo-Converted Fluorescent Signals in Live Cells.

Science.gov (United States)

Courtney, Jane; Woods, Elena; Scholz, Dimitri; Hall, William W; Gautier, Virginie W

2015-01-01

We introduce here MATtrack, an open source MATLAB-based computational platform developed to process multi-Tiff files produced by a photo-conversion time lapse protocol for live cell fluorescent microscopy. MATtrack automatically performs a series of steps required for image processing, including extraction and import of numerical values from Multi-Tiff files, red/green image classification using gating parameters, noise filtering, background extraction, contrast stretching and temporal smoothing. MATtrack also integrates a series of algorithms for quantitative image analysis enabling the construction of mean and standard deviation images, clustering and classification of subcellular regions and injection point approximation. In addition, MATtrack features a simple user interface, which enables monitoring of Fluorescent Signal Intensity in multiple Regions of Interest, over time. The latter encapsulates a region growing method to automatically delineate the contours of Regions of Interest selected by the user, and performs background and regional Average Fluorescence Tracking, and automatic plotting. Finally, MATtrack computes convenient visualization and exploration tools including a migration map, which provides an overview of the protein intracellular trajectories and accumulation areas. In conclusion, MATtrack is an open source MATLAB-based software package tailored to facilitate the analysis and visualization of large data files derived from real-time live cell fluorescent microscopy using photoconvertible proteins. It is flexible, user friendly, compatible with Windows, Mac, and Linux, and a wide range of data acquisition software. MATtrack is freely available for download at eleceng.dit.ie/courtney/MATtrack.zip.

Fluorescence resonance energy transfer (FRET-based subcellular visualization of pathogen-induced host receptor signaling

Directory of Open Access Journals (Sweden)

Zimmermann Timo

2009-11-01

Full Text Available Abstract Background Bacteria-triggered signaling events in infected host cells are key elements in shaping the host response to pathogens. Within the eukaryotic cell, signaling complexes are spatially organized. However, the investigation of protein-protein interactions triggered by bacterial infection in the cellular context is technically challenging. Here, we provide a methodological approach to exploit fluorescence resonance energy transfer (FRET to visualize pathogen-initiated signaling events in human cells. Results Live-cell microscopy revealed the transient recruitment of the Src family tyrosine kinase Hck upon bacterial engagement of the receptor carcinoembryonic antigen-related cell adhesion molecule 3 (CEACAM3. In cells expressing a CEACAM3 variant lacking the cytoplasmic domain, the Src homology 2 (SH2 domain of Hck (Hck-SH2 was not recruited, even though bacteria still bound to the receptor. FRET measurements on the basis of whole cell lysates revealed intimate binding between Hck-SH2 (using enhanced yellow fluorescent protein (YPet-Hck-SH2 and the tyrosine-phosphorylated enhanced cyan fluorescent protein-labeled cytoplasmic domain of wild-type CEACAM3 (CEACAM3 WT-CyPet and a flow cytometry-based FRET approach verified this association in intact cells. Using confocal microscopy and acceptor photobleaching, FRET between Hck-SH2 and CEACAM3 was localized to the sites of bacteria-host cell contact. Conclusion These data demonstrate not only the intimate binding of the SH2 domain of Hck to the tyrosine-phosphorylated cytoplasmic domain of CEACAM3 in intact cells, but furthermore, FRET measurements allow the subcellular localization of this process during bacterial infection. FRET-based assays are valuable tools to resolve bacteria-induced protein-protein interactions in the context of the intact host cell.

Whole-slide imaging is a robust alternative to traditional fluorescent microscopy for fluorescence in situ hybridization imaging using break-apart DNA probes.

Science.gov (United States)

Laurent, Camille; Guérin, Maxime; Frenois, François-Xavier; Thuries, Valérie; Jalabert, Laurence; Brousset, Pierre; Valmary-Degano, Séverine

2013-08-01

Fluorescence in situ hybridization is an indispensable technique used in routine pathology and for theranostic purposes. Because fluorescence in situ hybridization techniques require sophisticated microscopic workstations and long procedures of image acquisition with sometimes subjective and poorly reproducible results, we decided to test a whole-slide imaging system as an alternative approach. In this study, we used the latest generation of Pannoramic 250 Flash digital microscopes (P250 Flash digital microscopes; 3DHISTECH, Budapest, Hungary) to digitize fluorescence in situ hybridization slides of diffuse large B cells lymphoma cases for detecting MYC rearrangement. The P250 Flash digital microscope was found to be precise with better definition of split signals in cells containing MYC rearrangement with fewer truncated signals as compared to traditional fluorescence microscopy. This digital technique is easier thanks to the preview function, which allows almost immediate identification of the tumor area, and the panning and zooming functionalities as well as a shorter acquisition time. Moreover, fluorescence in situ hybridization analyses using the digital technique appeared to be more reproducible between pathologists. Finally, the digital technique also allowed prolonged conservation of photos. In conclusion, whole-slide imaging technologies represent rapid, robust, and highly sensitive methods for interpreting fluorescence in situ hybridization slides with break-apart probes. In addition, these techniques offer an easier way to interpret the signals and allow definitive storage of the images for pathology expert networks or e-learning databases. Copyright © 2013 Elsevier Inc. All rights reserved.

Time-synchronized continuous wave laser-induced fluorescence on an oscillatory xenon discharge.

Science.gov (United States)

MacDonald, N A; Cappelli, M A; Hargus, W A

2012-11-01

A novel approach to time-synchronizing laser-induced fluorescence measurements to an oscillating current in a 60 Hz xenon discharge lamp using a continuous wave laser is presented. A sample-hold circuit is implemented to separate out signals at different phases along a current cycle, and is followed by a lock-in amplifier to pull out the resulting time-synchronized fluorescence trace from the large background signal. The time evolution of lower state population is derived from the changes in intensity of the fluorescence excitation line shape resulting from laser-induced fluorescence measurements of the 6s(')[1/2](1)(0)-6p(')[3/2](2) xenon atomic transition at λ = 834.68 nm. Results show that the lower state population oscillates at twice the frequency of the discharge current, 120 Hz.

Fluorescence methods for analysis of interactions between Ca(2+) signaling, lysosomes, and endoplasmic reticulum.

Science.gov (United States)

Prole, David L; López-Sanjurjo, Cristina I; Tovey, Stephen C; Taylor, Colin W

2015-01-01

The endoplasmic reticulum (ER) is both the major source of intracellular Ca(2+) for cell signaling and the organelle that forms the most extensive contacts with the plasma membrane and other organelles. Lysosomes fulfill important roles in degrading cellular materials and in cholesterol handling, but they also contribute to Ca(2+) signaling by both releasing and sequestering Ca(2+). Interactions between ER and other Ca(2+)-transporting membranes, notably mitochondria and the plasma membrane, often occur at sites where the two membranes are closely apposed, allowing local Ca(2+) signaling between them. These interactions are often facilitated by scaffold proteins. Recent evidence suggests similar local interactions between ER and lysosomes. We describe simple fluorescence-based methods that allow the interplay between Ca(2+) signals, the ER, and lysosomes to be examined. Copyright © 2015 Elsevier Inc. All rights reserved.

Fiber optical assembly for fluorescence spectrometry

Science.gov (United States)

Carpenter, II, Robert W.; Rubenstein, Richard; Piltch, Martin; Gray, Perry

2010-12-07

A system for analyzing a sample for the presence of an analyte in a sample. The system includes a sample holder for containing the sample; an excitation source, such as a laser, and at least one linear array radially disposed about the sample holder. Radiation from the excitation source is directed to the sample, and the radiation induces fluorescent light in the sample. Each linear array includes a plurality of fused silica optical fibers that receive the fluorescent light and transmits a fluorescent light signal from the first end to an optical end port of the linear array. An end port assembly having a photo-detector is optically coupled to the optical end port. The photo-detector detects the fluorescent light signal and converts the fluorescent light signal into an electrical signal.

An aptamer-based fluorescence bio-sensor for chiral recognition of arginine enantiomers.

Science.gov (United States)

Yuan, Haiyan; Huang, Yunmei; Yang, Jidong; Guo, Yuan; Zeng, Xiaoqing; Zhou, Shang; Cheng, Jiawei; Zhang, Yuhui

2018-07-05

In this study, a novel aptamer - based fluorescence bio-sensor (aptamer-AuNps) was developed for chiral recognition of arginine (Arg) enantiomers based on aptamer and gold nanoparticles (AuNps). Carboxyfluorescein (FAM) labeled aptamers (Apt) were absorbed on AuNps and their fluorescence intensity could be significantly quenched by AuNps based on fluorescence resonance energy transfer (FRET). Once d-Arg or l-Arg were added into the above solution, the aptamer specifically bind to Arg enantiomers and released from AuNps, so the fluorescence intensity of d-Arg system and l-Arg system were all enhanced. The affinity of Apt to l-Arg is tighter to d-Arg, so the enhanced fluorescence signals of l-Arg system was stronger than d-Arg system. What's more, the enhanced fluorescence were directly proportional to the concentration of d-Arg and l-Arg ranging from 0-300 nM and 0-400 nM with related coefficients of 0.9939 and 0.9952, respectively. Furthermore, the method was successfully applied to detection l-Arg in human urine samples with satisfactory results. Eventually, a simple "OR" logic gate with d-Arg &l-Arg as inputs and AuNps aggregation state as outputs was fabricated, which can help us understand the chiral recognition process deeply. Copyright © 2018 Elsevier B.V. All rights reserved.

Mixed-signal instrumentation for large-signal device characterization and modelling

NARCIS (Netherlands)

Marchetti, M.

2013-01-01

This thesis concentrates on the development of advanced large-signal measurement and characterization tools to support technology development, model extraction and validation, and power amplifier (PA) designs that address the newly introduced third and fourth generation (3G and 4G) wideband

High-contrast fluorescence imaging based on the polarization dependence of the fluorescence enhancement using an optical interference mirror slide.

Science.gov (United States)

Yasuda, Mitsuru; Akimoto, Takuo

2015-01-01

High-contrast fluorescence imaging using an optical interference mirror (OIM) slide that enhances the fluorescence from a fluorophore located on top of the OIM surface is reported. To enhance the fluorescence and reduce the background light of the OIM, transverse-electric-polarized excitation light was used as incident light, and the transverse-magnetic-polarized fluorescence signal was detected. As a result, an approximate 100-fold improvement in the signal-to-noise ratio was achieved through a 13-fold enhancement of the fluorescence signal and an 8-fold reduction of the background light.

Time-synchronized continuous wave laser-induced fluorescence on an oscillatory xenon discharge

Energy Technology Data Exchange (ETDEWEB)

MacDonald, N. A.; Cappelli, M. A. [Stanford Plasma Physics Laboratory, Stanford University, Stanford, California 94305 (United States); Hargus, W. A. Jr. [Air Force Research Laboratory, Edwards AFB, California 93524 (United States)

2012-11-15

A novel approach to time-synchronizing laser-induced fluorescence measurements to an oscillating current in a 60 Hz xenon discharge lamp using a continuous wave laser is presented. A sample-hold circuit is implemented to separate out signals at different phases along a current cycle, and is followed by a lock-in amplifier to pull out the resulting time-synchronized fluorescence trace from the large background signal. The time evolution of lower state population is derived from the changes in intensity of the fluorescence excitation line shape resulting from laser-induced fluorescence measurements of the 6s{sup Prime }[1/2]{sub 1}{sup 0}-6p{sup Prime }[3/2]{sub 2} xenon atomic transition at {lambda}= 834.68 nm. Results show that the lower state population oscillates at twice the frequency of the discharge current, 120 Hz.

A set of enhanced green fluorescent protein concatemers for quantitative determination of nuclear localization signal strength.

Science.gov (United States)

Böhm, Jennifer; Thavaraja, Ramya; Giehler, Susanne; Nalaskowski, Marcus M

2017-09-15

Regulated transport of proteins between nucleus and cytoplasm is an important process in the eukaryotic cell. In most cases, active nucleo-cytoplasmic protein transport is mediated by nuclear localization signal (NLS) and/or nuclear export signal (NES) motifs. In this study, we developed a set of vectors expressing enhanced GFP (EGFP) concatemers ranging from 2 to 12 subunits (2xEGFP to 12xEGFP) for analysis of NLS strength. As shown by in gel GFP fluorescence analysis and αGFP Western blotting, EGFP concatemers are expressed as fluorescent full-length proteins in eukaryotic cells. As expected, nuclear localization of concatemeric EGFPs decreases with increasing molecular weight. By oligonucleotide ligation this set of EGFP concatemers can be easily fused to NLS motifs. After determination of intracellular localization of EGFP concatemers alone and fused to different NLS motifs we calculated the size of a hypothetic EGFP concatemer showing a defined distribution of EGFP fluorescence between nucleus and cytoplasm (n/c ratio = 2). Clear differences of the size of the hypothetic EGFP concatemer depending on the fused NLS motif were observed. Therefore, we propose to use the size of this hypothetic concatemer as quantitative indicator for comparing strength of different NLS motifs. Copyright © 2017 Elsevier Inc. All rights reserved.

A novel polymer probe for Zn(II) detection with ratiometric fluorescence signal

Science.gov (United States)

Diao, Haipeng; Guo, Lixia; Liu, Wen; Feng, Liheng

2018-05-01

A conjugated polymer probe comprised of fluorene, quinolone and benzothiazole units was designed and synthesized by the Suzuki coupling reaction. Through the studies of photophysical and thermal properties, the polymer displays blue-emitting feature and good thermal stability. A ratiometric fluorescence signal of the probe for Zn(II) was observed in ethanol with a new emission peak at 555 nm. The probe possesses a high selectivity and sensitivity for Zn(II) during familiar metal ions in ethanol. The detection limit of the probe for Zn (II) is up to 10-8 mol/L. The electron distributions of the polymer before and after bonding with Zn (II) were investigated by the Gaussian 09 software, which agreed with the experimental results. Noticeably, based on the color property of the probe with Zn(II), a series of color test paper were developed for visual detecting Zn(II) ions. This work helps to provide a platform or pattern for the development of polymer fluorescence probe in the chemosensor field.

Fluorescence guided lymph node biopsy in large animals using direct image projection device

Science.gov (United States)

Ringhausen, Elizabeth; Wang, Tylon; Pitts, Jonathan; Akers, Walter J.

2016-03-01

The use of fluorescence imaging for aiding oncologic surgery is a fast growing field in biomedical imaging, revolutionizing open and minimally invasive surgery practices. We have designed, constructed, and tested a system for fluorescence image acquisition and direct display on the surgical field for fluorescence guided surgery. The system uses a near-infrared sensitive CMOS camera for image acquisition, a near-infra LED light source for excitation, and DLP digital projector for projection of fluorescence image data onto the operating field in real time. Instrument control was implemented in Matlab for image capture, processing of acquired data and alignment of image parameters with the projected pattern. Accuracy of alignment was evaluated statistically to demonstrate sensitivity to small objects and alignment throughout the imaging field. After verification of accurate alignment, feasibility for clinical application was demonstrated in large animal models of sentinel lymph node biopsy. Indocyanine green was injected subcutaneously in Yorkshire pigs at various locations to model sentinel lymph node biopsy in gynecologic cancers, head and neck cancer, and melanoma. Fluorescence was detected by the camera system during operations and projected onto the imaging field, accurately identifying tissues containing the fluorescent tracer at up to 15 frames per second. Fluorescence information was projected as binary green regions after thresholding and denoising raw intensity data. Promising results with this initial clinical scale prototype provided encouraging results for the feasibility of optical projection of acquired luminescence during open oncologic surgeries.

A 13-element Ge detector for fluorescence EXAFS

International Nuclear Information System (INIS)

Cramer, S.P.; Tench, O.; Yocum, M.; George, G.N.

1988-01-01

At low concentrations, recording X-ray absorption spectra in fluorescence excitation mode is more sensitive than transmission mode. For dilute samples, the fluorescence signal is often obscured by scattered X-rays, and matrix and filter fluorescence. To discriminate against this background, while maintaining a large angular acceptance and high count rate capability, we have constructed a new detection system based on an array of intrinsic Ge detectors. The device uses 13 individuall 11 mm diameter Ge detectors, clustered in a 1:3:5:3:1 pattern on a common cryostat, combined with Soller slits and filters to reduce the background signals. Pulsed optical feedback preamplifiers are followed by Gaussian-shaping amplifiers having fast discriminators to register the incoming count rate (ICR). Correction for dead time using the ICR signal allowed operation in the vicinity of 75 kHz per channel, with a 1 μs shaping time at 6 keV. For lower count rate applications, an average resolution of 160 eV at 5.9 keV was obtained with 8 μs shaping. Recent experience with this detector at the Stanford Synchrotron Radiation Laboratory is presented. The performance is illustrated using spectra obtained from phosphorus compounds and a thin iridium foil. The performance of this device is compared with previous fluorescence detection schemes, such as those using filter/slit combinations or barrel monochromators. (orig.)

Multispectral open-air intraoperative fluorescence imaging.

Science.gov (United States)

Behrooz, Ali; Waterman, Peter; Vasquez, Kristine O; Meganck, Jeff; Peterson, Jeffrey D; Faqir, Ilias; Kempner, Joshua

2017-08-01

Intraoperative fluorescence imaging informs decisions regarding surgical margins by detecting and localizing signals from fluorescent reporters, labeling targets such as malignant tissues. This guidance reduces the likelihood of undetected malignant tissue remaining after resection, eliminating the need for additional treatment or surgery. The primary challenges in performing open-air intraoperative fluorescence imaging come from the weak intensity of the fluorescence signal in the presence of strong surgical and ambient illumination, and the auto-fluorescence of non-target components, such as tissue, especially in the visible spectral window (400-650 nm). In this work, a multispectral open-air fluorescence imaging system is presented for translational image-guided intraoperative applications, which overcomes these challenges. The system is capable of imaging weak fluorescence signals with nanomolar sensitivity in the presence of surgical illumination. This is done using synchronized fluorescence excitation and image acquisition with real-time background subtraction. Additionally, the system uses a liquid crystal tunable filter for acquisition of multispectral images that are used to spectrally unmix target fluorescence from non-target auto-fluorescence. Results are validated by preclinical studies on murine models and translational canine oncology models.

Quantitative analysis of phosphoinositide 3-kinase (PI3K) signaling using live-cell total internal reflection fluorescence (TIRF) microscopy.

Science.gov (United States)

Johnson, Heath E; Haugh, Jason M

2013-12-02

This unit focuses on the use of total internal reflection fluorescence (TIRF) microscopy and image analysis methods to study the dynamics of signal transduction mediated by class I phosphoinositide 3-kinases (PI3Ks) in mammalian cells. The first four protocols cover live-cell imaging experiments, image acquisition parameters, and basic image processing and segmentation. These methods are generally applicable to live-cell TIRF experiments. The remaining protocols outline more advanced image analysis methods, which were developed in our laboratory for the purpose of characterizing the spatiotemporal dynamics of PI3K signaling. These methods may be extended to analyze other cellular processes monitored using fluorescent biosensors. Copyright © 2013 John Wiley & Sons, Inc.

Large-signal modeling method for power FETs and diodes

Energy Technology Data Exchange (ETDEWEB)

Sun Lu; Wang Jiali; Wang Shan; Li Xuezheng; Shi Hui; Wang Na; Guo Shengping, E-mail: sunlu_1019@126.co [School of Electromechanical Engineering, Xidian University, Xi' an 710071 (China)

2009-06-01

Under a large signal drive level, a frequency domain black box model of the nonlinear scattering function is introduced into power FETs and diodes. A time domain measurement system and a calibration method based on a digital oscilloscope are designed to extract the nonlinear scattering function of semiconductor devices. The extracted models can reflect the real electrical performance of semiconductor devices and propose a new large-signal model to the design of microwave semiconductor circuits.

Large-signal modeling method for power FETs and diodes

International Nuclear Information System (INIS)

Sun Lu; Wang Jiali; Wang Shan; Li Xuezheng; Shi Hui; Wang Na; Guo Shengping

2009-01-01

Under a large signal drive level, a frequency domain black box model of the nonlinear scattering function is introduced into power FETs and diodes. A time domain measurement system and a calibration method based on a digital oscilloscope are designed to extract the nonlinear scattering function of semiconductor devices. The extracted models can reflect the real electrical performance of semiconductor devices and propose a new large-signal model to the design of microwave semiconductor circuits.

Large-signal stability analysis of PWM converters

Energy Technology Data Exchange (ETDEWEB)

Huynh, P.T. [Philips Labs., Briarcliff Manor, NY (United States); Cho, B.H. [Seoul National Univ. (Korea, Republic of). Dept. of Electrical Engineering

1995-12-31

Investigation of the effects of existing nonlinearities on the stability of PWM converters is performed. The bilinear structure, the duty cycle saturation, and the opamp saturation are the principal nonlinearities in PWM converters. These nonlinearities are incorporated in the large-signal analytical models of PWM converters, and the basic input-output stability theory is applied to analyze their stability. Design and optimization of the small-signal loop gains to counteract the undesirable nonlinear effects are also discussed.

Signals from fluorescent materials on the surface of silicon micro-strip sensors

CERN Document Server

Sperlich, Dennis; The ATLAS collaboration

2018-01-01

For the High-Luminosity Upgrade of the Large Hadron Collider at CERN, the ATLAS Inner Detector will be replaced with a new, all-silicon tracker (ITk). In order to minimise the amount of material in the ITk, circuit boards with readout electronics will be glued onto the active area of the sensor. Several adhesives, investigated to be used for the construction of detector modules, were found to become fluorescent when exposed to UV light. These adhesives could become a light source in the high-radiation environment of the ATLAS detector. The effect of fluorescent material covering the sensor surface in a high-radiation environment has been studied for a silicon micro-strip sensor using a micro-focused X-ray beam. By positioning the beam parallel to the sensor surfave and pointing it both inside the sensor and above the sensor surface inside the deposited glue, the sensor responses from direct hits and fluorescence can be compared with high precision. This contribution presents a setup to study the susceptibilit...

Large-scale solvothermal synthesis of fluorescent carbon nanoparticles

International Nuclear Information System (INIS)

Ku, Kahoe; Park, Jinwoo; Kim, Nayon; Kim, Woong; Lee, Seung-Wook; Chung, Haegeun; Han, Chi-Hwan

2014-01-01

The large-scale production of high-quality carbon nanomaterials is highly desirable for a variety of applications. We demonstrate a novel synthetic route to the production of fluorescent carbon nanoparticles (CNPs) in large quantities via a single-step reaction. The simple heating of a mixture of benzaldehyde, ethanol and graphite oxide (GO) with residual sulfuric acid in an autoclave produced 7 g of CNPs with a quantum yield of 20%. The CNPs can be dispersed in various organic solvents; hence, they are easily incorporated into polymer composites in forms such as nanofibers and thin films. Additionally, we observed that the GO present during the CNP synthesis was reduced. The reduced GO (RGO) was sufficiently conductive (σ ≈ 282 S m −1 ) such that it could be used as an electrode material in a supercapacitor; in addition, it can provide excellent capacitive behavior and high-rate capability. This work will contribute greatly to the development of efficient synthetic routes to diverse carbon nanomaterials, including CNPs and RGO, that are suitable for a wide range of applications. (paper)

Analysis of signal to background ratio in synchrotron radiation X-ray fluorescence

International Nuclear Information System (INIS)

Sakurai, Kenji; Gohshi, Yohichi; Iida, Atsuo.

1988-01-01

The signal to background (S/B) ratio in energy dispersive X-ray fluorescence using synchrotron radiation (SR) was quantitatively analyzed. The S/B ratio, which has been significantly improved by taking advantage of the polarized nature of SR, was found to be strongly dependent on geometrical factors of the measurement system. From the analysis on the origin of the scattered background, the dependence of the S/B ratio on the geometry was quantitatively explained, mainly by the polarization properties of SR. Experimental conditions could be optimized by adjusting the degree of polarization of the incident beam and the detector solid angle. (author)

Large scale analysis of signal reachability.

Science.gov (United States)

Todor, Andrei; Gabr, Haitham; Dobra, Alin; Kahveci, Tamer

2014-06-15

Major disorders, such as leukemia, have been shown to alter the transcription of genes. Understanding how gene regulation is affected by such aberrations is of utmost importance. One promising strategy toward this objective is to compute whether signals can reach to the transcription factors through the transcription regulatory network (TRN). Due to the uncertainty of the regulatory interactions, this is a #P-complete problem and thus solving it for very large TRNs remains to be a challenge. We develop a novel and scalable method to compute the probability that a signal originating at any given set of source genes can arrive at any given set of target genes (i.e., transcription factors) when the topology of the underlying signaling network is uncertain. Our method tackles this problem for large networks while providing a provably accurate result. Our method follows a divide-and-conquer strategy. We break down the given network into a sequence of non-overlapping subnetworks such that reachability can be computed autonomously and sequentially on each subnetwork. We represent each interaction using a small polynomial. The product of these polynomials express different scenarios when a signal can or cannot reach to target genes from the source genes. We introduce polynomial collapsing operators for each subnetwork. These operators reduce the size of the resulting polynomial and thus the computational complexity dramatically. We show that our method scales to entire human regulatory networks in only seconds, while the existing methods fail beyond a few tens of genes and interactions. We demonstrate that our method can successfully characterize key reachability characteristics of the entire transcriptions regulatory networks of patients affected by eight different subtypes of leukemia, as well as those from healthy control samples. All the datasets and code used in this article are available at bioinformatics.cise.ufl.edu/PReach/scalable.htm. © The Author 2014

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

2014-01-01

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. PMID:25071950

Fluorescence-enhanced optical imaging in large tissue volumes using a gain-modulated ICCD camera

International Nuclear Information System (INIS)

Godavarty, Anuradha; Eppstein, Margaret J; Zhang, Chaoyang; Theru, Sangeeta; Thompson, Alan B; Gurfinkel, Michael; Sevick-Muraca, Eva M

2003-01-01

A novel image-intensified charge-coupled device (ICCD) imaging system has been developed to perform 3D fluorescence tomographic imaging in the frequency-domain using near-infrared contrast agents. The imager is unique since it (i) employs a large tissue-mimicking phantom, which is shaped and sized to resemble a female breast and part of the extended chest-wall region, and (ii) enables rapid data acquisition in the frequency-domain by using a gain-modulated ICCD camera. Diffusion model predictions are compared to experimental measurements using two different referencing schemes under two different experimental conditions of perfect and imperfect uptake of fluorescent agent into a target. From these experimental measurements, three-dimensional images of fluorescent absorption were reconstructed using a computationally efficient variant of the approximate extended Kalman filter algorithm. The current work represents the first time that 3D fluorescence-enhanced optical tomographic reconstructions have been achieved from experimental measurements of the time-dependent light propagation on a clinically relevant breast-shaped tissue phantom using a gain-modulated ICCD camera

A Parallel Distributed-Memory Particle Method Enables Acquisition-Rate Segmentation of Large Fluorescence Microscopy Images.

Science.gov (United States)

Afshar, Yaser; Sbalzarini, Ivo F

2016-01-01

Modern fluorescence microscopy modalities, such as light-sheet microscopy, are capable of acquiring large three-dimensional images at high data rate. This creates a bottleneck in computational processing and analysis of the acquired images, as the rate of acquisition outpaces the speed of processing. Moreover, images can be so large that they do not fit the main memory of a single computer. We address both issues by developing a distributed parallel algorithm for segmentation of large fluorescence microscopy images. The method is based on the versatile Discrete Region Competition algorithm, which has previously proven useful in microscopy image segmentation. The present distributed implementation decomposes the input image into smaller sub-images that are distributed across multiple computers. Using network communication, the computers orchestrate the collectively solving of the global segmentation problem. This not only enables segmentation of large images (we test images of up to 10(10) pixels), but also accelerates segmentation to match the time scale of image acquisition. Such acquisition-rate image segmentation is a prerequisite for the smart microscopes of the future and enables online data compression and interactive experiments.

DNA abasic site-selective enhancement of sanguinarine fluorescence with a large emission shift.

Directory of Open Access Journals (Sweden)

Fei Wu

Full Text Available Small molecules that can specifically bind to a DNA abasic site (AP site have received much attention due to their importance in DNA lesion identification, drug discovery, and sensor design. Herein, the AP site binding behavior of sanguinarine (SG, a natural alkaloid, was investigated. In aqueous solution, SG has a short-wavelength alkanolamine emission band and a long-wavelength iminium emission band. At pH 8.3, SG experiences a fluorescence quenching for both bands upon binding to fully matched DNAs without the AP site, while the presence of the AP site induces a strong SG binding and the observed fluorescence enhancement for the iminium band are highly dependent on the nucleobases flanking the AP site, while the alkanolamine band is always quenched. The bases opposite the AP site also exert some modifications on the SG's emission behavior. It was found that the observed quenching for DNAs with Gs and Cs flanking the AP site is most likely caused by electron transfer between the AP site-bound excited-state SG and the nearby Gs. However, the flanking As and Ts that are not easily oxidized favor the enhanced emission. This AP site-selective enhancement of SG fluorescence accompanies a band conversion in the dominate emission from the alkanolamine to iminium band thus with a large emission shift of about 170 nm. Absorption spectra, steady-state and transient-state fluorescence, DNA melting, and electrolyte experiments confirm that the AP site binding of SG occurs and the stacking interaction with the nearby base pairs is likely to prevent the converted SG iminium form from contacting with water that is thus emissive when the AP site neighbors are bases other than guanines. We expect that this fluorophore would be developed as a promising AP site binder having a large emission shift.

A dual-color fluorescence-based platform to identify selective inhibitors of Akt signaling.

Directory of Open Access Journals (Sweden)

Aranzazú Rosado

Full Text Available BACKGROUND: Inhibition of Akt signaling is considered one of the most promising therapeutic strategies for many cancers. However, rational target-orientated approaches to cell based drug screens for anti-cancer agents have historically been compromised by the notorious absence of suitable control cells. METHODOLOGY/PRINCIPAL FINDINGS: In order to address this fundamental problem, we have developed BaFiso, a live-cell screening platform to identify specific inhibitors of this pathway. BaFiso relies on the co-culture of isogenic cell lines that have been engineered to sustain interleukin-3 independent survival of the parental Ba/F3 cells, and that are individually tagged with different fluorescent proteins. Whilst in the first of these two lines cell survival in the absence of IL-3 is dependent on the expression of activated Akt, the cells expressing constitutively-activated Stat5 signaling display IL-3 independent growth and survival in an Akt-independent manner. Small molecules can then be screened in these lines to identify inhibitors that rescue IL-3 dependence. CONCLUSIONS/SIGNIFICANCE: BaFiso measures differential cell survival using multiparametric live cell imaging and permits selective inhibitors of Akt signaling to be identified. BaFiso is a platform technology suitable for the identification of small molecule inhibitors of IL-3 mediated survival signaling.

Time-resolved spectroscopic imaging reveals the fundamentals of cellular NADH fluorescence.

Science.gov (United States)

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

2008-10-15

A time-resolved spectroscopic imaging system is built to study the fluorescence characteristics of nicotinamide adenine dinucleotide (NADH), an important metabolic coenzyme and endogenous fluorophore in cells. The system provides a unique approach to measure fluorescence signals in different cellular organelles and cytoplasm. The ratios of free over protein-bound NADH signals in cytosol and nucleus are slightly higher than those in mitochondria. The mitochondrial fluorescence contributes about 70% of overall cellular fluorescence and is not a completely dominant signal. Furthermore, NADH signals in mitochondria, cytosol, and the nucleus respond to the changes of cellular activity differently, suggesting that cytosolic and nuclear fluorescence may complicate the well-known relationship between mitochondrial fluorescence and cellular metabolism.

Large-Scale Transit Signal Priority Implementation

OpenAIRE

Lee, Kevin S.; Lozner, Bailey

2018-01-01

In 2016, the District Department of Transportation (DDOT) deployed Transit Signal Priority (TSP) at 195 intersections in highly urbanized areas of Washington, DC. In collaboration with a broader regional implementation, and in partnership with the Washington Metropolitan Area Transit Authority (WMATA), DDOT set out to apply a systems engineering–driven process to identify, design, test, and accept a large-scale TSP system. This presentation will highlight project successes and lessons learned.

An evanescent wave biosensor--Part I: Fluorescent signal acquisition from step-etched fiber optic probes.

Science.gov (United States)

Anderson, G P; Golden, J P; Ligler, F S

1994-06-01

A fiber-optic biosensor capable of remote continuous monitoring has recently been designed. To permit sensing at locations separate from the optoelectronic instrumentation, long optical fibers are utilized. An evanescent wave immuno-probe is prepared by removing the cladding near the distal end of the fiber and covalently attaching antibodies to the core. Probes with a radius unaltered from that of the original core inefficiently returned the signal produced upon binding the fluorescent-labelled antigen. To elucidate the limiting factors in signal acquisition, a series of fibers with increasingly reduced probe core radius was examined. The results were consistent with the V-number mismatch, the difference in mode carrying capacity between the clad and unclad fiber, being a critical factor in limiting signal coupling from the fiber probe. However, it was also delineated that conditions which conserve excitation power, such that power in the evanescent wave is optimized, must also be met to obtain a maximal signal. The threshold sensitivity for the optimal step-etched fiber probe was improved by over 20-fold in an immunoassay, although, it was demonstrated that signal acquisition decreased along the probe length, suggesting that a sensor region of uniform radius is not ideal.

Fluorescence detection of natural RNA using rationally designed "clickable" oligonucleotide probes

DEFF Research Database (Denmark)

Okholm, Anders; Kjems, Jørgen; Astakhova, Kira

2014-01-01

Herein a reliable approach to the design of effective fluorescent probes for RNA detection is described. The fluorescence signalling of hybridization by internally positioned polyaromatic hydrocarbons and rhodamine dyes was achieved with a low fluorescence background signal, high fluorescence qua...... quantum yields at ambient and elevated temperature, high selectivity and signal specificity of the probes when binding to miR-7 and circRNA targets....

Self-interference fluorescence microscopy with three-phase detection for depth-resolved confocal epi-fluorescence imaging.

Science.gov (United States)

Braaf, Boy; de Boer, Johannes F

2017-03-20

Three-dimensional confocal fluorescence imaging of in vivo tissues is challenging due to sample motion and limited imaging speeds. In this paper a novel method is therefore presented for scanning confocal epi-fluorescence microscopy with instantaneous depth-sensing based on self-interference fluorescence microscopy (SIFM). A tabletop epi-fluorescence SIFM setup was constructed with an annular phase plate in the emission path to create a spectral self-interference signal that is phase-dependent on the axial position of a fluorescent sample. A Mach-Zehnder interferometer based on a 3 × 3 fiber-coupler was developed for a sensitive phase analysis of the SIFM signal with three photon-counter detectors instead of a spectrometer. The Mach-Zehnder interferometer created three intensity signals that alternately oscillated as a function of the SIFM spectral phase and therefore encoded directly for the axial sample position. Controlled axial translation of fluorescent microsphere layers showed a linear dependence of the SIFM spectral phase with sample depth over axial image ranges of 500 µm and 80 µm (3.9 × Rayleigh range) for 4 × and 10 × microscope objectives respectively. In addition, SIFM was in good agreement with optical coherence tomography depth measurements on a sample with indocyanine green dye filled capillaries placed at multiple depths. High-resolution SIFM imaging applications are demonstrated for fluorescence angiography on a dye-filled capillary blood vessel phantom and for autofluorescence imaging on an ex vivo fly eye.

Optimization of a Fluorescence-Based Assay for Large-Scale Drug Screening against Babesia and Theileria Parasites.

Science.gov (United States)

Rizk, Mohamed Abdo; El-Sayed, Shimaa Abd El-Salam; Terkawi, Mohamed Alaa; Youssef, Mohamed Ahmed; El Said, El Said El Shirbini; Elsayed, Gehad; El-Khodery, Sabry; El-Ashker, Maged; Elsify, Ahmed; Omar, Mosaab; Salama, Akram; Yokoyama, Naoaki; Igarashi, Ikuo

2015-01-01

A rapid and accurate assay for evaluating antibabesial drugs on a large scale is required for the discovery of novel chemotherapeutic agents against Babesia parasites. In the current study, we evaluated the usefulness of a fluorescence-based assay for determining the efficacies of antibabesial compounds against bovine and equine hemoparasites in in vitro cultures. Three different hematocrits (HCTs; 2.5%, 5%, and 10%) were used without daily replacement of the medium. The results of a high-throughput screening assay revealed that the best HCT was 2.5% for bovine Babesia parasites and 5% for equine Babesia and Theileria parasites. The IC50 values of diminazene aceturate obtained by fluorescence and microscopy did not differ significantly. Likewise, the IC50 values of luteolin, pyronaridine tetraphosphate, nimbolide, gedunin, and enoxacin did not differ between the two methods. In conclusion, our fluorescence-based assay uses low HCT and does not require daily replacement of culture medium, making it highly suitable for in vitro large-scale drug screening against Babesia and Theileria parasites that infect cattle and horses.

Optimization of a Fluorescence-Based Assay for Large-Scale Drug Screening against Babesia and Theileria Parasites.

Directory of Open Access Journals (Sweden)

Mohamed Abdo Rizk

Full Text Available A rapid and accurate assay for evaluating antibabesial drugs on a large scale is required for the discovery of novel chemotherapeutic agents against Babesia parasites. In the current study, we evaluated the usefulness of a fluorescence-based assay for determining the efficacies of antibabesial compounds against bovine and equine hemoparasites in in vitro cultures. Three different hematocrits (HCTs; 2.5%, 5%, and 10% were used without daily replacement of the medium. The results of a high-throughput screening assay revealed that the best HCT was 2.5% for bovine Babesia parasites and 5% for equine Babesia and Theileria parasites. The IC50 values of diminazene aceturate obtained by fluorescence and microscopy did not differ significantly. Likewise, the IC50 values of luteolin, pyronaridine tetraphosphate, nimbolide, gedunin, and enoxacin did not differ between the two methods. In conclusion, our fluorescence-based assay uses low HCT and does not require daily replacement of culture medium, making it highly suitable for in vitro large-scale drug screening against Babesia and Theileria parasites that infect cattle and horses.

A Parallel Distributed-Memory Particle Method Enables Acquisition-Rate Segmentation of Large Fluorescence Microscopy Images

Science.gov (United States)

Afshar, Yaser; Sbalzarini, Ivo F.

2016-01-01

Modern fluorescence microscopy modalities, such as light-sheet microscopy, are capable of acquiring large three-dimensional images at high data rate. This creates a bottleneck in computational processing and analysis of the acquired images, as the rate of acquisition outpaces the speed of processing. Moreover, images can be so large that they do not fit the main memory of a single computer. We address both issues by developing a distributed parallel algorithm for segmentation of large fluorescence microscopy images. The method is based on the versatile Discrete Region Competition algorithm, which has previously proven useful in microscopy image segmentation. The present distributed implementation decomposes the input image into smaller sub-images that are distributed across multiple computers. Using network communication, the computers orchestrate the collectively solving of the global segmentation problem. This not only enables segmentation of large images (we test images of up to 1010 pixels), but also accelerates segmentation to match the time scale of image acquisition. Such acquisition-rate image segmentation is a prerequisite for the smart microscopes of the future and enables online data compression and interactive experiments. PMID:27046144

A Parallel Distributed-Memory Particle Method Enables Acquisition-Rate Segmentation of Large Fluorescence Microscopy Images.

Directory of Open Access Journals (Sweden)

Yaser Afshar

Full Text Available Modern fluorescence microscopy modalities, such as light-sheet microscopy, are capable of acquiring large three-dimensional images at high data rate. This creates a bottleneck in computational processing and analysis of the acquired images, as the rate of acquisition outpaces the speed of processing. Moreover, images can be so large that they do not fit the main memory of a single computer. We address both issues by developing a distributed parallel algorithm for segmentation of large fluorescence microscopy images. The method is based on the versatile Discrete Region Competition algorithm, which has previously proven useful in microscopy image segmentation. The present distributed implementation decomposes the input image into smaller sub-images that are distributed across multiple computers. Using network communication, the computers orchestrate the collectively solving of the global segmentation problem. This not only enables segmentation of large images (we test images of up to 10(10 pixels, but also accelerates segmentation to match the time scale of image acquisition. Such acquisition-rate image segmentation is a prerequisite for the smart microscopes of the future and enables online data compression and interactive experiments.

Enhanced speed in fluorescence imaging using beat frequency multiplexing

Science.gov (United States)

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

2016-03-01

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

The application of anti-ESAT-6 monoclonal antibody fluorescent probe in ex vivo near-infrared fluorescence imaging in mice with pulmonary tuberculosis.

Science.gov (United States)

Feng, Feng; Zhang, Haoling; Zhu, Zhaoqin; Li, Cong; Shi, Yuxin; Zhang, Zhiyong

2014-09-01

Here, we aimed to assess the feasibility of anti-ESAT-6 monoclonal antibody (mAb) coupling with IR783 and rhodamine fluorescent probe in the detection of ESAT-6 expression in tuberculosis tissue of mice using near-infrared fluorescence imaging. IR783 and rhodamine were conjugated to the anti-ESAT-6 mAb or IgG. Mice in the experimental group were injected with fluorescence-labeled mAb probe, and mice in the control group were injected with fluorescence-labeled non-specific IgG antibody. Twenty-four hours later, the lung tissue of mice was examined using ex vivo near-infrared fluorescence imaging. In addition, the contrast-to-noise ratio (CNR) was calculated by measuring the signal intensities of the pulmonary lesions, normal lung tissue and background noise. The frozen lung tissue section was examined under fluorescence microscopy and compared with hemoxylin and eosin (HE) staining. The ex vivo near-infrared fluorescence imaging showed that the fluorescence signal in the lung tuberculosis lesions in the experimental group was significantly enhanced, whereas there was only a weak fluorescence signal or even no fluorescence signal in the control group. CNR values were 64.40 ± 7.02 (n = 6) and 8.75 ± 3.87 (n = 6), respectively (t = 17.01, p fluorescence accumulation distribution detected under fluorescence microscopy was consistent with HE staining of the tuberculosis region. In conclusion, anti-ESAT-6 mAb fluorescent probe could target and be applied in specific ex vivo imaging of mice tuberculosis, and may be of further use in tuberculosis in living mice. Copyright © 2013 John Wiley & Sons, Ltd.

Creating infinite contrast in fluorescence microscopy by using lanthanide centered emission

DEFF Research Database (Denmark)

R. Carro-Temboury, Miguel; Arppe, Riikka Matleena; Hempel, Casper

2017-01-01

The popularity of fluorescence microscopy arises from the inherent mode of action, where the fluorescence emission from probes is used to visualize selected features on a presumed dark background. However, the background is rarely truly dark, and image processing and analysis is needed to enhance...... the fluorescent signal that is ascribed to the selected feature. The image acquisition is facilitated by using considerable illumination, bright probes at a relatively high concentration in order to make the fluorescent signal significantly more intense than the background signal. Here, we present two methods......, while method II resolves the fluorescent signal by subtracting a background calculated via the gradient. Both methods improve signal-to-background ratio significantly and we suggest that spectral imaging of lanthanide-centered emission can be used as a tool to obtain absolute contrast in bioimaging....

Dual-detection confocal fluorescence microscopy: fluorescence axial imaging without axial scanning.

Science.gov (United States)

Lee, Dong-Ryoung; Kim, Young-Duk; Gweon, Dae-Gab; Yoo, Hongki

2013-07-29

We propose a new method for high-speed, three-dimensional (3-D) fluorescence imaging, which we refer to as dual-detection confocal fluorescence microscopy (DDCFM). In contrast to conventional beam-scanning confocal fluorescence microscopy, where the focal spot must be scanned either optically or mechanically over a sample volume to reconstruct a 3-D image, DDCFM can obtain the depth of a fluorescent emitter without depth scanning. DDCFM comprises two photodetectors, each with a pinhole of different size, in the confocal detection system. Axial information on fluorescent emitters can be measured by the axial response curve through the ratio of intensity signals. DDCFM can rapidly acquire a 3-D fluorescent image from a single two-dimensional scan with less phototoxicity and photobleaching than confocal fluorescence microscopy because no mechanical depth scans are needed. We demonstrated the feasibility of the proposed method by phantom studies.

Nanosecond fluorescence spectroscopy

International Nuclear Information System (INIS)

Leskovar, B.

1985-03-01

This article is a summary of a short course lecture given in conjunction with the 1984 Nuclear Science Symposium. Measuring systems for nanosecond fluorescence spectroscopy using single-photon counting techniques are presented. These involve systems based on relaxation-type spark gap light pulser and synchronously pumped mode-locked dye lasers. Furthermore, typical characteristics and optimization of operating conditions of the critical components responsible for the system time resolution are discussed. A short comparison of the most important deconvolution methods for numerical analysis of experimental data is given particularly with respect to the signal-to-noise ratio of the fluorescence signal. 22 refs., 8 figs

Fluorescent dyes with large Stokes shifts for super-resolution optical microscopy of biological objects: a review

International Nuclear Information System (INIS)

Sednev, Maksim V; Belov, Vladimir N; Hell, Stefan W

2015-01-01

The review deals with commercially available organic dyes possessing large Stokes shifts and their applications as fluorescent labels in optical microscopy based on stimulated emission depletion (STED). STED microscopy breaks Abbe’s diffraction barrier and provides optical resolution beyond the diffraction limit. STED microscopy is non-invasive and requires photostable fluorescent markers attached to biomolecules or other objects of interest. Up to now, in most biology-related STED experiments, bright and photoresistant dyes with small Stokes shifts of 20–40 nm were used. The rapid progress in STED microscopy showed that organic fluorophores possessing large Stokes shifts are indispensable in multi-color super-resolution techniques. The ultimate result of the imaging relies on the optimal combination of a dye, the bio-conjugation procedure and the performance of the optical microscope. Modern bioconjugation methods, basics of STED microscopy, as well as structures and spectral properties of the presently available fluorescent markers are reviewed and discussed. In particular, the spectral properties of the commercial dyes are tabulated and correlated with the available depletion wavelengths found in STED microscopes produced by LEICA Microsytems, Abberior Instruments and Picoquant GmbH. (topical review)

Fluorescence spectroscopy of dental calculus

International Nuclear Information System (INIS)

Bakhmutov, D; Gonchukov, S; Sukhinina, A

2010-01-01

The aim of the present study was to investigate the fluorescence properties of dental calculus in comparison with the properties of adjacent unaffected tooth structure using both lasers and LEDs in the UV-visible range for fluorescence excitation. The influence of calculus color on the informative signal is demonstrated. The optimal spectral bands of excitation and registration of the fluorescence are determined

Fluorescence spectroscopy of dental calculus

Science.gov (United States)

Bakhmutov, D.; Gonchukov, S.; Sukhinina, A.

2010-05-01

The aim of the present study was to investigate the fluorescence properties of dental calculus in comparison with the properties of adjacent unaffected tooth structure using both lasers and LEDs in the UV-visible range for fluorescence excitation. The influence of calculus color on the informative signal is demonstrated. The optimal spectral bands of excitation and registration of the fluorescence are determined.

Fluorescence molecular tomography in the presence of background fluorescence

International Nuclear Information System (INIS)

Soubret, Antoine; Ntziachristos, Vasilis

2006-01-01

Fluorescence molecular tomography is an emerging imaging technique that resolves the bio-distribution of engineered fluorescent probes developed for in vivo reporting of specific cellular and sub-cellular targets. The method can detect fluorochromes in picomole amounts or less, imaged through entire animals, but the detection sensitivity and imaging performance drop in the presence of background, non-specific fluorescence. In this study, we carried out a theoretical and an experimental investigation on the effect of background fluorescence on the measured signal and on the tomographic reconstruction. We further examined the performance of three subtraction methods based on physical models of photon propagation, using experimental data on phantoms and small animals. We show that the data pre-processing with subtraction schemes can improve image quality and quantification when non-specific background florescence is present

Large signal simulation of photonic crystal Fano laser

DEFF Research Database (Denmark)

Zali, Aref Rasoulzadeh; Yu, Yi; Moravvej-Farshi, Mohammad Kazem

2017-01-01

be modulated at frequencies exceeding 1 THz which is much higher than its corresponding relaxation oscillation frequency. Large signal simulation of the Fano laser is also investigated based on pseudorandom bit sequence at 0.5 Tbit/s. It shows eye patterns are open at such high modulation frequency, verifying...

Dual turn-on fluorescence signal-based controlled release system for real-time monitoring of drug release dynamics in living cells and tumor tissues.

Science.gov (United States)

Kong, Xiuqi; Dong, Baoli; Song, Xuezhen; Wang, Chao; Zhang, Nan; Lin, Weiying

2018-01-01

Controlled release systems with capabilities for direct and real-time monitoring of the release and dynamics of drugs in living systems are of great value for cancer chemotherapy. Herein, we describe a novel dual turn-on fluorescence signal-based controlled release system ( CDox ), in which the chemotherapy drug doxorubicin ( Dox ) and the fluorescent dye ( CH ) are conjugated by a hydrazone moiety, a pH-responsive cleavable linker. CDox itself shows nearly no fluorescence as the fluorescence of CH and Dox is essentially quenched by the C=N isomerization and N-N free rotation. However, when activated under acidic conditions, CDox could be hydrolyzed to afford Dox and CH , resulting in dual turn-on signals with emission peaks at 595 nm and 488 nm, respectively. Notably, CDox exhibits a desirable controlled release feature as the hydrolysis rate is limited by the steric hindrance effect from both the Dox and CH moieties. Cytotoxicity assays indicate that CDox shows much lower cytotoxicity relative to Dox , and displays higher cell inhibition rate to cancer than normal cells. With the aid of the dual turn-on fluorescence at different wavelengths, the drug release dynamics of CDox in living HepG2 and 4T-1 cells was monitored in double channels in a real-time fashion. Importantly, two-photon fluorescence imaging of CDox in living tumor tissues was also successfully performed by high-definition 3D imaging. We expect that the unique controlled release system illustrated herein could provide a powerful means to investigate modes of action of drugs, which is critical for development of much more robust and effective chemotherapy drugs.

Ratiometric, visual, dual-signal fluorescent sensing and imaging of pH/copper ions in real samples based on carbon dots-fluorescein isothiocyanate composites.

Science.gov (United States)

Zhu, Xinxin; Jin, Hui; Gao, Cuili; Gui, Rijun; Wang, Zonghua

2017-01-01

In this article, a facile aqueous synthesis of carbon dots (CDs) was developed by using natural kelp as a new carbon source. Through hydrothermal carbonization of kelp juice, fluorescent CDs were prepared and the CDs' surface was modified with polyethylenimine (PEI). The PEI-modified CDs were conjugated with fluorescein isothiocyanate (FITC) to fabricate CDs-FITC composites. To exploit broad applications, the CDs-FITC composites were developed as fluorescent sensing or imaging platforms of pH and Cu 2+ . Analytical performances of the composites-based fluorescence (FL) sensors were evaluated, including visual FL imaging of pH in glass bottle, ratiometric FL sensing of pH in yogurt samples, visual FL latent fingerprint and leaf imaging detection of [Cu 2+ ], dual-signal FL sensing of [Cu 2+ ] in yogurt and human serum samples. Experimental results from ratiometric, visual, dual-signal FL sensing and imaging applications confirmed the high feasibility, accuracy, stabilization and simplicity of CDs-FITC composites-based FL sensors for the detection of pH and Cu 2+ ions in real samples. Copyright © 2016 Elsevier B.V. All rights reserved.

High-resolution methods for fluorescence retrieval from space

NARCIS (Netherlands)

Mazzoni, M.; Falorni, P.; Verhoef, W.

2010-01-01

The retrieval from space of a very weak fluorescence signal was studied in the O2A and O2B oxygen atmospheric absorption bands. The accuracy of the method was tested for the retrieval of the chlorophyll fluorescence and reflectance terms contributing to the sensor signal. The radiance at the top of

A study on a portable fluorescence imaging system

Science.gov (United States)

Chang, Han-Chao; Wu, Wen-Hong; Chang, Chun-Li; Huang, Kuo-Cheng; Chang, Chung-Hsing; Chiu, Shang-Chen

2011-09-01

The fluorescent reaction is that an organism or dye, excited by UV light (200-405 nm), emits a specific frequency of light; the light is usually a visible or near infrared light (405-900 nm). During the UV light irradiation, the photosensitive agent will be induced to start the photochemical reaction. In addition, the fluorescence image can be used for fluorescence diagnosis and then photodynamic therapy can be given to dental diseases and skin cancer, which has become a useful tool to provide scientific evidence in many biomedical researches. However, most of the methods on acquiring fluorescence biology traces are still stay in primitive stage, catching by naked eyes and researcher's subjective judgment. This article presents a portable camera to obtain the fluorescence image and to make up a deficit from observer competence and subjective judgment. Furthermore, the portable camera offers the 375nm UV-LED exciting light source for user to record fluorescence image and makes the recorded image become persuasive scientific evidence. In addition, when the raising the rate between signal and noise, the signal processing module will not only amplify the fluorescence signal up to 70 %, but also decrease the noise significantly from environmental light on bill and nude mouse testing.

LASSIE: the large analogue signal and scaling information environment for FAIR

International Nuclear Information System (INIS)

Hoffmann, T.; Braeuning, H.; Haseitl, R.

2012-01-01

At FAIR, the Facility for Antiproton and Ion Research, several new accelerators and storage rings such as the SIS-100, HESR, CR, the inter-connecting HEBT beam lines, S-FRS and experiments will be built. All of these installations are equipped with beam diagnostic devices and other components, which deliver time-resolved analogue signals to show status, quality and performance of the accelerators. These signals can originate from particle detectors such as ionization chambers and plastic scintillators, but also from adapted output signals of transformers, collimators, magnet functions, RF cavities and others. To visualize and precisely correlate the time axis of all input signals a dedicated FESA based data acquisition and analysis system named LASSIE, the Large Analogue Signal and Scaling Information Environment, is currently being developed. The main operation mode of LASSIE is currently pulse counting with latching VME scaler boards. Later enhancements for ADC, QDC, or TDC digitization in the future are foreseen. The concept, features and challenges of this large distributed data acquisition system are presented. (authors)

Multiphoton fluorescence lifetime imaging of chemotherapy distribution in solid tumors

Science.gov (United States)

Carlson, Marjorie; Watson, Adrienne L.; Anderson, Leah; Largaespada, David A.; Provenzano, Paolo P.

2017-11-01

Doxorubicin is a commonly used chemotherapeutic employed to treat multiple human cancers, including numerous sarcomas and carcinomas. Furthermore, doxorubicin possesses strong fluorescent properties that make it an ideal reagent for modeling drug delivery by examining its distribution in cells and tissues. However, while doxorubicin fluorescence and lifetime have been imaged in live tissue, its behavior in archival samples that frequently result from drug and treatment studies in human and animal patients, and murine models of human cancer, has to date been largely unexplored. Here, we demonstrate imaging of doxorubicin intensity and lifetimes in archival formalin-fixed paraffin-embedded sections from mouse models of human cancer with multiphoton excitation and multiphoton fluorescence lifetime imaging microscopy (FLIM). Multiphoton excitation imaging reveals robust doxorubicin emission in tissue sections and captures spatial heterogeneity in cells and tissues. However, quantifying the amount of doxorubicin signal in distinct cell compartments, particularly the nucleus, often remains challenging due to strong signals in multiple compartments. The addition of FLIM analysis to display the spatial distribution of excited state lifetimes clearly distinguishes between signals in distinct compartments such as the cell nuclei versus cytoplasm and allows for quantification of doxorubicin signal in each compartment. Furthermore, we observed a shift in lifetime values in the nuclei of transformed cells versus nontransformed cells, suggesting a possible diagnostic role for doxorubicin lifetime imaging to distinguish normal versus transformed cells. Thus, data here demonstrate that multiphoton FLIM is a highly sensitive platform for imaging doxorubicin distribution in normal and diseased archival tissues.

Frequency division multiplexed multi-color fluorescence microscope system

Science.gov (United States)

Le, Vu Nam; Yang, Huai Dong; Zhang, Si Chun; Zhang, Xin Rong; Jin, Guo Fan

2017-10-01

Grayscale camera can only obtain gray scale image of object, while the multicolor imaging technology can obtain the color information to distinguish the sample structures which have the same shapes but in different colors. In fluorescence microscopy, the current method of multicolor imaging are flawed. Problem of these method is affecting the efficiency of fluorescence imaging, reducing the sampling rate of CCD etc. In this paper, we propose a novel multiple color fluorescence microscopy imaging method which based on the Frequency division multiplexing (FDM) technology, by modulating the excitation lights and demodulating the fluorescence signal in frequency domain. This method uses periodic functions with different frequency to modulate amplitude of each excitation lights, and then combine these beams for illumination in a fluorescence microscopy imaging system. The imaging system will detect a multicolor fluorescence image by a grayscale camera. During the data processing, the signal obtained by each pixel of the camera will be processed with discrete Fourier transform, decomposed by color in the frequency domain and then used inverse discrete Fourier transform. After using this process for signals from all of the pixels, monochrome images of each color on the image plane can be obtained and multicolor image is also acquired. Based on this method, this paper has constructed and set up a two-color fluorescence microscope system with two excitation wavelengths of 488 nm and 639 nm. By using this system to observe the linearly movement of two kinds of fluorescent microspheres, after the data processing, we obtain a two-color fluorescence dynamic video which is consistent with the original image. This experiment shows that the dynamic phenomenon of multicolor fluorescent biological samples can be generally observed by this method. Compared with the current methods, this method can obtain the image signals of each color at the same time, and the color video's frame

Large-signal analysis of DC motor drive system using state-space averaging technique

International Nuclear Information System (INIS)

Bekir Yildiz, Ali

2008-01-01

The analysis of a separately excited DC motor driven by DC-DC converter is realized by using state-space averaging technique. Firstly, a general and unified large-signal averaged circuit model for DC-DC converters is given. The method converts power electronic systems, which are periodic time-variant because of their switching operation, to unified and time independent systems. Using the averaged circuit model enables us to combine the different topologies of converters. Thus, all analysis and design processes about DC motor can be easily realized by using the unified averaged model which is valid during whole period. Some large-signal variations such as speed and current relating to DC motor, steady-state analysis, large-signal and small-signal transfer functions are easily obtained by using the averaged circuit model

Iodinated oil-loaded, fluorescent mesoporous silica-coated iron oxide nanoparticles for magnetic resonance imaging/computed tomography/fluorescence trimodal imaging

Directory of Open Access Journals (Sweden)

Xue S

2014-05-01

Full Text Available Sihan Xue,1 Yao Wang,1 Mengxing Wang,2 Lu Zhang,1 Xiaoxia Du,2 Hongchen Gu,1 Chunfu Zhang1,31School of Biomedical Engineering and Med-X Research Institute, Shanghai Jiao Tong University, 2Shanghai Key Laboratory of Magnetic Resonance, Department of Physics, East China Normal University, 3State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, School of Biomedical Engineering, Shanghai Jiao Tong University, Shanghai, People’s Republic of ChinaAbstract: In this study, a novel magnetic resonance imaging (MRI/computed tomography (CT/fluorescence trifunctional probe was prepared by loading iodinated oil into fluorescent mesoporous silica-coated superparamagnetic iron oxide nanoparticles (i-fmSiO4@SPIONs. Fluorescent mesoporous silica-coated superparamagnetic iron oxide nanoparticles (fmSiO4@SPIONs were prepared by growing fluorescent dye-doped silica onto superparamagnetic iron oxide nanoparticles (SPIONs directed by a cetyltrimethylammonium bromide template. As prepared, fmSiO4@SPIONs had a uniform size, a large surface area, and a large pore volume, which demonstrated high efficiency for iodinated oil loading. Iodinated oil loading did not change the sizes of fmSiO4@SPIONs, but they reduced the MRI T2 relaxivity (r2 markedly. I-fmSiO4@SPIONs were stable in their physical condition and did not demonstrate cytotoxic effects under the conditions investigated. In vitro studies indicated that the contrast enhancement of MRI and CT, and the fluorescence signal intensity of i-fmSiO4@SPION aqueous suspensions and macrophages, were intensified with increased i-fmSiO4@SPION concentrations in suspension and cell culture media. Moreover, for the in vivo study, the accumulation of i-fmSiO4@SPIONs in the liver could also be detected by MRI, CT, and fluorescence imaging. Our study demonstrated that i-fmSiO4@SPIONs had great potential for MRI/C/fluorescence trimodal imaging.Keywords: multifunctional probe, SPIONs, mesoporous silica

Period doubling and chaos in large area Josephson junctions induced by rf signals

DEFF Research Database (Denmark)

Olsen, O. H.; Samuelsen, Mogens Rugholm

1985-01-01

The influence of an applied rf signal on the emitted radiation from a large area Josephson junction is examined. A model of the system is presented in the framework of the one-dimensional sine-Gordon equation. The model linearizes for small and large values of the amplitude of the applied signal...

Tools for the quantitative analysis of sedimentation boundaries detected by fluorescence optical analytical ultracentrifugation.

Directory of Open Access Journals (Sweden)

Huaying Zhao

Full Text Available Fluorescence optical detection in sedimentation velocity analytical ultracentrifugation allows the study of macromolecules at nanomolar concentrations and below. This has significant promise, for example, for the study of systems of high-affinity protein interactions. Here we describe adaptations of the direct boundary modeling analysis approach implemented in the software SEDFIT that were developed to accommodate unique characteristics of the confocal fluorescence detection system. These include spatial gradients of signal intensity due to scanner movements out of the plane of rotation, temporal intensity drifts due to instability of the laser and fluorophores, and masking of the finite excitation and detection cone by the sample holder. In an extensive series of experiments with enhanced green fluorescent protein ranging from low nanomolar to low micromolar concentrations, we show that the experimental data provide sufficient information to determine the parameters required for first-order approximation of the impact of these effects on the recorded data. Systematic deviations of fluorescence optical sedimentation velocity data analyzed using conventional sedimentation models developed for absorbance and interference optics are largely removed after these adaptations, resulting in excellent fits that highlight the high precision of fluorescence sedimentation velocity data, thus allowing a more detailed quantitative interpretation of the signal boundaries that is otherwise not possible for this system.

Large-Signal Code TESLA: Current Status and Recent Development

National Research Council Canada - National Science Library

Chernyavskiy, Igor A; Vlasov, Alexander N; Cooke, Simon J; Abe, David K; Levush, Baruch; Antonsen, Jr., Thomas M; Nguyen, Khanh T

2008-01-01

.... One such tool is the large-signal code TESLA, which was successfully applied for the modeling of single-beam and multiple-beam klystron devices at the Naval Research Laboratory and which is now used by number of U.S. companies...

Let's Exploit Available Knowledge on Vegetation Fluorescence

Science.gov (United States)

Magnani, Federico; Raddi, Sabrina; Mohammed, Gina; Middleton, Elizabeth M.

2014-01-01

. 2).We know, however, that the ratio is not constant, but changes widely in response to light, CO2, stomatal limitations, and extreme stress (4, 5). What’s more, we can make sense of such changes, thus extracting valuable information from the very scatter that is apparent in their data. However, this process will require the availability of more tailored instruments, such as the one planned for the FLEX mission. As already stressed by Guanter et al. (2), the spatial resolution of the Global Ozone Monitoring Experiment-2 sensor (40 × 80 km) makes it difficult to compare meaningfully the fluorescence signal with ground measurements, when only 60–70% of the footprint consists of the desired land-cover type (table S1 in ref. 2), suggesting that this could be largely responsible for the low signals observed in European grasslands. Moreover, the overpass time of the MetOp-A satellite (9:30 AM) implies that fluorescence is generally measured under light-limiting conditions, when fluorescence is only marginally affected by stomatal closure even under stress conditions. This result could explain the seasonal mismatch with daily GPP observed in natural ecosystems in the absence of irrigation (figure 4 in ref. 2). We hope, therefore, that this welcome contribution to this fast-advancing field will help demonstrate the potential of the new technique, and pave the way for more refined studies under both a technological and scientific point of view.

Applications of optical fiber to the remote fluorescence analysis

International Nuclear Information System (INIS)

Shin, Jang Soo; Kim, Duck Hueon; Lee, Soo Ho

1992-12-01

The laser fluorometer developed in 1987 has been used in real circumstances for trace uranium analysis. And, we have been trying to improve the instrument to be able to apply in analytical circumstances of remote measurement using optical fiber. The N 2 laser beam and the resulting fluorescence light could be successfully transmitted through a quartz-made optical fiber. The wavelength resolution and the fluorescence decay time resolution induced by pulsed N 2 laser were used to the uranium fluorescence analyses. The fluorescence of uranium in nitric acid medium was measured successfully using the system. The fluorescence signal was analysed using simplex method which is useful to deconvolute the mixed signals. An analytical method using thermal lens effect was developed. The method will be a complementary one for the fluorescence measurement. (Author)

Improving x-ray fluorescence signal for benchtop polychromatic cone-beam x-ray fluorescence computed tomography by incident x-ray spectrum optimization: a Monte Carlo study.

Science.gov (United States)

Manohar, Nivedh; Jones, Bernard L; Cho, Sang Hyun

2014-10-01

To develop an accurate and comprehensive Monte Carlo (MC) model of an experimental benchtop polychromatic cone-beam x-ray fluorescence computed tomography (XFCT) setup and apply this MC model to optimize incident x-ray spectrum for improving production/detection of x-ray fluorescence photons from gold nanoparticles (GNPs). A detailed MC model, based on an experimental XFCT system, was created using the Monte Carlo N-Particle (MCNP) transport code. The model was validated by comparing MC results including x-ray fluorescence (XRF) and scatter photon spectra with measured data obtained under identical conditions using 105 kVp cone-beam x-rays filtered by either 1 mm of lead (Pb) or 0.9 mm of tin (Sn). After validation, the model was used to investigate the effects of additional filtration of the incident beam with Pb and Sn. Supplementary incident x-ray spectra, representing heavier filtration (Pb: 2 and 3 mm; Sn: 1, 2, and 3 mm) were computationally generated and used with the model to obtain XRF/scatter spectra. Quasimonochromatic incident x-ray spectra (81, 85, 90, 95, and 100 keV with 10 keV full width at half maximum) were also investigated to determine the ideal energy for distinguishing gold XRF signal from the scatter background. Fluorescence signal-to-dose ratio (FSDR) and fluorescence-normalized scan time (FNST) were used as metrics to assess results. Calculated XRF/scatter spectra for 1-mm Pb and 0.9-mm Sn filters matched (r ≥ 0.996) experimental measurements. Calculated spectra representing additional filtration for both filter materials showed that the spectral hardening improved the FSDR at the expense of requiring a much longer FNST. In general, using Sn instead of Pb, at a given filter thickness, allowed an increase of up to 20% in FSDR, more prominent gold XRF peaks, and up to an order of magnitude decrease in FNST. Simulations using quasimonochromatic spectra suggested that increasing source x-ray energy, in the investigated range of 81-100 ke

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

Science.gov (United States)

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

2017-01-25

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

Large-area, low-noise, high-speed, photodiode-based fluorescence detectors with fast overdrive recovery

International Nuclear Information System (INIS)

Bickman, S.; DeMille, D.

2005-01-01

Two large-area, low-noise, high-speed fluorescence detectors have been built. One detector consists of a photodiode with an area of 28 mmx28 mm and a low-noise transimpedance amplifier. This detector has a input light-equivalent spectral noise density of less than 3 pW/√(Hz), can recover from a large scattered light pulse within 10 μs, and has a bandwidth of at least 900 kHz. The second detector consists of a 16-mm-diam avalanche photodiode and a low-noise transimpedance amplifier. This detector has an input light-equivalent spectral noise density of 0.08 pW/√(Hz), also can recover from a large scattered light pulse within 10 μs, and has a bandwidth of 1 MHz

Correlated cryo-fluorescence and cryo-electron microscopy with high spatial precision and improved sensitivity

International Nuclear Information System (INIS)

Schorb, Martin; Briggs, John A.G.

2014-01-01

Performing fluorescence microscopy and electron microscopy on the same sample allows fluorescent signals to be used to identify and locate features of interest for subsequent imaging by electron microscopy. To carry out such correlative microscopy on vitrified samples appropriate for structural cryo-electron microscopy it is necessary to perform fluorescence microscopy at liquid-nitrogen temperatures. Here we describe an adaptation of a cryo-light microscopy stage to permit use of high-numerical aperture objectives. This allows high-sensitivity and high-resolution fluorescence microscopy of vitrified samples. We describe and apply a correlative cryo-fluorescence and cryo-electron microscopy workflow together with a fiducial bead-based image correlation procedure. This procedure allows us to locate fluorescent bacteriophages in cryo-electron microscopy images with an accuracy on the order of 50 nm, based on their fluorescent signal. It will allow the user to precisely and unambiguously identify and locate objects and events for subsequent high-resolution structural study, based on fluorescent signals. - Highlights: • Workflow for correlated cryo-fluorescence and cryo-electron microscopy. • Cryo-fluorescence microscopy setup incorporating a high numerical aperture objective. • Fluorescent signals located in cryo-electron micrographs with 50 nm spatial precision

Correlated cryo-fluorescence and cryo-electron microscopy with high spatial precision and improved sensitivity

Energy Technology Data Exchange (ETDEWEB)

Schorb, Martin [Structural and Computational Biology Unit, European Molecular Biology Laboratory, D-69117 Heidelberg (Germany); Briggs, John A.G., E-mail: john.briggs@embl.de [Structural and Computational Biology Unit, European Molecular Biology Laboratory, D-69117 Heidelberg (Germany); Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, D-69117 Heidelberg (Germany)

2014-08-01

Performing fluorescence microscopy and electron microscopy on the same sample allows fluorescent signals to be used to identify and locate features of interest for subsequent imaging by electron microscopy. To carry out such correlative microscopy on vitrified samples appropriate for structural cryo-electron microscopy it is necessary to perform fluorescence microscopy at liquid-nitrogen temperatures. Here we describe an adaptation of a cryo-light microscopy stage to permit use of high-numerical aperture objectives. This allows high-sensitivity and high-resolution fluorescence microscopy of vitrified samples. We describe and apply a correlative cryo-fluorescence and cryo-electron microscopy workflow together with a fiducial bead-based image correlation procedure. This procedure allows us to locate fluorescent bacteriophages in cryo-electron microscopy images with an accuracy on the order of 50 nm, based on their fluorescent signal. It will allow the user to precisely and unambiguously identify and locate objects and events for subsequent high-resolution structural study, based on fluorescent signals. - Highlights: • Workflow for correlated cryo-fluorescence and cryo-electron microscopy. • Cryo-fluorescence microscopy setup incorporating a high numerical aperture objective. • Fluorescent signals located in cryo-electron micrographs with 50 nm spatial precision.

Hyperspectral solar-induced chlorophyll fluorescence of urban tree leaves: Analyses and applications

Science.gov (United States)

Van Wittenberghe, Shari

Solar energy is the primary energy source for life on Earth which is converted into chemical energy through photosynthesis by plants, algae and cyanobacteria, releasing fuel for the organisms' activities. To dissipate excess of absorbed light energy, plants emit chlorophyll (Chl) fluorescence (650-850 nm) from the same location where photosynthesis takes place. Hence, it provides information on the efficiency of primary energy conversion. From this knowledge, many applications on vegetation and crop stress monitoring could be developed, a necessity for our planet under threat of a changing global climate. Even though the Chl fluorescence signal is weak against the intense reflected radiation background, methods for retrieving the solar-induced Chl fluorescence have been refined over the last years, both at leaf and airborne scale. However, a lack of studies on solar-induced Chl fluorescence gives difficulties for the interpretation of the signal. Within this thesis, hyperspectral upward and downward solar-induced Chl fluorescence is measured at leaf level. Fluorescence yield (FY) is calculated as well as different ratios characterizing the emitted Chl fluorescence shape. The research in this PhD dissertation illustrates the influence of several factors on the solar-induced Chl fluorescence signal. For instance, both the intensity of FY and its spectral shape of urban tree leaves are able to change under influence of stress factors such as traffic air pollution. This shows how solar-induced Chl fluorescence could function as an early stress indicator for vegetation. Further, it is shown that the signal contains information on the ultrastructure of the photosynthetic apparatus. Also, it is proven that the leaf anatomical structure and related light scattering properties play a role in the partitioning between upward and downward Chl fluorescence emission. All these findings indicate how the Chl fluorescence spectrum is influenced by factors which also influence

Ultrasensitive fluorescence immunoassay for detection of ochratoxin A using catalase-mediated fluorescence quenching of CdTe QDs

Science.gov (United States)

Huang, Xiaolin; Zhan, Shengnan; Xu, Hengyi; Meng, Xianwei; Xiong, Yonghua; Chen, Xiaoyuan

2016-04-01

Herein, for the first time we report an improved competitive fluorescent enzyme linked immunosorbent assay (ELISA) for the ultrasensitive detection of ochratoxin A (OTA) by using hydrogen peroxide (H2O2)-induced fluorescence quenching of mercaptopropionic acid-modified CdTe quantum dots (QDs). In this immunoassay, catalase (CAT) was labeled with OTA as a competitive antigen to connect the fluorescence signals of the QDs with the concentration of the target. Through the combinatorial use of H2O2-induced fluorescence quenching of CdTe QDs as a fluorescence signal output and the ultrahigh catalytic activity of CAT to H2O2, our proposed method could be used to perform a dynamic linear detection of OTA ranging from 0.05 pg mL-1 to 10 pg mL-1. The half maximal inhibitory concentration was 0.53 pg mL-1 and the limit of detection was 0.05 pg mL-1. These values were approximately 283- and 300-folds lower than those of horseradish peroxidase (HRP)-based conventional ELISA, respectively. The reported method is accurate, highly reproducible, and specific against other mycotoxins in agricultural products as well. In summary, the developed fluorescence immunoassay based on H2O2-induced fluorescence quenching of CdTe QDs can be used for the rapid and highly sensitive detection of mycotoxins or haptens in food safety monitoring.Herein, for the first time we report an improved competitive fluorescent enzyme linked immunosorbent assay (ELISA) for the ultrasensitive detection of ochratoxin A (OTA) by using hydrogen peroxide (H2O2)-induced fluorescence quenching of mercaptopropionic acid-modified CdTe quantum dots (QDs). In this immunoassay, catalase (CAT) was labeled with OTA as a competitive antigen to connect the fluorescence signals of the QDs with the concentration of the target. Through the combinatorial use of H2O2-induced fluorescence quenching of CdTe QDs as a fluorescence signal output and the ultrahigh catalytic activity of CAT to H2O2, our proposed method could be used to

Particle Image Velocimetry Applications Using Fluorescent Dye-Doped Particles

Science.gov (United States)

Petrosky, Brian J.; Maisto, Pietro; Lowe, K. Todd; Andre, Matthieu A.; Bardet, Philippe M.; Tiemsin, Patsy I.; Wohl, Christopher J.; Danehy, Paul M.

2015-01-01

Polystyrene latex sphere particles are widely used to seed flows for velocimetry techniques such as Particle Image Velocimetry (PIV) and Laser Doppler Velocimetry (LDV). These particles may be doped with fluorescent dyes such that signals spectrally shifted from the incident laser wavelength may be detected via Laser Induced Fluorescence (LIF). An attractive application of the LIF signal is achieving velocimetry in the presence of strong interference from laser scatter, opening up new research possibilities very near solid surfaces or at liquid/gas interfaces. Additionally, LIF signals can be used to tag different fluid streams to study mixing. While fluorescence-based PIV has been performed by many researchers for particles dispersed in water flows, the current work is among the first in applying the technique to micron-scale particles dispersed in a gas. A key requirement for such an application is addressing potential health hazards from fluorescent dyes; successful doping of Kiton Red 620 (KR620) has enabled the use of this relatively safe dye for fluorescence PIV for the first time. In this paper, basic applications proving the concept of PIV using the LIF signal from KR620-doped particles are exhibited for a free jet and a twophase flow apparatus. Results indicate that while the fluorescence PIV techniques are roughly 2 orders of magnitude weaker than Mie scattering, they provide a viable method for obtaining data in flow regions previously inaccessible via standard PIV. These techniques have the potential to also complement Mie scattering signals, for example in multi-stream and/or multi-phase experiments.

New insight in the template decomposition process of large zeolite ZSM-5 crystals: an in situ UV-Vis/fluorescence micro-spectroscopy study

NARCIS (Netherlands)

Karwacki, L.|info:eu-repo/dai/nl/304824283; Weckhuysen, B.M.|info:eu-repo/dai/nl/285484397

2011-01-01

A combination of in situ UV-Vis and confocal fluorescence micro-spectroscopy was used to study the template decomposition process in large zeolite ZSM-5 crystals. Correlation of polarized light dependent UV-Vis absorption spectra with confocal fluorescence emission spectra in the 400–750 nm region

Fluorescence and Four-Wave Mixing in Electromagnetically Induced Transparency Windows

International Nuclear Information System (INIS)

Wang Zhi-Guo; Li Cheng; Zhang Zhao-Yang; Che Jun-Ling; Qin Meng-Zhe; He Jia-Nan; Zhang Yan-Peng

2013-01-01

We simultaneously compare the probe transmission, Four-Wave Mixing (FWM) and fluorescence signals with dressing effects in a four-level atomic system. The variation rules of three types of signals are exhibited by changing the frequency detuning and power of incident laser beams. The interplay between two ladder subsystems is investigated in the Y-type atomic system. In particular, the fluorescence signal with ultra-narrow linewidth is obtained due to being sheared twice by the electromagnetically induced transparency window. Such fluorescence with very high coherence and monochromaticity can be used for the quantum correlation and narrow linewidth laser

An accurate and simple large signal model of HEMT

DEFF Research Database (Denmark)

Liu, Qing

1989-01-01

A large-signal model of discrete HEMTs (high-electron-mobility transistors) has been developed. It is simple and suitable for SPICE simulation of hybrid digital ICs. The model parameters are extracted by using computer programs and data provided by the manufacturer. Based on this model, a hybrid...

Large-signal characterization of DDR silicon IMPATTs operating in millimeter-wave and terahertz regime

International Nuclear Information System (INIS)

Acharyya, Aritra; Banerjee, J. P.; Chakraborty, Jit; Das, Kausik; Datta, Subir; De, Pritam; Banerjee, Suranjana

2013-01-01

The authors have carried out the large-signal characterization of silicon-based double-drift region (DDR) impact avalanche transit time (IMPATT) devices designed to operate up to 0.5 THz using a large-signal simulation method developed by the authors based on non-sinusoidal voltage excitation. The effect of band-to-band tunneling as well as parasitic series resistance on the large-signal properties of DDR Si IMPATTs have also been studied at different mm-wave and THz frequencies. Large-signal simulation results show that DDR Si IMPATT is capable of delivering peak RF power of 633.69 mW with 7.95% conversion efficiency at 94 GHz for 50% voltage modulation, whereas peak RF power output and efficiency fall to 81.08 mW and 2.01% respectively at 0.5 THz for same voltage modulation. The simulation results are compared with the experimental results and are found to be in close agreement. (semiconductor devices)

A growth factor signaling cascade confined to circular ruffles in macrophages

Directory of Open Access Journals (Sweden)

Timothy P. Welliver

2012-06-01

The formation of macropinosomes requires large-scale movements of membranes and the actin cytoskeleton. Over several minutes, actin-rich surface ruffles transform into 1–5 µm diameter circular ruffles, which close at their distal margins, creating endocytic vesicles. Previous studies using fluorescent reporters of phosphoinositides and Rho-family GTPases showed that signals generated by macrophages in response to the growth factor Macrophage Colony-Stimulating Factor (M-CSF appeared transiently in domains of plasma membrane circumscribed by circular ruffles. To address the question of how signaling molecules are coordinated in such large domains of plasma membrane, this study analyzed the relative timing of growth factor-dependent signals as ruffles transformed into macropinosomes. Fluorescent protein chimeras expressed in macrophages were imaged by microscopy and quantified relative to circular ruffle formation and cup closure. The large size of macropinocytic cups allowed temporal resolution of the transitions in phosphoinositides and associated enzyme activities that organize cup closure. Circular ruffles contained transient and sequential spikes of phosphatidylinositol (4,5-bisphosphate (PI(4,5P2, phosphatidylinositol (3,4,5-trisphosphate (PIP3, diacylglycerol, PI(3,4P2, PI(3P and the activities of protein kinase C-α, Rac1, Ras and Rab5. The confinement of this signal cascade to circular ruffles indicated that diffusion barriers present in these transient structures focus feedback activation and deactivation of essential enzyme activities into restricted domains of plasma membrane.

Aptamer based fluorescent cocaine assay based on the use of graphene oxide and exonuclease III-assisted signal amplification

International Nuclear Information System (INIS)

Zhang, Yulin; Zhang, Guo-Jun; Sun, Zhongyue; Tang, Lina; Zhang, Hong

2016-01-01

The article reports an aptamer based assay for cocaine by employing graphene oxide and exonuclease III-assisted signal amplification. It is based on the following scheme and experimental steps: (1) Exo III can digest dsDNA with blunt or recessed 3-terminus, but it has limited activity to ssDNA or dsDNA with protruding 3-terminus; (2) GO can absorb the FAM-labeled ssDNA probe and quench the fluorescence of probe, while the affinity between FAM-labeled mononucleotide and GO is negligible; (3) Cocaine aptamer can be split into two flexible ssDNA pieces (Probe 1 and Probe 2) without significant perturbation of cocaine-binding abilities; (4) The triple complex consisting of Probe 1, Probe 2 and cocaine can be digested by Exo III with the similar efficiency as normal dsDNA. Cocaine aptamer is split into two flexible ssDNA pieces (Probe 2 and 3′-FAM-labeled Probe 1). Cocaine can mediate the cocaine aptamer fragments forming a triplex. The triple complex has unique characteristic with 3′-FAM-labeled blunt end at the Probe 1 and 3′-overhang end at Probe 2. If exonuclease III is added, it will catalyze the stepwise removal of fluorescein (FAM) labeled mononucleotides from the 3-hydroxy termini of the special triplex complex, resulting in liberation of cocaine. The cocaine released in this step can produce a new cleavage cycle, thereby leading to target recycling. Through such a cyclic bound-hydrolysis process, small amounts of cocaine can induce the cleavage of a large number of FAM-labeled probe 1. The cleaved FAM-labeled mononucleotides are not adsorbed on the surface of graphene oxide (GO), so a strong fluorescence signal enhancement is observed as the cocaine triggers enzymatic digestion. Under optimized conditions, the assay allows cocaine to be detected in the 1 to 500 nM concentration range with a detection limit of 0.1 nM. The method was applied to the determination of cocaine in spiked human plasma, with recoveries ranging from 92.0 to 111.8 % and RSD of <12

Plasmonically amplified fluorescence bioassay with microarray format

Science.gov (United States)

Gogalic, S.; Hageneder, S.; Ctortecka, C.; Bauch, M.; Khan, I.; Preininger, Claudia; Sauer, U.; Dostalek, J.

2015-05-01

Plasmonic amplification of fluorescence signal in bioassays with microarray detection format is reported. A crossed relief diffraction grating was designed to couple an excitation laser beam to surface plasmons at the wavelength overlapping with the absorption and emission bands of fluorophore Dy647 that was used as a label. The surface of periodically corrugated sensor chip was coated with surface plasmon-supporting gold layer and a thin SU8 polymer film carrying epoxy groups. These groups were employed for the covalent immobilization of capture antibodies at arrays of spots. The plasmonic amplification of fluorescence signal on the developed microarray chip was tested by using interleukin 8 sandwich immunoassay. The readout was performed ex situ after drying the chip by using a commercial scanner with high numerical aperture collecting lens. Obtained results reveal the enhancement of fluorescence signal by a factor of 5 when compared to a regular glass chip.

Efficient processing of fluorescence images using directional multiscale representations.

Science.gov (United States)

Labate, D; Laezza, F; Negi, P; Ozcan, B; Papadakis, M

2014-01-01

Recent advances in high-resolution fluorescence microscopy have enabled the systematic study of morphological changes in large populations of cells induced by chemical and genetic perturbations, facilitating the discovery of signaling pathways underlying diseases and the development of new pharmacological treatments. In these studies, though, due to the complexity of the data, quantification and analysis of morphological features are for the vast majority handled manually, slowing significantly data processing and limiting often the information gained to a descriptive level. Thus, there is an urgent need for developing highly efficient automated analysis and processing tools for fluorescent images. In this paper, we present the application of a method based on the shearlet representation for confocal image analysis of neurons. The shearlet representation is a newly emerged method designed to combine multiscale data analysis with superior directional sensitivity, making this approach particularly effective for the representation of objects defined over a wide range of scales and with highly anisotropic features. Here, we apply the shearlet representation to problems of soma detection of neurons in culture and extraction of geometrical features of neuronal processes in brain tissue, and propose it as a new framework for large-scale fluorescent image analysis of biomedical data.

Exciton-controlled fluorescence: application to hybridization-sensitive fluorescent DNA probe.

Science.gov (United States)

Okamoto, Akimitsu; Ikeda, Shuji; Kubota, Takeshi; Yuki, Mizue; Yanagisawa, Hiroyuki

2009-01-01

A hybridization-sensitive fluorescent probe has been designed for nucleic acid detection, using the concept of fluorescence quenching caused by the intramolecular excitonic interaction of fluorescence dyes. We synthesized a doubly thiazole orange-labeled nucleotide showing high fluorescence intensity for a hybrid with the target nucleic acid and effective quenching for the single-stranded state. This exciton-controlled fluorescent probe was applied to living HeLa cells using microinjection to visualize intracellular mRNA localization. Immediately after injection of the probe into the cell, fluorescence was observed from the probe hybridizing with the target RNA. This fluorescence rapidly decreased upon addition of a competitor DNA. Multicoloring of this probe resulted in the simple simultaneous detection of plural target nucleic acid sequences. This probe realized a large, rapid, reversible change in fluorescence intensity in sensitive response to the amount of target nucleic acid, and facilitated spatiotemporal monitoring of the behavior of intracellular RNA.

Synovitis in mice with inflammatory arthritis monitored with quantitative analysis of dynamic contrast-enhanced NIR fluorescence imaging using iRGD-targeted liposomes as fluorescence probes

Directory of Open Access Journals (Sweden)

Wu H

2018-03-01

Full Text Available Hao Wu,1,2,* Haohan Wu,1,2,* Yanni He,1 Zhen Gan,2 Zhili Xu,1,2 Meijun Zhou,1,2 Sai Liu,1,2 Hongmei Liu1 1Department of Ultrasonography, Guangdong Second Provincial General Hospital Affiliated to Southern Medical University, Guangzhou, China; 2Department of Ultrasonography, The Third Affiliated Hospital of Southern Medical University, Guangzhou, China *These authors contributed equally to this work Background: Rheumatoid arthritis (RA is a common inflammatory disorder characterized primarily by synovitis and pannus formation in multiple joints, causing joints destruction and irreversible disability in most cases. Early diagnosis and effective therapy monitoring of RA are of importance for achieving the favorable prognosis. Methods: We first prepared the targeted fluorescence probes, and then explored the feasibility of near-infrared (NIR fluorescence molecular imaging to detect and evaluate the RA via the targeted fluorescence probes by quantitative analysis in this study. Results: The targeted fluorescence probes (indocyanine green-liposomes decorated with iRGD peptide [iLPs] was successfully prepared. The quantitative analysis found that strong fluorescence signal was detected in inflamed paws and the fluorescence signal in iLPs group was 3.03-fold higher than that in non-targeted (indocyanine green-liposomes decorated without iRGD peptide [LPs] group (P<0.01 at 15 min after injection, whereas the fluorescence signal from iLPs signal can almost not be observed in the non-inflamed paws, showing the high sensitivity and accuracy for arthritis by the NIR fluorescence imaging based on iLPs. Conclusion: The NIR fluorescence imaging by iLPs may facilitate improved arthritis diagnosis and early assessment of the disease progression by providing an in vivo characterization of angiogenesis in inflammatory joint diseases. Keywords: rheumatoid arthritis, synovitis, diagnosis, near-infrared fluorescence imaging, iRGD-targeted probes

Intermolecular G-quadruplex structure-based fluorescent DNA detection system.

Science.gov (United States)

Zhou, Hui; Wu, Zai-Sheng; Shen, Guo-Li; Yu, Ru-Qin

2013-03-15

Adopting multi-donors to pair with one acceptor could improve the performance of fluorogenic detection probes. However, common dyes (e.g., fluorescein) in close proximity to each other would self-quench the fluorescence, and the fluorescence is difficult to restore. In this contribution, we constructed a novel "multi-donors-to-one acceptor" fluorescent DNA detection system by means of the intermolecular G-quadruplex (IGQ) structure-based fluorescence signal enhancement combined with the hairpin oligonucleotide. The novel IGQ-hairpin system was characterized using the p53 gene as the model target DNA. The proposed system showed an improved assay performance due to the introduction of IGQ-structure into fluorescent signaling probes, which could inhibit the background fluorescence and increase fluorescence restoration amplitude of fluoresceins upon target DNA hybridization. The proof-of-concept scheme is expected to provide new insight into the potential of G-quadruplex structure and promote the application of fluorescent oligonucleotide probes in fundamental research, diagnosis, and treatment of genetic diseases. Copyright © 2012 Elsevier B.V. All rights reserved.

Fluorescent Nanoprobes Dedicated to in Vivo Imaging: From Preclinical Validations to Clinical Translation

Directory of Open Access Journals (Sweden)

Isabelle Texier

2012-05-01

Full Text Available With the fast development, in the last ten years, of a large choice of set-ups dedicated to routine in vivo measurements in rodents, fluorescence imaging techniques are becoming essential tools in preclinical studies. Human clinical uses for diagnostic and image-guided surgery are also emerging. In comparison to low-molecular weight organic dyes, the use of fluorescent nanoprobes can improve both the signal sensitivity (better in vivo optical properties and the fluorescence biodistribution (passive “nano” uptake in tumours for instance. A wide range of fluorescent nanoprobes have been designed and tested in preclinical studies for the last few years. They will be reviewed and discussed considering the obstacles that need to be overcome for their potential everyday use in clinics. The conjugation of fluorescence imaging with the benefits of nanotechnology should open the way to new medical applications in the near future.

Ultraviolet signals in birds are special.

Science.gov (United States)

Hausmann, Franziska; Arnold, Kathryn E; Marshall, N Justin; Owens, Ian P F

2003-01-07

Recent behavioural experiments have shown that birds use ultraviolet (UV)-reflective and fluorescent plumage as cues in mate choice. It remains controversial, however, whether such UV signals play a special role in sexual communication, or whether they are part of general plumage coloration. We use a comparative approach to test for a general association between sexual signalling and either UV-reflective or fluorescent plumage. Among the species surveyed, 72% have UV colours and there is a significant positive association between UV reflectance and courtship displays. Among parrots (Psittaciformes), 68% of surveyed species have fluorescent plumage, and again there is a strong positive association between courtship displays and fluorescence. These associations are not artefacts of the plumage used in courtship displays, being generally more 'colourful' because there is no association between display and colours lacking UV reflectance or fluorescence. Equally, these associations are not phylogenetic artefacts because all results remain unchanged when families or genera, rather than species, are used as independent data points. We also find that, in parrots, fluorescent plumage is usually found adjacent to UV-reflective plumage. Using a simple visual model to examine one parrot, the budgerigar Melopsittacus undulatus, we show that the juxtaposition of UV-reflective and fluorescent plumage leads to a 25-fold increase in chromatic contrast to the budgerigar's visual system. Taken together, these results suggest that signals based on UV contrast are of special importance in the context of active sexual displays. We review briefly six hypotheses on why this may be the case: suitability for short-range signalling; high contrast with backgrounds; invisibility to predators; exploitation of pre-existing sensory biases; advertisement of feather structure; and amplification of behavioural signals.

Fluorescent Probes and Fluorescence (Microscopy Techniques — Illuminating Biological and Biomedical Research

Directory of Open Access Journals (Sweden)

Gregor P. C. Drummen

2012-11-01

Full Text Available Fluorescence, the absorption and re-emission of photons with longer wavelengths, is one of those amazing phenomena of Nature. Its discovery and utilization had, and still has, a major impact on biological and biomedical research, since it enables researchers not just to visualize normal physiological processes with high temporal and spatial resolution, to detect multiple signals concomitantly, to track single molecules in vivo, to replace radioactive assays when possible, but also to shed light on many pathobiological processes underpinning disease states, which would otherwise not be possible. Compounds that exhibit fluorescence are commonly called fluorochromes or fluorophores and one of these fluorescent molecules in particular has significantly enabled life science research to gain new insights in virtually all its sub-disciplines: Green Fluorescent Protein. Because fluorescent proteins are synthesized in vivo, integration of fluorescent detection methods into the biological system via genetic techniques now became feasible. Currently fluorescent proteins are available that virtually span the whole electromagnetic spectrum. Concomitantly, fluorescence imaging techniques were developed, and often progress in one field fueled innovation in the other. Impressively, the properties of fluorescence were utilized to develop new assays and imaging modalities, ranging from energy transfer to image molecular interactions to imaging beyond the diffraction limit with super-resolution microscopy. Here, an overview is provided of recent developments in both fluorescence imaging and fluorochrome engineering, which together constitute the “fluorescence toolbox” in life science research.

Fluorescence detection of esophageal neoplasia

Science.gov (United States)

Borisova, E.; Vladimirov, B.; Avramov, L.

2008-06-01

White-light endoscopy is well-established and wide used modality. However, despite the many technological advances that have been occurred, conventional endoscopy is suboptimal and usually detects advanced stage lesions. The limitations of standard endoscopy initiate development of spectroscopic techniques, additional to standard endoscopic equipment. One of the most sensitive approaches is fluorescence spectroscopy of gastrointestinal mucosa for neoplasia detection. In the recent study delta-aminolevulinic acid/Protoporphyrin IX (5-ALA/PpIX) is used as fluorescent marker for dysplasia and tumor detection in esophagus. The 5-ALA is administered per os six hours before measurements at dose 20 mg/kg weight. Excitation source has max of emission at 405 nm and light is delivered by the standard light guide of the endoscopic equipment. Through endoscopic instrumental channel a fiber is applied to return information about fluorescence to microspectrometer. Spectral features observed during endoscopic investigations could be distinct as the next regions: 450-630 nm region, where tissue autofluorescence is observed; 630-710 nm region, where fluorescence of PpIX is clearly pronounced; 530-580 nm region, where minima in the autofluorescence signal are observed, related to reabsorption of blood. The lack of fluorescence peaks in the red spectral area for normal mucosa is an indication for selective accumulation of 5-ALA/PpIX only in abnormal sites Very good correlation between fluorescence signals and histology examination of the lesions investigated is achieved.

5-ALA/PpIX fluorescence detection of gastrointestinal neoplasia

Science.gov (United States)

Borisova, Ekaterina G.; Vladimirov, Borislav; Terziev, Ivan; Ivanova, Radina; Avramov, Latchezar

2009-07-01

In the recent study delta-ALA/PpIX is used as fluorescent marker for dysplasia and tumor detection in esophagus, stomach and colon. ALA is administered per os six to eight (depending on the lesion location) hours before measurements at dose 20mg/kg weight. High-power light-emitting diode at 405 nm is used as an excitation source. Special opto-mechanical device is built for the LED to use the light guide of standard video-endoscopic system. Through endoscopic instrumental channel a fiber is applied to return information about fluorescence to microspectrometer. The fluorescence detected from tumor sites has very complex spectral origins. It consists of autofluorescence, fluorescence from exogenous fluorophores and re-absorption from the chromophores accumulated in the tissue investigated. Spectral features observed during endoscopic investigations could be distinct as the next regions: 450-630 nm region, where tissue autofluorescence is observed; 630-710 nm region, where fluorescence of PpIX is clearly pronounced; 530-580 nm region, where minima in the autofluorescence signal are observed, related to re-absorption of oxy-hemoglobin in this spectral area. Endogenous and exogenous fluorescence spectra are used to develop simple but effective algorithm, based on dimensionless ratio of the signals at 560 and 635 nm, for differentiation of normal/abnormal gastrointestinal tissues. Very good correlation between fluorescence signals and histology examination of the lesions investigated is achieved.

Resonant Scanning with Large Field of View Reduces Photobleaching and Enhances Fluorescence Yield in STED Microscopy.

Science.gov (United States)

Wu, Yong; Wu, Xundong; Lu, Rong; Zhang, Jin; Toro, Ligia; Stefani, Enrico

2015-10-01

Photobleaching is a major limitation of superresolution Stimulated Depletion Emission (STED) microscopy. Fast scanning has long been considered an effective means to reduce photobleaching in fluorescence microscopy, but a careful quantitative study of this issue is missing. In this paper, we show that the photobleaching rate in STED microscopy can be slowed down and the fluorescence yield be enhanced by scanning with high speed, enabled by using large field of view in a custom-built resonant-scanning STED microscope. The effect of scanning speed on photobleaching and fluorescence yield is more remarkable at higher levels of depletion laser irradiance, and virtually disappears in conventional confocal microscopy. With ≥6 GW∙cm(-2) depletion irradiance, we were able to extend the fluorophore survival time of Atto 647N and Abberior STAR 635P by ~80% with 8-fold wider field of view. We confirm that STED Photobleaching is primarily caused by the depletion light acting upon the excited fluorophores. Experimental data agree with a theoretical model. Our results encourage further increasing the linear scanning speed for photobleaching reduction in STED microscopy.

Double-gated spectral snapshots for biomolecular fluorescence

International Nuclear Information System (INIS)

Nakamura, Ryosuke; Hamada, Norio; Ichida, Hideki; Tokunaga, Fumio; Kanematsu, Yasuo

2007-01-01

A versatile method to take femtosecond spectral snapshots of fluorescence has been developed based on a double gating technique in the combination of an optical Kerr gate and an image intensifier as an electrically driven gate set in front of a charge-coupled device detector. The application of a conventional optical-Kerr-gate method is limited to molecules with the short fluorescence lifetime up to a few hundred picoseconds, because long-lifetime fluorescence itself behaves as a source of the background signal due to insufficiency of the extinction ratio of polarizers employed for the Kerr gate. By using the image intensifier with the gate time of 200 ps, we have successfully suppressed the background signal and overcome the application limit of optical-Kerr-gate method. The system performance has been demonstrated by measuring time-resolved fluorescence spectra for laser dye solution and the riboflavin solution as a typical sample of biomolecule

Thermal precipitation fluorescence assay for protein stability screening.

Science.gov (United States)

Fan, Junping; Huang, Bo; Wang, Xianping; Zhang, Xuejun C

2011-09-01

A simple and reliable method of protein stability assessment is desirable for high throughput expression screening of recombinant proteins. Here we described an assay termed thermal precipitation fluorescence (TPF) which can be used to compare thermal stabilities of recombinant protein samples directly from cell lysate supernatants. In this assay, target membrane proteins are expressed as recombinant fusions with a green fluorescence protein tag and solubilized with detergent, and the fluorescence signals are used to report the quantity of the fusion proteins in the soluble fraction of the cell lysate. After applying a heat shock, insoluble protein aggregates are removed by centrifugation. Subsequently, the amount of remaining protein in the supernatant is quantified by in-gel fluorescence analysis and compared to samples without a heat shock treatment. Over 60 recombinant membrane proteins from Escherichia coli were subject to this screening in the presence and absence of a few commonly used detergents, and the results were analyzed. Because no sophisticated protein purification is required, this TPF technique is suitable to high throughput expression screening of recombinant membrane proteins as well as soluble ones and can be used to prioritize target proteins based on their thermal stabilities for subsequent large scale expression and structural studies. Copyright © 2011 Elsevier Inc. All rights reserved.

Is the flower fluorescence relevant in biocommunication?

Science.gov (United States)

Iriel, Analía; Lagorio, María Gabriela

2010-10-01

Flower fluorescence has been previously proposed as a potential visual signal to attract pollinators. In this work, this point was addressed by quantitatively measuring the fluorescence quantum yield ( Φ f) for flowers of Bellis perennis (white, yellow, pink, and purple), Ornithogalum thyrsoides (petals and ovaries), Limonium sinuatum (white and yellow), Lampranthus productus (yellow), Petunia nyctaginiflora (white), Bougainvillea spectabilis (white and yellow), Antirrhinum majus (white and yellow), Eustoma grandiflorum (white and blue), Citrus aurantium (petals and stigma), and Portulaca grandiflora (yellow). The highest values were obtained for the ovaries of O. thyrsoides ( Φ f = 0.030) and for Citrus aurantium petals ( Φ f = 0.014) and stigma ( Φ f = 0.013). Emitted photons as fluorescence were compared with reflected photons. It was concluded that the fluorescence emission is negligible compared to the reflected light, even for the most fluorescent samples, and it may not be considered as an optical signal in biocommunication. The work was complemented with the calculation of quantum catches for each studied flower species to describe the visual sensitization of eye photoreceptors.

Microwave dynamic large signal waveform characterization of advanced InGaP HBT for power amplifiers

Energy Technology Data Exchange (ETDEWEB)

Zhao Lixin; Jin Zhi; Liu Xinyu, E-mail: zhaolixin@ime.ac.c [Institute of Microelectronics, Chinese Academy of Sciences, Beijing 100029 (China)

2009-12-15

In wireless mobile communications and wireless local area networks (WLAN), advanced InGaP HBT with power amplifiers are key components. In this paper, the microwave large signal dynamic waveform characteristics of an advanced InGaP HBT are investigated experimentally for 5.8 GHz power amplifier applications. The microwave large signal waveform distortions at various input power levels, especially at large signal level, are investigated and the reasons are analyzed. The output power saturation is also explained. These analyses will be useful for power amplifier designs. (semiconductor devices)

An organic dye with very large Stokes-shift and broad tunability of fluorescence: Potential two-photon probe for bioimaging and ultra-sensitive solid-state gas sensor

Energy Technology Data Exchange (ETDEWEB)

He, Tingchao; Tian, Xiaoqing; Lin, Xiaodong, E-mail: linxd@szu.edu.cn, E-mail: hdsun@ntu.edu.sg [College of Physics Science and Technology, Shenzhen University, Shenzhen 518060 (China); Wang, Yue; Zhao, Xin; Sun, Handong, E-mail: linxd@szu.edu.cn, E-mail: hdsun@ntu.edu.sg [Division of Physics and Applied Physics, and Centre for Disruptive Photonic Technologies (CDPT), School of Physical and Mathematical Sciences, Nanyang Technological University, 21 Nanyang Link, Singapore 637371 (Singapore); Gao, Yang; Grimsdale, Andrew C. [School of Materials Science and Engineering, Nanyang Technological University, Singapore 639798 (Singapore)

2016-01-04

Light-emitting nonlinear optical molecules, especially those with large Stokes shifts and broad tunability of their emission wavelength, have attracted considerable attention for various applications including biomedical imaging and fluorescent sensors. However, most fluorescent chromophores have only limited potential for such applications due to small Stokes shifts, narrow tunability of fluorescence emissions, and small optical nonlinearity in highly polar solvents. In this work, we demonstrate that a two-photon absorbing stilbene chromophore exhibits a large two-photon absorption action cross-section (ηδ = 320 GM) in dimethylsulfoxide (DMSO) and shows broad fluorescence tunability (125 nm) by manipulating the polarity of the surrounding medium. Importantly, a very large Stokes shift of up to 227 nm is achieved in DMSO. Thanks to these features, this chromophore can be utilized as a two-photon probe for bioimaging applications and in an ultrasensitive solid-state gas detector.

Origins of fluorescence in evolved bacteriophytochromes.

Science.gov (United States)

Bhattacharya, Shyamosree; Auldridge, Michele E; Lehtivuori, Heli; Ihalainen, Janne A; Forest, Katrina T

2014-11-14

Use of fluorescent proteins to study in vivo processes in mammals requires near-infrared (NIR) biomarkers that exploit the ability of light in this range to penetrate tissue. Bacteriophytochromes (BphPs) are photoreceptors that couple absorbance of NIR light to photoisomerization, protein conformational changes, and signal transduction. BphPs have been engineered to form NIR fluorophores, including IFP1.4, Wi-Phy, and the iRFP series, initially by replacement of Asp-207 by His. This position was suggestive because its main chain carbonyl is within hydrogen-bonding distance to pyrrole ring nitrogens of the biliverdin chromophore, thus potentially functioning as a crucial transient proton sink during photoconversion. To explain the origin of fluorescence in these phytofluors, we solved the crystal structures of IFP1.4 and a comparison non-fluorescent monomeric phytochrome DrCBDmon. Met-186 and Val-288 in IFP1.4 are responsible for the formation of a tightly packed hydrophobic hub around the biliverdin D ring. Met-186 is also largely responsible for the blue-shifted IFP1.4 excitation maximum relative to the parent BphP. The structure of IFP1.4 revealed decreased structural heterogeneity and a contraction of two surface regions as direct consequences of side chain substitutions. Unexpectedly, IFP1.4 with Asp-207 reinstalled (IFPrev) has a higher fluorescence quantum yield (∼9%) than most NIR phytofluors published to date. In agreement, fluorescence lifetime measurements confirm the exceptionally long excited state lifetimes, up to 815 ps, in IFP1.4 and IFPrev. Our research helps delineate the origin of fluorescence in engineered BphPs and will facilitate the wide-spread adoption of phytofluors as biomarkers. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Rectangular coordination polymer nanoplates: large-scale, rapid synthesis and their application as a fluorescent sensing platform for DNA detection.

Directory of Open Access Journals (Sweden)

Yingwei Zhang

Full Text Available In this paper, we report on the large-scale, rapid synthesis of uniform rectangular coordination polymer nanoplates (RCPNs assembled from Cu(II and 4,4'-bipyridine for the first time. We further demonstrate that such RCPNs can be used as a very effective fluorescent sensing platform for multiple DNA detection with a detection limit as low as 30 pM and a high selectivity down to single-base mismatch. The DNA detection is accomplished by the following two steps: (1 RCPN binds dye-labeled single-stranded DNA (ssDNA probe, which brings dye and RCPN into close proximity, leading to fluorescence quenching; (2 Specific hybridization of the probe with its target generates a double-stranded DNA (dsDNA which detaches from RCPN, leading to fluorescence recovery. It suggests that this sensing system can well discriminate complementary and mismatched DNA sequences. The exact mechanism of fluorescence quenching involved is elucidated experimentally and its use in a human blood serum system is also demonstrated successfully.

Remote sensing vegetation status by laser-induced fluorescence

International Nuclear Information System (INIS)

Günther, K.P.; Dahn, H.G.; Lüdeker, W.

1994-01-01

In November 1989 the EUREKA project LASFLEUR (EU 380) started as an European research effort to investigate the future application of far-field laser-induced plant fluorescence for synoptic, airborne environmental monitoring of vegetation. This report includes a brief introduction in a theoretically approach for the laser-induced fluorescence signals of leaves and their spectral and radiometric behaviour. In addition, a detailed description of the design and realization of the second generation of the far-field fluorescence lidar (DLidaR-2) is given with special regard to the optical and electronical setup, followed by a short explanation of the data processing. The main objectives of the far field measurements are to demonstrate the link between laser-induced fluorescence data and plant physiology and to show the reliability of remote single shot lidar measurements. The data sets include the typical daily cycles of the fluorescence for different global irradiation. As expected from biophysical models, the remotely sensed chlorophyll fluorescence is highly correlated with the carbon fixation rate, while the fluorescence ratio F685 / F730 is only dependent on the chlorophyll concentration. Drought stress measurement of evergreen oaks Quercus pubescens confirm the findings of healthy plants with regard to the fluorescence ratio F685 / F730 while the fluorescence signals of stressed plants show a different behavior than nonstressed plants. Additionally, the corresponding physiological data (porometer and PAM data) are presented. (author)

Atomic-fluorescence spectrophotometry

International Nuclear Information System (INIS)

Bakhturova, N.F.; Yudelevich, I.G.

1975-01-01

Atomic-fluorescence spectrophotometry, a comparatively new method for the analysis of trace quantities, has developed rapidly in the past ten years. Theoretical and experimental studies by many workers have shown that atomic-fluorescence spectrophotometry (AFS) is capable of achieving a better limit than atomic absorption for a large number of elements. The present review examines briefly the principles of atomic-fluorescence spectrophotometry and the types of fluorescent transition. The excitation sources, flame and nonflame atomizers, used in AFS are described. The limits of detection achieved up to the present, using flame and nonflame methods of atomization are given

Probe for intracellular concentrations of drugs: delayed fluorescence from acridine orange

International Nuclear Information System (INIS)

Wardman, P.; Dennis, M.F.; White, J.

1989-01-01

The aim of this work is to develop fluorescent probes that will indicate effective concentrations of therapeutic agents, or endogenous protectors, at important cellular sites. Acridine orange associates with nucleic acids and emits a 'delayed' fluorescence signal. This signal is quenched by oxidants such as oxygen, nitroaryl radiosensitizers, adriamycin and mitomycin-c, and reductants such as thiols, ascorbate and other radioprotectors. The quenching of the acridine orange delayed fluorescence reflects the effective concentration of these therapeutically-important oxidants and reductants near DNA. The relative concentration of basic radiosensitizers such as pimonidazole (Ro 03-8799) near the DNA is greater than that of misonidazole. Thiols quench the delayed fluorescence signal according to the degree of ionization of the thiol function; this may model the reactivity of thiols with guanine radical sites in DNA. Ascorbate and aminopyrine do not quench the delayed fluorescence from cells stained with acridine orange as these compounds are taken up by cells very inefficiently

Detection of Counterfeit Tequila by Fluorescence Spectroscopy

Directory of Open Access Journals (Sweden)

José Manuel de la Rosa Vázquez

2015-01-01

Full Text Available An ultraviolet (UV light induced fluorescence study to discriminate fake tequila from genuine ones is presented. A portable homemade system based on four light emitting diodes (LEDs from 255 to 405 nm and a miniature spectrometer was used. It has been shown that unlike fake and silver tequila, which produce weak fluorescence signal, genuine mixed, rested, and aged tequilas show high fluorescence emission in the range from 400 to 750 nm. The fluorescence intensity grows with aging in 100% agave tequila. Such fluorescence differences can even be observed with naked eyes. The presented results demonstrate that the fluorescence measurement could be a good method to detect counterfeit tequila.

Saturated excitation of Fluorescence to quantify excitation enhancement in aperture antennas

KAUST Repository

Aouani, Heykel

2012-07-23

Fluorescence spectroscopy is widely used to probe the electromagnetic intensity amplification on optical antennas, yet measuring the excitation intensity amplification is a challenge, as the detected fluorescence signal is an intricate combination of excitation and emission. Here, we describe a novel approach to quantify the electromagnetic amplification in aperture antennas by taking advantage of the intrinsic non linear properties of the fluorescence process. Experimental measurements of the fundamental f and second harmonic 2f amplitudes of the fluorescence signal upon excitation modulation are used to quantify the electromagnetic intensity amplification with plasmonic aperture antennas. © 2012 Optical Society of America.

Saturated excitation of Fluorescence to quantify excitation enhancement in aperture antennas

KAUST Repository

Aouani, Heykel; Hostein, Richard; Mahboub, Oussama; Devaux, Eloï se; Rigneault, Hervé ; Ebbesen, Thomas W.; Wenger, Jé rô me

2012-01-01

Fluorescence spectroscopy is widely used to probe the electromagnetic intensity amplification on optical antennas, yet measuring the excitation intensity amplification is a challenge, as the detected fluorescence signal is an intricate combination of excitation and emission. Here, we describe a novel approach to quantify the electromagnetic amplification in aperture antennas by taking advantage of the intrinsic non linear properties of the fluorescence process. Experimental measurements of the fundamental f and second harmonic 2f amplitudes of the fluorescence signal upon excitation modulation are used to quantify the electromagnetic intensity amplification with plasmonic aperture antennas. © 2012 Optical Society of America.

Analysis of hyperspectral fluorescence images for poultry skin tumor inspection

Science.gov (United States)

Kong, Seong G.; Chen, Yud-Ren; Kim, Intaek; Kim, Moon S.

2004-02-01

We present a hyperspectral fluorescence imaging system with a fuzzy inference scheme for detecting skin tumors on poultry carcasses. Hyperspectral images reveal spatial and spectral information useful for finding pathological lesions or contaminants on agricultural products. Skin tumors are not obvious because the visual signature appears as a shape distortion rather than a discoloration. Fluorescence imaging allows the visualization of poultry skin tumors more easily than reflectance. The hyperspectral image samples obtained for this poultry tumor inspection contain 65 spectral bands of fluorescence in the visible region of the spectrum at wavelengths ranging from 425 to 711 nm. The large amount of hyperspectral image data is compressed by use of a discrete wavelet transform in the spatial domain. Principal-component analysis provides an effective compressed representation of the spectral signal of each pixel in the spectral domain. A small number of significant features are extracted from two major spectral peaks of relative fluorescence intensity that have been identified as meaningful spectral bands for detecting tumors. A fuzzy inference scheme that uses a small number of fuzzy rules and Gaussian membership functions successfully detects skin tumors on poultry carcasses. Spatial-filtering techniques are used to significantly reduce false positives.

Fluorescence ELISA for sensitive detection of ochratoxin A based on glucose oxidase-mediated fluorescence quenching of CdTe QDs

International Nuclear Information System (INIS)

Liang, Yi; Huang, Xiaolin; Yu, Ruijin; Zhou, Yaofeng; Xiong, Yonghua

2016-01-01

The present study described a novel fluorescence enzyme-linked immunosorbent assay (ELISA) used to detect ochratoxin A (OTA) by using the glucose oxidase (GOx)-mediated fluorescence quenching of mercaptopropionic acid-capped CdTe quantum dots (MPA-QDs), in which GOx was used as an alternative to horseradish peroxidase (HRP) for the oxidization of glucose into hydrogen peroxide (H_2O_2) and gluconic acid. The MPA-QDs were used as a fluorescent signal output, whose fluorescence variation was extremely sensitive to the presence of H_2O_2 or hydrogen ions in the solution. Under the optimized conditions, the proposed fluorescence ELISA demonstrated a good linear detection of OTA in corn extract from 2.4 pg mL"−"1 to 625 pg mL"−"1 with a limit of detection of 2.2 pg mL"−"1, which was approximately 15-fold lower than that of conventional HRP-based ELISA. Our developed fluorescence immunoassay was also similar to HRP-based ELISA in terms of selectivity, accuracy, and reproducibility. In summary, this study was the first to use the GOx-mediated fluorescence quenching of QDs in immunoassay to detect OTA, offering a new possibility for the analysis of other mycotoxins and biomolecules. - Highlights: • A novel fluorescence ELISA was first developed for the detection of OTA by using GOx-mediated fluorescence quenching of QDs. • The pH- and H_2O_2-sensitive MPA-capped CdTe QDs were used as a fluorescent signal output to improve the detection sensitivity. • This novel method open up a different vision to detect other mycotoxins and biomolecules.

Small- and large-signal modeling of InP HBTs in transferred-substrate technology

DEFF Research Database (Denmark)

Johansen, Tom Keinicke; Rudolph, Matthias; Jensen, Thomas

2014-01-01

In this paper, the small- and large-signal modeling of InP heterojunction bipolar transistors (HBTs) in transferred substrate (TS) technology is investigated. The small-signal equivalent circuit parameters for TS-HBTs in two-terminal and three-terminal configurations are determined by employing...

Measuring fluorescence polarization with a dichrometer.

Science.gov (United States)

Sutherland, John C

2017-09-01

A method for obtaining fluorescence polarization data from an instrument designed to measure circular and linear dichroism is compared with a previously reported approach. The new method places a polarizer between the sample and a detector mounted perpendicular to the direction of the incident beam and results in determination of the fluorescence polarization ratio, whereas the previous method does not use a polarizer and yields the fluorescence anisotropy. A similar analysis with the detector located axially with the excitation beam demonstrates that there is no frequency modulated signal due to fluorescence polarization in the absence of a polarizer. Copyright © 2017. Published by Elsevier Inc.

Developing Fast Fluorescent Protein Voltage Sensors by Optimizing FRET Interactions.

Directory of Open Access Journals (Sweden)

Uhna Sung

Full Text Available FRET (Förster Resonance Energy Transfer-based protein voltage sensors can be useful for monitoring neuronal activity in vivo because the ratio of signals between the donor and acceptor pair reduces common sources of noise such as heart beat artifacts. We improved the performance of FRET based genetically encoded Fluorescent Protein (FP voltage sensors by optimizing the location of donor and acceptor FPs flanking the voltage sensitive domain of the Ciona intestinalis voltage sensitive phosphatase. First, we created 39 different "Nabi1" constructs by positioning the donor FP, UKG, at 8 different locations downstream of the voltage-sensing domain and the acceptor FP, mKO, at 6 positions upstream. Several of these combinations resulted in large voltage dependent signals and relatively fast kinetics. Nabi1 probes responded with signal size up to 11% ΔF/F for a 100 mV depolarization and fast response time constants both for signal activation (~2 ms and signal decay (~3 ms. We improved expression in neuronal cells by replacing the mKO and UKG FRET pair with Clover (donor FP and mRuby2 (acceptor FP to create Nabi2 probes. Nabi2 probes also had large signals and relatively fast time constants in HEK293 cells. In primary neuronal culture, a Nabi2 probe was able to differentiate individual action potentials at 45 Hz.

Azadioxatriangulenium: a long fluorescence lifetime fluorophore for large biomolecule binding assay

International Nuclear Information System (INIS)

Sørensen, Thomas Just; Thyrhaug, Erling; Szabelski, Mariusz; Gryczynski, Ignacy; Gryczynski, Zygmunt; Luchowski, Rafal; Laursen, Bo W

2013-01-01

Of the many optical bioassays available, sensing by fluorescence anisotropy has great advantages as it provides a sensitive, instrumentally simple, ratiometric method of detection. However, it is hampered by a severe limitation, as the emission lifetime of the label needs to be comparable to the correlation lifetime (tumbling time) of the biomolecule which is labelled. For proteins of moderate size this is on the order of 20–200 ns, which due to practical issues currently limits the choice of labels to the dansyl-type dyes and certain aromatic dyes. These have the significant drawback of UV/blue absorption and emission as well as an often significant solvent sensitivity. Here, we report the synthesis and characterization of a new fluorescent label for high molecular weight biomolecule assay based on the azadioxatriangulenium motif. The NHS ester of the long fluorescence lifetime, red-emitting fluorophore: azadioxatriangulenium (ADOTA-NHS) was conjugated to anti-rabbit Immunoglobulin G (antiIgG). The long fluorescence lifetime was exploited to determine the correlation time of the high molecular weight antibody and its complex with rabbit Immunoglobulin G (IgG) with steady-state fluorescence anisotropy and time-resolved methods: solution phase immuno-assay was performed following either steady-state or time-resolved fluorescence anisotropy. By performing a variable temperature experiment it was determined that the binding of the ligand resulted in an increase in correlation time of more than 75%, and an increase in the steady-state anisotropy of 18%. The results show that the triangulenium class of dyes can be used in anisotropy assay to detect binding events involving biomolecules of far larger size than what is possible with most other red-emitting organic dyes. (paper)

Azadioxatriangulenium: a long fluorescence lifetime fluorophore for large biomolecule binding assay

Science.gov (United States)

Just Sørensen, Thomas; Thyrhaug, Erling; Szabelski, Mariusz; Luchowski, Rafal; Gryczynski, Ignacy; Gryczynski, Zygmunt; Laursen, Bo W.

2013-06-01

Of the many optical bioassays available, sensing by fluorescence anisotropy has great advantages as it provides a sensitive, instrumentally simple, ratiometric method of detection. However, it is hampered by a severe limitation, as the emission lifetime of the label needs to be comparable to the correlation lifetime (tumbling time) of the biomolecule which is labelled. For proteins of moderate size this is on the order of 20-200 ns, which due to practical issues currently limits the choice of labels to the dansyl-type dyes and certain aromatic dyes. These have the significant drawback of UV/blue absorption and emission as well as an often significant solvent sensitivity. Here, we report the synthesis and characterization of a new fluorescent label for high molecular weight biomolecule assay based on the azadioxatriangulenium motif. The NHS ester of the long fluorescence lifetime, red-emitting fluorophore: azadioxatriangulenium (ADOTA-NHS) was conjugated to anti-rabbit Immunoglobulin G (antiIgG). The long fluorescence lifetime was exploited to determine the correlation time of the high molecular weight antibody and its complex with rabbit Immunoglobulin G (IgG) with steady-state fluorescence anisotropy and time-resolved methods: solution phase immuno-assay was performed following either steady-state or time-resolved fluorescence anisotropy. By performing a variable temperature experiment it was determined that the binding of the ligand resulted in an increase in correlation time of more than 75%, and an increase in the steady-state anisotropy of 18%. The results show that the triangulenium class of dyes can be used in anisotropy assay to detect binding events involving biomolecules of far larger size than what is possible with most other red-emitting organic dyes.

Confocal fluorescence techniques in industrial application

Science.gov (United States)

Eggeling, Christian; Gall, Karsten; Palo, Kaupo; Kask, Peet; Brand, Leif

2003-06-01

The FCS+plus family of evaluation tools for confocal fluorescence spectroscopy, which was developed during recent years, offers a comprehensive view to a series of fluorescence properties. Originating in fluorescence correlation spectroscopy (FCS) and using similar experimental equipment, a system of signal processing methods such as fluorescence intensity distribution analysis (FIDA) was created to analyze in detail the fluctuation behavior of fluorescent particles within a small area of detection. Giving simultaneous access to molecular parameters like concentration, translational and rotational diffusion, molecular brightness, and multicolor coincidence, this portfolio was enhanced by more traditional techniques of fluorescence lifetime as well as time-resolved anisotropy determination. The cornerstones of the FCS+plus methodology will be shortly described. The inhibition of a phosphatase enzyme activity gives a comprehensive industrial application that demonstrates FCS+plus' versatility and its potential for pharmaceutical drug discovery.

Measurement of MOSFET LF Noise Under Large Signal RF Excitation

NARCIS (Netherlands)

van der Wel, A.P.; Klumperink, Eric A.M.; Nauta, Bram

A new measurement technique is presented that allows measurement of MOSFET LF noise under large signal RF excitation. Measurements indicate that MOSFETS exhibit a reduction in LF noise when they are cycled from inversion to accummulation and that this reduction does not depend on the frequency of

Large signal S-parameters: modeling and radiation effects in microwave power transistors

International Nuclear Information System (INIS)

Graham, E.D. Jr.; Chaffin, R.J.; Gwyn, C.W.

1973-01-01

Microwave power transistors are usually characterized by measuring the source and load impedances, efficiency, and power output at a specified frequency and bias condition in a tuned circuit. These measurements provide limited data for circuit design and yield essentially no information concerning broadbanding possibilities. Recently, a method using large signal S-parameters has been developed which provides a rapid and repeatable means for measuring microwave power transistor parameters. These large signal S-parameters have been successfully used to design rf power amplifiers. Attempts at modeling rf power transistors have in the past been restricted to a modified Ebers-Moll procedure with numerous adjustable model parameters. The modified Ebers-Moll model is further complicated by inclusion of package parasitics. In the present paper an exact one-dimensional device analysis code has been used to model the performance of the transistor chip. This code has been integrated into the SCEPTRE circuit analysis code such that chip, package and circuit performance can be coupled together in the analysis. Using []his computational tool, rf transistor performance has been examined with particular attention given to the theoretical validity of large-signal S-parameters and the effects of nuclear radiation on device parameters. (auth)

Some secrets of fluorescent proteins: distinct bleaching in various mounting fluids and photoactivation of cyan fluorescent proteins at YFP-excitation.

Science.gov (United States)

Malkani, Naila; Schmid, Johannes A

2011-04-07

The use of spectrally distinct variants of green fluorescent protein (GFP) such as cyan or yellow mutants (CFP and YFP, respectively) is very common in all different fields of life sciences, e.g. for marking specific proteins or cells or to determine protein interactions. In the latter case, the quantum physical phenomenon of fluorescence resonance energy transfer (FRET) is exploited by specific microscopy techniques to visualize proximity of proteins. When we applied a commonly used FRET microscopy technique--the increase in donor (CFP)-fluorescence after bleaching of acceptor fluorophores (YFP), we obtained good signals in live cells, but very weak signals for the same samples after fixation and mounting in commercial microscopy mounting fluids. This observation could be traced back to much faster bleaching of CFP in these mounting media. Strikingly, the opposite effect of the mounting fluid was observed for YFP and also for other proteins such as Cerulean, TFP or Venus. The changes in photostability of CFP and YFP were not caused by the fixation but directly dependent on the mounting fluid. Furthermore we made the interesting observation that the CFP-fluorescence intensity increases by about 10-15% after illumination at the YFP-excitation wavelength--a phenomenon, which was also observed for Cerulean. This photoactivation of cyan fluorescent proteins at the YFP-excitation can cause false-positive signals in the FRET-microscopy technique that is based on bleaching of a yellow FRET acceptor. Our results show that photostability of fluorescent proteins differs significantly for various media and that CFP bleaches significantly faster in commercial mounting fluids, while the opposite is observed for YFP and some other proteins. Moreover, we show that the FRET microscopy technique that is based on bleaching of the YFP is prone to artifacts due to photoactivation of cyan fluorescent proteins under these conditions.

Fluorescence lifetime assays: current advances and applications in drug discovery.

Science.gov (United States)

Pritz, Stephan; Doering, Klaus; Woelcke, Julian; Hassiepen, Ulrich

2011-06-01

Fluorescence lifetime assays complement the portfolio of established assay formats available in drug discovery, particularly with the recent advances in microplate readers and the commercial availability of novel fluorescent labels. Fluorescence lifetime assists in lowering complexity of compound screening assays, affording a modular, toolbox-like approach to assay development and yielding robust homogeneous assays. To date, materials and procedures have been reported for biochemical assays on proteases, as well as on protein kinases and phosphatases. This article gives an overview of two assay families, distinguished by the origin of the fluorescence signal modulation. The pharmaceutical industry demands techniques with a robust, integrated compound profiling process and short turnaround times. Fluorescence lifetime assays have already helped the drug discovery field, in this sense, by enhancing productivity during the hit-to-lead and lead optimization phases. Future work will focus on covering other biochemical molecular modifications by investigating the detailed photo-physical mechanisms underlying the fluorescence signal.

Quenching-free fluorescence signal from plastic-fibres in proton dosimetry: understanding the influence of Čerenkov radiation

DEFF Research Database (Denmark)

Christensen, Jeppe Brage; Almhagen, Erik; Nyström, Håkan

2018-01-01

The origin of photons emitted in optical fibres under proton irradiation has been attributed to either entirely Čerenkov radiation or light consisting of fluorescence with a substantial amount of Čerenkov radiation. The source of the light emission is assessed in order to understand why the signal...... from optical fibres irradiated with protons is reportedly quenching-free. The present study uses the directional emittance of Čerenkov photons in 12 MeV and 20 MeV electron beams to validate a Monte Carlo model for simulating the emittance and transmission of Čerenkov radiation in optical fibres. We...

FLEX: an imaging spectrometer for measurement of vegetation fluorescence

Science.gov (United States)

Smorenburg, Kees; Visser, Huib; Court, Andrew; Stoll, Marc Ph.

2017-11-01

Detection of vegetation fluorescence gives information about plant functioning, stress and vitality. During the past decades several ground based laser fluorosensors have been developed to investigate these processes and to demonstrate the value of this technique. FLEX (= FLuorescense EXplorer) is a space mission to measure the fluorescence of vegetation on earth over large areas from space. Such a mission would greatly improve the understanding and enhance the capability to quantify e.g. the role of terrestrial vegetation in global carbon sequestration. Because the fluorescence signal, which is excited by solar irradiation is low with respect to the reflected sunlight the signal from a satellite is proposed to be measured in the solar Fraunhofer lines, where the reflection signal is much reduced. The heart of FLEX is a high resolution imaging spectrometer with 2 channels: channel 1 around the Fraunhofer lines at ‡ = 397 nm, ‡= 423 nm and/or ‡ = 434 nm and channel 2 around the Fraunhofer line at ‡ = 656 nm. The required spectral resolution will depend on the linewidth (0.02-0.3 nm). A first definition of the field of view is 8.4 degrees, leading from an 800 km satellite altitude to a swath of about 120 km. For detection a 1024x1024 pixel frame transfer CCD detector is proposed, with a pixel dimension of 13 x 13 ‡ mm2. The maximum footprint is about 500x500m2. The optical configuration contains a scan mirror for solar calibration, for pointing the FOV in swath direction and for freezing the observed ground scene up to a few seconds to increase the signal to noise performance. At this moment the concept of FLEX is elaborated in a feasibility study. Both the scientific and instrument requirements are updated and the concept is studied in detail. Besides a development plan for FLEX is made. In this paper the idea and the headlines of FLEX are described.

Si/SiC-based DD hetero-structure IMPATTs as MM-wave power-source: a generalized large-signal analysis

International Nuclear Information System (INIS)

Mukherjee, Moumita; Tripathy, P. R.; Pati, S. P.

2015-01-01

A full-scale, self-consistent, non-linear, large-signal model of double-drift hetero-structure IMPATT diode with general doping profile is derived. This newly developed model, for the first time, has been used to analyze the large-signal characteristics of hexagonal SiC-based double-drift IMPATT diode. Considering the fabrication feasibility, the authors have studied the large-signal characteristics of Si/SiC-based hetero-structure devices. Under small-voltage modulation (∼ 2%, i.e. small-signal conditions) results are in good agreement with calculations done using a linearised small-signal model. The large-signal values of the diode's negative conductance (5 × 10 6 S/m 2 ), susceptance (10.4 × 10 7 S/m 2 ), average breakdown voltage (207.6 V), and power generating efficiency (15%, RF power: 25.0 W at 94 GHz) are obtained as a function of oscillation amplitude (50% of DC breakdown voltage) for a fixed average current density. The large-signal calculations exhibit power and efficiency saturation for large-signal (> 50%) voltage modulation and thereafter decrease gradually with further increasing voltage-modulation. This generalized large-signal formulation is applicable for all types of IMPATT structures with distributed and narrow avalanche zones. The simulator is made more realistic by incorporating the space-charge effects, realistic field and temperature dependent material parameters in Si and SiC. The electric field snap-shots and the large-signal impedance and admittance of the diode with current excitation are expressed in closed loop form. This study will act as a guide for researchers to fabricate a high-power Si/SiC-based IMPATT for possible application in high-power MM-wave communication systems. (paper)

Phospholyl(borane) Amino Acids and Peptides: Stereoselective Synthesis and Fluorescent Properties with Large Stokes Shift.

Science.gov (United States)

Arribat, Mathieu; Rémond, Emmanuelle; Clément, Sébastien; Lee, Arie Van Der; Cavelier, Florine

2018-01-24

The synthesis of phospholyl(borane) amino acids was stereoselectively achieved by reaction of phospholide anion with iodo α-amino ester derived from l-aspartic acid or l-serine, followed by in situ complexation with borane. Phospholyl(borane) amino acids are easy to store and can be subjected to direct transformation into the corresponding free phospholyl, gold complex, oxide or sulfur derivatives as well as phospholinium salts, thus offering a variety of side chains. After selective deprotection of carboxylic function or amine, C- or N- peptide coupling with an alanine moiety proved the possible incorporation into peptides. Such phospholyl amino acid and peptide derivatives exhibit fluorescent properties with a large Stokes shift (160 nm) and fluorescence up to 535 nm, depending on the phosphole aromaticity and the chemical environment. These phospholyl(borane) amino acids constitute a new class of unnatural amino acids useful for structure-activities relationship studies and appear to be promising fluorophores for the development of labeled peptides.

Imaging a Large Sample with Selective Plane Illumination Microscopy Based on Multiple Fluorescent Microsphere Tracking

Science.gov (United States)

Ryu, Inkeon; Kim, Daekeun

2018-04-01

A typical selective plane illumination microscopy (SPIM) image size is basically limited by the field of view, which is a characteristic of the objective lens. If an image larger than the imaging area of the sample is to be obtained, image stitching, which combines step-scanned images into a single panoramic image, is required. However, accurately registering the step-scanned images is very difficult because the SPIM system uses a customized sample mount where uncertainties for the translational and the rotational motions exist. In this paper, an image registration technique based on multiple fluorescent microsphere tracking is proposed in the view of quantifying the constellations and measuring the distances between at least two fluorescent microspheres embedded in the sample. Image stitching results are demonstrated for optically cleared large tissue with various staining methods. Compensation for the effect of the sample rotation that occurs during the translational motion in the sample mount is also discussed.

Multi-spectral endogenous fluorescence imaging for bacterial differentiation

Science.gov (United States)

Chernomyrdin, Nikita V.; Babayants, Margarita V.; Korotkov, Oleg V.; Kudrin, Konstantin G.; Rimskaya, Elena N.; Shikunova, Irina A.; Kurlov, Vladimir N.; Cherkasova, Olga P.; Komandin, Gennady A.; Reshetov, Igor V.; Zaytsev, Kirill I.

2017-07-01

In this paper, the multi-spectral endogenous fluorescence imaging was implemented for bacterial differentiation. The fluorescence imaging was performed using a digital camera equipped with a set of visual bandpass filters. Narrowband 365 nm ultraviolet radiation passed through a beam homogenizer was used to excite the sample fluorescence. In order to increase a signal-to-noise ratio and suppress a non-fluorescence background in images, the intensity of the UV excitation was modulated using a mechanical chopper. The principal components were introduced for differentiating the samples of bacteria based on the multi-spectral endogenous fluorescence images.

Highly Selective Fluorescent Sensing of Proteins Based on a Fluorescent Molecularly Imprinted Nanosensor

Directory of Open Access Journals (Sweden)

Shuo Wang

2013-09-01

Full Text Available A fluorescent molecularly imprinted nanosensor was obtained by grafting imprinted polymer onto the surface of multi-wall carbon nanotubes and post-imprinting treatment with fluorescein isothiocyanate (FITC. The fluorescence of lysozyme-imprinted polymer (Lys-MIP was quenched more strongly by Lys than that of nonimprinted polymer (NIP, which indicated that the Lys-MIP could recognize Lys. The resulted imprinted material has the ability to selectively sense a target protein, and an imprinting factor of 3.34 was achieved. The Lys-MIP also showed selective detection for Lys among other proteins such as cytochrome C (Cyt C, hemoglobin (HB and bovine serum albumin (BSA due to the imprinted sites in the Lys-MIP. This approach combines the high selectivity of surface molecular imprinting technology and fluorescence, and converts binding events into detectable signals by monitoring fluorescence spectra. Therefore, it will have further applications for Lys sensing.

Automatic enhancement of skin fluorescence localization due to refractive index matching

Science.gov (United States)

Churmakov, Dmitry Y.; Meglinski, Igor V.; Piletsky, Sergey A.; Greenhalgh, Douglas A.

2004-07-01

Fluorescence diagnostic techniques are notable amongst many other optical methods, as they offer high sensitivity and non-invasive measurements of tissue properties. However, a combination of multiple scattering and physical heterogeneity of biological tissues hampers the interpretation of the fluorescence measurements. The analyses of the spatial distribution of endogenous and exogenous fluorophores excitations within tissues and their contribution to the detected signal localization are essential for many applications. We have developed a novel Monte Carlo technique that gives a graphical perception of how the excitation and fluorescence detected signal are localized in tissues. Our model takes into account spatial distribution of fluorophores and their quantum yields. We demonstrate that matching of the refractive indices of ambient medium and topical skin layer improves spatial localization of the detected fluorescence signal within the tissue. This result is consistent with the recent conclusion that administering biocompatible agents results in higher image contrast.

Fluorescence ELISA for sensitive detection of ochratoxin A based on glucose oxidase-mediated fluorescence quenching of CdTe QDs

Energy Technology Data Exchange (ETDEWEB)

Liang, Yi [State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047 (China); Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047 (China); Huang, Xiaolin [State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047 (China); Yu, Ruijin [College of Science, Northwest A& F University, Yangling, Shaanxi 712100 (China); Zhou, Yaofeng [State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047 (China); Xiong, Yonghua, E-mail: yhxiongchen@163.com [State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047 (China); Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047 (China)

2016-09-14

The present study described a novel fluorescence enzyme-linked immunosorbent assay (ELISA) used to detect ochratoxin A (OTA) by using the glucose oxidase (GOx)-mediated fluorescence quenching of mercaptopropionic acid-capped CdTe quantum dots (MPA-QDs), in which GOx was used as an alternative to horseradish peroxidase (HRP) for the oxidization of glucose into hydrogen peroxide (H{sub 2}O{sub 2}) and gluconic acid. The MPA-QDs were used as a fluorescent signal output, whose fluorescence variation was extremely sensitive to the presence of H{sub 2}O{sub 2} or hydrogen ions in the solution. Under the optimized conditions, the proposed fluorescence ELISA demonstrated a good linear detection of OTA in corn extract from 2.4 pg mL{sup −1} to 625 pg mL{sup −1} with a limit of detection of 2.2 pg mL{sup −1}, which was approximately 15-fold lower than that of conventional HRP-based ELISA. Our developed fluorescence immunoassay was also similar to HRP-based ELISA in terms of selectivity, accuracy, and reproducibility. In summary, this study was the first to use the GOx-mediated fluorescence quenching of QDs in immunoassay to detect OTA, offering a new possibility for the analysis of other mycotoxins and biomolecules. - Highlights: • A novel fluorescence ELISA was first developed for the detection of OTA by using GOx-mediated fluorescence quenching of QDs. • The pH- and H{sub 2}O{sub 2}-sensitive MPA-capped CdTe QDs were used as a fluorescent signal output to improve the detection sensitivity. • This novel method open up a different vision to detect other mycotoxins and biomolecules.

Catheter-based time-gated near-infrared fluorescence/OCT imaging system

Science.gov (United States)

Lu, Yuankang; Abran, Maxime; Cloutier, Guy; Lesage, Frédéric

2018-02-01

We developed a new dual-modality intravascular imaging system based on fast time-gated fluorescence intensity imaging and spectral domain optical coherence tomography (SD-OCT) for the purpose of interventional detection of atherosclerosis. A pulsed supercontinuum laser was used for fluorescence and OCT imaging. A double-clad fiber (DCF)- based side-firing catheter was designed and fabricated to have a 23 μm spot size at a 2.2 mm working distance for OCT imaging. Its single-mode core is used for OCT, while its inner cladding transports fluorescence excitation light and collects fluorescent photons. The combination of OCT and fluorescence imaging was achieved by using a DCF coupler. For fluorescence detection, we used a time-gated technique with a novel single-photon avalanche diode (SPAD) working in an ultra-fast gating mode. A custom-made delay chip was integrated in the system to adjust the delay between the excitation laser pulse and the SPAD gate-ON window. This technique allowed to detect fluorescent photons of interest while rejecting most of the background photons, thus leading to a significantly improved signal to noise ratio (SNR). Experiments were carried out in turbid media mimicking tissue with an indocyanine green (ICG) inclusion (1 mM and 100 μM) to compare the time-gated technique and the conventional continuous detection technique. The gating technique increased twofold depth sensitivity, and tenfold SNR at large distances. The dual-modality imaging capacity of our system was also validated with a silicone-based tissue-mimicking phantom.

Large-Signal Lyapunov-Based Stability Analysis of DC/AC Inverters and Inverter-Based Microgrids

Science.gov (United States)

Kabalan, Mahmoud

Microgrid stability studies have been largely based on small-signal linearization techniques. However, the validity and magnitude of the linearization domain is limited to small perturbations. Thus, there is a need to examine microgrids with large-signal nonlinear techniques to fully understand and examine their stability. Large-signal stability analysis can be accomplished by Lyapunov-based mathematical methods. These Lyapunov methods estimate the domain of asymptotic stability of the studied system. A survey of Lyapunov-based large-signal stability studies showed that few large-signal studies have been completed on either individual systems (dc/ac inverters, dc/dc rectifiers, etc.) or microgrids. The research presented in this thesis addresses the large-signal stability of droop-controlled dc/ac inverters and inverter-based microgrids. Dc/ac power electronic inverters allow microgrids to be technically feasible. Thus, as a prelude to examining the stability of microgrids, the research presented in Chapter 3 analyzes the stability of inverters. First, the 13 th order large-signal nonlinear model of a droop-controlled dc/ac inverter connected to an infinite bus is presented. The singular perturbation method is used to decompose the nonlinear model into 11th, 9th, 7th, 5th, 3rd and 1st order models. Each model ignores certain control or structural components of the full order model. The aim of the study is to understand the accuracy and validity of the reduced order models in replicating the performance of the full order nonlinear model. The performance of each model is studied in three different areas: time domain simulations, Lyapunov's indirect method and domain of attraction estimation. The work aims to present the best model to use in each of the three domains of study. Results show that certain reduced order models are capable of accurately reproducing the performance of the full order model while others can be used to gain insights into those three areas of

Time reversal optical tomography locates fluorescent targets in a turbid medium

Science.gov (United States)

Wu, Binlin; Cai, W.; Gayen, S. K.

2013-03-01

A fluorescence optical tomography approach that extends time reversal optical tomography (TROT) to locate fluorescent targets embedded in a turbid medium is introduced. It uses a multi-source illumination and multi-detector signal acquisition scheme, along with TR matrix formalism, and multiple signal classification (MUSIC) to construct pseudo-image of the targets. The samples consisted of a single or two small tubes filled with water solution of Indocyanine Green (ICG) dye as targets embedded in a 250 mm × 250 mm × 60 mm rectangular cell filled with Intralipid-20% suspension as the scattering medium. The ICG concentration was 1μM, and the Intralipid-20% concentration was adjusted to provide ~ 1-mm transport length for both excitation wavelength of 790 nm and fluorescence wavelength around 825 nm. The data matrix was constructed using the diffusely transmitted fluorescence signals for all scan positions, and the TR matrix was constructed by multiplying data matrix with its transpose. A pseudo spectrum was calculated using the signal subspace of the TR matrix. Tomographic images were generated using the pseudo spectrum. The peaks in the pseudo images provided locations of the target(s) with sub-millimeter accuracy. Concurrent transmission TROT measurements corroborated fluorescence-TROT findings. The results demonstrate that TROT is a fast approach that can be used to obtain accurate three-dimensional position information of fluorescence targets embedded deep inside a highly scattering medium, such as, a contrast-enhanced tumor in a human breast.

Construction of a bimolecular fluorescence complementation (BiFC ...

African Journals Online (AJOL)

Protein–protein interactions are essential for signal transduction in cells. Bimolecular fluorescence complementation (BiFC) is a novel technology that utilises green fluorescent proteins to visualize protein–protein interactions and subcellular protein localisation. BiFC based on pSATN vectors are a good system for ...

Strand displacement activated peroxidase activity of hemin for fluorescent DNA sensing.

Science.gov (United States)

Wang, Quanbo; Xu, Nan; Gui, Zhen; Lei, Jianping; Ju, Huangxian; Yan, Feng

2015-10-07

To efficiently regulate the catalytic activity of the peroxidase mimic hemin, this work designs a double-stranded DNA probe containing an intermolecular dimer of hemin, whose peroxidase activity can be activated by a DNA strand displacement reaction. The double-stranded probe is prepared by annealing two strands of hemin labelled DNA oligonucleotides. Using the fluorescent oxidation product of tyramine by H2O2 as a tracing molecule, the low peroxidase activity of the hemin dimer ensures a low fluorescence background. The strand displacement reaction of the target DNA dissociates the hemin dimer and thus significantly increases the catalytic activity of hemin to produce a large amount of dityramine for fluorescence signal readout. Based on the strand displacement regulated peroxidase activity, a simple and sensitive homogeneous fluorescent DNA sensing method is proposed. The detection can conveniently be carried out in a 96-well plate within 20 min with a detection limit of 0.18 nM. This method shows high specificity, which can effectively distinguish single-base mismatched DNA from perfectly matched target DNA. The DNA strand displacement regulated catalytic activity of hemin has promising application in the determination of various DNA analytes.

Large Scale Plasmonic nanoCones array For Spectroscopy Detection

KAUST Repository

Das, Gobind

2015-09-24

Advanced optical materials or interfaces are gaining attention for diagnostic applications. However, the achievement of large device interface as well as facile surface functionalization largely impairs their wide use. The present work is aimed to address different innovative aspects related to the fabrication of large area 3D plasmonic arrays, their direct and easy functionalization with capture elements and their spectroscopic verifications through enhanced Raman and enhanced fluorescence techniques. In detail we have investigated the effect of Au-based nanoCones array, fabricated by means of direct nanoimprint technique over large area (mm2), on protein capturing and on the enhancement in optical signal. A selective functionalization of gold surfaces was proposed by using a peptide (AuPi3) previously selected by phage display. In this regard, two different sequences, labeled with fluorescein and biotin, were chemisorbed on metallic surfaces. The presence of Au nanoCones array consents an enhancement in electric field on the apex of cone, enabling the detection of molecules. We have witnessed around 12-fold increase in fluorescence intensity and SERS enhancement factor around 1.75 ×105 with respect to the flat gold surface. Furthermore, a sharp decrease in fluorescence lifetime over nanoCones confirms the increase in radiative emission (i.e. an increase in photonics density at the apex of cones).

Large Scale Plasmonic nanoCones array For Spectroscopy Detection

KAUST Repository

Das, Gobind; Battista, Edmondo; Manzo, Gianluigi; Causa, Filippo; Netti, Paolo; Di Fabrizio, Enzo M.

2015-01-01

Advanced optical materials or interfaces are gaining attention for diagnostic applications. However, the achievement of large device interface as well as facile surface functionalization largely impairs their wide use. The present work is aimed to address different innovative aspects related to the fabrication of large area 3D plasmonic arrays, their direct and easy functionalization with capture elements and their spectroscopic verifications through enhanced Raman and enhanced fluorescence techniques. In detail we have investigated the effect of Au-based nanoCones array, fabricated by means of direct nanoimprint technique over large area (mm2), on protein capturing and on the enhancement in optical signal. A selective functionalization of gold surfaces was proposed by using a peptide (AuPi3) previously selected by phage display. In this regard, two different sequences, labeled with fluorescein and biotin, were chemisorbed on metallic surfaces. The presence of Au nanoCones array consents an enhancement in electric field on the apex of cone, enabling the detection of molecules. We have witnessed around 12-fold increase in fluorescence intensity and SERS enhancement factor around 1.75 ×105 with respect to the flat gold surface. Furthermore, a sharp decrease in fluorescence lifetime over nanoCones confirms the increase in radiative emission (i.e. an increase in photonics density at the apex of cones).

Construction of large signaling pathways using an adaptive perturbation approach with phosphoproteomic data.

Science.gov (United States)

Melas, Ioannis N; Mitsos, Alexander; Messinis, Dimitris E; Weiss, Thomas S; Rodriguez, Julio-Saez; Alexopoulos, Leonidas G

2012-04-01

Construction of large and cell-specific signaling pathways is essential to understand information processing under normal and pathological conditions. On this front, gene-based approaches offer the advantage of large pathway exploration whereas phosphoproteomic approaches offer a more reliable view of pathway activities but are applicable to small pathway sizes. In this paper, we demonstrate an experimentally adaptive approach to construct large signaling pathways from phosphoproteomic data within a 3-day time frame. Our approach--taking advantage of the fast turnaround time of the xMAP technology--is carried out in four steps: (i) screen optimal pathway inducers, (ii) select the responsive ones, (iii) combine them in a combinatorial fashion to construct a phosphoproteomic dataset, and (iv) optimize a reduced generic pathway via an Integer Linear Programming formulation. As a case study, we uncover novel players and their corresponding pathways in primary human hepatocytes by interrogating the signal transduction downstream of 81 receptors of interest and constructing a detailed model for the responsive part of the network comprising 177 species (of which 14 are measured) and 365 interactions.

The Potential Applications of Real-Time Monitoring of Water Quality in a Large Shallow Lake (Lake Taihu, China) Using a Chromophoric Dissolved Organic Matter Fluorescence Sensor

OpenAIRE

Niu, Cheng; Zhang, Yunlin; Zhou, Yongqiang; Shi, Kun; Liu, Xiaohan; Qin, Boqiang

2014-01-01

This study presents results from field surveys performed over various seasons in a large, eutrophic, shallow lake (Lake Taihu, China) using an in situ chromophoric dissolved organic matter (CDOM) fluorescence sensor as a surrogate for other water quality parameters. These measurements identified highly significant empirical relationships between CDOM concentration measured using the in situ fluorescence sensor and CDOM absorption, fluorescence, dissolved organic carbon (DOC), chemical oxygen ...

Endogenous and exogenous fluorescence of gastrointestinal tumors: initial clinical observations

Science.gov (United States)

Borisova, Ekaterina; Plamenova, Lilia; Keremedchiev, Momchil; Vladimirov, Borislav; Avramov, Latchezar

2013-03-01

The limitations of standard endoscopy for detection and evaluation of cancerous changes in gastrointestinal tract (GIT) are significant challenge and initiate development of new diagnostic modalities. Therefore many spectral and optical techniques are applied recently into the clinical practice for obtaining qualitatively and quantitatively new data from gastrointestinal neoplasia with different level of clinical applicability and diagnostic success. One of the most promising approaches is fluorescence detection using naturally existing fluorescent molecules or added fluorescent markers. Deltaaminolevulinic acid / protoporphyrin IX is applied for exogenous fluorescent tumor detection in the upper part of gastrointestinal tract. The 5-ALA is administered per os six hours before measurements at dose 20mg/kg weight. Highpower light-emitting diode at 405 nm is used as a source and the excitation light is passed through the light-guide of standard video-endoscopic system to obtain 2-D visualization. Both kinds of spectra - autofluorescence signals and protoporphyrin IX signal are recorded and stored using a fiber-optic microspectrometer, as in endoscopy instrumental channel a fiber is applied to return information about fluorescence signals. In such way 1-D detection and 2-D visualization of the lesions' fluorescence are received. The results from in vivo detection show significant differentiation between normal and abnormal tissues in 1-D spectroscopic regime, but only moderate discrimination in 2-D imaging.

Precision scan-imaging for paperboard quality inspection utilizing X-ray fluorescence

Science.gov (United States)

Norlin, B.; Reza, S.; Fröjdh, C.; Nordin, T.

2018-01-01

Paperboard is typically made up of a core of cellulose fibers [C6H10O5] and a coating layer of [CaCO3]. The uniformity of these layers is a critical parameter for the printing quality. Current quality control methods include chemistry based visual inspection methods as well as X-ray based methods to measure the coating thickness. In this work we combine the X-ray fluorescence signals from the Ca atoms (3.7 keV) in the coating and from a Cu target (8.0 keV) placed behind the paper to simultaneously measure both the coating and the fibers. Cu was selected as the target material since its fluorescence signal is well separated from the Ca signal while its fluorescence's still are absorbed sufficiently in the paper. A laboratory scale setup is built using stepper motors, a silicon drift detector based spectrometer and a collimated X-ray beam. The spectroscopic image is retrieved by scanning the paperboard surface and registering the fluorescence signals from Ca and Cu. The exposure time for this type of setups can be significantly improved by implementing spectroscopic imaging sensors. The material contents in the layers can then be retrieved from the absolute and relative intensities of these two signals.

Demonstration and comparison of tuned and detuned signal recycling in a large-scale gravitational wave detector

International Nuclear Information System (INIS)

Hild, S; Grote, H; Hewtison, M; Lueck, H; Smith, J R; Strain, K A; Willke, B; Danzmann, K

2007-01-01

The British/German gravitational wave detector GEO 600 located near Hannover in Germany is the first large-scale gravitational-wave detector using the advanced technique of signal recycling. Currently the instrument operates in detuned signal recycling mode. Several problems arise due to the fact that the signal recycling cavity changes amplitude and phase of all light fields (carrier and sidebands) present at the dark-port. In addition, in the case of detuned signal recycling this leads to unbalanced sideband fields at the detector output. The large amplitude modulation caused by this asymmetry does not carry any gravitational wave information, but might be the cause of saturation and nonlinearities on the main photodiode. We developed and demonstrated a new control method to realize tuned signal recycling operation in a large-scale gravitational wave detector. A detailed comparison of tuned and detuned signal recycling operation is given. The response function of the system (optical gain) was measured and compared, as was the size of amplitude modulation on the main photodiode. Some important noise couplings were measured and partly found to be strongly reduced in the case of tuned signal recycling operation

Efficient L1 regularization-based reconstruction for fluorescent molecular tomography using restarted nonlinear conjugate gradient.

Science.gov (United States)

Shi, Junwei; Zhang, Bin; Liu, Fei; Luo, Jianwen; Bai, Jing

2013-09-15

For the ill-posed fluorescent molecular tomography (FMT) inverse problem, the L1 regularization can protect the high-frequency information like edges while effectively reduce the image noise. However, the state-of-the-art L1 regularization-based algorithms for FMT reconstruction are expensive in memory, especially for large-scale problems. An efficient L1 regularization-based reconstruction algorithm based on nonlinear conjugate gradient with restarted strategy is proposed to increase the computational speed with low memory consumption. The reconstruction results from phantom experiments demonstrate that the proposed algorithm can obtain high spatial resolution and high signal-to-noise ratio, as well as high localization accuracy for fluorescence targets.

Design of General-purpose Industrial signal acquisition system in a large scientific device

Science.gov (United States)

Ren, Bin; Yang, Lei

2018-02-01

In order to measure the industrial signal of a large scientific device experiment, a set of industrial data general-purpose acquisition system has been designed. It can collect 4~20mA current signal and 0~10V voltage signal. Through the practical experiments, it shows that the system is flexible, reliable, convenient and economical, and the system has characters of high definition and strong anti-interference ability. Thus, the system fully meets the design requirements..

A new traffic control design method for large networks with signalized intersections

Science.gov (United States)

Leininger, G. G.; Colony, D. C.; Seldner, K.

1979-01-01

The paper presents a traffic control design technique for application to large traffic networks with signalized intersections. It is shown that the design method adopts a macroscopic viewpoint to establish a new traffic modelling procedure in which vehicle platoons are subdivided into main stream queues and turning queues. Optimization of the signal splits minimizes queue lengths in the steady state condition and improves traffic flow conditions, from the viewpoint of the traveling public. Finally, an application of the design method to a traffic network with thirty-three signalized intersections is used to demonstrate the effectiveness of the proposed technique.

Quantification of leakage from large unilamellar lipid vesicles by fluorescence correlation spectroscopy

DEFF Research Database (Denmark)

Kristensen, Kasper; Henriksen, Jonas Rosager; Andresen, Thomas Lars

2014-01-01

Fluorescence correlation spectroscopy (FCS) is a powerful experimental technique that in recent years has found numerous applications for studying biological phenomena. In this article, we scrutinize one of these applications, namely, FCS as a technique for studying leakage of fluorescent molecul...

Time-Resolved Fluorescence Immunoassay for C-Reactive Protein Using Colloidal Semiconducting Nanoparticles

Directory of Open Access Journals (Sweden)

Pekka Hänninen

2011-11-01

Full Text Available Besides the typical short-lived fluorescence with decay times in the nanosecond range, colloidal II/VI semiconductor nanoparticles dispersed in buffer also possess a long-lived fluorescence component with decay times in the microsecond range. Here, the signal intensity of the long-lived luminescence at microsecond range is shown to increase 1,000-fold for CdTe nanoparticles in PBS buffer. This long-lived fluorescence can be conveniently employed for time-gated fluorescence detection, which allows for improved signal-to-noise ratio and thus the use of low concentrations of nanoparticles. The detection principle is demonstrated with a time-resolved fluorescence immunoassay for the detection of C-reactive protein (CRP using CdSe-ZnS nanoparticles and green light excitation.

5-ALA induced fluorescent image analysis of actinic keratosis

Science.gov (United States)

Cho, Yong-Jin; Bae, Youngwoo; Choi, Eung-Ho; Jung, Byungjo

2010-02-01

In this study, we quantitatively analyzed 5-ALA induced fluorescent images of actinic keratosis using digital fluorescent color and hyperspectral imaging modalities. UV-A was utilized to induce fluorescent images and actinic keratosis (AK) lesions were demarcated from surrounding the normal region with different methods. Eight subjects with AK lesion were participated in this study. In the hyperspectral imaging modality, spectral analysis method was utilized for hyperspectral cube image and AK lesions were demarcated from the normal region. Before image acquisition, we designated biopsy position for histopathology of AK lesion and surrounding normal region. Erythema index (E.I.) values on both regions were calculated from the spectral cube data. Image analysis of subjects resulted in two different groups: the first group with the higher fluorescence signal and E.I. on AK lesion than the normal region; the second group with lower fluorescence signal and without big difference in E.I. between two regions. In fluorescent color image analysis of facial AK, E.I. images were calculated on both normal and AK lesions and compared with the results of hyperspectral imaging modality. The results might indicate that the different intensity of fluorescence and E.I. among the subjects with AK might be interpreted as different phases of morphological and metabolic changes of AK lesions.

High-Resolution Ultrasound-Switchable Fluorescence Imaging in Centimeter-Deep Tissue Phantoms with High Signal-To-Noise Ratio and High Sensitivity via Novel Contrast Agents.

Science.gov (United States)

Cheng, Bingbing; Bandi, Venugopal; Wei, Ming-Yuan; Pei, Yanbo; D'Souza, Francis; Nguyen, Kytai T; Hong, Yi; Yuan, Baohong

2016-01-01

For many years, investigators have sought after high-resolution fluorescence imaging in centimeter-deep tissue because many interesting in vivo phenomena-such as the presence of immune system cells, tumor angiogenesis, and metastasis-may be located deep in tissue. Previously, we developed a new imaging technique to achieve high spatial resolution in sub-centimeter deep tissue phantoms named continuous-wave ultrasound-switchable fluorescence (CW-USF). The principle is to use a focused ultrasound wave to externally and locally switch on and off the fluorophore emission from a small volume (close to ultrasound focal volume). By making improvements in three aspects of this technique: excellent near-infrared USF contrast agents, a sensitive frequency-domain USF imaging system, and an effective signal processing algorithm, for the first time this study has achieved high spatial resolution (~ 900 μm) in 3-centimeter-deep tissue phantoms with high signal-to-noise ratio (SNR) and high sensitivity (3.4 picomoles of fluorophore in a volume of 68 nanoliters can be detected). We have achieved these results in both tissue-mimic phantoms and porcine muscle tissues. We have also demonstrated multi-color USF to image and distinguish two fluorophores with different wavelengths, which might be very useful for simultaneously imaging of multiple targets and observing their interactions in the future. This work has opened the door for future studies of high-resolution centimeter-deep tissue fluorescence imaging.

Quantification of tumor fluorescence during intraoperative optical cancer imaging.

Science.gov (United States)

Judy, Ryan P; Keating, Jane J; DeJesus, Elizabeth M; Jiang, Jack X; Okusanya, Olugbenga T; Nie, Shuming; Holt, David E; Arlauckas, Sean P; Low, Phillip S; Delikatny, E James; Singhal, Sunil

2015-11-13

Intraoperative optical cancer imaging is an emerging technology in which surgeons employ fluorophores to visualize tumors, identify tumor-positive margins and lymph nodes containing metastases. This study compares instrumentation to measure tumor fluorescence. Three imaging systems (Spectropen, Glomax, Flocam) measured and quantified fluorescent signal-to-background ratios (SBR) in vitro, murine xenografts, tissue phantoms and clinically. Evaluation criteria included the detection of small changes in fluorescence, sensitivity of signal detection at increasing depths and practicality of use. In vitro, spectroscopy was superior in detecting incremental differences in fluorescence than luminescence and digital imaging (Ln[SBR] = 6.8 ± 0.6, 2.4 ± 0.3, 2.6 ± 0.1, p = 0.0001). In fluorescent tumor cells, digital imaging measured higher SBRs than luminescence (6.1 ± 0.2 vs. 4.3 ± 0.4, p = 0.001). Spectroscopy was more sensitive than luminometry and digital imaging in identifying murine tumor fluorescence (SBR = 41.7 ± 11.5, 5.1 ± 1.8, 4.1 ± 0.9, p = 0.0001), and more sensitive than digital imaging at detecting fluorescence at increasing depths (SBR = 7.0 ± 3.4 vs. 2.4 ± 0.5, p = 0.03). Lastly, digital imaging was the most practical and least time-consuming. All methods detected incremental differences in fluorescence. Spectroscopy was the most sensitive for small changes in fluorescence. Digital imaging was the most practical considering its wide field of view, background noise filtering capability, and sensitivity to increasing depth.

Identification of a functional nuclear export signal in the green fluorescent protein asFP499

International Nuclear Information System (INIS)

Mustafa, Huseyin; Strasser, Bernd; Rauth, Sabine; Irving, Robert A.; Wark, Kim L.

2006-01-01

The green fluorescent protein (GFP) asFP499 from Anemonia sulcata is a distant homologue of the GFP from Aequorea victoria. We cloned the asFP499 gene into a mammalian expression vector and showed that this protein was expressed in the human lymphoblast cell line Ramos RA1 and in the embryonic kidney 293T cell line (HEK 293T). In HEK 293T cells, asFP499 was localized mainly in the cytoplasm, suggesting that the protein was excluded from the nucleus. We identified 194 LRMEKLNI 201 as a candidate nuclear export signal in asFP499 and mutated the isoleucine at position 201 to an alanine. Unlike the wildtype form, the mutant protein was distributed throughout the cytoplasm and nucleus. This is First report of a GFP that contains a functional NES

Observation of self-assembled fluorescent beads by scanning near-field optical microscopy and atomic force microscopy

International Nuclear Information System (INIS)

Oh, Y.J.; Jo, W.; Kim, Min-Gon; Kyu Park, Hyun; Hyun Chung, Bong

2006-01-01

Optical response and topography of fluorescent latex beads both on flat self-assembled monolayer and on a micron-patterned surface with poly(dimethylsiloxane) are studied. Scanning near-field optical microscopy and atomic force microscopy were utilized together for detecting fluorescence and imaging topography of the patterned latex beads, respectively. As a result, the micro-patterned latex beads where a specific chemical binding occurred show a strong signal, whereas no signals are observed in the case of nonspecific binding. With fluorescein isothiocyanate (FITC), it is convenient to measure fluorescence signal from the patterned beads allowing us to monitor the small balls of fluorescent latex

Diagnostics of MCF plasmas using Lyman-α fluorescence excited by one or two photons

International Nuclear Information System (INIS)

Voslamber, D.

1998-11-01

Laser-induced Lyman-α fluorescence of the hydrogen isotopes is investigated with regard to diagnostic applications in magnetically confined fusion plasmas. A formal analysis is presented for two excitation schemes: one-photon and Doppler-free two-photon excitation. The analysis includes estimates of the expected experimental errors arising from the photon noise and from the sensitivity of the observed fluorescence signals to variations of the plasma and laser parameters. Both excitation schemes are suitable primarily for application in the plasma edge, but even in the plasma bulk of large machines they can still be applied in combination with a diagnostic neutral beam. The two-photon excitation scheme is particularly attractive because it involves absorption spectra that are resolved within the Doppler width. This implies a large diagnostic potential and in particular offers a way to measure the deuterium-tritium fuel mix in fusion reactors. (author)

Advanced statistical tools for SNP arrays : signal calibration, copy number estimation and single array genotyping

NARCIS (Netherlands)

Rippe, Ralph Christian Alexander

2012-01-01

Fluorescence bias in in signals from individual SNP arrays can be calibrated using linear models. Given the data, the system of equations is very large, so a specialized symbolic algorithm was developed. These models are also used to illustrate that genomic waves do not exist, but are merely an

Fluorescent Biosensors Based on Single-Molecule Counting.

Science.gov (United States)

Ma, Fei; Li, Ying; Tang, Bo; Zhang, Chun-Yang

2016-09-20

Biosensors for highly sensitive, selective, and rapid quantification of specific biomolecules make great contributions to biomedical research, especially molecular diagnostics. However, conventional methods for biomolecular assays often suffer from insufficient sensitivity and poor specificity. In some case (e.g., early disease diagnostics), the concentration of target biomolecules is too low to be detected by these routine approaches, and cumbersome procedures are needed to improve the detection sensitivity. Therefore, there is an urgent need for rapid and ultrasensitive analytical tools. In this respect, single-molecule fluorescence approaches may well satisfy the requirement and hold promising potential for the development of ultrasensitive biosensors. Encouragingly, owing to the advances in single-molecule microscopy and spectroscopy over past decades, the detection of single fluorescent molecule comes true, greatly boosting the development of highly sensitive biosensors. By in vitro/in vivo labeling of target biomolecules with proper fluorescent tags, the quantification of certain biomolecule at the single-molecule level is achieved. In comparison with conventional ensemble measurements, single-molecule detection-based analytical methods possess the advantages of ultrahigh sensitivity, good selectivity, rapid analysis time, and low sample consumption. Consequently, single-molecule detection may be potentially employed as an ideal analytical approach to quantify low-abundant biomolecules with rapidity and simplicity. In this Account, we will summarize our efforts for developing a series of ultrasensitive biosensors based on single-molecule counting. Single-molecule counting is a member of single-molecule detection technologies and may be used as a very simple and ultrasensitive method to quantify target molecules by simply counting the individual fluorescent bursts. In the fluorescent sensors, the signals of target biomolecules may be translated to the

Fluorescent nanocellulosic hydrogels based on graphene quantum dots for sensing laccase

International Nuclear Information System (INIS)

Ruiz-Palomero, Celia; Benítez-Martínez, Sandra; Soriano, M. Laura; Valcárcel, Miguel

2017-01-01

A novel low-cost fluorimetric platform based on sulfur, nitrogen-codoped graphene quantum dots immersed into nanocellulosic hydrogels is designed and applied in detecting the laccase enzyme. Although most of methods for detecting laccase are based on their catalytic activity, which is strongly dependent on environmental parameters, we report a sensitive and selective method based on the fluorescence response of hydrogels containing graphene quantum dots (GQDs) acting as luminophore towards laccase. The easily-prepared gel matrix not only improves the fluorescence signal of GQDs by avoiding their self-quenching but also stabilizes their fluorescence signal and improves their sensitivity towards laccase. Noncovalent interactions between the sensor and the analyte are believed to be causing this significant quenching without peak-shifts of GQD fluorescence via energy transfer. The selective extraction of laccase was proved in different shampoos as complex matrices achieving a detection limit of 0.048 U mL −1 and recoveries of 86.2–94.1%. As the unusual properties of nanocellulose and GQDs, the fluorescent sensor is simple, eco-friendly and cost-efficient. This straightforward strategy is able to detect and stabilize laccase, being an added-value for storage and recycling enzymes. - Highlights: • Fluorescent hydrogels were constructed by combining nanocellulose and graphene quantum dots. • The resulting hydrogels exhibited fluorescence quenching in presence of laccase. • Equilibrium in the optical signal of S,N-graphene quantum dots in presence of laccase was achieved faster within hydrogels. • The proposed method to determine laccase using fluorescent hydrogels was successfully applied in shampoo.

Fluorescence spectroscopy of gastrointestinal tumors using δ-ALA

Science.gov (United States)

Borisova, E. G.; Vladimirov, B. G.; Angelov, I. G.; Avramov, L. A.

2007-03-01

In the recent study delta-aminolevulinic acid/Protoporphyrin IX (δ-ALA/PpIX) is used as fluorescent marker for dysplasia and tumor detection in esophagus and stomach. The δ-ALA is administered per os six hours before measurements at dose 20mg/kg weight. High-power light-emitting diode at 405 nm is used as an excitation source. Special opto-mechanical device is built to use the light guide of standard video-endoscopic system (Olimpus Corp.). Through endoscopic instrumental channel a fiber is applied to return information about fluorescence to microspectrometer (USB4000, OceanOptics Inc.). The fluorescence detected from tumor sites has very complex spectral origins. It consists of autofluorescence, fluorescence from exogenous fluorophores and re-absorption from the chromophores accumulated in the tissue investigated. Mucosa autofluorescence lies at 450-600 nm region. The fluorescence of PpIX is clearly pronounced at the 630-710 nm region. Deep minima in the tumor fluorescence signals are observed in the region 540-575 nm, related to hemoglobin re-absorption. Such high hemoglobin content is an indication of the tumors neovascularisation and it is clearly pronounced in all dysplastic and tumor sites investigated. The lack of fluorescence peaks in the red spectral area for normal mucosa is an indication for selective accumulation of δ-ALA/PpIX only in abnormal sites and gives high contrast when lesion borders are determined from clinicians during video observation in the process of diagnostic procedure. Very good correlation between fluorescence signals and histology examination results of the lesions investigated is achieved.

ICT-Isomerization-Induced Turn-On Fluorescence Probe with a Large Emission Shift for Mercury Ion: Application in Combinational Molecular Logic.

Science.gov (United States)

Bhatta, Sushil Ranjan; Mondal, Bijan; Vijaykumar, Gonela; Thakur, Arunabha

2017-10-02

A unique turn-on fluorescent device based on a ferrocene-aminonaphtholate derivative specific for Hg 2+ cation was developed. Upon binding with Hg 2+ ion, the probe shows a dramatic fluorescence enhancement (the fluorescence quantum yield increases 58-fold) along with a large red shift of 68 nm in the emission spectrum. The fluorescence enhancement with a red shift may be ascribed to the combinational effect of C═N isomerization and an extended intramolecular charge transfer (ICT) mechanism. The response was instantaneous with a detection limit of 2.7 × 10 -9 M. Upon Hg 2+ recognition, the ferrocene/ferrocenium redox peak was anodically shifted by ΔE 1/2 = 72 mV along with a "naked eye" color change from faint yellow to pale orange for this metal cation. Further, upon protonation of the imine nitrogen, the present probe displays a high fluorescence output due to suppression of the C═N isomerization process. Upon deprotonation using strong base, the fluorescence steadily decreases, which indicates that H + and OH - can be used to regulate the off-on-off fluorescence switching of the present probe. Density functional theory studies revealed that the addition of acid leads to protonation of the imine N (according to natural bond orbital analysis), and the resulting iminium proton forms a strong H-bond (2.307 Å) with one of the triazole N atoms to form a five-membered ring, which makes the molecule rigid; hence, enhancement of the ICT process takes place, thereby leading to a fluorescence enhancement with a red shift. The unprecedented combination of H + , OH - , and Hg 2+ ions has been used to generate a molecular system exhibiting the INHIBIT-OR combinational logic operation.

Distributed Sensor Particles for Remote Fluorescence Detection of Trace Analytes: UXO/CW; TOPICAL

International Nuclear Information System (INIS)

SINGH, ANUP K.; GUPTA, ALOK; MULCHANDANI, ASHOK; CHEN, WILFRED; BHATIA, RIMPLE B.; SCHOENIGER, JOSEPH S.; ASHLEY, CAROL S.; BRINKER, C. JEFFREY; HANCE, BRADLEY G.; SCHMITT, RANDAL L.; JOHNSON, MARK S.; HARGIS JR. PHILIP J.; SIMONSON, ROBERT J.

2001-01-01

This report summarizes the development of sensor particles for remote detection of trace chemical analytes over broad areas, e.g residual trinitrotoluene from buried landmines or other unexploded ordnance (UXO). We also describe the potential of the sensor particle approach for the detection of chemical warfare (CW) agents. The primary goal of this work has been the development of sensor particles that incorporate sample preconcentration, analyte molecular recognition, chemical signal amplification, and fluorescence signal transduction within a ''grain of sand''. Two approaches for particle-based chemical-to-fluorescence signal transduction are described: (1) enzyme-amplified immunoassays using biocompatible inorganic encapsulants, and (2) oxidative quenching of a unique fluorescent polymer by TNT

Optimization of a Fluorescence-Based Assay for Large-Scale Drug Screening against Babesia and Theileria Parasites

OpenAIRE

Rizk, Mohamed Abdo; El-Sayed, Shimaa Abd El-Salam; Terkawi, Mohamed Alaa; Youssef, Mohamed Ahmed; El Said, El Said El Shirbini; Elsayed, Gehad; El-Khodery, Sabry; El-Ashker, Maged; Elsify, Ahmed; Omar, Mosaab; Salama, Akram; Yokoyama, Naoaki; Igarashi, Ikuo

2015-01-01

A rapid and accurate assay for evaluating antibabesial drugs on a large scale is required for the discovery of novel chemotherapeutic agents against Babesia parasites. In the current study, we evaluated the usefulness of a fluorescence-based assay for determining the efficacies of antibabesial compounds against bovine and equine hemoparasites in in vitro cultures. Three different hematocrits (HCTs; 2.5%, 5%, and 10%) were used without daily replacement of the medium. The results of a high-thr...

Large-signal modulation characteristics of a GaN-based micro-LED for Gbps visible-light communication

Science.gov (United States)

Tian, Pengfei; Wu, Zhengyuan; Liu, Xiaoyan; Fang, Zhilai; Zhang, Shuailong; Zhou, Xiaolin; Liu, Kefu; Liu, Ming-Gang; Chen, Shu-Jhih; Lee, Chia-Yu; Cong, Chunxiao; Hu, Laigui; Qiu, Zhi-Jun; Zheng, Lirong; Liu, Ran

2018-04-01

The large-signal modulation characteristics of a GaN-based micro-LED have been studied for Gbps visible-light communication. With an increasing signal modulation depth the modulation bandwidth decreases, which matches up with the increase in the sum of the signal rise time and fall time. By simulating the band diagram and the carrier recombination rate of the micro-LED under large-signal modulation, carrier recombination and the carrier sweep-out effect are analyzed and found to be the dominant mechanisms behind the variation of modulation bandwidth. These results give further insight into improving the modulation bandwidth for high-speed visible-light communication.

Large-Signal Code TESLA: Improvements in the Implementation and in the Model

National Research Council Canada - National Science Library

Chernyavskiy, Igor A; Vlasov, Alexander N; Anderson, Jr., Thomas M; Cooke, Simon J; Levush, Baruch; Nguyen, Khanh T

2006-01-01

We describe the latest improvements made in the large-signal code TESLA, which include transformation of the code to a Fortran-90/95 version with dynamical memory allocation and extension of the model...

Fluorescent standards for photodynamic therapy

Science.gov (United States)

Belko, N.; Kavalenka, S.; Samtsov, M.

2016-08-01

Photodynamic therapy is an evolving technique for treatment of various oncological diseases. This method employs photosensitizers - species that lead to death of tumor cells after the photoactivation. For further development and novel applications of photodynamic therapy new photosensitizers are required. After synthesis of a new photosensitizer it is important to know its concentration in different biological tissues after its administration and distribution. The concentration is frequently measured by the extraction method, which has some disadvantages, e.g. it requires many biological test subjects that are euthanized during the measurement. We propose to measure the photosensitizer concentration in tissue by its fluorescence. For this purpose fluorescent standards were developed. The standards are robust and simple to produce; their fluorescence signal does not change with time. The fluorescence intensity of fluorescent standards seems to depend linearly on the dye concentration. A set of standards thus allow the calibration of a spectrometer. Finally, the photosensitizer concentration can be determined by the fluorescence intensity after comparing the corresponding spectrum with spectra of the set of fluorescent standards. A biological test subject is not euthanized during this kind of experiment. We hope this more humane technique can be used in future instead of the extraction method.

Fluorescence single-molecule counting assays for protein quantification using epi-fluorescence microscopy with quantum dots labeling

International Nuclear Information System (INIS)

Jiang Dafeng; Liu Chunxia; Wang Lei; Jiang Wei

2010-01-01

A single-molecule counting approach for quantifying the antibody affixed to a surface using quantum dots and epi-fluorescence microscopy is presented. Modifying the glass substrates with carboxyl groups provides a hydrophilic surface that reacts with amine groups of an antibody to allow covalent immobilization of the antibody. Nonspecific adsorption of single molecules on the modified surfaces was first investigated. Then, quantum dots were employed to form complexes with surface-immobilized antibody molecules and used as fluorescent probes for single-molecule imaging. Epi-fluorescence microscopy was chosen as the tool for single-molecule fluorescence detection here. The generated fluorescence signals were taken by an electron multiplying charge-coupled device and were found to be proportional to the sample concentrations. Under optimal conditions, a linear response range of 5.0 x 10 -14 -3.0 x 10 -12 mol L -1 was obtained between the number of single molecules and sample concentration via a single-molecule counting approach.

Ultraviolet signals in birds are special.

OpenAIRE

Hausmann, Franziska; Arnold, Kathryn E; Marshall, N Justin; Owens, Ian P F

2003-01-01

Recent behavioural experiments have shown that birds use ultraviolet (UV)-reflective and fluorescent plumage as cues in mate choice. It remains controversial, however, whether such UV signals play a special role in sexual communication, or whether they are part of general plumage coloration. We use a comparative approach to test for a general association between sexual signalling and either UV-reflective or fluorescent plumage. Among the species surveyed, 72% have UV colours and there is a si...

Tomato seeds maturity detection system based on chlorophyll fluorescence

Science.gov (United States)

Li, Cuiling; Wang, Xiu; Meng, Zhijun

2016-10-01

Chlorophyll fluorescence intensity can be used as seed maturity and quality evaluation indicator. Chlorophyll fluorescence intensity of seed coats is tested to judge the level of chlorophyll content in seeds, and further to judge the maturity and quality of seeds. This research developed a detection system of tomato seeds maturity based on chlorophyll fluorescence spectrum technology, the system included an excitation light source unit, a fluorescent signal acquisition unit and a data processing unit. The excitation light source unit consisted of two high power LEDs, two radiators and two constant current power supplies, and it was designed to excite chlorophyll fluorescence of tomato seeds. The fluorescent signal acquisition unit was made up of a fluorescence spectrometer, an optical fiber, an optical fiber scaffolds and a narrowband filter. The data processing unit mainly included a computer. Tomato fruits of green ripe stage, discoloration stage, firm ripe stage and full ripe stage were harvested, and their seeds were collected directly. In this research, the developed tomato seeds maturity testing system was used to collect fluorescence spectrums of tomato seeds of different maturities. Principal component analysis (PCA) method was utilized to reduce the dimension of spectral data and extract principal components, and PCA was combined with linear discriminant analysis (LDA) to establish discriminant model of tomato seeds maturity, the discriminant accuracy was greater than 90%. Research results show that using chlorophyll fluorescence spectrum technology is feasible for seeds maturity detection, and the developed tomato seeds maturity testing system has high detection accuracy.

Absorption and fluorescence properties of chromophoric dissolved organic matter: implications for the monitoring of water quality in a large subtropical reservoir.

Science.gov (United States)

Liu, Xiaohan; Zhang, Yunlin; Shi, Kun; Zhu, Guangwei; Xu, Hai; Zhu, Mengyuan

2014-12-01

The development of techniques for real-time monitoring of water quality is of great importance for effectively managing inland water resources. In this study, we first analyzed the absorption and fluorescence properties in a large subtropical reservoir and then used a chromophoric dissolved organic matter (CDOM) fluorescence monitoring sensor to predict several water quality parameters including the total nitrogen (TN), total phosphorus (TP), chemical oxygen demand (COD), dissolved organic carbon (DOC), and CDOM fluorescence parallel factor analysis (PARAFAC) components in the reservoir. The CDOM absorption coefficient at 254 nm (a(254)), the humic-like component (C1), and the tryptophan-like component (C3) decreased significantly along a gradient from the northwest to the lake center, northeast, southwest, and southeast region in the reservoir. However, no significant spatial difference was found for the tyrosine-like component (C2), which contributed only four marked peaks. A highly significant linear correlation was found between the a(254) and CDOM concentration measured using the CDOM fluorescence sensor (r(2) = 0.865, n = 76, p CDOM concentrations could act as a proxy for the CDOM absorption coefficient measured in the laboratory. Significant correlations were also found between the CDOM concentration and TN, TP, COD, DOC, and the maximum fluorescence intensity of C1, suggesting that the real-time monitoring of CDOM concentrations could be used to predict these water quality parameters and trace the humic-like fluorescence substance in clear aquatic ecosystems with DOC CDOM fluorescence sensor is a useful tool for on-line water quality monitoring if the empirical relationship between the CDOM concentration measured using the CDOM fluorescence sensor and the water quality parameters is calibrated and validated.

Azadioxatriangulenium (ADOTA+): A long fluorescence lifetime fluorophore for large biomolecule binding assay

Science.gov (United States)

Sørensen, Thomas Just; Thyrhaug, Erling; Szabelski, Mariusz; Luchowski, Rafal; Gryczynski, Ignacy; Gryczynski, Zygmunt; Laursen, Bo W.

2013-01-01

Of the many optical bioassays available, sensing by fluorescence anisotropy have great advantages as it provides a sensitive, instrumentally simple, ratiometric method of detection. However, it is hampered by a severe limitation as the emission lifetime of the label needs to be comparable to the correlation lifetime (tumbling time) of the biomolecule which is labelled. For proteins of moderate size this is in the order of 20–200 ns, which due to practical issues currently limits the choice of labels to the dansyl-type dyes and certain aromatics dyes. These have the significant drawback of UV/blue absorption and emission as well as an often significant solvent sensitivity. Here, we report the synthesis and characterization of a new fluorescent label for high molecular weight biomolecules assay based on the azadioxatriangulenium motif. The NHS ester of the long fluorescence lifetime, red emitting fluorophore: azadioxatriangulenium (ADOTA-NHS) was conjugated to anti-rabbit Immunoglobulin G (antiIgG). The long fluorescence lifetime was exploited to determine the correlation time of the high molecular weight antibody and its complex with rabbit Immuniglobulin G (IgG) with steady-state fluorescence anisotropy and time-resolved methods: solution phase immuno-assay was performed following either steady-state or time-resolved fluorescence anisotropy. By performing a variable temperature experiment it was determined that the binding of the ligand resulted in an increase in correlation time by more than 75 %, and a change in the steady-state anisotropy increase of 18%. The results show that the triangulenium class of dyes can be used in anisotropy assay for detecting binding events involving biomolecules of far larger size than what is possible with the other red emitting organic dyes. PMID:24058730

Development of laser-induced fluorescence for precombustion diagnostics in spark-ignition engines

Energy Technology Data Exchange (ETDEWEB)

Neij, H.

1998-11-01

Motivated by a desire to understand and optimize combustion in spark-ignition (SI) engines, laser techniques have been developed for measurement of fuel and residual gas, respectively, in the precombustion mixture of an operating SI engine. The primary objective was to obtain two-dimensional, quantitative data in the vicinity of the spark gap at the time of ignition. A laser-induced fluorescence (LIF) technique was developed for fuel visualization in engine environments. Since the fluorescence signal from any commercial gasoline fuel would be unknown to its origin, with an unpredictable dependence on collisional partners, pressure and temperature, a non-fluorescent base fuel - isooctane - was used. For LIF detection, a fluorescent species was added to the fuel. An additive not commonly used in this context - 3-pentanone - was chosen based on its suitable vaporization characteristics and fluorescent properties. The LIF technique was applied to an optically accessible research engine. By calibration, the fluorescence signal from the additive was converted to fuel-to-air equivalence ratio ({phi}). The accuracy and precision of the acquired data were assessed. A statistical evaluation revealed that the spatially averaged equivalence ratio around the spark plug had a significant impact on the combustion event. The strong correlation between these two quantities suggested that the early combustion was sensitive to large-scale inhomogeneities in the precombustion mixture. A similar LIF technique, using acetone as a fluorescent additive in methane, was applied to a combustion cell for ion current evaluation. The local equivalence ratio around the spark gap at the time of ignition was extracted from LIF data. Useful relations were identified between different ion current parameters and the local equivalence ratio, although the impact of the flow field, the fuel type, and the electrode geometry were identified as areas for future research. A novel fuel - dimethyl ether (DME

Controllable ultra-narrow fluorescence and six-wave mixing under double electromagnetically induced transparency

International Nuclear Information System (INIS)

Wang, Z G; Zhang, Z Y; Che, J L; Zhang, Y Z; Li, C B; Zheng, H B; Zhang, Y P

2013-01-01

We report the first observation of six-wave mixing (SWM) and fluorescence signals in an electromagnetically induced transparency (EIT) window. Several remarkable advantages are described. First, multiple bright and dark states are simultaneously observed due to enhancement or suppression of the SWM signal. Second, ultra-narrow fluorescence, much narrower than the EIT window, is experimentally obtained. Third, the ultra-narrow fluorescence can also generate Autler–Townes splitting on scanning the coupling beam. Fourth, a double-peak EIT window is obtained using the nest-dressing scheme. Such studies concerning SWM and fluorescence have applications in optical switching, multi-channel communication and narrowband and long-range quantum communication. (letter)

A Low-Cost, High-Performance System for Fluorescence Lateral Flow Assays

Directory of Open Access Journals (Sweden)

Linda G. Lee

2013-10-01

Full Text Available We demonstrate a fluorescence lateral flow system that has excellent sensitivity and wide dynamic range. The illumination system utilizes an LED, plastic lenses and plastic and colored glass filters for the excitation and emission light. Images are collected on an iPhone 4. Several fluorescent dyes with long Stokes shifts were evaluated for their signal and nonspecific binding in lateral flow. A wide range of values for the ratio of signal to nonspecific binding was found, from 50 for R-phycoerythrin (R-PE to 0.15 for Brilliant Violet 605. The long Stokes shift of R-PE allowed the use of inexpensive plastic filters rather than costly interference filters to block the LED light. Fluorescence detection with R-PE and absorbance detection with colloidal gold were directly compared in lateral flow using biotinylated bovine serum albumen (BSA as the analyte. Fluorescence provided linear data over a range of 0.4–4,000 ng/mL with a 1,000-fold signal change while colloidal gold provided non-linear data over a range of 16–4,000 ng/mL with a 10-fold signal change. A comparison using human chorionic gonadotropin (hCG as the analyte showed a similar advantage in the fluorescent system. We believe our inexpensive yet high-performance platform will be useful for providing quantitative and sensitive detection in a point-of-care setting.

Sun-induced fluorescence - a new probe of photosynthesis: First maps from the imaging spectrometer HyPlant.

Science.gov (United States)

Rascher, U; Alonso, L; Burkart, A; Cilia, C; Cogliati, S; Colombo, R; Damm, A; Drusch, M; Guanter, L; Hanus, J; Hyvärinen, T; Julitta, T; Jussila, J; Kataja, K; Kokkalis, P; Kraft, S; Kraska, T; Matveeva, M; Moreno, J; Muller, O; Panigada, C; Pikl, M; Pinto, F; Prey, L; Pude, R; Rossini, M; Schickling, A; Schurr, U; Schüttemeyer, D; Verrelst, J; Zemek, F

2015-12-01

Variations in photosynthesis still cause substantial uncertainties in predicting photosynthetic CO2 uptake rates and monitoring plant stress. Changes in actual photosynthesis that are not related to greenness of vegetation are difficult to measure by reflectance based optical remote sensing techniques. Several activities are underway to evaluate the sun-induced fluorescence signal on the ground and on a coarse spatial scale using space-borne imaging spectrometers. Intermediate-scale observations using airborne-based imaging spectroscopy, which are critical to bridge the existing gap between small-scale field studies and global observations, are still insufficient. Here we present the first validated maps of sun-induced fluorescence in that critical, intermediate spatial resolution, employing the novel airborne imaging spectrometer HyPlant. HyPlant has an unprecedented spectral resolution, which allows for the first time quantifying sun-induced fluorescence fluxes in physical units according to the Fraunhofer Line Depth Principle that exploits solar and atmospheric absorption bands. Maps of sun-induced fluorescence show a large spatial variability between different vegetation types, which complement classical remote sensing approaches. Different crop types largely differ in emitting fluorescence that additionally changes within the seasonal cycle and thus may be related to the seasonal activation and deactivation of the photosynthetic machinery. We argue that sun-induced fluorescence emission is related to two processes: (i) the total absorbed radiation by photosynthetically active chlorophyll; and (ii) the functional status of actual photosynthesis and vegetation stress. © 2015 John Wiley & Sons Ltd.

Quantitative Studies of Antimicrobial Peptide Pore Formation in Large Unilamellar Vesicles by Fluorescence Correlation Spectroscopy (FCS)

DEFF Research Database (Denmark)

Kristensen, Kasper; Henriksen, Jonas Rosager; Andresen, Thomas Lars

2013-01-01

In spite of intensive research efforts over the past decades, the mechanisms by which membrane-active antimicrobial peptides interact with phospholipid membranes are not yet fully elucidated. New tools that can be used to characterize antimicrobial peptide-lipid membrane interactions are therefore...... to quantify leakage from large unilamellar vesicles is associated with a number of experimental pitfalls. Based on theoretical and experimental considerations, we discuss how to properly design experiments to avoid these pitfalls. Subsequently, we apply fluorescence correlation spectroscopy to quantify...

High-quality substrate for fluorescence enhancement using agarose-coated silica opal film.

Science.gov (United States)

Xu, Ming; Li, Juan; Sun, Liguo; Zhao, Yuanjin; Xie, Zhuoying; Lv, Linli; Zhao, Xiangwei; Xiao, Pengfeng; Hu, Jing; Lv, Mei; Gu, Zhongze

2010-08-01

To improve the sensitivity of fluorescence detection in biochip, a new kind of substrates was developed by agarose coating on silica opal film. In this study, silica opal film was fabricated on glass substrate using the vertical deposition technique. It can provide stronger fluorescence signals and thus improve the detection sensitivity. After coating with agarose, the hybrid film could provide a 3D support for immobilizing sample. Comparing with agarose-coated glass substrate, the agarose-coated opal substrates could selectively enhance particular fluorescence signals with high sensitivity when the stop band of the silica opal film in the agarose-coated opal substrate overlapped the fluorescence emission wavelength. A DNA hybridization experiment demonstrated that fluorescence intensity of special type of agarose-coated opal substrates was about four times that of agarose-coated glass substrate. These results indicate that the optimized agarose-coated opal substrate can be used for improving the sensitivity of fluorescence detection with high quality and selectivity.

Spectral and physiological information from chlorophyll fluorescence signals in the detection of pine damage

Energy Technology Data Exchange (ETDEWEB)

Meinander, O. [Finnish Meteorological Inst., Helsinki (Finland). Air Quality Dept.; Somersalo, S. [Helsinki Univ., Helsinki (Finland). Dept. of Plant Biology

1995-12-31

Photosynthesis is often among the first targets of the air pollution stress of plants. As chlorophyll fluorescence is a process competing with photosynthetic electron transport it can be employed to study the potential photosynthetic capacity and to detect damage to the photosynthetic apparatus. Many previous studies have shown that chlorophyll fluorescence can be a powerful tool in the detection of forest damage. In this preliminary study, singular value analysis of the fluorescence induction curves was used together with the traditional way of analyzing fluorescence measurements. The experimental data were collected from ozone and carbon dioxide fumigated Scots pine saplings. (author)

Spectral and physiological information from chlorophyll fluorescence signals in the detection of pine damage

Energy Technology Data Exchange (ETDEWEB)

Meinander, O [Finnish Meteorological Inst., Helsinki (Finland). Air Quality Dept.; Somersalo, S [Helsinki Univ., Helsinki (Finland). Dept. of Plant Biology

1996-12-31

Photosynthesis is often among the first targets of the air pollution stress of plants. As chlorophyll fluorescence is a process competing with photosynthetic electron transport it can be employed to study the potential photosynthetic capacity and to detect damage to the photosynthetic apparatus. Many previous studies have shown that chlorophyll fluorescence can be a powerful tool in the detection of forest damage. In this preliminary study, singular value analysis of the fluorescence induction curves was used together with the traditional way of analyzing fluorescence measurements. The experimental data were collected from ozone and carbon dioxide fumigated Scots pine saplings. (author)

Structural design of intrinsically fluorescent oxysterols

DEFF Research Database (Denmark)

Nåbo, Lina J; Modzel, Maciej; Krishnan, Kathiresan

2018-01-01

Oxysterols are oxidized derivatives of cholesterol with many important biological functions. Trafficking of oxysterols in and between cells is not well studied, largely due to the lack of appropriate oxysterol analogs. Intrinsically fluorescent oxysterols present a new route towards direct...... observation of intracellular oxysterol trafficking by fluorescence microscopy. We characterize the fluorescence properties of the existing fluorescent 25-hydroxycholesterol analog 25-hydroxycholestatrienol, and propose a new probe with an extended conjugated system. The location of both probes inside...

The potential applications of real-time monitoring of water quality in a large shallow lake (Lake Taihu, China) using a chromophoric dissolved organic matter fluorescence sensor.

Science.gov (United States)

Niu, Cheng; Zhang, Yunlin; Zhou, Yongqiang; Shi, Kun; Liu, Xiaohan; Qin, Boqiang

2014-06-30

This study presents results from field surveys performed over various seasons in a large, eutrophic, shallow lake (Lake Taihu, China) using an in situ chromophoric dissolved organic matter (CDOM) fluorescence sensor as a surrogate for other water quality parameters. These measurements identified highly significant empirical relationships between CDOM concentration measured using the in situ fluorescence sensor and CDOM absorption, fluorescence, dissolved organic carbon (DOC), chemical oxygen demand (COD) and total phosphorus (TP) concentrations. CDOM concentration expressed in quinine sulfate equivalent units, was highly correlated with the CDOM absorption coefficient (r(2) = 0.80, p CDOM concentration measured using the in situ fluorescence sensor could act as a substitute for the CDOM absorption coefficient and fluorescence measured in the laboratory. Similarly, CDOM concentration was highly correlated with DOC concentration (r(2) = 0.68, p CDOM fluorescence sensor measurements could be a proxy for DOC concentration. In addition, significant positive correlations were found between laboratory CDOM absorption coefficients and COD (r(2) = 0.83, p CDOM fluorescence sensor.

Fluorescence-Raman Dual Modal Endoscopic System for Multiplexed Molecular Diagnostics

Science.gov (United States)

Jeong, Sinyoung; Kim, Yong-Il; Kang, Homan; Kim, Gunsung; Cha, Myeong Geun; Chang, Hyejin; Jung, Kyung Oh; Kim, Young-Hwa; Jun, Bong-Hyun; Hwang, Do Won; Lee, Yun-Sang; Youn, Hyewon; Lee, Yoon-Sik; Kang, Keon Wook; Lee, Dong Soo; Jeong, Dae Hong

2015-03-01

Optical endoscopic imaging, which was recently equipped with bioluminescence, fluorescence, and Raman scattering, allows minimally invasive real-time detection of pathologies on the surface of hollow organs. To characterize pathologic lesions in a multiplexed way, we developed a dual modal fluorescence-Raman endomicroscopic system (FRES), which used fluorescence and surface-enhanced Raman scattering nanoprobes (F-SERS dots). Real-time, in vivo, and multiple target detection of a specific cancer was successful, based on the fast imaging capability of fluorescence signals and the multiplex capability of simultaneously detected SERS signals using an optical fiber bundle for intraoperative endoscopic system. Human epidermal growth factor receptor 2 (HER2) and epidermal growth factor receptor (EGFR) on the breast cancer xenografts in a mouse orthotopic model were successfully detected in a multiplexed way, illustrating the potential of FRES as a molecular diagnostic instrument that enables real-time tumor characterization of receptors during routine endoscopic procedures.

Large-Signal DG-MOSFET Modelling for RFID Rectification

Directory of Open Access Journals (Sweden)

R. Rodríguez

2016-01-01

Full Text Available This paper analyses the undoped DG-MOSFETs capability for the operation of rectifiers for RFIDs and Wireless Power Transmission (WPT at microwave frequencies. For this purpose, a large-signal compact model has been developed and implemented in Verilog-A. The model has been numerically validated with a device simulator (Sentaurus. It is found that the number of stages to achieve the optimal rectifier performance is inferior to that required with conventional MOSFETs. In addition, the DC output voltage could be incremented with the use of appropriate mid-gap metals for the gate, as TiN. Minor impact of short channel effects (SCEs on rectification is also pointed out.

Development of confocal X-ray fluorescence (XRF) microscopy at the Cornell high energy synchrotron source

International Nuclear Information System (INIS)

Woll, A.R.; Huang, R.; Mass, J.; Bisulca, C.; Bilderback, D.H.; Gruner, S.; Gao, N.

2006-01-01

A confocal X-ray fluorescence microscope was built at the Cornell High Energy Synchrotron Source (CHESS) to obtain compositional depth profiles of historic paintings. The microscope consists of a single-bounce, borosilicate monocapillary optic to focus the incident beam onto the painting and a commercial borosilicate polycapillary lens to collect the fluorescent X-rays. The resolution of the microscope was measured by scanning a variety of thin metal films through this confocal volume while monitoring the fluorescence signal. The capabilities of the technique were then probed using test paint microstructures with up to four distinct layers, each having a thickness in the range of 10-80 microns. Results from confocal XRF were compared with those from stand-alone XRF and visible light microscopy of the paint cross-sections. A large area, high-resolution scanner is currently being built to perform 3D scans on moderately sized paintings. (orig.)

Limitations of fluorescence spectroscopy to characterize organic matter in engineered systems

Science.gov (United States)

Korak, J.

2017-12-01

Fluorescence spectroscopy has been widely used to characterize dissolved organic matter (DOM) in engineered systems, such as drinking water, municipal wastewater and industrial water treatment. While fluorescence data collected in water treatment applications has led to the development of strong empirical relationships between fluorescence responses and process performance, the use of fluorescence to infer changes in the underlying organic matter chemistry is often oversimplified and applied out of context. Fluorescence only measures a small fraction of DOM as fluorescence quantum yields are less than 5% for many DOM sources. Relying on fluorescence as a surrogate for DOM presence, character or reactivity may not be appropriate for systems where small molecular weight, hydrophilic constituents unlikely to fluoresce are important. In addition, some methods rely on interpreting fluorescence signals at different excitation wavelengths as a surrogate for operationally-defined humic- and fulvic-acids in lieu of traditional XAD fractionation techniques, but these approaches cannot be supported by other lines of evidence considering natural abundance and fluorescence quantum yields of these fractions. These approaches also conflict with parallel factor analysis (PARAFAC), a statistical approach that routinely identifies fluorescence components with dual excitation behavior. Lastly, methods developed for natural systems are often applied out of context to engineered systems. Fluorescence signals characteristic of phenols or indoles are often interpreted as indicators for biological activity in natural systems due to fluorescent amino acids and peptides, but this interpretation is may not be appropriate in engineering applications where non-biological sources of phenolic functional groups may be present. This presentation explores common fluorescence interpretation approaches, discusses the limitations and provides recommendations related to engineered systems.

Plasmon enhanced silver quantum cluster fluorescence for biochemical applications

DEFF Research Database (Denmark)

Bernard, S.; Kutter, J.P.; Mogensen, Klaus Bo

2014-01-01

Fluorescence microscopy of individual silver quantum clusters on the surface of silver nanoparticles reveals strong photoactivated emission under blue light excitation [1-4]. In this work, silver nanoparticles are produced by annealing silver thin films deposited on a glass substrate and silver q...... purposes. It was found, that in presence of a strong nucleophile (such as CN-), silver quantum clusters are dissolved into non-fluorescing AgCN complexes, resulting in a fast and observable decrease of the fluorescent signal....

Fluorescence detection system for microfluidic droplets

Science.gov (United States)

Chen, Binyu; Han, Xiaoming; Su, Zhen; Liu, Quanjun

2018-05-01

In microfluidic detection technology, because of the universality of optical methods in laboratory, optical detection is an attractive solution for microfluidic chip laboratory equipment. In addition, the equipment with high stability and low cost can be realized by integrating appropriate optical detection technology on the chip. This paper reports a detection system for microfluidic droplets. Photomultiplier tubes (PMT) is used as a detection device to improve the sensitivity of detection. This system improves the signal to noise ratio by software filtering and spatial filter. The fluorescence intensity is proportional to the concentration of the fluorescence and intensity of the laser. The fluorescence micro droplets of different concentrations can be distinguished by this system.

Photobleaching correction in fluorescence microscopy images

International Nuclear Information System (INIS)

Vicente, Nathalie B; Diaz Zamboni, Javier E; Adur, Javier F; Paravani, Enrique V; Casco, Victor H

2007-01-01

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

Dual-Color Fluorescence Imaging of Magnetic Nanoparticles in Live Cancer Cells Using Conjugated Polymer Probes

Science.gov (United States)

Sun, Minjie; Sun, Bin; Liu, Yun; Shen, Qun-Dong; Jiang, Shaojun

2016-01-01

Rapid growth in biological applications of nanomaterials brings about pressing needs for exploring nanomaterial-cell interactions. Cationic blue-emissive and anionic green-emissive conjugated polymers are applied as dual-color fluorescence probes to the surface of negatively charged magnetic nanoparticles through sequentially electrostatic adsorption. These conjugated polymers have large extinction coefficients and high fluorescence quantum yield (82% for PFN and 62% for ThPFS). Thereby, one can visualize trace amount (2.7 μg/mL) of fluorescence-labeled nanoparticles within cancer cells by confocal laser scanning microscopy. Fluorescence labeling by the conjugated polymers is also validated for quantitative determination of the internalized nanoparticles in each individual cell by flow cytometry analysis. Extensive overlap of blue and green fluorescence signals in the cytoplasm indicates that both conjugated polymer probes tightly bind to the surface of the nanoparticles during cellular internalization. The highly charged and fluorescence-labeled nanoparticles non-specifically bind to the cell membranes, followed by cellular uptake through endocytosis. The nanoparticles form aggregates inside endosomes, which yields a punctuated staining pattern. Cellular internalization of the nanoparticles is dependent on the dosage and time. Uptake efficiency can be enhanced three-fold by application of an external magnetic field. The nanoparticles are low cytotoxicity and suitable for simultaneously noninvasive fluorescence and magnetic resonance imaging application. PMID:26931282

Method and device for X-ray fluorescence analysis

International Nuclear Information System (INIS)

Jagoutz, E.; Palme, C.

1978-01-01

In the x-ray fluorescence analyzer the useful signal can be completely separated from the spurious signals, and especially the pulse can be determined. For this purpose the output of the radiation detector is connected with a multichannel pulse height discriminator. The measured signal determined in the pulse heigth discriminator may be indicated by a visual display or processed by a computer (coincidence circuits). (DG) [de

Labeling RNAs in Live Cells Using Malachite Green Aptamer Scaffolds as Fluorescent Probes.

Science.gov (United States)

Yerramilli, V Siddartha; Kim, Kyung Hyuk

2018-03-16

RNAs mediate many different processes that are central to cellular function. The ability to quantify or image RNAs in live cells is very useful in elucidating such functions of RNA. RNA aptamer-fluorogen systems have been increasingly used in labeling RNAs in live cells. Here, we use the malachite green aptamer (MGA), an RNA aptamer that can specifically bind to malachite green (MG) dye and induces it to emit far-red fluorescence signals. Previous studies on MGA showed a potential for the use of MGA for genetically tagging other RNA molecules in live cells. However, these studies also exhibited low fluorescence signals and high background noise. Here we constructed and tested RNA scaffolds containing multiple tandem repeats of MGA as a strategy to increase the brightness of the MGA aptamer-fluorogen system as well as to make the system fluoresce when tagging various RNA molecules, in live cells. We demonstrate that our MGA scaffolds can induce fluorescence signals by up to ∼20-fold compared to the basal level as a genetic tag for other RNA molecules. We also show that our scaffolds function reliably as genetically encoded fluorescent tags for mRNAs of fluorescent proteins and other RNA aptamers.

Sheet Fluorescence and Annular Analysis of Ultracold Neutral Plasmas

International Nuclear Information System (INIS)

Castro, J.; Gao, H.; Killian, T. C.

2009-01-01

Annular analysis of fluorescence imaging measurements on Ultracold Neutral Plasmas (UNPs) is demonstrated. Spatially-resolved fluorescence imaging of the strontium ions produces a spectrum that is Doppler-broadened due to the thermal ion velocity and shifted due to the ion expansion velocity. The fluorescence excitation beam is spatially narrowed into a sheet, allowing for localized analysis of ion temperatures within a volume of the plasma with small density variation. Annular analysis of fluorescence images permits an enhanced signal-to-noise ratio compared to previous fluorescence measurements done in strontium UNPs. Using this technique and analysis, plasma ion temperatures are measured and shown to display characteristics of plasmas with strong coupling such as disorder induced heating and kinetic energy oscillations.

Monte Carlo simulation of zinc protoporphyrin fluorescence in the retina

Science.gov (United States)

Chen, Xiaoyan; Lane, Stephen

2010-02-01

We have used Monte Carlo simulation of autofluorescence in the retina to determine that noninvasive detection of nutritional iron deficiency is possible. Nutritional iron deficiency (which leads to iron deficiency anemia) affects more than 2 billion people worldwide, and there is an urgent need for a simple, noninvasive diagnostic test. Zinc protoporphyrin (ZPP) is a fluorescent compound that accumulates in red blood cells and is used as a biomarker for nutritional iron deficiency. We developed a computational model of the eye, using parameters that were identified either by literature search, or by direct experimental measurement to test the possibility of detecting ZPP non-invasively in retina. By incorporating fluorescence into Steven Jacques' original code for multi-layered tissue, we performed Monte Carlo simulation of fluorescence in the retina and determined that if the beam is not focused on a blood vessel in a neural retina layer or if part of light is hitting the vessel, ZPP fluorescence will be 10-200 times higher than background lipofuscin fluorescence coming from the retinal pigment epithelium (RPE) layer directly below. In addition we found that if the light can be focused entirely onto a blood vessel in the neural retina layer, the fluorescence signal comes only from ZPP. The fluorescence from layers below in this second situation does not contribute to the signal. Therefore, the possibility that a device could potentially be built and detect ZPP fluorescence in retina looks very promising.

Instrument for fluorescence sensing of circulating cells with diffuse light in mice in vivo.

Science.gov (United States)

Zettergren, Eric; Vickers, Dwayne; Runnels, Judith; Murthy, Shashi K; Lin, Charles P; Niedre, Mark

2012-03-01

Accurate quantification of circulating cell populations in mice is important in many areas of preclinical biomedical research. Normally, this is done either by extraction and analysis of small blood samples or, more recently, by using microscopy-based in vivo fluorescence flow cytometry. We describe a new technological approach to this problem using detection of diffuse fluorescent light from relatively large blood vessels in vivo. The diffuse fluorescence flow cytometer (DFFC) uses a laser to illuminate a mouse limb and an array of optical fibers coupled to a high-sensitivity photomultiplier tube array operating in photon counting mode to detect weak fluorescence signals from cells. We first demonstrate that the DFFC instrument is capable of detecting fluorescent microspheres and Vybrant-DiD-labeled cells in a custom-made optical flow phantom with similar size, optical properties, linear flow rates, and autofluorescence as a mouse limb. We also present preliminary data demonstrating that the DFFC is capable of detecting circulating cells in nude mice in vivo. In principle, this device would allow interrogation of the whole blood volume of a mouse in minutes, with sensitivity improvement by several orders of magnitude compared to current approaches. © 2012 Society of Photo-Optical Instrumentation Engineers (SPIE).

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...... 800, is uniformly embedded in a dielectric host that coats the nanoantennae. The nanoantennae act to enhance the dye absorption. In turn, emission from the dye drives the plasmon resonance of the antennae; the nanoantennae act to enhance the fluorescence signal and change the angular distribution...... of emission. These effects depend upon the overlap of the plasmon resonance with the excitation wavelength and the fluorescence emission band. A decreased fluorescence lifetime is observed along with highly polarized emission that displays the characteristics of the nanoantenna's dipole mode. Being able...

A High-Performance Fluorescence Immunoassay Based on the Relaxation of Quenching, Exemplified by Detection of Cardiac Troponin I

Directory of Open Access Journals (Sweden)

Seung-Wan Kim

2016-05-01

Full Text Available The intramolecular fluorescence self-quenching phenomenon is a major drawback in developing high-performance fluorometric biosensors which use common fluorophores as signal generators. We propose two strategies involving liberation of the fluorescent molecules by means of enzymatic fragmentation of protein or dehybridization of double-stranded DNA. In the former, bovine serum albumin (BSA was coupled with the fluorescent BODIPY dye (Red BSA, and then immobilized on a solid surface. When the insolubilized Red BSA was treated with proteinase K (10 units/mL for 30 min, the fluorescent signal was significantly increased (3.5-fold compared to the untreated control. In the second case, fluorophore-tagged DNA probes were linked to gold nanoparticles by hybridization with capture DNA strands densely immobilized on the surface. The quenched fluorescence signal was recovered (3.7-fold by thermal dehybridization, which was induced with light of a specific wavelength (e.g., 530 nm for less than 1 min. We next applied the Red BSA self-quenching relaxation technique employing enzymatic fragmentation to a high-performance immunoassay of cardiac troponin I (cTnI in a microtiter plate format. The detection limit was 0.19 ng/mL cTnI, and the fluorescent signal was enhanced approximately 4.1-fold compared with the conventional method of direct measurement of the fluorescent signal from a non-fragmented fluorophore-labeled antibody.

Nonlinear Vibration Signal Tracking of Large Offshore Bridge Stayed Cable Based on Particle Filter

Directory of Open Access Journals (Sweden)

Ye Qingwei

2015-12-01

Full Text Available The stayed cables are key stress components of large offshore bridge. The fault detection of stayed cable is very important for safe of large offshore bridge. A particle filter model and algorithm of nonlinear vibration signal are used in this paper. Firstly, the particle filter model of stayed cable of large offshore bridge is created. Nonlinear dynamic model of the stayed-cable and beam coupling system is dispersed in temporal dimension by using the finite difference method. The discrete nonlinear vibration equations of any cable element are worked out. Secondly, a state equation of particle filter is fitted by least square algorithm from the discrete nonlinear vibration equations. So the particle filter algorithm can use the accurate state equations. Finally, the particle filter algorithm is used to filter the vibration signal of bridge stayed cable. According to the particle filter, the de-noised vibration signal can be tracked and be predicted for a short time accurately. Many experiments are done at some actual bridges. The simulation experiments and the actual experiments on the bridge stayed cables are all indicating that the particle filter algorithm in this paper has good performance and works stably.

Fluorescence imaging as a diagnostic of M-band x-ray drive condition in hohlraum with fluorescent Si targets

International Nuclear Information System (INIS)

Li, Qi; Hu, Zhimin; Yao, Li; Huang, Chengwu; Yuan, Zheng; Zhao, Yang; Xiong, Gang; Qing, Bo; Lv, Min; Zhu, Tuo; Deng, Bo; Li, Jin; Wei, Minxi; Zhan, Xiayu; Li, Jun; Yang, Yimeng; Su, Chunxiao; Yang, Guohong; Zhang, Jiyan; Li, Sanwei

2017-01-01

Fluorescence imaging of surrogate Si-doped CH targets has been used to provide a measurement for drive condition of high-energy x-ray (i.e. M-band x-ray) drive symmetry upon the capsule in hohlraum on Shenguang-II laser facility. A series of experiments dedicated to the study of photo-pumping and fluorescence effect in Si-plasma are presented. To investigate the feasibility of fluorescence imaging in Si-plasma, an silicon plasma in Si-foil target is pre-formed at ground state by the soft x-ray from a half-hohlraum, which is then photo-pumped by the K-shell lines from a spatially distinct laser-produced Si-plasma. The resonant Si photon pump is used to improve the fluorescence signal and cause visible image in the Si-foil. Preliminary fluorescence imaging of Si-ball target is performed in both Si-doped and pure Au hohlraum. The usual capsule at the center of the hohlraum is replaced with a solid Si-doped CH-ball (Si-ball). Since the fluorescence is proportional to the photon pump upon the Si-plasma, high-energy x-ray drive symmetry is equal to the fluorescence distribution of the Si-ball. (paper)

Fluorescent sensing with Fresnel microlenses for optofluidic systems

Science.gov (United States)

Siudzińska, Anna; Miszczuk, Andrzej; Marczak, Jacek; Komorowska, Katarzyna

2017-05-01

The concept of fluorescent sensing in a microchannel equipped with focusing light Fresnel lenses has been demonstrated. The concept employs a line or array of Fresnel lenses generating a line or array of focused light spots within a microfluidic channel, to increase the sensitivity of fluorescent signal detection in the system. We have presented efficient methods of master mold fabrication based on the lithography method and focused ion beam milling. The flexible microchannel was fabricated by an imprint process with new thiolene-epoxy resin with a good ability to replicate even submicron-size features. For final imprinted lenses, the measured background to peak signal level shows more than nine times the increase in brightness at the center of the focal spot for the green part of the spectrum (532 nm). The effectiveness of the microlenses in fluorescent-marked Escherichia coli bacteria was confirmed in a basic fluoroscope experiment, showing the increase of the sensitivity of the detection by the order of magnitude.

A fluorescent biosensing platform based on the polydopamine nanospheres intergrating with Exonuclease III-assisted target recycling amplification.

Science.gov (United States)

Qiang, Weibing; Wang, Xi; Li, Wei; Chen, Xiang; Li, Hui; Xu, Danke

2015-09-15

Rapid, cost-effective, sensitive and specific analysis of biomolecules is important in the modern healthcare system. Here, a fluorescent biosensing platform based on the polydopamine nanospheres (PDANS) intergrating with Exonuclease III (Exo III) was developed. Due to the interaction between the ssDNA and the PDANS, the fluorescence of 6-carboxyfluorescein (FAM) labelled in the probe would been quenched by PDANS through FRET. While, in the present of the target DNA, the probe DNA would hybridize with the target DNA to form the double-strand DNA complex. Thus, Exo III could catalyze the stepwise removal of mononucleotides from 3'-terminus in the probe DNA, releasing the target DNA. As the FAM was released from the probe DNA, the fluorescence would no longer been quenched, led to the signal on. As one target DNA molecule could undergo a number of cycles to trigger the degradation of abundant probe DNA, Exo III-assisted target recycling would led to the amplification of the signal. The detection limit for DNA was 5 pM, which was 20 times lower than that without Exo III. And the assay time was largely shortened due to the faster signal recovery kinetics. What is more, this target recycling strategy was also applied to conduct an aptamer-based biosensing platform. The fluorescence intensity was also enhanced for the assay of adenosine triphosphate (ATP). For the Exo III-assisted target recycling amplification, DNA and ATP were fast detected with high sensitivity and selectivity. This work provides opportunities to develop simple, rapid, economical, and sensitive biosensing platforms for biomedical diagnostics. Copyright © 2015 Elsevier B.V. All rights reserved.

The Potential Applications of Real-Time Monitoring of Water Quality in a Large Shallow Lake (Lake Taihu, China) Using a Chromophoric Dissolved Organic Matter Fluorescence Sensor

Science.gov (United States)

Niu, Cheng; Zhang, Yunlin; Zhou, Yongqiang; Shi, Kun; Liu, Xiaohan; Qin, Boqiang

2014-01-01

This study presents results from field surveys performed over various seasons in a large, eutrophic, shallow lake (Lake Taihu, China) using an in situ chromophoric dissolved organic matter (CDOM) fluorescence sensor as a surrogate for other water quality parameters. These measurements identified highly significant empirical relationships between CDOM concentration measured using the in situ fluorescence sensor and CDOM absorption, fluorescence, dissolved organic carbon (DOC), chemical oxygen demand (COD) and total phosphorus (TP) concentrations. CDOM concentration expressed in quinine sulfate equivalent units, was highly correlated with the CDOM absorption coefficient (r2 = 0.80, p CDOM concentration measured using the in situ fluorescence sensor could act as a substitute for the CDOM absorption coefficient and fluorescence measured in the laboratory. Similarly, CDOM concentration was highly correlated with DOC concentration (r2 = 0.68, p CDOM fluorescence sensor measurements could be a proxy for DOC concentration. In addition, significant positive correlations were found between laboratory CDOM absorption coefficients and COD (r2 = 0.83, p CDOM fluorescence sensor. PMID:24984060

Simultaneous electrical and mechanical resonance drive for large signal amplification of micro resonators

KAUST Repository

Hasan, M. H.

2018-01-12

Achieving large signal-noise ratio using low levels of excitation signal is key requirement for practical applications of micro and nano electromechanical resonators. In this work, we introduce the double electromechanical resonance drive concept to achieve an order-of-magnitude dynamic signal amplification in micro resonators. The concept relies on simultaneously activating the micro-resonator mechanical and electrical resonance frequencies. We report an input voltage amplification up to 15 times for a micro-resonator when its electrical resonance is tuned to match the mechanical resonance that leads to dynamic signal amplification in air (Quality factor enhancement). Furthermore, using a multi-frequency excitation technique, input voltage and vibrational amplification of up to 30 times were shown for the same micro-resonator while relaxing the need to match its mechanical and electrical resonances.

Simultaneous electrical and mechanical resonance drive for large signal amplification of micro resonators

KAUST Repository

Hasan, M. H.; Alsaleem, F. M.; Jaber, Nizar; Hafiz, Md Abdullah Al; Younis, Mohammad I.

2018-01-01

Achieving large signal-noise ratio using low levels of excitation signal is key requirement for practical applications of micro and nano electromechanical resonators. In this work, we introduce the double electromechanical resonance drive concept to achieve an order-of-magnitude dynamic signal amplification in micro resonators. The concept relies on simultaneously activating the micro-resonator mechanical and electrical resonance frequencies. We report an input voltage amplification up to 15 times for a micro-resonator when its electrical resonance is tuned to match the mechanical resonance that leads to dynamic signal amplification in air (Quality factor enhancement). Furthermore, using a multi-frequency excitation technique, input voltage and vibrational amplification of up to 30 times were shown for the same micro-resonator while relaxing the need to match its mechanical and electrical resonances.

Quantitative laser-induced fluorescence measurements of nitric oxide in a heavy-duty Diesel engine

NARCIS (Netherlands)

Verbiezen, K.; Klein-Douwel, R. J. H.; van Viet, A. P.; Donkerbroek, A. J.; Meerts, W. L.; Dam, N. J.; ter Meulen, J. J.

2007-01-01

We present quantitative, in-cylinder, UV-laser-induced fluorescence measurements of nitric oxide in a heavy-duty Diesel engine. Processing of the raw fluorescence signals includes a detailed correction, based on additional measurements, for the effect of laser beam and fluorescence attenuation, and

A PDMS-based cylindrical hybrid lens for enhanced fluorescence detection in microfluidic systems.

Science.gov (United States)

Lin, Bor-Shyh; Yang, Yu-Ching; Ho, Chong-Yi; Yang, Han-Yu; Wang, Hsiang-Yu

2014-02-13

Microfluidic systems based on fluorescence detection have been developed and applied for many biological and chemical applications. Because of the tiny amount of sample in the system; the induced fluorescence can be weak. Therefore, most microfluidic systems deploy multiple optical components or sophisticated equipment to enhance the efficiency of fluorescence detection. However, these strategies encounter common issues of complex manufacturing processes and high costs. In this study; a miniature, cylindrical and hybrid lens made of polydimethylsiloxane (PDMS) to improve the fluorescence detection in microfluidic systems is proposed. The hybrid lens integrates a laser focusing lens and a fluorescence collecting lens to achieve dual functions and simplify optical setup. Moreover, PDMS has advantages of low-cost and straightforward fabrication compared with conventional optical components. The performance of the proposed lens is first examined with two fluorescent dyes and the results show that the lens provides satisfactory enhancement for fluorescence detection of Rhodamine 6G and Nile Red. The overall increments in collected fluorescence signal and detection sensitivity are more than 220% of those without lens, and the detection limits of Rhodamine 6G and Nile red are lowered to 0.01 μg/mL and 0.05 μg/mL, respectively. The hybrid lens is further applied to the detection of Nile red-labeled Chlorella vulgaris cells and it increases both signal intensity and detection sensitivity by more than 520%. The proposed hybrid lens also dramatically reduces the variation in detected signal caused by the deviation in incident angle of excitation light.

Integrated three-dimensional optical MEMS for chip-based fluorescence detection

Science.gov (United States)

Hung, Kuo-Yung; Tseng, Fan-Gang; Khoo, Hwa-Seng

2009-04-01

This paper presents a novel fluorescence sensing chip for parallel protein microarray detection in the context of a 3-in-1 protein chip system. This portable microchip consists of a monolithic integration of CMOS-based avalanche photo diodes (APDs) combined with a polymer micro-lens, a set of three-dimensional (3D) inclined mirrors for separating adjacent light signals and a low-noise transformer-free dc-dc boost mini-circuit to power the APDs (ripple below 1.28 mV, 0-5 V input, 142 V and 12 mA output). We fabricated our APDs using the planar CMOS process so as to facilitate the post-CMOS integration of optical MEMS components such as the lenses. The APD arrays were arranged in unique circular patterns appropriate for detecting the specific fluorescently labelled protein spots in our study. The array-type APDs were designed so as to compensate for any alignment error as detected by a positional error signal algorithm. The condenser lens was used as a structure for light collection to enhance the fluorescent signals by about 25%. This element also helped to reduce the light loss due to surface absorption. We fabricated an inclined mirror to separate two adjacent fluorescent signals from different specimens. Excitation using evanescent waves helped reduce the interference of the excitation light source. This approach also reduced the number of required optical lenses and minimized the complexity of the structural design. We achieved detection floors for anti-rabbit IgG and Cy5 fluorescent dye as low as 0.5 ng/µl (~3.268 nM). We argue that the intrinsic nature of point-to-point and batch-detection methods as showcased in our chip offers advantages over the serial-scanning approach used in traditional scanner systems. In addition, our system is low cost and lightweight.

Using Dual Fluorescence Reporting Genes to Establish an In Vivo Imaging Model of Orthotopic Lung Adenocarcinoma in Mice.

Science.gov (United States)

Lai, Cheng-Wei; Chen, Hsiao-Ling; Yen, Chih-Ching; Wang, Jiun-Long; Yang, Shang-Hsun; Chen, Chuan-Mu

2016-12-01

Lung adenocarcinoma is characterized by a poor prognosis and high mortality worldwide. In this study, we purposed to use the live imaging techniques and a reporter gene that generates highly penetrative near-infrared (NIR) fluorescence to establish a preclinical animal model that allows in vivo monitoring of lung cancer development and provides a non-invasive tool for the research on lung cancer pathogenesis and therapeutic efficacy. A human lung adenocarcinoma cell line (A549), which stably expressed the dual fluorescence reporting gene (pCAG-iRFP-2A-Venus), was used to generate subcutaneous or orthotopic lung cancer in nude mice. Cancer development was evaluated by live imaging via the NIR fluorescent signals from iRFP, and the signals were verified ex vivo by the green fluorescence of Venus from the gross lung. The tumor-bearing mice received miR-16 nucleic acid therapy by intranasal administration to demonstrate therapeutic efficacy in this live imaging system. For the subcutaneous xenografts, the detection of iRFP fluorescent signals revealed delicate changes occurring during tumor growth that are not distinguishable by conventional methods of tumor measurement. For the orthotopic xenografts, the positive correlation between the in vivo iRFP signal from mice chests and the ex vivo green fluorescent signal from gross lung tumors and the results of the suppressed tumorigenesis by miR-16 treatment indicated that lung tumor size can be accurately quantified by the emission of NIR fluorescence. In addition, orthotopic lung tumor localization can be accurately visualized using iRFP fluorescence tomography in vivo, thus revealing the trafficking of lung tumor cells. We introduced a novel dual fluorescence lung cancer model that provides a non-invasive option for preclinical research via the use of NIR fluorescence in live imaging of lung.

A fluorescence detection of D-penicillamine based on Cu(2+)-induced fluorescence quenching system of protein-stabilized gold nanoclusters.

Science.gov (United States)

Wang, Peng; Li, Bang Lin; Li, Nian Bing; Luo, Hong Qun

2015-01-25

In this contribution, a luminescent gold nanoclusters which were synthesized by bovine serum albumin as novel fluorescent probes were successfully utilized for the determination of D-penicillamine for the first time. Cupric ion was employed to quench the strong fluorescence of the gold nanoclusters, whereas the addition of D-penicillamine caused obvious restoration of fluorescence intensity of the Cu(2+)-gold nanoclusters system. Under optimum conditions, the increment in fluorescence intensity of Cu(2+)-gold nanoclusters system caused by D-penicillamine was linearly proportional to the concentration of D-penicillamine in the range of 2.0×10(-5)-2.39×10(-4) M. The detection limit for D-penicillamine was 5.4×10(-6) M. With the off-on fluorescence signal at 650 nm approaching the near-infrared region, the present sensor for D-penicillamine detection had high sensitivity and low spectral interference. Furthermore, the novel gold nanoclusters-based fluorescent sensor has been applied to the determination of D-penicillamine in real biological samples with satisfactory results. Copyright © 2014 Elsevier B.V. All rights reserved.

Design of remote laser-induced fluorescence system's acquisition circuit

Science.gov (United States)

Wang, Guoqing; Lou, Yue; Wang, Ran; Yan, Debao; Li, Xin; Zhao, Xin; Chen, Dong; Zhao, Qi

2017-10-01

Laser-induced fluorescence system(LIfS) has been found its significant application in identifying one kind of substance from another by its properties even it's thimbleful, and becomes useful in plenty of fields. Many superior works have reported LIfS' theoretical analysis , designs and uses. However, the usual LIPS is always constructed in labs to detect matter quite closely, for the system using low-power laser as excitation source and charge coupled device (CCD) as detector. Promoting the detectivity of LIfS is of much concern to spread its application. Here, we take a high-energy narrow-pulse laser instead of commonly used continuous wave laser to operate sample, thus we can get strong fluorescent. Besides, photomultiplier (PMT) with high sensitivity is adopted in our system to detect extremely weak fluorescence after a long flight time from the sample to the detector. Another advantage in our system, as the fluorescence collected into spectroscopy, multiple wavelengths of light can be converted to the corresponding electrical signals with the linear array multichannel PMT. Therefore, at the cost of high-powered incentive and high-sensitive detector, a remote LIFS is get. In order to run this system, it is of importance to turn light signal to digital signal which can be processed by computer. The pulse width of fluorescence is deeply associated with excitation laser, at the nanosecond(ns) level, which has a high demand for acquisition circuit. We design an acquisition circuit including, I/V conversion circuit, amplifying circuit and peak-holding circuit. The simulation of circuit shows that peak-holding circuit can be one effective approach to reducing difficulty of acquisition circuit.

Boronic acid recognition of non-interacting carbohydrates for biomedical applications: increasing fluorescence signals of minimally interacting aldoses and sucralose.

Science.gov (United States)

Resendez, Angel; Halim, Md Abdul; Singh, Jasmeet; Webb, Dominic-Luc; Singaram, Bakthan

2017-11-22

To address carbohydrates that are commonly used in biomedical applications with low binding affinities for boronic acid based detection systems, two chemical modification methods were utilized to increase sensitivity. Modified carbohydrates were analyzed using a two component fluorescent probe based on boronic acid-appended viologen-HPTS (4,4'-o-BBV). Carbohydrates normally giving poor signals (fucose, l-rhamnose, xylose) were subjected to sodium borohydride (NaBH 4 ) reduction in ambient conditions for 1 h yielding the corresponding sugar alcohols from fucose, l-rhamnose and xylose in essentially quantitative yields. Compared to original aldoses, apparent binding affinities were increased 4-25-fold. The chlorinated sweetener and colon permeability marker sucralose (Splenda), otherwise undetectable by boronic acids, was dechlorinated to a detectable derivative by reactive oxygen and hydroxide intermediates by the Fenton reaction or by H 2 O 2 and UV light. This method is specific to sucralose as other common sugars, such as sucrose, do not contain any carbon-chlorine bonds. Significant fluorescence response was obtained for chemically modified sucralose with the 4,4'-o-BBV-HPTS probe system. This proof of principle can be applied to biomedical applications, such as gut permeability, malabsorption, etc.

Ns-scaled time-gated fluorescence lifetime imaging for forensic document examination

Science.gov (United States)

Zhong, Xin; Wang, Xinwei; Zhou, Yan

2018-01-01

A method of ns-scaled time-gated fluorescence lifetime imaging (TFLI) is proposed to distinguish different fluorescent substances in forensic document examination. Compared with Video Spectral Comparator (VSC) which can examine fluorescence intensity images only, TFLI can detect questioned documents like falsification or alteration. TFLI system can enhance weak signal by accumulation method. The two fluorescence intensity images of the interval delay time tg are acquired by ICCD and fitted into fluorescence lifetime image. The lifetimes of fluorescence substances are represented by different colors, which make it easy to detect the fluorescent substances and the sequence of handwritings. It proves that TFLI is a powerful tool for forensic document examination. Furthermore, the advantages of TFLI system are ns-scaled precision preservation and powerful capture capability.

Hybridization chain reaction amplification for highly sensitive fluorescence detection of DNA with dextran coated microarrays.

Science.gov (United States)

Chao, Jie; Li, Zhenhua; Li, Jing; Peng, Hongzhen; Su, Shao; Li, Qian; Zhu, Changfeng; Zuo, Xiaolei; Song, Shiping; Wang, Lianhui; Wang, Lihua

2016-07-15

Microarrays of biomolecules hold great promise in the fields of genomics, proteomics, and clinical assays on account of their remarkably parallel and high-throughput assay capability. However, the fluorescence detection used in most conventional DNA microarrays is still limited by sensitivity. In this study, we have demonstrated a novel universal and highly sensitive platform for fluorescent detection of sequence specific DNA at the femtomolar level by combining dextran-coated microarrays with hybridization chain reaction (HCR) signal amplification. Three-dimensional dextran matrix was covalently coated on glass surface as the scaffold to immobilize DNA recognition probes to increase the surface binding capacity and accessibility. DNA nanowire tentacles were formed on the matrix surface for efficient signal amplification by capturing multiple fluorescent molecules in a highly ordered way. By quantifying microscopic fluorescent signals, the synergetic effects of dextran and HCR greatly improved sensitivity of DNA microarrays, with a detection limit of 10fM (1×10(5) molecules). This detection assay could recognize one-base mismatch with fluorescence signals dropped down to ~20%. This cost-effective microarray platform also worked well with samples in serum and thus shows great potential for clinical diagnosis. Copyright © 2016 Elsevier B.V. All rights reserved.

Breaking the color barrier - a multi-selective antibody reporter offers innovative strategies of fluorescence detection.

Science.gov (United States)

Gallo, Eugenio; Jarvik, Jonathan W

2017-08-01

A novel bi-partite fluorescence platform exploits the high affinity and selectivity of antibody scaffolds to capture and activate small-molecule fluorogens. In this report, we investigated the property of multi-selectivity activation by a single antibody against diverse cyanine family fluorogens. Our fluorescence screen identified three cell-impermeant fluorogens, each with unique emission spectra (blue, green and red) and nanomolar affinities. Most importantly, as a protein fusion tag to G-protein-coupled receptors, the antibody biosensor retained full activity - displaying bright fluorogen signals with minimal background on live cells. Because fluorogen-activating antibodies interact with their target ligands via non-covalent interactions, we were able to perform advanced multi-color detection strategies on live cells, previously difficult or impossible with conventional reporters. We found that by fine-tuning the concentrations of the different color fluorogen molecules in solution, a user may interchange the fluorescence signal (onset versus offset), execute real-time signal exchange via fluorogen competition, measure multi-channel fluorescence via co-labeling, and assess real-time cell surface receptor traffic via pulse-chase experiments. Thus, here we inform of an innovative reporter technology based on tri-color signal that allows user-defined fluorescence tuning in live-cell applications. © 2017. Published by The Company of Biologists Ltd.

Beveled fiber-optic probe couples a ball lens for improving depth-resolved fluorescence measurements of layered tissue: Monte Carlo simulations

International Nuclear Information System (INIS)

Jaillon, Franck; Zheng Wei; Huang Zhiwei

2008-01-01

In this study, we evaluate the feasibility of designing a beveled fiber-optic probe coupled with a ball lens for improving depth-resolved fluorescence measurements of epithelial tissue using Monte Carlo (MC) simulations. The results show that by using the probe configuration with a beveled tip collection fiber and a flat tip excitation fiber associated with a ball lens, discrimination of fluorescence signals generated in different tissue depths is achievable. In comparison with a flat-tip collection fiber, the use of a large bevel angled collection fiber enables a better differentiation between the shallow and deep tissue layers by changing the excitation-collection fiber separations. This work suggests that the beveled fiber-optic probe coupled with a ball lens has the potential to facilitate depth-resolved fluorescence measurements of epithelial tissues

Clinical application of fluorescence in situ hybridization for prenatal diagnosis

Directory of Open Access Journals (Sweden)

Shu-fang JIANG

2012-07-01

Full Text Available Objective To establish and optimize the procedures of fluorescence in situ hybridization（FISH）, and evaluate its clinical value in rapid prenatal diagnosis of fetal numerical abnormality of chromosomes 21, 18, 13, X, Y. Methods Amniotic fluid or fetal blood was sampled by routine invasive procedures. After the amniotic fluid cells or fetal blood cells were separated and sequentially processed with hypotonic solution, fixation solution, smear and high temperature, they were hybridized in situ with two panels of specific fluorescence probes to detect numerical abnormality of chromosomes 21, 18, 13, X, Y. All the samples were also cultured and analyzed for their karyotype by conventional methods. Results When it was used as a diagnostic criterion of chromosomal number that the fluorescence signals were observed in ≥90% cells, GLP 13/GLP 21 probe panel showed 2 green/2 red fluorescence signals and CSP18/CSP X/CSP Y probe panel showed 2 blue/2 yellow (female or 2 blue/1 yellow/1 red fluorescence signals (male under normal condition. The test reports of all 196 cases were sent out in 72-96 hours, and 7 cases of Down syndrome, 2 cases of trisomy 18 and 1 case of sex chromosomal numerical abnormality were detected, which were accordant with karyotype analysis results reported one month later. Conclusions FISH has potential for clinical application, and is applicable to rapid prenatal diagnosis of fetal numerical abnormality of chromosomes 21, 18, 13, X, Y. The rapid FISH, together with conventional karyotyping, offer a valuable means for prenatal diagnosis of fetal aneuploidies.

Fluorogen-activating proteins: beyond classical fluorescent proteins

Directory of Open Access Journals (Sweden)

Shengnan Xu

2018-05-01

Full Text Available Fluorescence imaging is a powerful technique for the real-time noninvasive monitoring of protein dynamics. Recently, fluorogen activating proteins (FAPs/fluorogen probes for protein imaging were developed. Unlike the traditional fluorescent proteins (FPs, FAPs do not fluoresce unless bound to their specific small-molecule fluorogens. When using FAPs/fluorogen probes, a washing step is not required for the removal of free probes from the cells, thus allowing rapid and specific detection of proteins in living cells with high signal-to-noise ratio. Furthermore, with different fluorogens, living cell multi-color proteins labeling system was developed. In this review, we describe about the discovery of FAPs, the design strategy of FAP fluorogens, the application of the FAP technology and the advances of FAP technology in protein labeling systems. KEY WORDS: Fluorogen activating proteins, Fluorogens, Genetically encoded sensors, Fluorescence imaging, Molecular imaging

Evaluation of a fluorescence-based method for antibabesial drug screening.

Science.gov (United States)

Guswanto, Azirwan; Sivakumar, Thillaiampalam; Rizk, Mohamed Abdo; Elsayed, Shimaa Abd Elsalam; Youssef, Mohamed Ahmed; ElSaid, ElSaid El Shirbini; Yokoyama, Naoaki; Igarashi, Ikuo

2014-08-01

In vitro evaluation of chemotherapeutic agents against Babesia and Theileria parasites has become routine, and the effectiveness of these chemicals is usually determined by comparing the parasitemia dynamics of untreated and treated parasites. Although microscopy is widely used to calculate parasitemia, several disadvantages are associated with this technique. The present study evaluated a fluorescence-based method using SYBR green I stain (SG I) to screen antibabesial agents in in vitro cultures of Babesia bovis. The linearity between relative fluorescence units (RFU) and parasitemia was found to be well correlated with a 0.9944 goodness-of-fit (r(2)) value. Subsequently, 50% inhibitory concentration (IC50) values were calculated for 3 antiprotozoan agents, diminazene aceturate, nimbolide, and gedunin, by this method. For diminazene aceturate and nimbolide, the IC(50)s determined by the fluorescence-based method (408 nM and 8.13 μM, respectively) and microscopy (400.3 nM and 9.4 μM, respectively) were in agreement. Furthermore, the IC50 of gedunin determined by the fluorescence-based method (19 μM) was similar to the recently described microscopy-based value (21.7 μM) for B. bovis. Additionally, the Z' factor (0.80 to 0.90), signal-to-noise (S/N) ratio (44.15 to 87.64), coefficient of variation at the maximum signal (%CVmax) (0.50 to 2.85), and coefficient of variation at the minimum signal (%CVmin) (1.23 to 2.21) calculated for the fluorescence method using diminazene aceturate were comparable to those previously determined in malaria research for this assay. These findings suggest that the fluorescence-based method might be useful for antibabesial drug screening and may have potential to be developed into a high-throughput screening (HTS) assay. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Microbubble embedded with upconversion nanoparticles as a bimodal contrast agent for fluorescence and ultrasound imaging

International Nuclear Information System (INIS)

Jin, Birui; Lin, Min; You, Minli; Xu, Feng; Lu, Tianjian; Zong, Yujin; Wan, Mingxi; Duan, Zhenfeng

2015-01-01

Bimodal imaging offers additional imaging signal thus finds wide spread application in clinical diagnostic imaging. Fluorescence/ultrasound bimodal imaging contrast agent using fluorescent dyes or quantum dots for fluorescence signal has emerged as a promising method, which however requires visible light or UV irradiation resulting in photobleaching, photoblinking, auto-fluorescence and limited tissue penetration depth. To surmount these problems, we developed a novel bimodal contrast agent using layer-by-layer assembly of upconversion nanoparticles onto the surface of microbubbles. The resulting microbubbles with average size of 2 μm provide enhanced ultrasound echo for ultrasound imaging and upconversion emission upon near infrared irradiation for fluorescence imaging. The developed bimodal contrast agent holds great potential to be applied in ultrasound target technique for targeted diseases diagnostics and therapy. (paper)

Visualization of mcr mRNA in a methanogen by fluorescence in situ hybridization with an oligonucleotide probe and two-pass tyramide signal amplification (two-pass TSA-FISH).

Science.gov (United States)

Kubota, Kengo; Ohashi, Akiyoshi; Imachi, Hiroyuki; Harada, Hideki

2006-09-01

Two-pass tyramide signal amplification-fluorescence in situ hybridization (two-pass TSA-FISH) with a horseradish peroxidase (HRP)-labeled oligonucleotide probe was applied to detect prokaryotic mRNA. In this study, mRNA of a key enzyme for methanogenesis, methyl coenzyme M reductase (mcr), in Methanococcus vannielii was targeted. Applicability of mRNA-targeted probes to in situ hybridization was verified by Clone-FISH. It was observed that sensitivity of two-pass TSA-FISH was significantly higher than that of TSA-FISH, which was further increased by the addition of dextran sulphate in TSA working solution. Signals from two-pass TSA-FISH were more reliable compared to the weak, spotty signals yielded by TSA-FISH.

Click chemistry for the conservation of cellular structures and fluorescent proteins: ClickOx.

Science.gov (United States)

Löschberger, Anna; Niehörster, Thomas; Sauer, Markus

2014-05-01

Reactive oxygen species (ROS), including hydrogen peroxide, are known to cause structural damage not only in living, but also in fixed, cells. Copper-catalyzed azide-alkyne cycloaddition (click chemistry) is known to produce ROS. Therefore, fluorescence imaging of cellular structures, such as the actin cytoskeleton, remains challenging when combined with click chemistry protocols. In addition, the production of ROS substantially weakens the fluorescence signal of fluorescent proteins. This led us to develop ClickOx, which is a new click chemistry protocol for improved conservation of the actin structure and better conservation of the fluorescence signal of green fluorescent protein (GFP)-fusion proteins. Herein we demonstrate that efficient oxygen removal by addition of an enzymatic oxygen scavenger system (ClickOx) considerably reduces ROS-associated damage during labeling of nascent DNA with ATTO 488 azide by Cu(I)-catalyzed click chemistry. Standard confocal and super-resolution fluorescence images of phalloidin-labeled actin filaments and GFP/yellow fluorescent protein-labeled cells verify the conservation of the cytoskeleton microstructure and fluorescence intensity, respectively. Thus, ClickOx can be used advantageously for structure preservation in conventional and most notably in super-resolution microscopy methods. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Developing LED UV fluorescence sensors for online monitoring DOM and predicting DBPs formation potential during water treatment.

Science.gov (United States)

Li, Wen-Tao; Jin, Jing; Li, Qiang; Wu, Chen-Fei; Lu, Hai; Zhou, Qing; Li, Ai-Min

2016-04-15

Online monitoring dissolved organic matter (DOM) is urgent for water treatment management. In this study, high performance size exclusion chromatography with multi-UV absorbance and multi-emission fluorescence scans were applied to spectrally characterize samples from 16 drinking water sources across Yangzi River and Huai River Watersheds. The UV absorbance indices at 254 nm and 280 nm referred to the same DOM components and concentration, and the 280 nm UV light could excite both protein-like and humic-like fluorescence. Hence a novel UV fluorescence sensor was developed out using only one UV280 light-emitting diode (LED) as light source. For all samples, enhanced coagulation was mainly effective for large molecular weight biopolymers; while anion exchange further substantially removed humic substances. During chlorination tests, UVA280 and UVA254 showed similar correlations with yields of disinfection byproducts (DBPs); the humic-like fluorescence obtained from LED sensors correlated well with both trihalomethanes and haloacetic acids yields, while the correlation between protein-like fluorescence and trihalomethanes was relatively poor. Anion exchange exhibited more reduction of DBPs yields as well as UV absorbance and fluorescence signals than enhanced coagulation. The results suggest that the LED UV fluorescence sensors are very promising for online monitoring DOM and predicting DBPs formation potential during water treatment. Copyright © 2016 Elsevier Ltd. All rights reserved.

Fluorescence suppression using wavelength modulated Raman spectroscopy in fiber-probe-based tissue analysis.

Science.gov (United States)

Praveen, Bavishna B; Ashok, Praveen C; Mazilu, Michael; Riches, Andrew; Herrington, Simon; Dholakia, Kishan

2012-07-01

In the field of biomedical optics, Raman spectroscopy is a powerful tool for probing the chemical composition of biological samples. In particular, fiber Raman probes play a crucial role for in vivo and ex vivo tissue analysis. However, the high-fluorescence background typically contributed by the auto fluorescence from both a tissue sample and the fiber-probe interferes strongly with the relatively weak Raman signal. Here we demonstrate the implementation of wavelength-modulated Raman spectroscopy (WMRS) to suppress the fluorescence background while analyzing tissues using fiber Raman probes. We have observed a significant signal-to-noise ratio enhancement in the Raman bands of bone tissue, which have a relatively high fluorescence background. Implementation of WMRS in fiber-probe-based bone tissue study yielded usable Raman spectra in a relatively short acquisition time (∼30  s), notably without any special sample preparation stage. Finally, we have validated its capability to suppress fluorescence on other tissue samples such as adipose tissue derived from four different species.

Fluorescence-Doped Particles for Simultaneous Temperature and Velocity Imaging

Science.gov (United States)

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

2012-01-01

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

Fluorescent quenching immune chromatographic strips with quantum dots and upconversion nanoparticles as fluorescent donors for visual detection of sulfaquinoxaline in foods of animal origin

International Nuclear Information System (INIS)

Hu, Gaoshuang; Sheng, Wei; Li, Jingmin; Zhang, Yan; Wang, Junping; Wang, Shuo

2017-01-01

In this study, two novel fluorescence quenching immune chromatographic strips (FQICS) were developed to detect sulfaquinoxaline (SQX) in foods of animal origin. These proposed FQICSs were based on fluorescence resonance energy transfer (FRET) from fluorescence donors (quantum dots or upconversion nanoparticles) to fluorescence acceptors (colloidal gold nanoparticles). Compared with traditional colloidal gold-based immune chromatographic strips (ICS), these FQICSs showed positive correlation between the fluorescent signals and the targets, and allowed user to get test results from weak fluorescent signals. The visual detection limits of these two FQICSs were both 1 ng mL −1 in standard solution and 8 μg kg −1 in samples, while the visual detection limit of the colloidal gold-based ICS was 10 ng mL −1 in standard solution and 80 μg kg −1 in samples. Besides, the results we obtained by the use of FQICS showed high agreement with those obtained by the use of commercial ELISA kits, indicating the good accuracy of these strips. As a conclusion, these proposed FQICS based on quantum dots and upconversion nanoparticles can be applied in sensitive, rapid and on-site detection of SQX in foods of animal origin. - Highlights: • Two novel FQICS based on FRET were developed for the first time. • QDs and UCNPs were used as fluorescent donors in the FQICS. • The proposed FQICS showed low LOD compared with traditional ICS. • The proposed FQICS were applied in real samples analysis. • The proposed FQICS were verified by commercial ELISA kits.

Research on a Small Signal Stability Region Boundary Model of the Interconnected Power System with Large-Scale Wind Power

Directory of Open Access Journals (Sweden)

Wenying Liu

2015-03-01

Full Text Available For the interconnected power system with large-scale wind power, the problem of the small signal stability has become the bottleneck of restricting the sending-out of wind power as well as the security and stability of the whole power system. Around this issue, this paper establishes a small signal stability region boundary model of the interconnected power system with large-scale wind power based on catastrophe theory, providing a new method for analyzing the small signal stability. Firstly, we analyzed the typical characteristics and the mathematic model of the interconnected power system with wind power and pointed out that conventional methods can’t directly identify the topological properties of small signal stability region boundaries. For this problem, adopting catastrophe theory, we established a small signal stability region boundary model of the interconnected power system with large-scale wind power in two-dimensional power injection space and extended it to multiple dimensions to obtain the boundary model in multidimensional power injection space. Thirdly, we analyzed qualitatively the topological property’s changes of the small signal stability region boundary caused by large-scale wind power integration. Finally, we built simulation models by DIgSILENT/PowerFactory software and the final simulation results verified the correctness and effectiveness of the proposed model.

Benzothiadiazole Derivatives as Fluorescence Imaging Probes: Beyond Classical Scaffolds.

Science.gov (United States)

Neto, Brenno A D; Carvalho, Pedro H P R; Correa, Jose R

2015-06-16

This Account describes the origins, features, importance, and trends of the use of fluorescent small-molecule 2,1,3-benzothiadiazole (BTD) derivatives as a new class of bioprobes applied to bioimaging analyses of several (live and fixed) cell types. BTDs have been successfully used as probes for a plethora of biological analyses for only a few years, and the impressive responses obtained by using this important class of heterocycle are fostering the development of new fluorescent BTDs and expanding the biological applications of such derivatives. The first use of a fluorescent small-molecule BTD derivative as a selective cellular probe dates back to 2010, and since then impressive advances have been described by us and others. The well-known limitations of classical scaffolds urged the development of new classes of bioprobes. Although great developments have been achieved by using classical scaffolds such as coumarins, BODIPYs, fluoresceins, rhodamines, cyanines, and phenoxazines, there is still much to be done, and BTDs aim to succeed where these dyes have shown their limitations. Important organelles and cell components such as nuclear DNA, mitochondria, lipid droplets, and others have already been successfully labeled by fluorescent small-molecule BTD derivatives. New technological systems that use BTDs as the fluorophores for bioimaging experiments have been described in recent scientific literature. The successful application of BTDs as selective bioprobes has led some groups to explore their potential for use in studying membrane pores or tumor cells under hypoxic conditions. Finally, BTDs have also been used as fluorescent tags to investigate the action mechanism of some antitumor compounds. The attractive photophysical data typically observed for π-extended BTD derivatives is fostering interest in the use of this new class of bioprobes. Large Stokes shifts, large molar extinction coefficients, high quantum yields, high stability when stored in solution or

WellReader: a MATLAB program for the analysis of fluorescence and luminescence reporter gene data.

Science.gov (United States)

Boyer, Frédéric; Besson, Bruno; Baptist, Guillaume; Izard, Jérôme; Pinel, Corinne; Ropers, Delphine; Geiselmann, Johannes; de Jong, Hidde

2010-05-01

Fluorescent and luminescent reporter gene systems in combination with automated microplate readers allow real-time monitoring of gene expression on the population level at high precision and sampling density. This generates large amounts of data for the analysis of which computer tools are missing to date. We have developed WellReader, a MATLAB program for the analysis of fluorescent and luminescent reporter gene data. WellReader allows the user to load the output files of microplate readers, remove outliers, correct for background effects and smooth and fit the data. Moreover, it computes biologically relevant quantities from the measured signals, notably promoter activities and protein concentrations, and compares the resulting expression profiles of different genes under different conditions. WellReader is available under a LGPL licence at http://prabi1.inrialpes.fr/trac/wellreader.

Development of a fluorescent microscope combined with a real-time autoradiography system

International Nuclear Information System (INIS)

Rai, Hiroki; Kanno, Satomi; Hayashi, Yoshitake; Nihei, Naoto; Nakanishi, Tomoko M.

2008-01-01

For combination with microscope, we developed real-time autoradiography system for micro-scale analysis with adjustment of the CsI(Ti) scintillator thickness for higher resolution and applying tapered fiber optic plate for magnification of autoradiograph image. We combined real-time autoradiography system with an inverted fluorescent microscope so that an autoradiograph image as well as fluorescent image, bright-field image can be acquired at the same time. In the case of observation of sliced soybean stalk traced 45 CaCl, the fluorescent and bright-field image was acquired which magnified to 50 times, the autoradiograph image of 45 Ca distribution in the tissue was acquired in almost same scale. The new microscopic system which can acquire autoradiograph image of labeled signals (low molecular weight) is expected to develop the signal transduction study and gene expression, combined with fluorescent protein techniques such as GFP etc. (author)

Corrections of residual fluorescence distortions for a glancing-emergence-angle x-ray-absorption technique

International Nuclear Information System (INIS)

Brewe, D.L.; Pease, D.M.; Budnick, J.I.

1994-01-01

Distortions appear in x-ray-absorption spectra obtained by monitoring the fluorescence from thick samples with concentrated absorbing species. The glancing-emergence-angle technique for obtaining spectra from this type of sample eliminates distortions from the measured spectra by monitoring the fluorescence leaving the sample at a small angle relative to the sample surface. This technique is limited by the small signal available from the inherently limited detector solid angle. In addition, no precise estimate of the required restriction on maximum emergent angle θ max has been available. We have calculated residual extended x-ray-absorption fine structure distortions as a function of θ max , and performed experimental tests of the calculations. These calculations provide a means to estimate the required detector geometry for negligible distortions, or alternatively, allow the use of a larger θ max , increasing the available signal, with the remaining residual distortions removed by application of the calculations. The calculations are also applicable to other detector geometries, and account for detectors subtending a large solid angle by an integration over the subtended angle. This represents an improvement over previous calculations. The application to more general detector configurations is also discussed

Femtosecond, two-photon laser-induced-fluorescence imaging of atomic oxygen in an atmospheric-pressure plasma jet

Science.gov (United States)

Schmidt, Jacob B.; Sands, Brian L.; Kulatilaka, Waruna D.; Roy, Sukesh; Scofield, James; Gord, James R.

2015-06-01

Femtosecond, two-photon-absorption laser-induced-fluorescence (fs-TALIF) spectroscopy is employed to measure space- and time-resolved atomic-oxygen distributions in a nanosecond, repetitively pulsed, externally grounded, atmospheric-pressure plasma jet flowing helium with a variable oxygen admixture. The high-peak-intensity, low-average-energy femtosecond pulses result in increased TALIF signal with reduced photolytic inferences. This allows 2D imaging of absolute atomic-oxygen number densities ranging from 5.8   ×   1015 to 2.0   ×   1012cm-3 using a cooled CCD with an external intensifier. Xenon is used for signal and imaging-system calibrations to quantify the atomic-oxygen fluorescence signal. Initial results highlight a transition in discharge morphology from annular to filamentary, corresponding with a change in plasma chemistry from ozone to atomic oxygen production, as the concentration of oxygen in the feed gas is changed at a fixed voltage-pulse-repetition rate. In this configuration, significant concentrations of reactive oxygen species may be remotely generated by sustaining an active discharge beyond the confines of the dielectric capillary, which may benefit applications that require large concentrations of reactive oxygen species such as material processing or biomedical devices.

NMR signal analysis in the large COMPASS $^{14}$NH$_{3}$ target

CERN Document Server

Koivuniemi, J; Hess, C; Kisselev, Y U; Meyer, W; Radtke, E; Reicherz, G; Doshita, N; Iwata, T; Kondo, K; Michigami, T

2009-01-01

In the large COMPASS polarized proton target the 1508 cm$^{3}$ of irradiated granular ammonia is polarized with dynamic nuclear polarization method using 4 mm microwaves in 2.5 T eld. The nuclear polarization up to 90 - 93 % is determined with cw NMR. The properties of the observed ammonia proton signals are described and spin thermodynamics in high elds is presented. Also the second moment of the NMR line is estimated.

Plasma characterization using terahertz-wave-enhanced fluorescence

International Nuclear Information System (INIS)

Liu Jingle; Zhang, X.-C.

2010-01-01

We demonstrate that the terahertz-wave-enhanced fluorescence emission from excited atoms or molecules can be employed in the characterization of laser-induced gas plasmas. The electron relaxation time and plasma density were deduced through applying the electron impact excitation/ionization and electron-ion recombination processes to the measured time-dependent enhanced fluorescence. The electron collision dynamics of nitrogen plasma excited at different gas pressures and laser pulse energies have been systematically investigated. This plasma characterization method provides picosecond temporal resolution and enables omnidirectional optical signal collection.

Quantification of fluorescence angiography in a porcine model

DEFF Research Database (Denmark)

Nerup, Nikolaj; Andersen, Helene Schou; Ambrus, Rikard

2017-01-01

PURPOSE: There is no consensus on how to quantify indocyanine green (ICG) fluorescence angiography. The aim of the present study was to establish and gather validity evidence for a method of quantifying fluorescence angiography, to assess organ perfusion. METHODS: Laparotomy was performed on seven...... pigs, with two regions of interest (ROIs) marked. ICG and neutron-activated microspheres were administered and the stomach was illuminated in the near-infrared range, parallel to continuous recording of fluorescence signal. Tissue samples from the ROIs were sent for quantification of microspheres...... to calculate the regional blood flow. A software system was developed to assess the fluorescent recordings quantitatively, and each quantitative parameter was compared with the regional blood flow. The parameter with the strongest correlation was then compared with results from an independently developed...

Antiphase dual-color correlation in a reactant-product pair imparts ultrasensitivity in reaction-linked double-photoswitching fluorescence imaging.

Science.gov (United States)

Wan, Wei; Zhu, Ming-Qiang; Tian, Zhiyuan; Li, Alexander D Q

2015-04-08

A pair of reversible photochemical reactions correlates their reactant and product specifically, and such a correlation uniquely distinguishes their correlated signal from others that are not linked by this reversible reaction. Here a nanoparticle-shielded fluorophore is photodriven to undergo structural dynamics, alternating between a green-fluorescence state and a red-fluorescence state. As time elapses, the fluorophore can be in either state but not both at the same time. Thus, the red fluorescence is maximized while the green fluorescence is minimized and vice versa. Such an antiphase dual-color (AD) corelationship between the red and green fluorescence maxima as well as between their minima can be exploited to greatly improve the signal-to-noise ratio, thus enhancing the ultimate detection limit. Potential benefits of this correlation include elimination of all interferences originating from single-color dyes and signal amplification of AD photoswitching molecules by orders of magnitude.

Digital communication through intermolecular fluorescence modulation.

Science.gov (United States)

Raymo, F M; Giordani, S

2001-06-14

[see reaction]. Ultraminiaturized processors incorporating molecular components can be developed only after devising efficient strategies to communicate signals at the molecular level. We have demonstrated that a three-state molecular switch responds to ultraviolet light, visible light, and H+, attenuating the emission intensity of a fluorescent probe. Intermolecular communication is responsible for the transduction of three input signals into a single optical output. The behavior of the communicating ensemble of molecules corresponds to that of a logic circuit incorporating seven gates.

Signals of doubly-charged Higgsinos at the CERN Large Hadron Collider

International Nuclear Information System (INIS)

Demir, Durmus A.; Frank, Mariana; Turan, Ismail; Huitu, Katri; Rai, Santosh Kumar

2008-01-01

Several supersymmetric models with extended gauge structures, motivated by either grand unification or by neutrino mass generation, predict light doubly-charged Higgsinos. In this work we study productions and decays of doubly-charged Higgsinos present in left-right supersymmetric models, and show that they invariably lead to novel collider signals not found in the minimal supersymmetric model or in any of its extensions motivated by the μ problem or even in extra dimensional theories. We investigate their distinctive signatures at the Large Hadron Collider in both pair- and single-production modes, and show that they are powerful tools in determining the underlying model via the measurements at the Large Hadron Collider experiments.

The Potential Applications of Real-Time Monitoring of Water Quality in a Large Shallow Lake (Lake Taihu, China Using a Chromophoric Dissolved Organic Matter Fluorescence Sensor

Directory of Open Access Journals (Sweden)

Cheng Niu

2014-06-01

Full Text Available This study presents results from field surveys performed over various seasons in a large, eutrophic, shallow lake (Lake Taihu, China using an in situ chromophoric dissolved organic matter (CDOM fluorescence sensor as a surrogate for other water quality parameters. These measurements identified highly significant empirical relationships between CDOM concentration measured using the in situ fluorescence sensor and CDOM absorption, fluorescence, dissolved organic carbon (DOC, chemical oxygen demand (COD and total phosphorus (TP concentrations. CDOM concentration expressed in quinine sulfate equivalent units, was highly correlated with the CDOM absorption coefficient (r2 = 0.80, p < 0.001, fluorescence intensities (Ex./Em. 370/460 nm (r2 = 0.91, p < 0.001, the fluorescence index (r2 = 0.88, p < 0.001 and the humification index (r2 = 0.78, p < 0.001, suggesting that CDOM concentration measured using the in situ fluorescence sensor could act as a substitute for the CDOM absorption coefficient and fluorescence measured in the laboratory. Similarly, CDOM concentration was highly correlated with DOC concentration (r2 = 0.68, p < 0.001, indicating that in situ CDOM fluorescence sensor measurements could be a proxy for DOC concentration. In addition, significant positive correlations were found between laboratory CDOM absorption coefficients and COD (r2 = 0.83, p < 0.001, TP (r2 = 0.82, p < 0.001 concentrations, suggesting a potential further application for the real-time monitoring of water quality using an in situ CDOM fluorescence sensor.

Measurement of the Low Frequency Noise of MOSFETs under Large Signal RF Excitation

NARCIS (Netherlands)

van der Wel, A.P.; Klumperink, Eric A.M.; Nauta, Bram

2002-01-01

A measurement technique [1] is presented that allows measurement of MOSFET low frequency (LF) noise under large signal RF (Radio Frequency) excitation. Measurements indicate that MOSFETS exhibit a reduction in LF noise when they are cycled from inversion to accummulation and that this reduction does

Responses of sun-induced chlorophyll fluorescence to biological and environmental variations measured with a versatile Fluorescence Auto-Measurement Equipment (FAME)

Science.gov (United States)

Gu, L.

2017-12-01

In this study, we examine responses of sun-induced chlorophyll fluorescence to biological and environmental variations measured with a versatile Fluorescence Auto-Measurement Equipment (FAME). FAME was developed to automatically and continuously measure chlorophyll fluorescence (F) of a leaf, plant or canopy in both laboratory and field environments, excited by either artificial light source or sunlight. FAME is controlled by a datalogger and allows simultaneous measurements of environmental variables complementary to the F signals. A built-in communication system allows FAME to be remotely monitored and data-downloaded. Radiance and irradiance calibrations can be done online. FAME has been applied in a variety of environments, allowing an investigation of biological and environmental controls on F emission.

Fluorenyl benzothiadiazole and benzoselenadiazole near-IR fluorescent probes for two-photon fluorescence imaging (Conference Presentation)

Science.gov (United States)

Belfield, Kevin D.; Yao, Sheng; Kim, Bosung; Yue, Xiling

2016-03-01

Imaging biological samples with two-photon fluorescence (2PF) microscopy has the unique advantage of resulting high contrast 3D resolution subcellular image that can reach up to several millimeters depth. 2PF probes that absorb and emit at near IR region need to be developed. Two-photon excitation (2PE) wavelengths are less concerned as 2PE uses wavelengths doubles the absorption wavelength of the probe, which means 2PE wavelengths for probes even with absorption at visible wavelength will fall into NIR region. Therefore, probes that fluoresce at near IR region with high quantum yields are needed. A series of dyes based on 5-thienyl-2, 1, 3-benzothiadiazole and 5-thienyl-2, 1, 3-benzoselenadiazole core were synthesized as near infrared two-photon fluorophores. Fluorescence maxima wavelengths as long as 714 nm and fluorescence quantum yields as high as 0.67 were achieved. The fluorescence quantum yields of the dyes were nearly constant, regardless of solvents polarity. These diazoles exhibited large Stokes shift (GM), and high two-photon fluorescence figure of merit (FM , 1.04×10-2 GM). Cells incubated on a 3D scaffold with one of the new probes (encapsulated in Pluronic micelles) exhibited bright fluorescence, enabling 3D two-photon fluorescence imaging to a depth of 100 µm.

The Pseudo signal peptide of the corticotropin-releasing factor receptor type 2A prevents receptor oligomerization.

Science.gov (United States)

Teichmann, Anke; Rutz, Claudia; Kreuchwig, Annika; Krause, Gerd; Wiesner, Burkhard; Schülein, Ralf

2012-08-03

N-terminal signal peptides mediate the interaction of native proteins with the translocon complex of the endoplasmic reticulum membrane and are cleaved off during early protein biogenesis. The corticotropin-releasing factor receptor type 2a (CRF(2(a))R) possesses an N-terminal pseudo signal peptide, which represents a so far unique domain within the large protein family of G protein-coupled receptors (GPCRs). In contrast to a conventional signal peptide, the pseudo signal peptide remains uncleaved and consequently forms a hydrophobic extension at the N terminus of the receptor. The functional consequence of the presence of the pseudo signal peptide is not understood. Here, we have analyzed the significance of this domain for receptor dimerization/oligomerization in detail. To this end, we took the CRF(2(a))R and the homologous corticotropin-releasing factor receptor type 1 (CRF(1)R) possessing a conventional cleaved signal peptide and conducted signal peptide exchange experiments. Using single cell and single molecule imaging methods (fluorescence resonance energy transfer and fluorescence cross-correlation spectroscopy, respectively) as well as biochemical experiments, we obtained two novel findings; we could show that (i) the CRF(2(a))R is expressed exclusively as a monomer, and (ii) the presence of the pseudo signal peptide prevents its oligomerization. Thus, we have identified a novel functional domain within the GPCR protein family, which plays a role in receptor oligomerization and which may be useful to study the functional significance of this process in general.

Fluorescence detection of dental calculus

International Nuclear Information System (INIS)

Gonchukov, S; Sukhinina, A; Vdovin, Yu; Biryukova, T

2010-01-01

This work is devoted to the optimization of fluorescence dental calculus diagnostics in optical spectrum. The optimal wavelengths for fluorescence excitation and registration are determined. Two spectral ranges 620 – 645 nm and 340 – 370 nm are the most convenient for supra- and subgingival calculus determination. The simple implementation of differential method free from the necessity of spectrometer using was investigated. Calculus detection reliability in the case of simple implementation is higher than in the case of spectra analysis at optimal wavelengths. The use of modulated excitation light and narrowband detection of informative signal allows us to decrease essentially its diagnostic intensity even in comparison with intensity of the low level laser dental therapy

Fluorescence detection of dental calculus

Science.gov (United States)

Gonchukov, S.; Biryukova, T.; Sukhinina, A.; Vdovin, Yu

2010-11-01

This work is devoted to the optimization of fluorescence dental calculus diagnostics in optical spectrum. The optimal wavelengths for fluorescence excitation and registration are determined. Two spectral ranges 620 - 645 nm and 340 - 370 nm are the most convenient for supra- and subgingival calculus determination. The simple implementation of differential method free from the necessity of spectrometer using was investigated. Calculus detection reliability in the case of simple implementation is higher than in the case of spectra analysis at optimal wavelengths. The use of modulated excitation light and narrowband detection of informative signal allows us to decrease essentially its diagnostic intensity even in comparison with intensity of the low level laser dental therapy.

Fluorescent optical liquid-level sensor

International Nuclear Information System (INIS)

Weiss, Jonathan D.

2000-01-01

An optical method of detecting a liquid level is presented that uses fluorescence radiation generated in an impurity-doped glass or plastic slab. In operation, the slab is inserted into the liquid and pump light is coupled into it so that the light is guided by the slab-air interface above the liquid and escapes into the liquid just below its surface. Since the fluorescence is generated only in that section of the slab above the liquid, the fluorescence power will monotonically decrease with increasing liquid level. Thus, a relationship can be established between any signal proportional to it and the liquid level. Because optical fibers link the pump source and the detector of fluorescence radiation to the sensor, no electrical connections are needed in or near the liquid. Their absence vastly decreases the hazard associated with placing a liquid-level sensor in a potentially explosive environment. A laboratory prototype, consisting of a methyl styrene slab doped with an organic dye, has been built and successfully tested in water. Its response to liquid level when pumped by a tunable argon-ion laser at 476, 488, and 496 nm, and by a blue LED, is presented and shown to be consistent with theory. The fluorescence spectra, optical efficiency, temperature, and other effects are also presented and discussed. (c) 2000 Society of Photo-Optical Instrumentation Engineers

Super-nonlinear fluorescence microscopy for high-contrast deep tissue imaging

Science.gov (United States)

Wei, Lu; Zhu, Xinxin; Chen, Zhixing; Min, Wei

2014-02-01

Two-photon excited fluorescence microscopy (TPFM) offers the highest penetration depth with subcellular resolution in light microscopy, due to its unique advantage of nonlinear excitation. However, a fundamental imaging-depth limit, accompanied by a vanishing signal-to-background contrast, still exists for TPFM when imaging deep into scattering samples. Formally, the focusing depth, at which the in-focus signal and the out-of-focus background are equal to each other, is defined as the fundamental imaging-depth limit. To go beyond this imaging-depth limit of TPFM, we report a new class of super-nonlinear fluorescence microscopy for high-contrast deep tissue imaging, including multiphoton activation and imaging (MPAI) harnessing novel photo-activatable fluorophores, stimulated emission reduced fluorescence (SERF) microscopy by adding a weak laser beam for stimulated emission, and two-photon induced focal saturation imaging with preferential depletion of ground-state fluorophores at focus. The resulting image contrasts all exhibit a higher-order (third- or fourth- order) nonlinear signal dependence on laser intensity than that in the standard TPFM. Both the physical principles and the imaging demonstrations will be provided for each super-nonlinear microscopy. In all these techniques, the created super-nonlinearity significantly enhances the imaging contrast and concurrently extends the imaging depth-limit of TPFM. Conceptually different from conventional multiphoton processes mediated by virtual states, our strategy constitutes a new class of fluorescence microscopy where high-order nonlinearity is mediated by real population transfer.

The measurement of X-rays radiation temperature with a new developed filter-fluorescence spectroscopy

International Nuclear Information System (INIS)

Zhang Chuanfei; Lin Libin; Lou Fuhong; Peng Taiping

2001-01-01

The author introduces how to measure the energy spectra of X-rays by filter-fluorescence spectroscopy. The design principle and structure of new-developed double diaphragms and filter-fluorescence spectroscopy with 5 channels are depicted. The parameters of optimized spectroscopy by numerical method are given. The filter-fluorescence spectroscopy designed according as Rousseau balance principle improves signal-noises ratio

Fluorescence Detection and Discrimination of ss- and ds-DNA with a Water Soluble Oligopyrene Derivative

Directory of Open Access Journals (Sweden)

Gaoquan Shi

2009-06-01

Full Text Available A novel water-soluble cationic conjugated oligopyrene derivative, oligo(N1,N1,N1,N4,N4,N4-hexamethyl-2-(4-(pyren-1-yl butane-1,4-diaminium bromide (OHPBDB, was synthesized by a combination of chemical and electrochemical synthesis techniques. Each oligomer chain has five pyrene derivative repeating units and brings 10 positive charges. OHPBDB showed high and rapid fluorescence quenching in aqueous media upon addition of trace amounts of single-stranded (ss and double-stranded (ds DNA. The Stern-Volmer constants for ss- and ds-DNA were measured to be as high as 1.3 × 108 mol-1·L and 1.2 × 108 mol-1·L, respectively. On the other hand, distinct fluorescence enhancement of OHPBDB upon addition of large amount of ss-DNA or ds-DNA was observed. Furthermore, ss-DNA showed much stronger fluorescence enhancement than that of ds-DNA, thus yielding a clear and simple signal useful for the discrimination between ss- and ds-DNA in aqueous media.

Non-invasive In Vivo Fluorescence Optical Imaging of Inflammatory MMP Activity Using an Activatable Fluorescent Imaging Agent.

Science.gov (United States)

Schwenck, Johannes; Maier, Florian C; Kneilling, Manfred; Wiehr, Stefan; Fuchs, Kerstin

2017-05-08

This paper describes a non-invasive method for imaging matrix metalloproteinases (MMP)-activity by an activatable fluorescent probe, via in vivo fluorescence optical imaging (OI), in two different mouse models of inflammation: a rheumatoid arthritis (RA) and a contact hypersensitivity reaction (CHR) model. Light with a wavelength in the near infrared (NIR) window (650 - 950 nm) allows a deeper tissue penetration and minimal signal absorption compared to wavelengths below 650 nm. The major advantages using fluorescence OI is that it is cheap, fast and easy to implement in different animal models. Activatable fluorescent probes are optically silent in their inactivated states, but become highly fluorescent when activated by a protease. Activated MMPs lead to tissue destruction and play an important role for disease progression in delayed-type hypersensitivity reactions (DTHRs) such as RA and CHR. Furthermore, MMPs are the key proteases for cartilage and bone degradation and are induced by macrophages, fibroblasts and chondrocytes in response to pro-inflammatory cytokines. Here we use a probe that is activated by the key MMPs like MMP-2, -3, -9 and -13 and describe an imaging protocol for near infrared fluorescence OI of MMP activity in RA and control mice 6 days after disease induction as well as in mice with acute (1x challenge) and chronic (5x challenge) CHR on the right ear compared to healthy ears.

Detection of Salmonella typhi utilizing bioconjugated fluorescent polymeric nanoparticles

Energy Technology Data Exchange (ETDEWEB)

Jain, Swati, E-mail: swatijain.iitd@gmail.com; Chattopadhyay, Sruti, E-mail: sruticiitd@gmail.com; Jackeray, Richa; Abid, Zainul; Singh, Harpal, E-mail: harpal2000@yahoo.com [Centre for Biomedical Engineering, Indian Institute of Technology-Delhi (India)

2016-05-15

Present work demonstrates effective utilization of functionalized polymeric fluorescent nanoparticles as biosensing probe for the detection of Salmonella typhi bacteria on modified polycarbonate (PC) filters in about 3 h. Antibody modified-PC membranes were incubated with contaminated bacterial water for selective capturing which were detected by synthesized novel bioconjugate probe. Core–shell architecture of polymeric nanoparticles endows them with aqueous stabilization and keto-enolic functionalities making them usable for covalently linking S. typhi antibodies without any crosslinker or activator. Bradford analysis revealed that one nanoparticle has an average of 3.51 × 10{sup −19} g or 21 × 10{sup 4} bound S. typhi Ab molecules. Analysis of the regions of interest (ROI) in fluorescent micrographs of modified fluoroimmunoassay showed higher detection sensitivity of 5 × 10{sup 2} cells/mL due to signal amplification unlike conventional naked dye FITC-Ab conjugate. Fluorescence of pyrene dye remained same on immobilization of biomolecules and nanoparticles showed stable fluorescent intensity under prolong exposure to laser owing to protective polymeric layer allowing accurate identification of bacteria. Surface-functionalized PC matrix and fluorescent label NPs permit covalent interactions among biomolecules enhancing signal acquisitions showing higher detection efficiency as compared to conventional microtiter plate-based system. Our novel immunoassay has the potential to be explored as rapid detection method for identifying S. typhi contaminations in water.Graphical Abstract.

Detection of Salmonella typhi utilizing bioconjugated fluorescent polymeric nanoparticles

International Nuclear Information System (INIS)

Jain, Swati; Chattopadhyay, Sruti; Jackeray, Richa; Abid, Zainul; Singh, Harpal

2016-01-01

Present work demonstrates effective utilization of functionalized polymeric fluorescent nanoparticles as biosensing probe for the detection of Salmonella typhi bacteria on modified polycarbonate (PC) filters in about 3 h. Antibody modified-PC membranes were incubated with contaminated bacterial water for selective capturing which were detected by synthesized novel bioconjugate probe. Core–shell architecture of polymeric nanoparticles endows them with aqueous stabilization and keto-enolic functionalities making them usable for covalently linking S. typhi antibodies without any crosslinker or activator. Bradford analysis revealed that one nanoparticle has an average of 3.51 × 10"−"1"9 g or 21 × 10"4 bound S. typhi Ab molecules. Analysis of the regions of interest (ROI) in fluorescent micrographs of modified fluoroimmunoassay showed higher detection sensitivity of 5 × 10"2 cells/mL due to signal amplification unlike conventional naked dye FITC-Ab conjugate. Fluorescence of pyrene dye remained same on immobilization of biomolecules and nanoparticles showed stable fluorescent intensity under prolong exposure to laser owing to protective polymeric layer allowing accurate identification of bacteria. Surface-functionalized PC matrix and fluorescent label NPs permit covalent interactions among biomolecules enhancing signal acquisitions showing higher detection efficiency as compared to conventional microtiter plate-based system. Our novel immunoassay has the potential to be explored as rapid detection method for identifying S. typhi contaminations in water.Graphical Abstract

Wide-field fluorescent microscopy and fluorescent imaging flow cytometry on a cell-phone.

Science.gov (United States)

Zhu, Hongying; Ozcan, Aydogan

2013-04-11

Fluorescent microscopy and flow cytometry are widely used tools in biomedical research and clinical diagnosis. However these devices are in general relatively bulky and costly, making them less effective in the resource limited settings. To potentially address these limitations, we have recently demonstrated the integration of wide-field fluorescent microscopy and imaging flow cytometry tools on cell-phones using compact, light-weight, and cost-effective opto-fluidic attachments. In our flow cytometry design, fluorescently labeled cells are flushed through a microfluidic channel that is positioned above the existing cell-phone camera unit. Battery powered light-emitting diodes (LEDs) are butt-coupled to the side of this microfluidic chip, which effectively acts as a multi-mode slab waveguide, where the excitation light is guided to uniformly excite the fluorescent targets. The cell-phone camera records a time lapse movie of the fluorescent cells flowing through the microfluidic channel, where the digital frames of this movie are processed to count the number of the labeled cells within the target solution of interest. Using a similar opto-fluidic design, we can also image these fluorescently labeled cells in static mode by e.g. sandwiching the fluorescent particles between two glass slides and capturing their fluorescent images using the cell-phone camera, which can achieve a spatial resolution of e.g. - 10 μm over a very large field-of-view of - 81 mm(2). This cell-phone based fluorescent imaging flow cytometry and microscopy platform might be useful especially in resource limited settings, for e.g. counting of CD4+ T cells toward monitoring of HIV+ patients or for detection of water-borne parasites in drinking water.

In-depth characterization of the fluorescent signal of HyPer, a probe for hydrogen peroxide, in bacteria exposed to external oxidative stress.

Science.gov (United States)

Lim, Joseph B; Barker, Kimberly A; Huang, Beijing K; Sikes, Hadley D

2014-11-01

Genetically encoded, fluorescent biosensors have been developed to probe the activities of various signaling molecules inside cells ranging from changes in intracellular ion concentrations to dynamics of lipid second messengers. HyPer is a member of this class of biosensors and is the first to dynamically respond to hydrogen peroxide (H2O2), a reactive oxygen species that functions as a signaling molecule. However, detailed characterization of HyPer's signal is not currently available within the context of bacteria exposed to external oxidative stress, which occurs in the immunological response of higher organisms against invasive pathogenic bacteria. Here, we performed this characterization, specifically in Escherichia coli exposed to external H2O2. We found that the temporal behavior of the signal does not correspond exactly to peroxide concentration in the system as a function of time and expression of the sensor decreases the peroxide scavenging activity of the cell. We also determined the effects of cell number, both before and after normalization of externally added H2O2 to the number of cells. Finally, we report quantitative characteristics of HyPer's signal in this context, including the dynamic range of the signal, the signal-to-noise ratio, and the half saturation constant. These parameters show statistically meaningful differences in signal between two commonly used strains of E. coli, demonstrating how signal can vary with strain. Taken together, our results establish a systematic, quantitative framework for researchers seeking to better understand the role of H2O2 in the immunological response against bacteria, and for understanding potential differences in the details of HyPer's quantitative performance across studies. Copyright © 2014 Elsevier B.V. All rights reserved.

Monitoring super-volcanoes: geophysical and geochemical signals at Yellowstone and other large caldera systems.

Science.gov (United States)

Lowenstern, Jacob B; Smith, Robert B; Hill, David P

2006-08-15

Earth's largest calderas form as the ground collapses during immense volcanic eruptions, when hundreds to thousands of cubic kilometres of magma are explosively withdrawn from the Earth's crust over a period of days to weeks. Continuing long after such great eruptions, the resulting calderas often exhibit pronounced unrest, with frequent earthquakes, alternating uplift and subsidence of the ground, and considerable heat and mass flux. Because many active and extinct calderas show evidence for repetition of large eruptions, such systems demand detailed scientific study and monitoring. Two calderas in North America, Yellowstone (Wyoming) and Long Valley (California), are in areas of youthful tectonic complexity. Scientists strive to understand the signals generated when tectonic, volcanic and hydrothermal (hot ground water) processes intersect. One obstacle to accurate forecasting of large volcanic events is humanity's lack of familiarity with the signals leading up to the largest class of volcanic eruptions. Accordingly, it may be difficult to recognize the difference between smaller and larger eruptions. To prepare ourselves and society, scientists must scrutinize a spectrum of volcanic signals and assess the many factors contributing to unrest and toward diverse modes of eruption.

Simultaneous neuron- and astrocyte-specific fluorescent marking

Energy Technology Data Exchange (ETDEWEB)

Schulze, Wiebke [Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Hayata-Takano, Atsuko [Molecular Research Center for Children' s Mental Development, United Graduate School of Child Development, Osaka University, Kanazawa University, Hamamatsu University School of Medicine, Chiba University and University of Fukui, 2-2 Yamadaoka, Suita, Osaka 565-0871 (Japan); Kamo, Toshihiko [Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Nakazawa, Takanobu, E-mail: takanobunakazawa-tky@umin.ac.jp [iPS Cell-based Research Project on Brain Neuropharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Nagayasu, Kazuki [iPS Cell-based Research Project on Brain Neuropharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Kasai, Atsushi; Seiriki, Kaoru [Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Interdisciplinary Program for Biomedical Sciences, Institute for Academic Initiatives, Osaka University, 1-1 Yamadaoka, Suita, Osaka 565-0871 (Japan); Shintani, Norihito [Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Ago, Yukio [Laboratory of Medicinal Pharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Farfan, Camille [Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); and others

2015-03-27

Systematic and simultaneous analysis of multiple cell types in the brain is becoming important, but such tools have not yet been adequately developed. Here, we aimed to generate a method for the specific fluorescent labeling of neurons and astrocytes, two major cell types in the brain, and we have developed lentiviral vectors to express the red fluorescent protein tdTomato in neurons and the enhanced green fluorescent protein (EGFP) in astrocytes. Importantly, both fluorescent proteins are fused to histone 2B protein (H2B) to confer nuclear localization to distinguish between single cells. We also constructed several expression constructs, including a tandem alignment of the neuron- and astrocyte-expression cassettes for simultaneous labeling. Introducing these vectors and constructs in vitro and in vivo resulted in cell type-specific and nuclear-localized fluorescence signals enabling easy detection and distinguishability of neurons and astrocytes. This tool is expected to be utilized for the simultaneous analysis of changes in neurons and astrocytes in healthy and diseased brains. - Highlights: • We develop a method for the specific fluorescent labeling of neurons and astrocytes. • Neuron-specific labeling is achieved using Scg10 and synapsin promoters. • Astrocyte-specific labeling is generated using the minimal GFAP promoter. • Nuclear localization of fluorescent proteins is achieved with histone 2B protein.

Simultaneous neuron- and astrocyte-specific fluorescent marking

International Nuclear Information System (INIS)

Schulze, Wiebke; Hayata-Takano, Atsuko; Kamo, Toshihiko; Nakazawa, Takanobu; Nagayasu, Kazuki; Kasai, Atsushi; Seiriki, Kaoru; Shintani, Norihito; Ago, Yukio; Farfan, Camille

2015-01-01

Systematic and simultaneous analysis of multiple cell types in the brain is becoming important, but such tools have not yet been adequately developed. Here, we aimed to generate a method for the specific fluorescent labeling of neurons and astrocytes, two major cell types in the brain, and we have developed lentiviral vectors to express the red fluorescent protein tdTomato in neurons and the enhanced green fluorescent protein (EGFP) in astrocytes. Importantly, both fluorescent proteins are fused to histone 2B protein (H2B) to confer nuclear localization to distinguish between single cells. We also constructed several expression constructs, including a tandem alignment of the neuron- and astrocyte-expression cassettes for simultaneous labeling. Introducing these vectors and constructs in vitro and in vivo resulted in cell type-specific and nuclear-localized fluorescence signals enabling easy detection and distinguishability of neurons and astrocytes. This tool is expected to be utilized for the simultaneous analysis of changes in neurons and astrocytes in healthy and diseased brains. - Highlights: • We develop a method for the specific fluorescent labeling of neurons and astrocytes. • Neuron-specific labeling is achieved using Scg10 and synapsin promoters. • Astrocyte-specific labeling is generated using the minimal GFAP promoter. • Nuclear localization of fluorescent proteins is achieved with histone 2B protein

Fluorescent detection of C-reactive protein using polyamide beads

Science.gov (United States)

Jagadeesh, Shreesha; Chen, Lu; Aitchison, Stewart

2016-03-01

Bacterial infection causes Sepsis which is one of the leading cause of mortality in hospitals. This infection can be quantified from blood plasma using C - reactive protein (CRP). A quick diagnosis at the patient's location through Point-of- Care (POC) testing could give doctors the confidence to prescribe antibiotics. In this paper, the development and testing of a bead-based procedure for CRP quantification is described. The size of the beads enable them to be trapped in wells without the need for magnetic methods of immobilization. Large (1.5 mm diameter) Polyamide nylon beads were used as the substrate for capturing CRP from pure analyte samples. The beads captured CRP either directly through adsorption or indirectly by having specific capture antibodies on their surface. Both methods used fluorescent imaging techniques to quantify the protein. The amount of CRP needed to give a sufficient fluorescent signal through direct capture method was found suitable for identifying bacterial causes of infection. Similarly, viral infections could be quantified by the more sensitive indirect capture method. This bead-based assay can be potentially integrated as a disposable cartridge in a POC device due to its passive nature and the small quantities needed.

Site-specific multipoint fluorescence measurement system with end-capped optical fibers.

Science.gov (United States)

Song, Woosub; Moon, Sucbei; Lee, Byoung-Cheol; Park, Chul-Seung; Kim, Dug Young; Kwon, Hyuk Sang

2011-07-10

We present the development and implementation of a spatially and spectrally resolved multipoint fluorescence correlation spectroscopy (FCS) system utilizing multiple end-capped optical fibers and an inexpensive laser source. Specially prepared end-capped optical fibers placed in an image plane were used to both collect fluorescence signals from the sample and to deliver signals to the detectors. The placement of independently selected optical fibers on the image plane was done by monitoring the end-capped fiber tips at the focus using a CCD, and fluorescence from specific positions of a sample were collected by an end-capped fiber, which could accurately represent light intensities or spectral data without incurring any disturbance. A fast multipoint spectroscopy system with a time resolution of ∼1.5 ms was then implemented using a prism and an electron multiplying charge coupled device with a pixel binning for the region of interest. The accuracy of our proposed system was subsequently confirmed by experimental results, based on an FCS analysis of microspheres in distilled water. We expect that the proposed multipoint site-specific fluorescence measurement system can be used as an inexpensive fluorescence measurement tool to study many intracellular and molecular dynamics in cell biology. © 2011 Optical Society of America

Multiwavelength time-resolved detection of fluorescence during the inflow of indocyanine green into the adult's brain

Science.gov (United States)

Gerega, Anna; Milej, Daniel; Weigl, Wojciech; Botwicz, Marcin; Zolek, Norbert; Kacprzak, Michal; Wierzejski, Wojciech; Toczylowska, Beata; Mayzner-Zawadzka, Ewa; Maniewski, Roman; Liebert, Adam

2012-08-01

Optical technique based on diffuse reflectance measurement combined with indocyanine green (ICG) bolus tracking is extensively tested as a method for clinical assessment of brain perfusion in adults at the bedside. Methodology of multiwavelength and time-resolved detection of fluorescence light excited in the ICG is presented and advantages of measurements at multiple wavelengths are discussed. Measurements were carried out: 1. on a physical homogeneous phantom to study the concentration dependence of the fluorescence signal, 2. on the phantom to simulate the dynamic inflow of ICG at different depths, and 3. in vivo on surface of the human head. Pattern of inflow and washout of ICG in the head of healthy volunteers after intravenous injection of the dye was observed for the first time with time-resolved instrumentation at multiple emission wavelengths. The multiwavelength detection of fluorescence signal confirms that at longer emission wavelengths, probability of reabsorption of the fluorescence light by the dye itself is reduced. Considering different light penetration depths at different wavelengths, and the pronounced reabsorption at longer wavelengths, the time-resolved multiwavelength technique may be useful in signal decomposition, leading to evaluation of extra- and intracerebral components of the measured signals.

Spatial discrimination against background with different optical systems for collection of fluorescence in laser-excited atomic fluorescence spectrometry with a graphite tube electrothermal atomizer.

Science.gov (United States)

Yuzefovsky, A I; Lonardo, R F; Michel, R G

1995-07-01

A single 90 degrees off-axis ellipsoidal mirror fragment was used in a dispersive detection system for electrothermal atomization laser-excited atomic fluorescence spectrometry. The performance of the new optical arrangement was compared with those of optical arrangements that employed a plane mirror in combination with biconvex or plano-convex lenses. All the optical arrangements collected fluorescence in a scheme called front surface illustration. BEAM-4, an optical ray tracing program, was used for calculations of spatial ray distributions and optical collection efficiency for the various optical configurations. Experimentally, the best collection efficiency was obtained by use of the ellipsoidal mirror, in qualitative agreement with simulations done by use of the BEAM-4 software. The best detection limit for cobalt with the new optical arrangement was 20 fg, which was a factor of 5 better than that obtained with conventional optical arrangements with otherwise the same instrumentation. The signal-to-background ratio and the fluorescence collection efficiency were also studied as a function of position of the optical components for the various optical arrangements. For both cobalt and phosphorus, the signal-to-background ratio with the new optical arrangement remained stable within 10-20% during +/- 8 mm shifts in the position of the detection system from the focal plane of the optics. Overall, the new optical arrangement offered high collection efficiency, excellent sensitivity, and facile optical alignment due to efficient spatial separation between the fluorescence signal and the background radiation. The advantages of the new optical arrangement were particularly important during measurements in the presence of high levels of blackbody radiation.

First-in-human evaluation of a hybrid modality that allows combined radio- and (near-infrared) fluorescence tracing during surgery

Energy Technology Data Exchange (ETDEWEB)

Berg, Nynke S. van den [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology (Netherlands); The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Urology, Amsterdam (Netherlands); Simon, Herve [Eurorad S.A., Eckbolsheim (France); Kleinjan, Gijs H. [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology (Netherlands); The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Nuclear Medicine, Amsterdam (Netherlands); Engelen, Thijs [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology (Netherlands); The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Head and Neck Surgery and Oncology, Amsterdam (Netherlands); Bunschoten, Anton; Welling, Mick M. [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology (Netherlands); Tijink, Bernard M. [The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Head and Neck Surgery and Oncology, Amsterdam (Netherlands); Horenblas, Simon [The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Urology, Amsterdam (Netherlands); Chambron, Jacques [The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Nuclear Medicine, Amsterdam (Netherlands); Leeuwen, Fijs W.B. van [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology (Netherlands); The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Urology, Amsterdam (Netherlands); The Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, Department of Head and Neck Surgery and Oncology, Amsterdam (Netherlands)

2015-10-15

The clinical introduction of the hybrid tracer indocyanine green (ICG)-{sup 99m}Tc-nanocolloid, composed of a radioactive and a near-infrared (NIR) fluorescence component, has created the need for surgical (imaging) modalities that allow for simultaneous detection of both signals. This study describes the first-in-human use of a prototype opto-nuclear probe during sentinel node (SN) biopsy using ICG-{sup 99m}Tc-nanocolloid. To allow for fluorescence tracing, a derivative of the conventional gamma probe technology was generated in which two optical fibers were integrated to allow for excitation (785 nm) and emission signal collection (> 810 nm). The ability of this opto-nuclear probe to detect the fluorescence signal of the hybrid tracer ICG-{sup 99m}Tc-nanocolloid was firstly determined ex vivo in (non)SNs samples obtained from 41 patients who underwent hybrid tracer-based SN biopsy in the head and neck or urogenital area. In an in vivo proof-of-concept study in nine of these 41 patients, SNs were localized using combined gamma and fluorescence tracing with the opto-nuclear probe. Fluorescence tracing was performed in a similar manner as gamma tracing and under ambient light conditions. Ex vivo, the gamma tracing option of the opto-nuclear probe correctly identified the SN in all 150 evaluated (non)SN samples. Ex vivo fluorescence tracing in the low-sensitivity mode correctly identified 71.7 % of the samples. This increased to 98.9 % when fluorescence tracing was performed in the high-sensitivity mode. In vivo fluorescence tracing (high-sensitivity mode) accurately identified the SNs in all nine patients (20 SNs evaluated; 100 %). This study demonstrates the first-in-human evaluation of a hybrid modality capable of detecting both gamma and fluorescence signals during a surgical procedure. Fluorescence tracing could be performed in ambient light. (orig.)

Construction of a bimolecular fluorescence complementation (BiFC ...

African Journals Online (AJOL)

DR. NJ TONUKARI

2012-08-02

Aug 2, 2012 ... Accepted 8 June, 2012. Protein–protein interactions are essential for signal transduction in cells. ... BiFC is a novel technology that is used for identifying .... occasional fluorescence was observed, it was very weak ... Light field.

Smartphone Cortex Controlled Real-Time Image Processing and Reprocessing for Concentration Independent LED Induced Fluorescence Detection in Capillary Electrophoresis.

Science.gov (United States)

Szarka, Mate; Guttman, Andras

2017-10-17

We present the application of a smartphone anatomy based technology in the field of liquid phase bioseparations, particularly in capillary electrophoresis. A simple capillary electrophoresis system was built with LED induced fluorescence detection and a credit card sized minicomputer to prove the concept of real time fluorescent imaging (zone adjustable time-lapse fluorescence image processor) and separation controller. The system was evaluated by analyzing under- and overloaded aminopyrenetrisulfonate (APTS)-labeled oligosaccharide samples. The open source software based image processing tool allowed undistorted signal modulation (reprocessing) if the signal was inappropriate for the actual detection system settings (too low or too high). The novel smart detection tool for fluorescently labeled biomolecules greatly expands dynamic range and enables retrospective correction for injections with unsuitable signal levels without the necessity to repeat the analysis.

Fluorescent Nanodiamonds in Biomedical Applications.

Science.gov (United States)

Mitura, Katarzyna Anna; Włodarczyk, Elżbieta

2018-04-18

Nanoparticles have an extended surface and a large surface area, which is the ratio of the size of the surfacearea to the volume. A functionalized surface can give rise to more modifications and therefore allows this nanomaterial to have new properties. Fluorescent molecules contain fluorophore, which is capable of being excited via the absorption of light energy at a specific wavelength and subsequently emitting radiation energy of a longer wavelength. A chemically modified surface of nanodiamond (ND; by carboxylation) demonstrated biocompatibility with DNA, cytochrome C, and antigens. In turn, fluorescent nanodiamonds (FNDs) belong to a group of new nanomaterials. Their surface can be modified by joining functional groups such as carboxyl, hydroxyl, or amino, after which they can be employed as a fluorescence agent. Their fluorescent properties result from defects in the crystal lattice. FNDs reach dimensions of 4-100 nm, have attributes such as photostability, long fluorescence lifetimes (10 ns), and fluorescence emission between 600 and 700 nm. They are also nontoxic, chemically inert, biocompatible, and environmentally harmless. The main purpose of this article was to present the medical applications of various types of modified NDs.

Measurement of cell volume changes by fluorescence self-quenching

DEFF Research Database (Denmark)

Hamann, Steffen; Kiilgaard, J.F.; Litman, Thomas

2002-01-01

At high concentrations, certain fluorophores undergo self-quenching, i.e., fluorescence intensity decreases with increasing fluorophore concentration. Accordingly, the self-quenching properties can be used for measuring water volume changes in lipid vesicles. In cells, quantitative determination...... concentrations of the fluorophore calcein suitable for measurement of changes in cell water volume by self-quenching. The relationship between calcein fluorescence intensity, when excited at 490 nm (its excitation maximum), and calcein concentration was investigated in vitro and in various cultured cell types...... to a decrease in calcein fluorescence with high signal-to-noise ratio (>15). Similar results were obtained with the fluorophore BCECF when excited at its isosbestic wavelength (436 nm). The present results demonstrate the usefulness of fluorescence self-quenching to measure rapid changes in cell water volume....

A simple optical fiber device for quantitative fluorescence microscopy of single living cells

OpenAIRE

van Graft, M.; van Graft, Marja; Oosterhuis, B.; Oosterhuis, Bernard; van der Werf, Kees; de Grooth, B.G.; Greve, Jan

1993-01-01

simple and relatively inexpensive system is described for obtaining quantitative fluorescence measurements on single living cells loaded with a fluorescent probe to study cell physiological processes. The light emitted from the fluorescent cells is captured by and transported through an optical fiber. After passage through appropriate filters the light is measured using a photomultiplier tube. The optical fiber is mounted in one of the microscope outlets. Signals derived from the photomultipl...

Molecule-binding dependent assembly of split aptamer and γ-cyclodextrin: A sensitive excimer signaling approach for aptamer biosensors

International Nuclear Information System (INIS)

Jin, Fen; Lian, Yan; Li, Jishan; Zheng, Jing; Hu, Yaping; Liu, Jinhua; Huang, Jin; Yang, Ronghua

2013-01-01

Graphical abstract: Adenosine-binding aptamer was splitted into two fragments P2 and P3 which labeled pyrene molecules, mainly produce monomer signal. γ-CD cavity brings P2 and P3 in close proximity, allowing for weak excimer emission. In the presence of target, P2 and P3 are expected to bind ATP and form an aptamer/target complex, leads to large increase of the pyrene excimer fluorescence. -- Highlights: •We assembled split aptamer and γ-cyclodextrin fluorescence biosensors for ATP detection. •The biosensor increased quantum yield and emission lifetime of the excimer. •Time-resolved fluorescence is effective for ATP assay in complicated environment. -- Abstract: A highly sensitive and selective fluorescence aptamer biosensors for the determination of adenosine triphosphate (ATP) was developed. Binding of a target with splitting aptamers labeled with pyrene molecules form stable pyrene dimer in the γ-cyclodextrin (γ-CD) cavity, yielding a strong excimer emission. We have found that inclusion of pyrene dimer in γ-cyclodextrin cavity not only exhibits additive increases in quantum yield and emission lifetime of the excimer, but also facilitates target-induced fusion of the splitting aptamers to form the aptamer/target complex. As proof-of-principle, the approach was applied to fluorescence detection of adenosine triphosphate. With an anti-ATP aptamer, the approach exhibits excimer fluorescence response toward ATP with a maximum signal-to-background ratio of 32.1 and remarkably low detection limit of 80 nM ATP in buffer solution. Moreover, due to the additive fluorescence lifetime of excimer induced by γ-cyclodextrin, time-resolved measurements could be conveniently used to detect as low as 0.5 μM ATP in blood serum quantitatively

Fluorescence Enhancement on Large Area Self-Assembled Plasmonic-3D Photonic Crystals.

Science.gov (United States)

Chen, Guojian; Wang, Dongzhu; Hong, Wei; Sun, Lu; Zhu, Yongxiang; Chen, Xudong

2017-03-01

Discontinuous plasmonic-3D photonic crystal hybrid structures are fabricated in order to evaluate the coupling effect of surface plasmon resonance and the photonic stop band. The nanostructures are prepared by silver sputtering deposition on top of hydrophobic 3D photonic crystals. The localized surface plasmon resonance of the nanostructure has a symbiotic relationship with the 3D photonic stop band, leading to highly tunable characteristics. Fluorescence enhancements of conjugated polymer and quantum dot based on these hybrid structures are studied. The maximum fluorescence enhancement for the conjugated polymer of poly(5-methoxy-2-(3-sulfopropoxy)-1,4-phenylenevinylene) potassium salt by a factor of 87 is achieved as compared with that on a glass substrate due to the enhanced near-field from the discontinuous plasmonic structures, strong scattering effects from rough metal surface with photonic stop band, and accelerated decay rates from metal-coupled excited state of the fluorophore. It is demonstrated that the enhancement induced by the hybrid structures has a larger effective distance (optimum thickness ≈130 nm) than conventional plasmonic systems. It is expected that this approach has tremendous potential in the field of sensors, fluorescence-imaging, and optoelectronic applications. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

An image fiber based fluorescent probe with associated signal processing scheme for biomedical diagnostics

International Nuclear Information System (INIS)

Vaishakh, M; Murukeshan, V M; Seah, L K

2008-01-01

A dual-modality image fiber based fluorescent probe that can be used for depth sensitive imaging and suppression of fluorescent emissions with nanosecond lifetime difference is proposed and illustrated in this paper. The system can give high optical sectioning and employs an algorithm for obtaining phase sensitive images. The system can find main application in in vivo biomedical diagnostics for detecting biochemical changes for distinguishing malignant tissue from healthy tissue

Dual fluorescence of single LH2 antenna nanorings

International Nuclear Information System (INIS)

Freiberg, A.; Raetsep, M.; Timpmann, K.; Trinkunas, G.

2004-01-01

A dual nature of fluorescence from LH2 pigment-protein complexes, which is a part of the light harvesting system of purple bacteria, is confirmed by fluorescence-lifetime dependence on recording wavelength and spectrally selective spectroscopy. An analysis based on the Holstein molecular crystal model, modified by allowing diagonal disorder, suggests coexistence of large- and small-radius self-trapped excitons, which serve as the origin of the dual fluorescence

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

Science.gov (United States)

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

2014-07-09

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

Waveguide evanescent field fluorescence microscopy & its application in cell biology

Science.gov (United States)

Hassanzadeh, Abdollah

are captured and collected during the experiment, permitting time lapse analysis. As a proof of concept, we have monitored the response of cells on the waveguide surface to an external lethal agent. Imaging analysis showed very low photobleaching. Therefore photobleaching can be neglected during the experiments. The effects of secondary patterns (inhomogeneities) in the grating and scratches and inhomogeneities in the wave guiding film on the fluorescence background, ultra-thin film and cell-substrate contact regions image were investigated. In conclusion, we developed and established WEFF microscopy and have visualized and quantified solid thin films thicknesses and cell-substrate contact regions. The achieved low scattering results in an improved signal-to-noise ratio and increased sensitivity. Photobleaching and phototoxicity are largely reduced compared to other microscopy techniques. Therefore imaging can be carried out over extended periods and having better temporal resolution without sample damage, such as effect of external agents on the cell-substrate contact regions. Keywords. Waveguide Evanescent Field Fluorescence Microscopy, Evanescent Field, Ion-exchanged Waveguides, TE modes, TM modes, Electromagnetic Field Distribution, Fluorescence, Microscopy, Optical Waveguides, Imaging, Interface, Triton X-100, Focal Contacts, Close Contacts, Cell-Substrate Contact Regions, Cell-Substrate Separation Distances, Photobleaching, Phototoxicity, Cell-Substrate Interaction, Langmuir-Blodgett Films, Phase Separated Lipid Films, SGG 11 Glass, Resolution, Total Internal Reflection, Total Internal Reflection Fluorescence Microscopy, Osteoblast Cells, Grating Coupling, Prism Coupling, Grating.

All Fiber-Coupled OH Planar Laser-Induced-Fluorescence (OH-PLIF)-Based Two-Dimensional Thermometry.

Science.gov (United States)

Hsu, Paul S; Jiang, Naibo; Patnaik, Anil K; Katta, Vish; Roy, Sukesh; Gord, James R

2018-04-01

Two-color, planar laser-induced fluorescence (PLIF)-based two-dimensional (2D) thermometry techniques for reacting flows, which are typically developed in the laboratory conditions, face a stiff challenge in their practical implementation in harsh environments such as combustion rigs. In addition to limited optical access, the critical experimental conditions (i.e., uncontrolled humidity, vibration, and large thermal gradients) often restrict sensitive laser system operation and cause difficulties maintaining beam-overlap. Thus, an all fiber-coupled, two-color OH-PLIF system has been developed, employing two long optical fibers allowing isolation of the laser and signal-collection systems. Two OH-excitation laser beams (∼283 nm and ∼286 nm) are delivered through a common 6 m long, 400 µm core, deep ultraviolet (UV)-enhanced multimode fiber. The fluorescence signal (∼310 nm) is collected by a 3 m long, UV-grade imaging fiber. Proof-of-principle temperature measurements are demonstrated in atmospheric pressure, near adiabatic, CH 4 /O 2 /N 2 jet flames. The effects of the excitation pulse interval on fiber transmission are investigated. The proof-of-principle measurements show significant promise for thermometry in harsh environments such as gas turbine engine tests.

European Space Agency's Fluorescence Explorer Mission: Concept and Applications

Science.gov (United States)

Mohammed, G.; Moreno, J. F.; Goulas, Y.; Huth, A.; Middleton, E.; Miglietta, F.; Nedbal, L.; Rascher, U.; Verhoef, W.; Drusch, M.

2012-12-01

The Fluorescence Explorer (FLEX) is a dedicated satellite for the detection and measurement of solar-induced fluorescence (SIF). It is one of two candidate missions currently under evaluation by ESA for deployment in its Earth Explorer 8 program, with Phase A/B1 assessments now underway. FLEX is planned as a tandem mission with ESA's core mission Sentinel-3, and would carry an instrument, FLORIS, optimized for discrimination of the fluorescence signal in terrestrial vegetation. The FLEX mission would be the first to be focussed upon optimization of SIF detection in terrestrial vegetation, and using finer spatial resolution than is available with current satellites. It would open up a novel avenue for monitoring photosynthetic function from space, with diverse potential applications. Plant photosynthetic tissues absorbing sunlight in the wavebands of photosynthetically active radiation (400 to 700 nm) emit fluorescence in the form of red and far-red light. This signal confers a small but measurable contribution to apparent reflectance spectra, and with appropriate analysis it may be detected and quantified. Over the last 15-20 years, techniques for SIF detection have progressed from contact or near-contact methods using single leaves to remote techniques using airborne sensors and towers over plant canopies. Ongoing developments in instrumentation, atmospheric correction procedures, signal extraction techniques, and utilization of the SIF signal itself are all critical aspects of progress in this area. The FLEX mission would crystallize developments to date into a state-of-the-art pioneering mission targeting actual photosynthetic function. This compares to existing methods which address only potential function. Thus, FLEX could serve to provide real-time data on vegetation health and stress status, and inputs for parameterization of photosynthetic models (e.g. with measures of light-use efficiency). SIF might be correlated or modelled to photosynthetic rates or

Morphological spot counting from stacked images for automated analysis of gene copy numbers by fluorescence in situ hybridization.

Science.gov (United States)

Grigoryan, Artyom M; Dougherty, Edward R; Kononen, Juha; Bubendorf, Lukas; Hostetter, Galen; Kallioniemi, Olli

2002-01-01

Fluorescence in situ hybridization (FISH) is a molecular diagnostic technique in which a fluorescent labeled probe hybridizes to a target nucleotide sequence of deoxyribose nucleic acid. Upon excitation, each chromosome containing the target sequence produces a fluorescent signal (spot). Because fluorescent spot counting is tedious and often subjective, automated digital algorithms to count spots are desirable. New technology provides a stack of images on multiple focal planes throughout a tissue sample. Multiple-focal-plane imaging helps overcome the biases and imprecision inherent in single-focal-plane methods. This paper proposes an algorithm for global spot counting in stacked three-dimensional slice FISH images without the necessity of nuclei segmentation. It is designed to work in complex backgrounds, when there are agglomerated nuclei, and in the presence of illumination gradients. It is based on the morphological top-hat transform, which locates intensity spikes on irregular backgrounds. After finding signals in the slice images, the algorithm groups these together to form three-dimensional spots. Filters are employed to separate legitimate spots from fluorescent noise. The algorithm is set in a comprehensive toolbox that provides visualization and analytic facilities. It includes simulation software that allows examination of algorithm performance for various image and algorithm parameter settings, including signal size, signal density, and the number of slices.

Signal improvement in multiphoton microscopy by reflection with simple mirrors near the sample

Science.gov (United States)

Rehberg, Markus; Krombach, Fritz; Pohl, Ulrich; Dietzel, Steffen

2010-03-01

In conventional fluorescence or confocal microscopy, emitted light is generated not only in the focal plane but also above and below. The situation is different in multiphoton-induced fluorescence and multiphoton-induced higher harmonic generation. Here, restriction of signal generation to a single focal point permits that all emitted photons can contribute to image formation if collected, regardless of their path through the specimen. Often, the intensity of the emitted light is rather low in biological specimens. We present a method to significantly increase the fraction of photons collected by an epi (backward) detector by placing a simple mirror, an aluminum-coated coverslip, directly under the sample. Samples investigated include fluorescent test slides, collagen gels, and thin-layered, intact mouse skeletal muscles. Quantitative analysis revealed an intensity increase of second- and third-harmonic generated signal in skeletal muscle of nine- and sevenfold respectively, and of fluorescent signal in test slides of up to twofold. Our approach thus allows significant signal improvement also for situations were a forward detection is impossible, e.g., due to the anatomy of animals in intravital microscopy.

Ensemble empirical mode decomposition based fluorescence spectral noise reduction for low concentration PAHs

Science.gov (United States)

Wang, Shu-tao; Yang, Xue-ying; Kong, De-ming; Wang, Yu-tian

2017-11-01

A new noise reduction method based on ensemble empirical mode decomposition (EEMD) is proposed to improve the detection effect for fluorescence spectra. Polycyclic aromatic hydrocarbons (PAHs) pollutants, as a kind of important current environmental pollution source, are highly oncogenic. Using the fluorescence spectroscopy method, the PAHs pollutants can be detected. However, instrument will produce noise in the experiment. Weak fluorescent signals can be affected by noise, so we propose a way to denoise and improve the detection effect. Firstly, we use fluorescence spectrometer to detect PAHs to obtain fluorescence spectra. Subsequently, noises are reduced by EEMD algorithm. Finally, the experiment results show the proposed method is feasible.

Fluorescence background subtraction technique for hybrid fluorescence molecular tomography/x-ray computed tomography imaging of a mouse model of early stage lung cancer.

Science.gov (United States)

Ale, Angelique; Ermolayev, Vladimir; Deliolanis, Nikolaos C; Ntziachristos, Vasilis

2013-05-01

The ability to visualize early stage lung cancer is important in the study of biomarkers and targeting agents that could lead to earlier diagnosis. The recent development of hybrid free-space 360-deg fluorescence molecular tomography (FMT) and x-ray computed tomography (XCT) imaging yields a superior optical imaging modality for three-dimensional small animal fluorescence imaging over stand-alone optical systems. Imaging accuracy was improved by using XCT information in the fluorescence reconstruction method. Despite this progress, the detection sensitivity of targeted fluorescence agents remains limited by nonspecific background accumulation of the fluorochrome employed, which complicates early detection of murine cancers. Therefore we examine whether x-ray CT information and bulk fluorescence detection can be combined to increase detection sensitivity. Correspondingly, we research the performance of a data-driven fluorescence background estimator employed for subtraction of background fluorescence from acquisition data. Using mice containing known fluorochromes ex vivo, we demonstrate the reduction of background signals from reconstructed images and sensitivity improvements. Finally, by applying the method to in vivo data from K-ras transgenic mice developing lung cancer, we find small tumors at an early stage compared with reconstructions performed using raw data. We conclude with the benefits of employing fluorescence subtraction in hybrid FMT-XCT for early detection studies.

Fluorescent single walled nanotube/silica composite materials

Science.gov (United States)

Dattelbaum, Andrew M.; Gupta, Gautam; Duque, Juan G.; Doorn, Stephen K.; Hamilton, Christopher E.; DeFriend Obrey, Kimberly A.

2013-03-12

Fluorescent composites of surfactant-wrapped single-walled carbon nanotubes (SWNTs) were prepared by exposing suspensions of surfactant-wrapped carbon nanotubes to tetramethylorthosilicate (TMOS) vapor. Sodium deoxycholate (DOC) and sodium dodecylsulphate (SDS) were the surfactants. No loss in emission intensity was observed when the suspension of DOC-wrapped SWNTs were exposed to the TMOS vapors, but about a 50% decrease in the emission signal was observed from the SDS-wrapped SWNTs nanotubes. The decrease in emission was minimal by buffering the SDS/SWNT suspension prior to forming the composite. Fluorescent xerogels were prepared by adding glycerol to the SWNT suspensions prior to TMOS vapor exposure, followed by drying the gels. Fluorescent aerogels were prepared by replacing water in the gels with methanol and then exposing them to supercritical fluid drying conditions. The aerogels can be used for gas sensing.

Conditional-sampling spectrograph detection system for fluorescence measurements of individual airborne biological particles

Science.gov (United States)

Nachman, Paul; Pinnick, R. G.; Hill, Steven C.; Chen, Gang; Chang, Richard K.; Mayo, Michael W.; Fernandez, Gilbert L.

1996-03-01

We report the design and operation of a prototype conditional-sampling spectrograph detection system that can record the fluorescence spectra of individual, micrometer-sized aerosols as they traverse an intense 488-nm intracavity laser beam. The instrument's image-intensified CCD detector is gated by elastic scattering or by undispersed fluorescence from particles that enter the spectrograph's field of view. It records spectra only from particles with preselected scattering-fluorescence levels (a fiber-optic-photomultiplier subsystem provides the gating signal). This conditional-sampling procedure reduces data-handling rates and increases the signal-to-noise ratio by restricting the system's exposures to brief periods when aerosols traverse the beam. We demonstrate these advantages by reliably capturing spectra from individual fluorescent microspheres dispersed in an airstream. The conditional-sampling procedure also permits some discrimination among different types of particles, so that spectra may be recorded from the few interesting particles present in a cloud of background aerosol. We demonstrate such discrimination by measuring spectra from selected fluorescent microspheres in a mixture of two types of microspheres, and from bacterial spores in a mixture of spores and nonfluorescent kaolin particles.

Multi-spectral and fluorescence diffuse optical tomography of breast cancer

Science.gov (United States)

Corlu, Alper

Multi-spectral and fluorescence diffuse optical tomography (DOT) techniques are explored and applied to image human breast cancer in vivo. Image reconstruction algorithms that utilize first and second order gradient information are described in detail. Breast DOT requires large computational memory and long run times. To this end, parallel computation techniques were developed appropriate to each reconstruction algorithm. A parallel plate DOT instrument developed for breast cancer imaging is described. The system relies heavily on continuous-wave (CW) transmission measurements and utilizes frequency domain (FD) measurements on the reemission side. However, traditional DOT image reconstruction methods based on CW measurements fail to separate tissue absorption and scattering uniquely. In this manuscript, multi-spectral DOT is shown to be capable of minimizing cross-talk and retrieving spectral parameters almost uniquely when the measurement wavelengths are optimized. A theoretical framework to select optimum wavelengths is provided, and tested with computer simulations. Results from phantom spectroscopy experiments and in vivo patient measurements support the notion that multi-spectral methods are superior to traditional DOT image reconstruction schemes. The same breast DOT instrument is improved and utilized to obtain the first in vivo images of human breast cancer based on fluorescence DOT (FDOT). To this end the fluorophore Indocyanine Green (ICG) is injected intravenously and fluorescence excitation and detection are accomplished in the soft-compression, parallel-plane, transmission geometry using laser sources at 786 nm and spectrally filtered CCD detection. Careful phantom and in vivo measurements are carried on to assure that the signals are due to ICG fluorescence, rather than tissue autofluorescence and excitation light leakage. An in vivo measurement protocol is designed to maximize the ICG contrast by acquiring full fluorescence tomographic scan during

Laser-Induced Photofragmentation Fluorescence Imaging of Alkali Compounds in Flames.

Science.gov (United States)

Leffler, Tomas; Brackmann, Christian; Aldén, Marcus; Li, Zhongshan

2017-06-01

Laser-induced photofragmentation fluorescence has been investigated for the imaging of alkali compounds in premixed laminar methane-air flames. An ArF excimer laser, providing pulses of wavelength 193 nm, was used to photodissociate KCl, KOH, and NaCl molecules in the post-flame region and fluorescence from the excited atomic alkali fragment was detected. Fluorescence emission spectra showed distinct lines of the alkali atoms allowing for efficient background filtering. Temperature data from Rayleigh scattering measurements together with simulations of potassium chemistry presented in literature allowed for conclusions on the relative contributions of potassium species KOH and KCl to the detected signal. Experimental approaches for separate measurements of these components are discussed. Signal power dependence and calculated fractions of dissociated molecules indicate the saturation of the photolysis process, independent on absorption cross-section, under the experimental conditions. Quantitative KCl concentrations up to 30 parts per million (ppm) were evaluated from the fluorescence data and showed good agreement with results from ultraviolet absorption measurements. Detection limits for KCl photofragmentation fluorescence imaging of 0.5 and 1.0 ppm were determined for averaged and single-shot data, respectively. Moreover, simultaneous imaging of KCl and NaCl was demonstrated using a stereoscope with filters. The results indicate that the photofragmentation method can be employed for detailed studies of alkali chemistry in laboratory flames for validation of chemical kinetic mechanisms crucial for efficient biomass fuel utilization.

The enhanced cyan fluorescent protein: a sensitive pH sensor for fluorescence lifetime imaging.

Science.gov (United States)

Poëa-Guyon, Sandrine; Pasquier, Hélène; Mérola, Fabienne; Morel, Nicolas; Erard, Marie

2013-05-01

pH is an important parameter that affects many functions of live cells, from protein structure or function to several crucial steps of their metabolism. Genetically encoded pH sensors based on pH-sensitive fluorescent proteins have been developed and used to monitor the pH of intracellular compartments. The quantitative analysis of pH variations can be performed either by ratiometric or fluorescence lifetime detection. However, most available genetically encoded pH sensors are based on green and yellow fluorescent proteins and are not compatible with multicolor approaches. Taking advantage of the strong pH sensitivity of enhanced cyan fluorescent protein (ECFP), we demonstrate here its suitability as a sensitive pH sensor using fluorescence lifetime imaging. The intracellular ECFP lifetime undergoes large changes (32 %) in the pH 5 to pH 7 range, which allows accurate pH measurements to better than 0.2 pH units. By fusion of ECFP with the granular chromogranin A, we successfully measured the pH in secretory granules of PC12 cells, and we performed a kinetic analysis of intragranular pH variations in living cells exposed to ammonium chloride.

The fluorescence theatre: a cost-effective device using theatre gels for fluorescent protein and dye screening.

Science.gov (United States)

Heil, John R; Nordeste, Ricardo F; Charles, Trevor C

2011-04-01

Here we report a simple cost-effective device for screening colonies on plates for expression of the monomeric red fluorescent protein mRFP1 and the fluorescent dye Nile red. This device can be built from any simple light source, in our case a Quebec Colony Counter, and cost-effective theatre gels. The device can be assembled in as little as 20 min, and it produces excellent results when screening a large number of colonies.

Control of the blue fluorescent protein with advanced evolutionary pulse shaping

International Nuclear Information System (INIS)

Tkaczyk, Eric R.; Mauring, Koit; Tkaczyk, Alan H.; Krasnenko, Veera; Ye, Jing Yong; Baker, James R.; Norris, Theodore B.

2008-01-01

We demonstrate optical coherent control of the two-photon fluorescence of the blue fluorescent protein (BFP), which is of interest in investigations of protein-protein interactions. In addition to biological relevance, BFP represents an interesting target for coherent control from a chemical perspective due to its many components of highly nonexponential fluorescence decay and low quantum yield resulting from excited state isomerization. Using a genetic algorithm with a multiplicative (rather than ratiometric) fitness parameter, we are able to control the ratio of BFP fluorescence to second-harmonic generation without a considerable drop in the maximized signal. The importance of linear chirp and power-scaling on the discrimination process is investigated in detail

Signaling in large-scale neural networks

DEFF Research Database (Denmark)

Berg, Rune W; Hounsgaard, Jørn

2009-01-01

We examine the recent finding that neurons in spinal motor circuits enter a high conductance state during functional network activity. The underlying concomitant increase in random inhibitory and excitatory synaptic activity leads to stochastic signal processing. The possible advantages of this m......We examine the recent finding that neurons in spinal motor circuits enter a high conductance state during functional network activity. The underlying concomitant increase in random inhibitory and excitatory synaptic activity leads to stochastic signal processing. The possible advantages...... of this metabolically costly organization are analyzed by comparing with synaptically less intense networks driven by the intrinsic response properties of the network neurons....

Assessment of drinking water quality at the tap using fluorescence spectroscopy.

Science.gov (United States)

Heibati, Masoumeh; Stedmon, Colin A; Stenroth, Karolina; Rauch, Sebastien; Toljander, Jonas; Säve-Söderbergh, Melle; Murphy, Kathleen R

2017-11-15

Treated drinking water may become contaminated while travelling in the distribution system on the way to consumers. Elevated dissolved organic matter (DOM) at the tap relative to the water leaving the treatment plant is a potential indicator of contamination, and can be measured sensitively, inexpensively and potentially on-line via fluorescence and absorbance spectroscopy. Detecting elevated DOM requires potential contamination events to be distinguished from natural fluctuations in the system, but how much natural variation to expect in a stable distribution system is unknown. In this study, relationships between DOM optical properties, microbial indicator organisms and trace elements were investigated for households connected to a biologically-stable drinking water distribution system. Across the network, humic-like fluorescence intensities showed limited variation (RSD = 3.5-4.4%), with half of measured variation explained by interactions with copper. After accounting for quenching by copper, fluorescence provided a very stable background signal (RSD infiltration of soil water would be detectable. Smaller infiltrations would be detectable in the case of contamination by sewage with a strong tryptophan-like fluorescence signal. These findings indicate that DOM fluorescence is a sensitive indicator of water quality changes in drinking water networks, as long as potential interferents are taken into account. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

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.

Large Signal Model of a Four-quadrant AC to DC Converter for Accelerator Magnets

CERN Document Server

De la Calle, R; Rinaldi, L; Völker, F V

2001-01-01

This paper presents the large signal model of a four-quadrant AC to DC converter, which is expected to be used in the area of particle accelerators. The system’s first stage is composed of a three-phase boost PWM (Pulse Width Modulated) rectifier with DSP (Digital Signal Processing) based power factor correction (PFC) and output voltage regulation. The second stage is a full-bridge PWM inverter that allows fast four-quadrant operation. The structure is fully reversible, and an additional resistance (brake chopper) is not needed to dissipate the energy when the beam deflection magnet acts as generator.

Mixture-fraction imaging at 1  kHz using femtosecond laser-induced fluorescence of krypton.

Science.gov (United States)

Richardson, Daniel R; Jiang, Naibo; Stauffer, Hans U; Kearney, Sean P; Roy, Sukesh; Gord, James R

2017-09-01

Femtosecond, two-photon-absorption laser-induced-fluorescence (TALIF) imaging measurements of krypton (Kr) are demonstrated to study mixing in gaseous flows. A measurement approach is presented in which observed Kr TALIF signals are 7 times stronger than the current state-of-the-art methodology. Fluorescence emission is compared for different gas pressures and excitation wavelengths, and the strongest fluorescence signals were observed when the excitation wavelength was tuned to 212.56 nm. Using this optimized excitation scheme, 1-kHz, single-laser-shot visualizations of unsteady flows and two-dimensional measurements of mixture fraction and scalar dissipation rate of a Kr-seeded jet are demonstrated.

Fluorescence spectroscopy of soil pellets : The use of CP/PARAFAC.

Science.gov (United States)

Mounier, Stéphane; Nicolodeli, Gustavo; Redon, Roland; Hacherouf, Kalhed; Milori, Debora M. B. P.

2014-05-01

Fluorescence spectroscopy is one of the most sensitive techniques available for analytical purposes. It is relatively easy to implement, phenomenologically straightforward and well investigated. Largely non-invasive and fast, so that it can be useful for environmental applications. Fluorescence phenomenon is highly probable in molecular systems containing atoms with lone pairs of electrons such as C=O, aromatic, phenolic, quinone and more rigid unsaturated conjugated systems. These functional groups are present in humic substances (HS) from soils (Senesi, 1990; N. Senesi et al., 1991) and represent the main fluorophors of Soil Organic Matter (SOM). The extension of the conjugated electronic system, the level of heteroatom substitution and type and number of substituting groups under the aromatic rings strongly affect the intensity and wavelength of molecular fluorescence. However, to analyse the SOM it is generally done a chemical extraction that allows measuring the fluorescence response of the liquid extract. To avoid this fractionation of the SOM, Milori et al. (2006) proposed the application of laser induced fluorescence spectroscopy (LIFS) in whole soil. This work intends to assess the technical feasibility of 3D fluorescence spectroscopy using lamp for excitation to analyse solids opaque samples prepared with different substances. Seventy four (74) solid samples were prepared from different mixtures of boric acid (BA), humic substance acid and tryptophan (TRP) powder. The compounds were mixture and a pellet was done by using pressure (8 ton). The pellets were measured using a spectrofluorimeter HITACHI F4500, and a 3D fluorescence tensor was done from emission spectra (200-600 nm) with excitation range from 200 to 500 nm. The acquisition parameters were: step at 5 nm, scan speed at 2400 nm.min-1, response time at 0.1 s, excitation and emission slits at 5 nm and photomultiplier voltage at 700 V. Furthermore, measures of Laser-induced Fluorescence were

Smart optical probes for near-infrared fluorescence imaging of Alzheimer's disease pathology

International Nuclear Information System (INIS)

Raymond, Scott B.; Bacskai, Brian J.; Skoch, Jesse; Hills, Ivory D.; Swager, Timothy M.; Nesterov, Evgueni E.

2008-01-01

Near-infrared fluorescent probes for amyloid-beta (Aβ) are an exciting option for molecular imaging in Alzheimer's disease research and may translate to clinical diagnostics. However, Aβ-targeted optical probes often suffer from poor specificity and slow clearance from the brain. We are designing smart optical probes that emit characteristic fluorescence signal only when bound to Aβ. We synthesized a family of dyes and tested Aβ-binding sensitivity with fluorescence spectroscopy and tissue-staining. Select compounds exhibited Aβ-dependent changes in fluorescence quantum yield, lifetime, and emission spectra that may be imaged microscopically or in vivo using new lifetime and spectral fluorescence imaging techniques. Smart optical probes that turn on when bound to Aβ will improve amyloid detection and may enable quantitative molecular imaging in vivo. (orig.)

Chemical point detection using differential fluorescence from molecularly imprinted polymers

Science.gov (United States)

Pestov, Dmitry; Anderson, John E.; Nelson, Jean; Tepper, Gary C.

2004-12-01

Fluorescence represents one of the most attractive approaches for chemical sensing due to the abundant light produced by most fluorophores, resulting in excellent detection sensitivity. However, the broad and overlapping emission spectra of target and background species have made it difficult to perform species identification in a field instrument because of the need to perform spectral decomposition and analysis. This paper describes a new chemical sensing strategy based on differential fluorescence measurements from molecularly imprinted polymers, which eliminates the need to perform any spectral analysis. Species identification is accomplished by measuring the differential light output from a pair of polymers-one imprinted to a target species and the other identical, but not imprinted. The imprinted polymer selectively concentrates the target molecule and controls the energy (wavelength) of the emitted fluorescence signal and the differential output eliminates common mode signals associated with non-specific background interference. Because no spectral analysis is required, the sensors can be made extremely small and require very little power. Preliminary performance parameters from a prototype sensor are presented and discussed.

Received signal strength in large-scale wireless relay sensor network: a stochastic ray approach

NARCIS (Netherlands)

Hu, L.; Chen, Y.; Scanlon, W.G.

2011-01-01

The authors consider a point percolation lattice representation of a large-scale wireless relay sensor network (WRSN) deployed in a cluttered environment. Each relay sensor corresponds to a grid point in the random lattice and the signal sent by the source is modelled as an ensemble of photons that

Testing the utility of fluorescent proteins in Mimulus lewisii by an Agrobacterium-mediated transient assay.

Science.gov (United States)

Ding, Baoqing; Yuan, Yao-Wu

2016-04-01

The Agrobacterium -mediated transient expression assay by leaf infiltration in Mimulus lewisii is robust. Fluorescent proteins EGFP, EYFP and DsRed give bright fluorescence signals in the infiltrated tissue. Mimulus lewisii is an emerging developmental genetic model system. Recently developed genomic and genetic resources and a stable transformation protocol have greatly facilitated the identification and functional characterization of genes controlling the development of ecologically important floral traits using this species. To further expedite gene and protein function analyses in M. lewisii, we adopted and simplified the Agrobacterium-mediated transient gene expression method routinely used in tobacco plants. With the validated transient assay, we examined the performance of fluorescent proteins EGFP, EYFP and DsRed in M. lewisii. All three proteins gave bright fluorescence signals when transiently expressed in agroinfiltrated leaves. Furthermore, we demonstrated the utility of fluorescent proteins in M. lewisii by showing the nuclear localization of Reduced Carotenoid Pigmentation 1 (RCP1), a recently discovered R2R3-MYB transcription factor that regulates carotenoid pigmentation during flower development. Both the transient assay and the fluorescent proteins are valuable additions to the M. lewisii toolbox, making this emerging genetic and developmental model system even more powerful.

Optical Aptamer Probes of Fluorescent Imaging to Rapid Monitoring of Circulating Tumor Cell

Directory of Open Access Journals (Sweden)

Ji Yeon Hwang

2016-11-01

Full Text Available Fluorescence detecting of exogenous EpCAM (epithelial cell adhesion molecule or muc1 (mucin1 expression correlated to cancer metastasis using nanoparticles provides pivotal information on CTC (circulating tumor cell occurrence in a noninvasive tool. In this study, we study a new skill to detect extracellular EpCAM/muc1 using quantum dot-based aptamer beacon (QD-EpCAM/muc1 ALB (aptamer linker beacon. The QD-EpCAM/muc1 ALB was designed using QDs (quantum dots and probe. The EpCAM/muc1-targeting aptamer contains a Ep-CAM/muc1 binding sequence and BHQ1 (black hole quencher 1 or BHQ2 (black hole quencher2. In the absence of target EpCAM/muc1, the QD-EpCAM/muc1 ALB forms a partial duplex loop-like aptamer beacon and remained in quenched state because the BHQ1/2 quenches the fluorescence signal-on of the QD-EpCAM/muc1 ALB. The binding of EpCAM/muc1 of CTC to the EpCAM/muc1 binding aptamer sequence of the EpCAM/muc1-targeting oligonucleotide triggered the dissociation of the BHQ1/2 quencher and subsequent signal-on of a green/red fluorescence signal. Furthermore, acute inflammation was stimulated by trigger such as caerulein in vivo, which resulted in increased fluorescent signal of the cy5.5-EpCAM/muc1 ALB during cancer metastasis due to exogenous expression of EpCAM/muc1 in Panc02-implanted mouse model.

Thermal maturity of Tasmanites microfossils from confocal laser scanning fluorescence microscopy

Science.gov (United States)

Hackley, Paul C.; Kus, Jolanta

2015-01-01

We report here, for the first time, spectral properties of Tasmanites microfossils determined by confocal laser scanning fluorescence microscopy (CLSM, using Ar 458 nm excitation). The Tasmanites occur in a well-characterized natural maturation sequence (Ro 0.48–0.74%) of Devonian shale (n = 3 samples) from the Appalachian Basin. Spectral property λmax shows excellent agreement (r2 = 0.99) with extant spectra from interlaboratory studies which used conventional fluorescence microscopy techniques. This result suggests spectral measurements from CLSM can be used to infer thermal maturity of fluorescent organic materials in geologic samples. Spectra of regions with high fluorescence intensity at fold apices and flanks in individual Tasmanites are blue-shifted relative to less-deformed areas in the same body that have lower fluorescence intensity. This is interpreted to result from decreased quenching moiety concentration at these locations, and indicates caution is needed in the selection of measurement regions in conventional fluorescence microscopy, where it is common practice to select high intensity regions for improved signal intensity and better signal to noise ratios. This study also documents application of CLSM to microstructural characterization of Tasmanites microfossils. Finally, based on an extant empirical relation between conventional λmax values and bitumen reflectance, λmax values from CLSM of Tasmanites microfossils can be used to calculate a bitumen reflectance equivalent value. The results presented herein can be used as a basis to broaden the future application of CLSM in the geological sciences into hydrocarbon prospecting and basin analysis.

Laser-induced fluorescence for medical diagnostics

International Nuclear Information System (INIS)

Andersson Engels, S.

1989-12-01

Laser-induced fluorescence as a tool for tissue diagnostics is discussed. Both spectrally and time-resolved fluorescence signals are studied to optimize the demarcation of diseased lesions from normal tissue. The presentation is focused on two fields of application: the identification of malignant tumours and atherosclerotic plaques. Tissue autofluorescence as well as fluorescence from administered drugs have been utilized in diseased tissue diagnosis. The fluorescence criterion for tissue diagnosis is, as far as possible, chosen to be independent of unknown fluorescence parameters, which are not correlated to the type of tissue investigated. Both a dependence on biological parameters, such as light absorption in blood, and instrumental characteristics, such as excitation pulse fluctuations and detection geometry, can be minimized. Several chemical compounds have been studied in animal experiments after intraveneous injection to verify their capacity as malignant tumour marking drugs under laser excitation and fluorescence detection. Another objective of these studies was to improve our understanding of the mechanism and chemistry behind the retention of the various drugs in tissue. The properties of a chemical which maximize its selective retention in tumours are discussed. In order to utilize this diagnostic modality, three different clinically adapted sets of instrumentation have been developed and are presented. Two of the systems are nitrogen-laser-based fluorosensors; one is a point-monitoring system with full spectral resolution and the other one is an imaging system with up to four simultaneously recorded images in different spectral bands. The third system is a low-cost point-monitoring mercury-lamp-based fluoroscence emission as well as reflection characteristics of tissue. (author)

Stage-specific fluorescence intensity of GFP and mCherry during sporulation In Bacillus Subtilis

Directory of Open Access Journals (Sweden)

Bailey Kirra

2010-11-01

Full Text Available Abstract Background Fluorescent proteins are powerful molecular biology tools that have been used to study the subcellular dynamics of proteins within live cells for well over a decade. Two fluorescent proteins commonly used to enable dual protein labelling are GFP (green and mCherry (red. Sporulation in the Gram positive bacterium Bacillus subtilis has been studied for many years as a paradigm for understanding the molecular basis for differential gene expression. As sporulation initiates, cells undergo an asymmetric division leading to differential gene expression in the small prespore and large mother cell compartments. Use of two fluorescent protein reporters permits time resolved examination of differential gene expression either in the same compartments or between compartments. Due to the spectral properties of GFP and mCherry, they are considered an ideal combination for co-localisation and co-expression experiments. They can also be used in combination with fluorescent DNA stains such as DAPI to correlate protein localisation patterns with the developmental stage of sporulation which can be linked to well characterised changes in DNA staining patterns. Findings While observing the recruitment of the transcription machinery into the forespore of sporulating Bacillus subtilis, we noticed the occurrence of stage-specific fluorescence intensity differences between GFP and mCherry. During vegetative growth and the initial stages of sporulation, fluorescence from both GFP and mCherry fusions behaved similarly. During stage II-III of sporulation we found that mCherry fluorescence was considerably diminished, whilst GFP signals remained clearly visible. This fluorescence pattern reversed during the final stage of sporulation with strong mCherry and low GFP fluorescence. These trends were observed in reciprocal tagging experiments indicating a direct effect of sporulation on fluorescent protein fluorophores. Conclusions Great care should be taken

Sensitive and selective tumor imaging with novel and highly activatable fluorescence probes

International Nuclear Information System (INIS)

Urano, Yasuteru

2008-01-01

Selective and sensitive tumor imaging in vivo is one of the most requested methodologies in medical sciences. Although several imaging modalities have been developed including positron emission tomography (PET) and magnetic resonance (MR) imaging for the detection of tumors, none of these modalities can activate the signals upon being accumulated or uptaken to tumor sites. Among these modalities, only optical fluorescence imaging has a marked advantage, that is, their signals can be dramatically increased upon detecting some biological features. In this short review, I will introduce some recent strategies for activatable optical fluorescence imaging of tumors, and discuss their advantages over other modalities. (author)

Beta-induced fluorescence detection in liquid chromatography

International Nuclear Information System (INIS)

Malcolme-Lawes, D.J.; Massey, S.; Warwick, P.

1981-01-01

A theoretical analysis of beta-induced fluorescence is used to determine the factors which influence the sensitivity of the technique as applied to liquid chromatography. Equations are presented for detector response and for signal-to-noise ratios and the theoretical response for a typical detector is compared with experimentally determined values. (author)

A High Density Low Cost Digital Signal Processing Module for Large Scale Radiation Detectors

International Nuclear Information System (INIS)

Tan, Hui; Hennig, Wolfgang; Walby, Mark D.; Breus, Dimitry; Harris, Jackson T.; Grudberg, Peter M.; Warburton, William K.

2013-06-01

A 32-channel digital spectrometer PIXIE-32 is being developed for nuclear physics or other radiation detection applications requiring digital signal processing with large number of channels at relatively low cost. A single PIXIE-32 provides spectrometry and waveform acquisition for 32 input signals per module whereas multiple modules can be combined into larger systems. It is based on the PCI Express standard which allows data transfer rates to the host computer of up to 800 MB/s. Each of the 32 channels in a PIXIE-32 module accepts signals directly from a detector preamplifier or photomultiplier. Digitally controlled offsets can be individually adjusted for each channel. Signals are digitized in 12-bit, 50 MHz multi-channel ADCs. Triggering, pile-up inspection and filtering of the data stream are performed in real time, and pulse heights and other event data are calculated on an event-by event basis. The hardware architecture, internal and external triggering features, and the spectrometry and waveform acquisition capability of the PIXIE- 32 as well as its capability to distribute clock and triggers among multiple modules, are presented. (authors)

Combinatorial mutagenesis of the voltage-sensing domain enables the optical resolution of action potentials firing at 60 Hz by a genetically encoded fluorescent sensor of membrane potential.

Science.gov (United States)

Piao, Hong Hua; Rajakumar, Dhanarajan; Kang, Bok Eum; Kim, Eun Ha; Baker, Bradley J

2015-01-07

ArcLight is a genetically encoded fluorescent voltage sensor using the voltage-sensing domain of the voltage-sensing phosphatase from Ciona intestinalis that gives a large but slow-responding optical signal in response to changes in membrane potential (Jin et al., 2012). Fluorescent voltage sensors using the voltage-sensing domain from other species give faster yet weaker optical signals (Baker et al., 2012; Han et al., 2013). Sequence alignment of voltage-sensing phosphatases from different species revealed conserved polar and charged residues at 7 aa intervals in the S1-S3 transmembrane segments of the voltage-sensing domain, suggesting potential coil-coil interactions. The contribution of these residues to the voltage-induced optical signal was tested using a cassette mutagenesis screen by flanking each transmembrane segment with unique restriction sites to allow for the testing of individual mutations in each transmembrane segment, as well as combinations in all four transmembrane segments. Addition of a counter charge in S2 improved the kinetics of the optical response. A double mutation in the S4 domain dramatically reduced the slow component of the optical signal seen in ArcLight. Combining that double S4 mutant with the mutation in the S2 domain yielded a probe with kinetics voltage-sensing domain could potentially lead to fluorescent sensors capable of optically resolving neuronal inhibition and subthreshold synaptic activity. Copyright © 2015 the authors 0270-6474/15/350372-15$15.00/0.

Two-photon excited UV fluorescence for protein crystal detection

International Nuclear Information System (INIS)

Madden, Jeremy T.; DeWalt, Emma L.; Simpson, Garth J.

2011-01-01

Complementary measurements using SONICC and TPE-UVF allow the sensitive and selective detection of protein crystals. Two-photon excited ultraviolet fluorescence (TPE-UVF) microscopy is explored for sensitive protein-crystal detection as a complement to second-order nonlinear optical imaging of chiral crystals (SONICC). Like conventional ultraviolet fluorescence (UVF), TPE-UVF generates image contrast based on the intrinsic fluorescence of aromatic residues, generally producing higher fluorescence emission within crystals than the mother liquor by nature of the higher local protein concentration. However, TPE-UVF has several advantages over conventional UVF, including (i) insensitivity to optical scattering, allowing imaging in turbid matrices, (ii) direct compatibility with conventional optical plates and windows by using visible light for excitation, (iii) elimination of potentially damaging out-of-plane UV excitation, (iv) improved signal to noise through background reduction from out-of-plane excitation and (v) relatively simple integration into instrumentation developed for SONICC

Fluorescent Nanoparticle Uptake for Brain Tumor Visualization

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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.

Second and third generation voltage-sensitive fluorescent proteins for monitoring membrane potential

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Amelie Perron

2009-06-01

Full Text Available Over the last decade, optical neuroimaging methods have been enriched by engineered biosensors derived from fluorescent protein (FP reporters fused to protein detectors that convert physiological signals into changes of intrinsic FP fluorescence. These FP-based indicators are genetically encoded, and hence targetable to specific cell populations within networks of heterologous cell types. Among this class of biosensors, the development of optical probes for membrane potential is both highly desirable and challenging. A suitable FP voltage sensor would indeed be a valuable tool for monitoring the activity of thousands of individual neurons simultaneously in a non-invasive manner. Previous prototypic genetically-encoded FP voltage indicators achieved a proof of principle but also highlighted several difficulties such as poor cell surface targeting and slow kinetics. Recently, we developed a new series of FRET-based Voltage-Sensitive Fluorescent Proteins (VSFPs, referred to as VSFP2s, with efficient targeting to the plasma membrane and high responsiveness to membrane potential signaling in excitable cells. In addition to these FRET-based voltage sensors, we also generated a third series of probes consisting of single FPs with response kinetics suitable for the optical imaging of fast neuronal signals. These newly available genetically-encoded reporters for membrane potential will be instrumental for future experimental approaches directed toward the understanding of neuronal network dynamics and information processing in the brain. Here, we review the development and current status of these novel fluorescent probes.

Fully time-resolved near-field scanning optical microscopy fluorescence imaging

International Nuclear Information System (INIS)

Kwak, Eun-Soo; Vanden Bout, David A.

2003-01-01

Time-correlated single photon counting has been coupled with near-field scanning optical microscopy (NSOM) to record complete fluorescence lifetime decays at each pixel in an NSOM image. The resulting three-dimensional data sets can be binned in the time dimension to create images of photons at particular time delays or images of the fluorescence lifetime. Alternatively, regions of interest identified in the topography and fluorescence images can be used to bin the data in the spatial dimensions resulting in high signal to noise fluorescence decays of particular regions of the sample. The technique has been demonstrated on films of poly(vinylalcohol), doped with the fluorescent dye, cascade blue (CB). The CB segregates into small circular regions of high concentration within the films during the drying process. The lifetime imaging shows that the spots have slightly faster excited state decays due to quenching of the luminescence as a result of the higher concentration. The technique is also used to image the fluorescence lifetime of an annealed film of poly(dihexylfluorene). The samples show high contrast in the total intensity fluorescence image, but the lifetime image reveals the sample to be extremely uniform

An improved large signal model of InP HEMTs

Science.gov (United States)

Li, Tianhao; Li, Wenjun; Liu, Jun

2018-05-01

An improved large signal model for InP HEMTs is proposed in this paper. The channel current and charge model equations are constructed based on the Angelov model equations. Both the equations for channel current and gate charge models were all continuous and high order drivable, and the proposed gate charge model satisfied the charge conservation. For the strong leakage induced barrier reduction effect of InP HEMTs, the Angelov current model equations are improved. The channel current model could fit DC performance of devices. A 2 × 25 μm × 70 nm InP HEMT device is used to demonstrate the extraction and validation of the model, in which the model has predicted the DC I–V, C–V and bias related S parameters accurately. Project supported by the National Natural Science Foundation of China (No. 61331006).

Ultra-high throughput real-time instruments for capturing fast signals and rare events

Science.gov (United States)

Buckley, Brandon Walter

Wide-band signals play important roles in the most exciting areas of science, engineering, and medicine. To keep up with the demands of exploding internet traffic, modern data centers and communication networks are employing increasingly faster data rates. Wide-band techniques such as pulsed radar jamming and spread spectrum frequency hopping are used on the battlefield to wrestle control of the electromagnetic spectrum. Neurons communicate with each other using transient action potentials that last for only milliseconds at a time. And in the search for rare cells, biologists flow large populations of cells single file down microfluidic channels, interrogating them one-by-one, tens of thousands of times per second. Studying and enabling such high-speed phenomena pose enormous technical challenges. For one, parasitic capacitance inherent in analog electrical components limits their response time. Additionally, converting these fast analog signals to the digital domain requires enormous sampling speeds, which can lead to significant jitter and distortion. State-of-the-art imaging technologies, essential for studying biological dynamics and cells in flow, are limited in speed and sensitivity by finite charge transfer and read rates, and by the small numbers of photo-electrons accumulated in short integration times. And finally, ultra-high throughput real-time digital processing is required at the backend to analyze the streaming data. In this thesis, I discuss my work in developing real-time instruments, employing ultrafast optical techniques, which overcome some of these obstacles. In particular, I use broadband dispersive optics to slow down fast signals to speeds accessible to high-bit depth digitizers and signal processors. I also apply telecommunication multiplexing techniques to boost the speeds of confocal fluorescence microscopy. The photonic time stretcher (TiSER) uses dispersive Fourier transformation to slow down analog signals before digitization and

Environmentally Sensitive Fluorescent Sensors Based on Synthetic Peptides

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Laurence Choulier

2010-03-01

Full Text Available Biosensors allow the direct detection of molecular analytes, by associating a biological receptor with a transducer able to convert the analyte-receptor recognition event into a measurable signal. We review recent work aimed at developing synthetic fluorescent molecular sensors for a variety of analytes, based on peptidic receptors labeled with environmentally sensitive fluorophores. Fluorescent indicators based on synthetic peptides are highly interesting alternatives to protein-based sensors, since they can be synthesized chemically, are stable, and can be easily modified in a site-specific manner for fluorophore coupling and for immobilization on solid supports.

Quenching of p-Cyanophenylalanine Fluorescence by Various Anions.

Science.gov (United States)

Pazos, Ileana M; Roesch, Rachel M; Gai, Feng

2013-03-20

To expand the spectroscopic utility of the non-natural amino acid p -cyanophenylalanine (Phe CN ), we examine the quenching efficiencies of a series of commonly encountered anions toward its fluorescence. We find that iodide exhibits an unusually large Stern-Volmer quenching constant, making it a convenient choice in Phe CN fluorescence quenching studies. Indeed, using the villin headpiece subdomain as a testbed we demonstrate that iodide quenching of Phe CN fluorescence offers a convenient means to reveal protein conformational heterogeneity. Furthermore, we show that the amino group of Phe CN strongly quenches its fluorescence, suggesting that Phe CN could be used as a local pH sensor.

In situ analysis of elemental depth distributions in thin films by combined evaluation of synchrotron x-ray fluorescence and diffraction

International Nuclear Information System (INIS)

Mainz, R.; Klenk, R.

2011-01-01

In this work we present a method for the in situ analysis of elemental depth distributions in thin films using a combined evaluation of synchrotron x-ray fluorescence and energy-dispersive x-ray diffraction signals. We recorded diffraction and fluorescence signals simultaneously during the reactive annealing of thin films. By means of the observed diffraction signals, the time evolution of phases in the thin films during the annealing processes can be determined. We utilized this phase information to parameterize the depth distributions of the elements in the films. The time-dependent fluorescence signals were then taken to determine the parameters representing the parameterized depth distributions. For this latter step, we numerically calculated the fluorescence intensities for a given set of depth distributions. These calculations handle polychromatic excitation and arbitrary functions of depth distributions and take into account primary and secondary fluorescence. Influences of lateral non-uniformities of the films, as well as the accuracy limits of the method, are investigated. We apply the introduced method to analyze the evolution of elemental depth distributions and to quantify the kinetic parameters during a synthesis process of CuInS 2 thin films via the reactive annealing of Cu-In precursors in a sulfur atmosphere.

Fluorescence-based calculus detection using a 405-nm excitation wavelength

Science.gov (United States)

Brede, O.; Schelle, F.; Krueger, S.; Oehme, B.; Dehn, C.; Frentzen, M.; Braun, A.

2011-03-01

The aim of this study was to assess the difference of fluorescence signals of cement and calculus using a 405 nm excitation wavelength. A total number of 20 freshly extracted teeth was used. The light source used for this study was a blue LED with a wavelength of 405nm. For each tooth the spectra of calculus and cementum were measured separately. Fluorescence light was collimated into an optical fibre and spectrally analyzed using an echelle spectrometer (aryelle 200, Lasertechnik Berlin, Germany) with an additionally bandpass (fgb 67, Edmund Industrial Optics, Karlsruhe, Germany). From these 40 measurements the median values were calculated over the whole spectrum, leading to two different median spectra, one for calculus and one for cementum. For further statistical analysis we defined 8 areas of interest (AOI) in wavelength regions, showing remarkable differences in signal strength. In 7 AOIs the intensity of the calculus spectrum differed statistically significant from the intensity of the cementum spectrum (p calculus and cement between 600nm and 700nm. Thus, we can conclude that fluorescence of calculus shows a significant difference to the fluorescence of cement. A differentiation over the intensity is possible as well as over the spectrum. Using a wavelength of 405nm, it is possible to distinguish between calculus and cement. These results could be used for further devices to develop a method for feedback controlled calculus removal.

Generation of a new spectral format, the lifetime synchronous spectrum (LiSS), using phase-resolved fluorescence spectroscopy

International Nuclear Information System (INIS)

Shaver, J.M.; McGown, L.B.

1994-01-01

A new fluorescence spectral format is introduced in which fluorescence lifetime is shown as a function of synchronously scanned wavelength to generate a Lifetime Synchronous Spectrum (LiSS). Lifetimes are determined in the frequency domain with the use of Phase-Resolved Fluorescence Spectroscopy (PRFS) to obtain the phase of the fluorescence signal. Theory and construction of the LiSS are presented and experimental results are shown for solutions of single components and simple binary and ternary mixtures. These results show how the lifetime information in the LiSS augments the steady-state intensity information of a standard synchronous spectrum, providing unique information for identification of components and resolution of overlapping spectral peaks. The LiSS technique takes advantage of noise reduction inherent in the extraction of lifetime from PRFS in addition to standard spectral smoothing techniques. The precision of phase determination through PRFS is found to be comparable to that of direct phase measurements at normal fluorescence intensities and superior for low-intensity signals

A dark green fluorescent protein as an acceptor for measurement of Förster resonance energy transfer.

Science.gov (United States)

Murakoshi, Hideji; Shibata, Akihiro C E; Nakahata, Yoshihisa; Nabekura, Junichi

2015-10-15

Measurement of Förster resonance energy transfer by fluorescence lifetime imaging microscopy (FLIM-FRET) is a powerful method for visualization of intracellular signaling activities such as protein-protein interactions and conformational changes of proteins. Here, we developed a dark green fluorescent protein (ShadowG) that can serve as an acceptor for FLIM-FRET. ShadowG is spectrally similar to monomeric enhanced green fluorescent protein (mEGFP) and has a 120-fold smaller quantum yield. When FRET from mEGFP to ShadowG was measured using an mEGFP-ShadowG tandem construct with 2-photon FLIM-FRET, we observed a strong FRET signal with low cell-to-cell variability. Furthermore, ShadowG was applied to a single-molecule FRET sensor to monitor a conformational change of CaMKII and of the light oxygen voltage (LOV) domain in HeLa cells. These sensors showed reduced cell-to-cell variability of both the basal fluorescence lifetime and response signal. In contrast to mCherry- or dark-YFP-based sensors, our sensor allowed for precise measurement of individual cell responses. When ShadowG was applied to a separate-type Ras FRET sensor, it showed a greater response signal than did the mCherry-based sensor. Furthermore, Ras activation and translocation of its effector ERK2 into the nucleus could be observed simultaneously. Thus, ShadowG is a promising FLIM-FRET acceptor.

Acoustically levitated droplets: a contactless sampling method for fluorescence studies.

Science.gov (United States)

Leiterer, Jork; Grabolle, Markus; Rurack, Knut; Resch-Genger, Ute; Ziegler, Jan; Nann, Thomas; Panne, Ulrich

2008-01-01

Acoustic levitation is used as a new tool to study concentration-dependent processes in fluorescence spectroscopy. With this technique, small amounts of liquid and solid samples can be measured without the need for sample supports or containers, which often limits signal acquisition and can even alter sample properties due to interactions with the support material. We demonstrate that, because of the small sample volume, fluorescence measurements at high concentrations of an organic dye are possible without the limitation of inner-filter effects, which hamper such experiments in conventional, cuvette-based measurements. Furthermore, we show that acoustic levitation of liquid samples provides an experimentally simple way to study distance-dependent fluorescence modulations in semiconductor nanocrystals. The evaporation of the solvent during levitation leads to a continuous increase of solute concentration and can easily be monitored by laser-induced fluorescence.

3-D Image Analysis of Fluorescent Drug Binding

Directory of Open Access Journals (Sweden)

M. Raquel Miquel

2005-01-01

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

Inner filter correction of dissolved organic matter fluorescence

DEFF Research Database (Denmark)

Kothawala, D.N.,; Murphy, K.R.; Stedmon, Colin

2013-01-01

The fluorescence of dissolved organic matter (DOM) is suppressed by a phenomenon of self-quenching known as the inner filter effect (IFE). Despite widespread use of fluorescence to characterize DOM in surface waters, the advantages and constraints of IFE correction are poorly defined. We assessed...... the effectiveness of a commonly used absorbance-based approach (ABA), and a recently proposed controlled dilution approach (CDA) to correct for IFE. Linearity between corrected fluorescence and total absorbance (ATotal; the sum of absorbance at excitation and emission wavelengths) across the full excitation......-emission matrix (EEM) in dilution series of four samples indicated both ABA and CDA were effective to an absorbance of at least 1.5 in a 1 cm cell, regardless of wavelength positioning. In regions of the EEMs where signal to background noise (S/N) was low, CDA correction resulted in more variability than ABA...

Fluorescent Endoscopy of Tumors in Upper Part of Gastrointestinal Tract

Science.gov (United States)

Borisova, E.; Vladimirov, B.; Angelov, I.; Avramov, L.

2007-04-01

In the recent study delta-aminolevulinic acid/Protoporphyrin IX (5-ALA/PpIX) is used as fluorescent marker for dysplasia and tumor detection in esophagus and stomach. The 5-ALA is administered per os six hours before measurements at dose 20mg/kg weight. High-power light-emitting diode at 405 nm is used as an excitation source. Special opto-mechanical device is built for LED to use the light guide of standard video-endoscopic system (Olimpus Corp.). Through endoscopic instrumental channel a fiber is applied to return information about fluorescence to microspectrometer (USB4000, OceanOptics Inc.). Very good correlation between fluorescence signals and histology examination of the lesions investigated is achieved.

Large-signal, dynamic simulation of the slowpoke-3 nuclear heating reactor

International Nuclear Information System (INIS)

Tseng, C.M.; Lepp, R.M.

1983-07-01

A 2 MWt nuclear reactor, called SLOWPOKE-3, is being developed at the Chalk River Nuclear Laboratories (CRNL). This reactor, which is cooled by natural circulation, is designed to produce hot water for commercial space heating and perhaps generate some electricity in remote locations where the costs of alternate forms of energy are high. A large-signal, dynamic simulation of this reactor, without closed-loop control, was developed and implemented on a hybrid computer, using the basic equations of conservation of mass, energy and momentum. The natural circulation of downcomer flow in the pool was simulated using a special filter, capable of modelling various flow conditions. The simulation was then used to study the intermediate and long-term transient response of SLOWPOKE-3 to large disturbances, such as loss of heat sink, loss of regulation, daily load following, and overcooling of the reactor coolant. Results of the simulation show that none of these disturbances produce hazardous transients

Use of green fluorescent protein fusions to analyse the N- and C-terminal signal peptides of GPI-anchored cell wall proteins in Candida albicans.

Science.gov (United States)

Mao, Yuxin; Zhang, Zimei; Wong, Brian

2003-12-01

Glycophosphatidylinositol (GPI)-anchored proteins account for 26-35% of the Candida albicans cell wall. To understand the signals that regulate these proteins' cell surface localization, green fluorescent protein (GFP) was fused to the N- and C-termini of the C. albicans cell wall proteins (CWPs) Hwp1p, Als3p and Rbt5p. C. albicans expressing all three fusion proteins were fluorescent at the cell surface. GFP was released from membrane fractions by PI-PLC and from cell walls by beta-glucanase, which implied that GFP was GPI-anchored to the plasma membrane and then covalently attached to cell wall glucans. Twenty and 25 amino acids, respectively, from the N- and C-termini of Hwp1p were sufficient to target GFP to the cell surface. C-terminal substitutions that are permitted by the omega rules (G613D, G613N, G613S, G613A, G615S) did not interfere with GFP localization, whereas some non-permitted substitutions (G613E, G613Q, G613R, G613T and G615Q) caused GFP to accumulate in intracellular ER-like structures and others (G615C, G613N/G615C and G613D/G615C) did not. These results imply that (i) GFP fusions can be used to analyse the N- and C-terminal signal peptides of GPI-anchored CWPs, (ii) the omega amino acid in Hwp1p is G613, and (iii) C can function at the omega+2 position in C. albicans GPI-anchored proteins.

Fluorescent nanodiamonds and their use in biomedical research

Science.gov (United States)

Suarez-Kelly, Lorena P.; Rampersaud, Isaac V.; Moritz, Charles E.; Campbell, Amanda R.; Hu, Zhiwei; Alkahtani, Masfer H.; Alghannam, Fahad S.; Hemmer, Phillip; Carson, William E.; Rampersaud, Arfaan A.

2016-03-01

Nanodiamonds containing color-centers produce non-quenching fluorescence that is easily detected. This makes them useful for cellular, proteomic and genomic applications. However, fluorescent nanodiamonds have yet to become popular in the biomedical research community as labeling reagents. We discuss production of nanodiamonds with distinct color-centers and assess their biocompatibility and techniques for bioconjugation. Fluorescent diamonds were fabricated by electron irradiation of high-pressure, high-temperature micron-sized diamonds which generated diamonds with vacancy-related defects (V). These diamonds were annealed to create nitrogen vacancy (NV)-centers then following a milling step were fractionated into nanoparticle sizes of 30, 60, and 95 nm. Optical characterization of Vand NV-center diamonds demonstrated fluorescence in two distinct green and red channels, respectively. In vitro studies demonstrated that these nanodiamonds are biocompatible and readily taken up by murine macrophage cells. Quantification of NV-center nanodiamond uptake by flow cytometry, showed that uptake was independent of nanodiamond size. Confocal microscopy demonstrated that NV-center nanodiamonds accumulate within the cytoplasm of these cells. NV-center nanodiamonds were then conjugated with streptavidin using a short polyethylene chain as linker. Conjugation was confirmed via a catalytic assay employing biotinylated-horseradish peroxidase. We present a technique for large-scale production of biocompatible conjugated V- or NV-center nanodiamonds. Functional testing is essential for standardization of fluorescent nanodiamond bioconjugates and quality control. Large-scale production of bioconjugated fluorescent nanodiamonds is crucial to their development as novel tools for biological and medical applications.

Calix[2]pyreno[2]pyrrole as a Fluorescence Chemical Probe for Polynitroaromatics

International Nuclear Information System (INIS)

Park, Kyung Hwa; Yoo, Jae Duk; Lee, Chang Hee; Ka, Jae Won

2012-01-01

We have demonstrated that the new, readily synthesized and well characterized calix pyreno pyrrole fluorescence molecular probe can detect polynitroaromatic compounds with high affinity. In addition, this highly fluorescent neutral molecular receptor also exhibits enhanced binding affinity towards TNT which is associated with the formation of a pi-complex. The dynamic nature of the current system may enable it to serve as an excellent scaffold for electron-deficient guest molecular binding. Studies for other neutral molecules including metal ions are under in active progress. The elevated selectivity and sensitivity for specific analytes are the core requirements for an ideal chemical probes. A signal induced by guest binding must be sensitive enough so that accurate real time monitoring could be satisfactorily achieved. Introduction of signaling units at suitable places in which they can directly interact with the recognition events is critical in designing chemosensors. Among various signaling events, fluorescence changes are often applied for the sensitive detection of various analytes. An easy modulation of the photochemical properties of the signaling units is an additional advantage in compatibility and applications. Chemosensors for the detection of explosives becomes important due to their immediate applications in remedation of explosive manufacturing sites, homeland security or forensic sciences. With that in mind, numerous methods for the explosive detection have been reported in recent years. Nevertheless, some detection methods sometimes require sophisticated instrumentation which is not easy for on-site testing

Bulky Counterions: Enhancing the Two-Photon Excited Fluorescence of Gold Nanoclusters.

Science.gov (United States)

Bertorelle, Franck; Moulin, Christophe; Soleilhac, Antonin; Comby-Zerbino, Clothilde; Dugourd, Philippe; Russier-Antoine, Isabelle; Brevet, Pierre-François; Antoine, Rodolphe

2018-01-19

Increasing fluorescence quantum yields of ligand-protected gold nanoclusters has attracted wide research interest. The strategy consisting in using bulky counterions has been found to dramatically enhance the fluorescence. In this Communication, we push forward this concept to the nonlinear optical regime. We show that by an appropriate choice of bulky counterions and of solvent, a 30-fold increase in two-photon excited fluorescence (TPEF) signal at ≈600 nm for gold nanoclusters can be obtained. This would correspond to a TPEF cross-section in the range of 0.1 to 1 GM. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Steady state and time-resolved fluorescence spectroscopy of quinine sulfate dication bound to sodium dodecylsulfate micelles: Fluorescent complex formation

Energy Technology Data Exchange (ETDEWEB)

Joshi, Sunita; Pant, Debi D., E-mail: ddpant@pilani.bits-pilani.ac.in

2014-01-15

Interaction of quinine sulfate dication (QSD) with anionic, sodium dodecylsulphate (SDS) surfactant has been studied at different premicellar, micellar and postmicellar concentrations in aqueous phase using steady state, time-resolved fluorescence and fluorescence anisotropy techniques. At premicellar concentrations of SDS, the decrease in absorbance, appearance of an extra fluorescence band at lower wavelengths and tri-exponential decay behavior of fluorescence, are attributed to complex formation between QSD molecules and surfactant monomers. At postmicellar concentrations the red shift in fluorescence spectrum, increase in quantum yield and increase in fluorescence lifetimes are attributed to incorporation of solute molecules to micelles. At lower concentrations of SDS, a large shift in fluorescence is observed on excitation at the red edge of absorption spectrum and this is explained in terms of distribution of ion pairs of different energies in the ground state and the observed fluorescence lifetime behavior corroborates with this model. The temporal fluorescence anisotropy decay of QSD in SDS micelles allowed determination of restriction on the motion of the fluorophore. All the different techniques used in this study reveal that the photophysics of QSD is very sensitive to the microenvironments of SDS micelles and QSD molecules reside at the water-micelle interface. -- Highlights: • Probe molecule is very sensitive to microenvironment of micelles. • Highly fluorescent ion-pair formation has been observed. • Modulated photophysics of probe molecule in micellar solutions has been observed. • Probe molecules strongly bind with micelles and reside at probe–micelle interface.

Micelle-assisted signaling of peracetic acid by the oxidation of pyreneboronic acid via monomer-excimer switching.

Science.gov (United States)

Choi, Jiyoung; Lee, Hyo Jin; Cho, Min Jeoung; Chang, Suk-Kyu

2015-08-15

A simple fluorescent probe for the industrial oxidant peracetic acid (PAA) was investigated. PAA-assisted oxidative conversion of pyrene-1-boronic acid into 1-hydroxypyrene was used as the signaling tool. Pyreneboronic acid was found to display selective signaling behavior, being more responsive to PAA than to other commonly used practical oxidants such as H2O2 and HOCl. The changes in pyrene monomer fluorescence to excimer were used in the quantitative analysis of PAA. When using the surfactant hexadecyltrimethylammonium bromide as a micellar additive, the signaling of PAA was markedly enhanced. Selective fluorescence signaling of PAA by pyrene-1-boronic acid with a detection limit of 1.5×10(-6)M in aqueous environment was successfully achieved. Copyright © 2015 Elsevier B.V. All rights reserved.

Development of a dielectrophoresis-assisted surface plasmon resonance fluorescence biosensor for detection of bacteria

Science.gov (United States)

Kuroda, Chiaki; Iizuka, Ryota; Ohki, Yoshimichi; Fujimaki, Makoto

2018-05-01

To detect biological substances such as bacteria speedily and accurately, a dielectrophoresis-assisted surface plasmon resonance (SPR) fluorescence biosensor is being developed. Using Escherichia coli as a target organism, an appropriate voltage frequency to collect E. coli cells on indium tin oxide quadrupole electrodes by dielectrophoresis is analyzed. Then, E. coli is stained with 4‧,6-diamidino-2-phenylindole (DAPI). To clearly detect fluorescence signals from DAPI-stained E. coli cells, the sensor is optimized so that we can excite SPR on Al electrodes by illuminating 405 nm photons. As a result, the number of fluorescence signals is increased on the electrodes by the application of a low-frequency voltage. This indicates that E. coli cells with a lower permittivity than the surrounding water are collected by negative dielectrophoresis onto the electrodes where the electric field strength is lowest.

Intrinsic fluorescence biomarkers in cells treated with chemopreventive drugs

Science.gov (United States)

Kirkpatrick, Nathaniel D.; Brands, William R.; Zou, Changping; Brewer, Molly A.; Utzinger, Urs

2005-03-01

Non-invasive monitoring of cellular metabolism offers promising insights into areas ranging from biomarkers for drug activity to cancer diagnosis. Fluorescence spectroscopy can be utilized in order to exploit endogenous fluorophores, typically metabolic co-factors nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD), and estimate the redox status of the sample. Fluorescence spectroscopy was applied to follow metabolic changes in epithelial ovarian cells as well as bladder epithelial cancer cells during treatment with a chemopreventive drug that initiates cellular quiescence. Fluorescence signals consistent with NADH, FAD, and tryptophan were measured to monitor cellular activity, redox status, and protein content. Cells were treated with varying concentrations of N-4-(hydroxyphenyl) retinamide (4-HPR) and measured in a stable environment with a sensitive fluorescence spectrometer. A subset of measurements was completed on a low concentration of cells to demonstrate feasibility for medical application such as in bladder or ovary washes. Results suggest that all of the cells responded with similar dose dependence but started at different estimated redox ratio baseline levels correlating with cell cycle, growth inhibition, and apoptosis assays. NADH and tryptophan related fluorescence changed significantly while FAD related fluorescence remained unaltered. Fluorescence data collected from approximately 1000 - 2000 cells, comparable to a bladder or ovary wash, was measurable and useful for future experiments. This study suggests that future intrinsic biomarker measurements may need to be most sensitive to changes in NADH and tryptophan related fluorescence while using FAD related fluorescence to help estimate the baseline redox ratio and predict response to chemopreventive agents.

Smartphone-based fluorescence spectroscopy device aiding in preliminary skin screening

Science.gov (United States)

Sahoo, Aparajita; Wahi, Akshat; Das, Anshuman

2018-02-01

Preliminary diagnosis of closely resembling skin conditions can be highly subjective for dermatologists. In ambiguous cases, it often leads to performing invasive procedures like biopsies. Different skin conditions, however, have varying concentrations of fluorophores (like collagen, NADH) and chromophores (like melanin, hemoglobin) which can alter their fluorescence spectra. We demonstrate a handheld, portable, smartphone-based spectrometer that leverages these alterations in skin autofluorescence spectra for rapid screening of skin conditions. This methodology involves excitation of affected skin areas with ultraviolet (UV-A) 385 nm light, capturing the generated fluorescence spectra and sending the data wirelessly to a companion mobile application for data storage, analysis and visualization. By collecting the fluorescence spectral signals from healthy and unhealthy skin conditions, we establish that the signals collected using this portable device can be used to develop a classification method to help in differentially diagnosing these conditions. It shows promise as a useful skin screening tool for both dermatologists and primary health care workers. This device can enable quick, non-invasive and a more objective preliminary examination. We envision the device to be especially useful in primary healthcare centers of developing countries where availability of dermatologists is limited.

Ratiometric fluorescent receptors for both Zn2+ and H2PO4(-) ions based on a pyrenyl-linked triazole-modified homooxacalix[3]arene: a potential molecular traffic signal with an R-S latch logic circuit.

Science.gov (United States)

Ni, Xin-long; Zeng, Xi; Redshaw, Carl; Yamato, Takehiko

2011-07-15

A ratiometric fluorescent receptor with a C(3) symmetric structure based on a pyrene-linked triazole-modified homooxacalix[3]arene (L) was synthesized and characterized. This system exhibited an interesting ratiometric detection signal output for targeting cations and anions through switching the excimer emission of pyrene from the "on-off" to the "off-on" type in neutral solution. (1)H NMR titration results suggested that the Zn(2+) center of receptor L·Zn(2+) provided an excellent pathway of organizing anion binding groups for optimal host-guest interactions. It is thus believed that this receptor has potential application in sensing, detection, and recognition of both Zn(2+) and H(2)PO(4)(-) ions with different optical signals. In addition, the fluorescence emission changes by the inputs of Zn(2+) and H(2)PO(4)(-) ions can be viewed as a combinational R-S latch logic circuit at the molecular level.

Graphitic Carbon Nitride Nanosheets-Based Ratiometric Fluorescent Probe for Highly Sensitive Detection of H2O2 and Glucose.

Science.gov (United States)

Liu, Jin-Wen; Luo, Ying; Wang, Yu-Min; Duan, Lu-Ying; Jiang, Jian-Hui; Yu, Ru-Qin

2016-12-14

Graphitic carbon nitride (g-C 3 N 4 ) nanosheets, an emerging graphene-like carbon-based nanomaterial with high fluorescence and large specific surface areas, hold great potential for biosensor applications. Current g-C 3 N 4 nanosheets based fluorescent biosensors majorly rely on single fluorescent intensity reading through fluorescence quenching interactions between the nanosheets and metal ions. Here we report for the first time the development of a novel g-C 3 N 4 nanosheets-based ratiometric fluorescence sensing strategy for highly sensitive detection of H 2 O 2 and glucose. With o-phenylenediamine (OPD) oxidized by H 2 O 2 in the presence of horseradish peroxidase (HRP), the oxidization product can assemble on the g-C 3 N 4 nanosheets through hydrogen bonding and π-π stacking, which effectively quenches the fluorescence of g-C 3 N 4 while delivering a new emission peak. The ratiometric signal variations enable robust and sensitive detection of H 2 O 2 . On the basis of the glucose converting into H 2 O 2 through the catalysis of glucose oxidase, the g-C 3 N 4 -based ratiometric fluorescence sensing platform is also exploited for glucose assay. The developed strategy is demonstrated to give a detection limit of 50 nM for H 2 O 2 and 0.4 μM for glucose, at the same time, it has been successfully used for glucose levels detection in human serum. This strategy may provide a cost-efficient, robust, and high-throughput platform for detecting various species involving H 2 O 2 -generation reactions for biomedical applications.

Rapid Cellular Phenotyping of Human Pluripotent Stem Cell-Derived Cardiomyocytes using a Genetically Encoded Fluorescent Voltage Sensor

Directory of Open Access Journals (Sweden)

Jordan S. Leyton-Mange

2014-02-01

Full Text Available In addition to their promise in regenerative medicine, pluripotent stem cells have proved to be faithful models of many human diseases. In particular, patient-specific stem cell-derived cardiomyocytes recapitulate key features of several life-threatening cardiac arrhythmia syndromes. For both modeling and regenerative approaches, phenotyping of stem cell-derived tissues is critical. Cellular phenotyping has largely relied upon expression of lineage markers rather than physiologic attributes. This is especially true for cardiomyocytes, in part because electrophysiological recordings are labor intensive. Likewise, most optical voltage indicators suffer from phototoxicity, which damages cells and degrades signal quality. Here we present the use of a genetically encoded fluorescent voltage indicator, ArcLight, which we demonstrate can faithfully report transmembrane potentials in human stem cell-derived cardiomyocytes. We demonstrate the application of this fluorescent sensor in high-throughput, serial phenotyping of differentiating cardiomyocyte populations and in screening for drug-induced cardiotoxicity.

Quantitative nitric oxide measurements by means of laser-induced fluorescence in a heavy-duty Diesel engine

NARCIS (Netherlands)

Verbiezen, K.; Vliet, van A.P.; Klein-Douwel, R.J.H.; Ganippa, L.C.; Bougie, H.J.T.; Meerts, W.L.; Dam, N.J.; Meulen, ter J.J.

2005-01-01

Quantitative in-cylinder laser-induced fluorescence measurements ofnitric oxide in a heavy-duty Diesel engine are presented. Special attention is paid to experimental techniques to assess the attenuation of the laser beam and the fluorescence signal by the cylinder contents.This attenuation can be

Signal Processing for Neuroscientists, A Companion Volume Advanced Topics, Nonlinear Techniques and Multi-Channel Analysis

CERN Document Server

van Drongelen, Wim

2010-01-01

The popularity of signal processing in neuroscience is increasing, and with the current availability and development of computer hardware and software, it is anticipated that the current growth will continue. Because electrode fabrication has improved and measurement equipment is getting less expensive, electrophysiological measurements with large numbers of channels are now very common. In addition, neuroscience has entered the age of light, and fluorescence measurements are fully integrated into the researcher's toolkit. Because each image in a movie contains multiple pixels, these measureme

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

Directory of Open Access Journals (Sweden)

Nan Guo

2014-10-01

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

Fluorescence Molecular Tomography: Principles and Potential for Pharmaceutical Research

Directory of Open Access Journals (Sweden)

Florian Stuker

2011-04-01

Full Text Available Fluorescence microscopic imaging is widely used in biomedical research to study molecular and cellular processes in cell culture or tissue samples. This is motivated by the high inherent sensitivity of fluorescence techniques, the spatial resolution that compares favorably with cellular dimensions, the stability of the fluorescent labels used and the sophisticated labeling strategies that have been developed for selectively labeling target molecules. More recently, two and three-dimensional optical imaging methods have also been applied to monitor biological processes in intact biological organisms such as animals or even humans. These whole body optical imaging approaches have to cope with the fact that biological tissue is a highly scattering and absorbing medium. As a consequence, light propagation in tissue is well described by a diffusion approximation and accurate reconstruction of spatial information is demanding. While in vivo optical imaging is a highly sensitive method, the signal is strongly surface weighted, i.e., the signal detected from the same light source will become weaker the deeper it is embedded in tissue, and strongly depends on the optical properties of the surrounding tissue. Derivation of quantitative information, therefore, requires tomographic techniques such as fluorescence molecular tomography (FMT, which maps the three-dimensional distribution of a fluorescent probe or protein concentration. The combination of FMT with a structural imaging method such as X-ray computed tomography (CT or Magnetic Resonance Imaging (MRI will allow mapping molecular information on a high definition anatomical reference and enable the use of prior information on tissue’s optical properties to enhance both resolution and sensitivity. Today many of the fluorescent assays originally developed for studies in cellular systems have been successfully translated for experimental studies in animals. The opportunity of monitoring molecular

A Conjugate of Pentamethine Cyanine and 18F as a Positron Emission Tomography/Near-Infrared Fluorescence Probe for Multimodality Tumor Imaging

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Fei-Fei An

2017-06-01

Full Text Available The novel synthesis of a dual-modality, pentamethine cyanine (Cy5 fluorescent, 18F positron emission tomography (PET imaging probe is reported. The probe shows a large extinction coefficient and large quantum yield in the biologically transparent, near-infrared window (650–900 nm for in vivo fluorescent imaging. This fluorophore bears the isotope, 18F, giving a 18F-PET/near-infrared fluorescent (NIRF, bi-modal imaging probe, that combines the long-term stability of NIRF and the unlimited penetration depth of PET imaging. The bi-modal probe is labeled with 18F in a quick, one-step reaction, which is important in working with the rapid decay of 18F. The bi-modal probe bears a free carboxyl group, highlighting a PET/NIRF synthon that can be conjugated onto many advanced biomolecules for biomarker-specific in vivo dual-modal PET/NIR tumor imaging, confocal histology, and utility in multi-fluorophore, fluorescence-guided surgery. Its potential in vivo biocompatibility is explored in a quick proof-of-principal in vivo study. The dye is delivered to A549 xenograft flank-tumors to generate PET and NIRF signals at the tumor site. The tumor distribution is confirmed in ex vivo gamma counting and imaging. Pentamethine cyanine (Cy5 has the ability to preferentially accumulate in tumor xenografts. We substitute the PET/NIRF probe for Cy5, and explore this phenomenon.

Facile method for CLSM imaging unfunctionalized Au nanoparticles through fluorescent channels

International Nuclear Information System (INIS)

Yuan Lan; Wei Wei; Li Juan; Sun, Zhiwei; Wang Hongfang; Zhang Xiuzhi; Chen Yueyue

2009-01-01

The microscopic visualization of metal nanoparticles has become a useful tool for the investigation of their applications in cell labeling and the study of their bio-effects. In the current study, we have developed a facile method with confocal laser scanning microscope (CLSM) to observe unfunctionalized Au nanoparticles through fluorescent channels. The sharp reflected signal and photostable property of the metal nanoparticles makes the present method very ideal for fluorescent co-localization, real-time imaging, and further quantitative analysis.

A label-free, fluorescence based assay for microarray

Science.gov (United States)

Niu, Sanjun

DNA chip technology has drawn tremendous attention since it emerged in the mid 90's as a method that expedites gene sequencing by over 100-fold. DNA chip, also called DNA microarray, is a combinatorial technology in which different single-stranded DNA (ssDNA) molecules of known sequences are immobilized at specific spots. The immobilized ssDNA strands are called probes. In application, the chip is exposed to a solution containing ssDNA of unknown sequence, called targets, which are labeled with fluorescent dyes. Due to specific molecular recognition among the base pairs in the DNA, the binding or hybridization occurs only when the probe and target sequences are complementary. The nucleotide sequence of the target is determined by imaging the fluorescence from the spots. The uncertainty of background in signal detection and statistical error in data analysis, primarily due to the error in the DNA amplification process and statistical distribution of the tags in the target DNA, have become the fundamental barriers in bringing the technology into application for clinical diagnostics. Furthermore, the dye and tagging process are expensive, making the cost of DNA chips inhibitive for clinical testing. These limitations and challenges make it difficult to implement DNA chip methods as a diagnostic tool in a pathology laboratory. The objective of this dissertation research is to provide an alternative approach that will address the above challenges. In this research, a label-free assay is designed and studied. Polystyrene (PS), a commonly used polymeric material, serves as the fluorescence agent. Probe ssDNA is covalently immobilized on polystyrene thin film that is supported by a reflecting substrate. When this chip is exposed to excitation light, fluorescence light intensity from PS is detected as the signal. Since the optical constants and conformations of ssDNA and dsDNA (double stranded DNA) are different, the measured fluorescence from PS changes for the same

Solvent induced fluorescence enhancement of graphene oxide studied by ultrafast spectroscopy

Science.gov (United States)

Zhao, Litao; Chen, Jinquan; He, Xiaoxiao; Yu, Xiantong; Yan, Shujun; Zhang, Sanjun; Pan, Haifeng; Xu, Jianhua

2018-05-01

Femtosecond transient absorption (TA) spectroscopy combined with picosecond time resolved fluorescence (TRF) were used to reveal the fluorescence kinetics of graphene oxide (GO) in water, ethanol and water-ethanol mixtures. Size-independent fluorescence of GO were observed in water, and pH-dependent fluorescence spectra could be fitted well by a triple emission relaxation with peaks around 440 nm, 500 nm, and 590 nm respectively. The results indicate that polycyclic aromatic hydrocarbons (PAHs) linked by oxygen-containing functional groups dominate GO's fluorescence emission. GO's fluorescence quantum yield was measured to be 2.8% in ethanol but 1.2% in water. The three decay components fluorescence decay, as well as the transient absorption dynamics with an offset, confirmed this solvent induced fluorescence enhancement. GO's Raman spectral signals showed that GO in ethanol has a smaller average size of PAHs than that of GO in water. Therefore, besides other enhancement effects reported in literatures, we proposed that solvents could also change the size of PAHs, resulting in a photoluminescence enhancement. Our experimental data demonstrates that GO's quantum yield could be up to 2.8% in water and 8.4% in ethanol and this observation may help ones to improve GO's photoluminescence efficiency as well as its applications in solution.

A Graphene Oxide-Based Fluorescent Aptasensor for the Turn-on Detection of CCRF-CEM.

Science.gov (United States)

Tan, Jie; Lai, Zongqiang; Zhong, Liping; Zhang, Zhenghua; Zheng, Rong; Su, Jing; Huang, Yong; Huang, Panpan; Song, Hui; Yang, Nuo; Zhou, Sufang; Zhao, Yongxiang

2018-04-01

A convenient, low-cost, and highly sensitive fluorescent aptasensor for detection of leukemia has been developed based on graphene oxide-aptamer complex (GO-apt). Graphene oxide (GO) can absorb carboxyfluorescein-labeled Sgc8 aptamer (FAM-apt) by π-π stacking and quench the fluorescence through fluorescence resonance energy transfer (FRET). In the absence of Sgc8 target cell CCRF-CEM, the fluorescence is almost all quenched. Conversely, when the CCRF-CEM cells are added, the quenched fluorescence can be recovered rapidly and significantly. Therefore, based on the change of fluorescence signals, we can detect the number of CCRF-CEM cells in a wide range from 1 × 10 2 to 1 × 10 7  cells/mL with a limit of detection (LOD) of 10 cells/mL. Therefore, this strategy of graphene oxide-based fluorescent aptasensor may be promising for the detection of cancer.

Self-Tuning Linear Quadratic Supervisory Regulation of a Diesel Generator using Large-Signal State Estimation

DEFF Research Database (Denmark)

Knudsen, Jesper Viese; Bendtsen, Jan Dimon; Andersen, Palle

2016-01-01

In this paper, a self-tuning linear quadratic supervisory regulator using a large-signal state estimator for a diesel driven generator set is proposed. The regulator improves operational efficiency, in comparison to current implementations, by (i) automating the initial tuning process and (ii...... throughout the operating range of the diesel generator....

Fabrication of fluorescent chitosan-containing microcapsules

Directory of Open Access Journals (Sweden)

Zhang R.

2013-08-01

Full Text Available Intense emission peaks of Eu(DBM3Phen (DBM and Phen are dibenzoylmethane and 1,10-phenanthroline, respectively in the microcapsules containing molecules of quaternary ammonium chitosan (QACS and sodium alginate are observed. The microcapsules are assembled by using CaCO3 particles as template cores by the layer-by-layer (LbL technique. Observation of microcapsules by the fluorescence mode and the transmission mode in the confocal laser scanning microscopy shows that the microcapsules are intact after core decomposition. Fluorescence under ultraviolet irradiation comes directly from the Eu(DBM3Phen. Homogeneous assembly of Eu(DBM3Phen can be deduced due to the homogeneous fluorescence of the microcapsules in the fluorescence micrographs. The microcapsules show adherence to solid substrates due to large quantities of hydroxyl groups of QACS. AFM measurements of dried hollow microcapsules with only 4 bilayers of (CS/SA fabricated with Eu(DBM3Phen show the intact shell with a thickness of 3.0 nm. Regarding the biocompatible natural polysaccharides and the intense fluorescence emission, the microcapsules in this work might be of great importance in potential application in drug delivery and bioassay.

Investigation of Membrane Receptors' Oligomers Using Fluorescence Resonance Energy Transfer and Multiphoton Microscopy in Living Cells

Science.gov (United States)

Mishra, Ashish K.

Investigating quaternary structure (oligomerization) of macromolecules (such as proteins and nucleic acids) in living systems (in vivo) has been a great challenge in biophysics, due to molecular diffusion, fluctuations in several biochemical parameters such as pH, quenching of fluorescence by oxygen (when fluorescence methods are used), etc. We studied oligomerization of membrane receptors in living cells by means of Fluorescence (Forster) Resonance Energy Transfer (FRET) using fluorescent markers and two photon excitation fluorescence micro-spectroscopy. Using suitable FRET models, we determined the stoichiometry and quaternary structure of various macromolecular complexes. The proteins of interest for this work are : (1) sigma-1 receptor and (2) rhodopsin, are described as below. (1) Sigma-1 receptors are molecular chaperone proteins, which also regulate ion channels. S1R seems to be involved in substance abuse, as well as several diseases such as Alzheimer's. We studied S1R in the presence and absence of its ligands haloperidol (an antagonist) and pentazocine +/- (an agonist), and found that at low concentration they reside as a mixture of monomers and dimers and that they may form higher order oligomers at higher concentrations. (2) Rhodopsin is a prototypical G protein coupled receptor (GPCR) and is directly involved in vision. GPCRs form a large family of receptors that participate in cell signaling by responding to external stimuli such as drugs, thus being a major drug target (more than 40% drugs target GPCRs). Their oligomerization has been largely controversial. Understanding this may help to understand the functional role of GPCRs oligomerization, and may lead to the discovery of more drugs targeting GPCR oligomers. It may also contribute toward finding a cure for Retinitis Pigmentosa, which is caused by a mutation (G188R) in rhodopsin, a disease which causes blindness and has no cure so far. Comparing healthy rhodopsin's oligomeric structure with that

Fluorescence-based optimization of human bitter taste receptor expression in Saccharomyces cerevisiae

International Nuclear Information System (INIS)

Sugawara, Taishi; Ito, Keisuke; Shiroishi, Mitsunori; Tokuda, Natsuko; Asada, Hidetsugu; Yurugi-Kobayashi, Takami; Shimamura, Tatsuro; Misaka, Takumi; Nomura, Norimichi; Murata, Takeshi; Abe, Keiko; Iwata, So

2009-01-01

Human TAS2 receptors (hTAS2Rs) perceive bitter tastants, but few studies have explored the structure-function relationships of these receptors. In this paper, we report our trials on the large-scale preparations of hTAS2Rs for structural analysis. Twenty-five hTAS2Rs were expressed using a GFP-fusion yeast system in which the constructs and the culture conditions (e.g., the signal sequence, incubation time and temperature after induction) were optimized by measuring GFP fluorescence. After optimization, five hTAS2Rs (hTAS2R7, hTAS2R8, hTAS2R16, hTAS2R41, and hTAS2R48) were expressed at levels greater than 1 mg protein/L of culture, which is a preferable level for purification and crystallization. Among these five bitter taste receptors, hTAS2R41 exhibited the highest detergent solubilization efficiency of 87.1% in n-dodecyl-β-D-maltopyranoside (DDM)/cholesteryl hemisuccinate (CHS). Fluorescence size-exclusion chromatography showed that hTAS2R41 exhibited monodispersity in DDM/CHS without aggregates, suggesting that hTAS2R41 is a good target for future crystallization trials.

Absorption and fluorescence spectroscopic characterization of BLUF domain of AppA from Rhodobacter sphaeroides

Science.gov (United States)

Zirak, P.; Penzkofer, A.; Schiereis, T.; Hegemann, P.; Jung, A.; Schlichting, I.

2005-08-01

The BLUF domain of the transcriptional anti-repressor protein AppA from the non-sulfur anoxyphototrophic purple bacterium Rhodobacter sphaeroides was characterized by absorption and emission spectroscopy. The BLUF domain constructs AppA 148 (consisting of amino-acid residues 1-148) and AppA 126 (amino-acid residues 1-126) are investigated. The cofactor of the investigated domains is found to consist of a mixture of the flavins riboflavin, FMN, and FAD. The dark-adapted domains exist in two different active receptor conformations (receptor states) with different sub-nanosecond fluorescence lifetimes (BLUF r,f and BLUF r,sl) and a small non-interacting conformation (BLUF nc). The active receptor conformations are transformed to putative signalling states (BLUF s,f and BLUF s,sl) of low fluorescence efficiency and picosecond fluorescence lifetime by blue-light excitation (light-adapted domains). In the dark at room temperature both signalling states recover back to the initial receptor states with a time constant of about 17 min. A quantum yield of signalling state formation of about 25% was determined by intensity dependent transmission measurements. A photo-cycle scheme is presented including photo-induced charge transfer complex formation, charge recombination, and protein binding pocket reorganisation.

Absorption and fluorescence spectroscopic characterization of BLUF domain of AppA from Rhodobacter sphaeroides

International Nuclear Information System (INIS)

Zirak, P.; Penzkofer, A.; Schiereis, T.; Hegemann, P.; Jung, A.; Schlichting, I.

2005-01-01

The BLUF domain of the transcriptional anti-repressor protein AppA from the non-sulfur anoxyphototrophic purple bacterium Rhodobacter sphaeroides was characterized by absorption and emission spectroscopy. The BLUF domain constructs AppA 148 (consisting of amino-acid residues 1-148) and AppA 126 (amino-acid residues 1-126) are investigated. The cofactor of the investigated domains is found to consist of a mixture of the flavins riboflavin, FMN, and FAD. The dark-adapted domains exist in two different active receptor conformations (receptor states) with different sub-nanosecond fluorescence lifetimes (BLUF r,f and BLUF r,sl ) and a small non-interacting conformation (BLUF nc ). The active receptor conformations are transformed to putative signalling states (BLUF s,f and BLUF s,sl ) of low fluorescence efficiency and picosecond fluorescence lifetime by blue-light excitation (light-adapted domains). In the dark at room temperature both signalling states recover back to the initial receptor states with a time constant of about 17 min. A quantum yield of signalling state formation of about 25% was determined by intensity dependent transmission measurements. A photo-cycle scheme is presented including photo-induced charge transfer complex formation, charge recombination, and protein binding pocket reorganisation

Microplate-compatible total internal reflection fluorescence microscopy for receptor pharmacology

Science.gov (United States)

Chen, Minghan; Zaytseva, Natalya V.; Wu, Qi; Li, Min; Fang, Ye

2013-05-01

We report the use of total internal reflection fluorescence (TIRF) microscopy for analyzing receptor pharmacology and the development of a microplate-compatible TIRF imaging system. Using stably expressed green fluorescence protein tagged β2-adrenergic receptor as the reporter, we found that the activation of different receptors results in distinct kinetic signatures of the TIRF intensity of cells. These TIRF signatures closely resemble the characteristics of their respective label-free dynamic mass redistribution signals in the same cells. This suggests that TIRF in microplate can be used for profiling and screening drugs.

Method for detecting binding events using micro-X-ray fluorescence spectrometry

Science.gov (United States)

Warner, Benjamin P.; Havrilla, George J.; Mann, Grace

2010-12-28

Method for detecting binding events using micro-X-ray fluorescence spectrometry. Receptors are exposed to at least one potential binder and arrayed on a substrate support. Each member of the array is exposed to X-ray radiation. The magnitude of a detectable X-ray fluorescence signal for at least one element can be used to determine whether a binding event between a binder and a receptor has occurred, and can provide information related to the extent of binding between the binder and receptor.

A ratiometric fluorescent quantum dots based biosensor for organophosphorus pesticides detection by inner-filter effect.

Science.gov (United States)

Yan, Xu; Li, Hongxia; Han, Xiaosong; Su, Xingguang

2015-12-15

In this work, we develop a novel and sensitive sensor for the detection of organophosphorus pesticides based on the inner-filter effect (IFE) between gold nanoparticles (AuNPs) and ratiometric fluorescent quantum dots (RF-QDs). The RF-QDs has been designed by hybridizing two differently colored CdTe QDs, in which the red emissive QDs entrapped in the silica sphere acting as the reference signal, and the green emissive QDs covalently attached on the silica surface serving as the response signal.The fluorescence of RF-QDs could be quenched by AuNPs based on IFE. Protamine could effectively turn on the fluorescence due to the electrostatic attraction between protamine and AuNPs. Trypsin can easily hydrolyze protamine, leading to the quench of the fluorescence. Then, the fluorescence could be recovered again by the addition of parathion-methyl (PM) which could inhibit the activity of trypsin. By measuring the fluorescence of RF-QDs, the inhibition efficiency of PM to trypsin activity was evaluated. Under the optimized conditions, the inhibition efficiency was proportional to the logarithm of PM concentration in the range of 0.04-400 ng mL(-1), with a detection limit of 0.018 ng mL(-1). Furthermore, the simple and convenient method had been used for PM detection in environmental and agricultural samples with satisfactory results. Copyright © 2015 Elsevier B.V. All rights reserved.

Scanless multitarget-matching multiphoton excitation fluorescence microscopy

Directory of Open Access Journals (Sweden)

Junpeng Qiu

2018-03-01

Full Text Available Using the combination of a reflective blazed grating and a reflective phase-only diffractive spatial light modulator (SLM, scanless multitarget-matching multiphoton excitation fluorescence microscopy (SMTM-MPM was achieved. The SLM shaped an incoming mode-locked, near-infrared Ti:sapphire laser beam into an excitation pattern with addressable shapes and sizes that matched the samples of interest in the field of view. Temporal and spatial focusing were simultaneously realized by combining an objective lens and a blazed grating. The fluorescence signal from illuminated areas was recorded by a two-dimensional sCMOS camera. Compared with a conventional temporal focusing multiphoton microscope, our microscope achieved effective use of the laser power and decreased photodamage with higher axial resolution.

AUTOMATED CELL SEGMENTATION WITH 3D FLUORESCENCE MICROSCOPY IMAGES.

Science.gov (United States)

Kong, Jun; Wang, Fusheng; Teodoro, George; Liang, Yanhui; Zhu, Yangyang; Tucker-Burden, Carol; Brat, Daniel J

2015-04-01

A large number of cell-oriented cancer investigations require an effective and reliable cell segmentation method on three dimensional (3D) fluorescence microscopic images for quantitative analysis of cell biological properties. In this paper, we present a fully automated cell segmentation method that can detect cells from 3D fluorescence microscopic images. Enlightened by fluorescence imaging techniques, we regulated the image gradient field by gradient vector flow (GVF) with interpolated and smoothed data volume, and grouped voxels based on gradient modes identified by tracking GVF field. Adaptive thresholding was then applied to voxels associated with the same gradient mode where voxel intensities were enhanced by a multiscale cell filter. We applied the method to a large volume of 3D fluorescence imaging data of human brain tumor cells with (1) small cell false detection and missing rates for individual cells; and (2) trivial over and under segmentation incidences for clustered cells. Additionally, the concordance of cell morphometry structure between automated and manual segmentation was encouraging. These results suggest a promising 3D cell segmentation method applicable to cancer studies.

Kinetics of bacterial fluorescence staining with 3,3'-diethylthiacyanine.

Science.gov (United States)

Thomas, Marlon S; Nuñez, Vicente; Upadhyayula, Srigokul; Zielins, Elizabeth R; Bao, Duoduo; Vasquez, Jacob M; Bahmani, Baharak; Vullev, Valentine I

2010-06-15

For more than a century, colorimetric and fluorescence staining have been the foundation of a broad range of key bioanalytical techniques. The dynamics of such staining processes, however, still remains largely unexplored. We investigated the kinetics of fluorescence staining of two gram-negative and two gram-positive species with 3,3'-diethylthiacyanine (THIA) iodide. An increase in the THIA fluorescence quantum yield, induced by the bacterial dye uptake, was the principal reason for the observed emission enhancement. The fluorescence quantum yield of THIA depended on the media viscosity and not on the media polarity, which suggested that the microenvironment of the dye molecules taken up by the cells was restrictive. The kinetics of fluorescence staining did not manifest a statistically significant dependence neither on the dye concentration, nor on the cell count. In the presence of surfactant additives, however, the fluorescence-enhancement kinetic patterns manifested species specificity with statistically significant discernibility.

Applying fluorescence correlation spectroscopy to investigate peptide-induced membrane disruption

DEFF Research Database (Denmark)

Kristensen, Kasper; Henriksen, Jonas Rosager; Andresen, Thomas Lars

2017-01-01

to quantify leakage of fluorescent molecules of different sizes from large unilamellar lipid vesicles, thereby providing a tool for estimating the size of peptide-induced membrane disruptions. If fluorescently labeled lipids are incorporated into the membranes of the vesicles, FCS can also be used to obtain...

Automated detection of fluorescent cells in in-resin fluorescence sections for integrated light and electron microscopy.

Science.gov (United States)

Delpiano, J; Pizarro, L; Peddie, C J; Jones, M L; Griffin, L D; Collinson, L M

2018-04-26

Integrated array tomography combines fluorescence and electron imaging of ultrathin sections in one microscope, and enables accurate high-resolution correlation of fluorescent proteins to cell organelles and membranes. Large numbers of serial sections can be imaged sequentially to produce aligned volumes from both imaging modalities, thus producing enormous amounts of data that must be handled and processed using novel techniques. Here, we present a scheme for automated detection of fluorescent cells within thin resin sections, which could then be used to drive automated electron image acquisition from target regions via 'smart tracking'. The aim of this work is to aid in optimization of the data acquisition process through automation, freeing the operator to work on other tasks and speeding up the process, while reducing data rates by only acquiring images from regions of interest. This new method is shown to be robust against noise and able to deal with regions of low fluorescence. © 2018 The Authors. Journal of Microscopy published by JohnWiley & Sons Ltd on behalf of Royal Microscopical Society.

Characterization of excited-state reactions with instant spectra of fluorescence kinetics

International Nuclear Information System (INIS)

Tomin, Vladimir I.; Ushakou, Dzmitryi V.

2015-01-01

Comprehensible knowledge of the excited-state proton transfer processes in organic compounds is overwhelmingly important not only for physics, but also chemistry and Life Sciences, since they play a key role in main processes of photosynthesis and functioning of biological organisms. Moreover compounds with Excited-State Intramolecular Proton Transfer (ESIPT) are in the focus of the interest of scientists throughout the world, because dual fluorescence spectra of such objects corresponding to two forms of molecular structure (normal and photoproduct) are very sensitive to characteristics of molecular microenvironment. This property allows to use such substances as fluorescent probes for diverse applications in chemistry and Life Sciences. But at the same time studying of proton transfer processes is not simple, because this process is characterized by extremely fast times (on picoseconds time scale and less order) and very often contribution of reverse reactions is essentially complicates an interpretation of observed properties of dual fluorescence. Hence, understanding of a role of reversible reactions is crucial for a comprehensive description of all processes accompanying excited state reactions. We discuss new approach for treatment ESIPT reaction on the basis of experimentally measured instant spectra of dual fluorescence and temporal behavior of ratiometric signal of normal to tautomer form intensities. Simple analytical expressions show in transparent way how to distinguish a degree of reverse reaction contribution to ratiometric signal. A validation of the approach under consideration is fulfilled with two different flavonols – 3-hydroxyflavone and 4′-(Dimethylamino)-3-hydroxyflavone – representing two extreme cases in affecting reversible reaction on dual emission. A comparing of new approach and traditional method when we analyze kinetics of separate the N* and T* fluorescence bands decays, has been carried out. - Highlights: • The excited

Characterization of excited-state reactions with instant spectra of fluorescence kinetics

Energy Technology Data Exchange (ETDEWEB)

Tomin, Vladimir I., E-mail: tomin@apsl.edu.pl; Ushakou, Dzmitryi V.

2015-10-15

Comprehensible knowledge of the excited-state proton transfer processes in organic compounds is overwhelmingly important not only for physics, but also chemistry and Life Sciences, since they play a key role in main processes of photosynthesis and functioning of biological organisms. Moreover compounds with Excited-State Intramolecular Proton Transfer (ESIPT) are in the focus of the interest of scientists throughout the world, because dual fluorescence spectra of such objects corresponding to two forms of molecular structure (normal and photoproduct) are very sensitive to characteristics of molecular microenvironment. This property allows to use such substances as fluorescent probes for diverse applications in chemistry and Life Sciences. But at the same time studying of proton transfer processes is not simple, because this process is characterized by extremely fast times (on picoseconds time scale and less order) and very often contribution of reverse reactions is essentially complicates an interpretation of observed properties of dual fluorescence. Hence, understanding of a role of reversible reactions is crucial for a comprehensive description of all processes accompanying excited state reactions. We discuss new approach for treatment ESIPT reaction on the basis of experimentally measured instant spectra of dual fluorescence and temporal behavior of ratiometric signal of normal to tautomer form intensities. Simple analytical expressions show in transparent way how to distinguish a degree of reverse reaction contribution to ratiometric signal. A validation of the approach under consideration is fulfilled with two different flavonols – 3-hydroxyflavone and 4′-(Dimethylamino)-3-hydroxyflavone – representing two extreme cases in affecting reversible reaction on dual emission. A comparing of new approach and traditional method when we analyze kinetics of separate the N* and T* fluorescence bands decays, has been carried out. - Highlights: • The excited

Fluorescent Pluronic nanodots for in vivo two-photon imaging

International Nuclear Information System (INIS)

Maurin, Mathieu; Vurth, Laeticia; Vial, Jean-Claude; Baldeck, Patrice; Stephan, Olivier; Marder, Seth R; Sanden, Boudewijn Van der

2009-01-01

We report the synthesis of new nanosized fluorescent probes based on bio-compatible polyethylene-polypropylene glycol (Pluronic) materials. In aqueous solution, mini-emulsification of Pluronic with a high fluorescent di-stryl benzene-modified derivative, exhibiting a two-photon absorption cross section as high as 2500 Goeppert-Mayer units at 800 nm, leads to nanoparticles exhibiting a hydrodynamic radius below 100 nm. We have demonstrated that these new probes with luminescence located in the spectral region of interest for bio-imaging (the yellow part of the visible spectrum) allow deep (500 μm) bio-imaging of the mice brain vasculature. The dose injected during our experiments is ten times lower when compared to the classical commercial rhodamine-B isothicyanate-Dextran system but gives similar results to homogeneous blood plasma staining. The mean fluorescent signal intensity stayed constant during more than 1 h.

Simultaneous resolution of spectral and temporal properties of UV and visible fluorescence using single-photon counting with a position-sensitive detector

International Nuclear Information System (INIS)

Kelly, L.A.; Trunk, J.G.; Polewski, K.; Sutherland, J.C.

1995-01-01

A new fluorescence spectrometer has been assembled at the U9B beamline of the National Synchrotron Light Source to allow simultaneous multiwavelength and time-resolved fluorescence detection, as well as spatial imaging of the sample fluorescence. The spectrometer employs monochromatized, tunable UV and visible excitation light from a synchrotron bending magnet and an imaging spectrograph equipped with a single-photon sensitive emission detector. The detector is comprised of microchannel plates in series, with a resistive anode for encoding the position of the photon-derived current. The centroid position of the photon-induced electron cascade is derived in a position analyzer from the four signals measured at the corners of the resistive anode. Spectral information is obtained by dispersing the fluorescence spectrum across one dimension of the detector photocathode. Timing information is obtained by monitoring the voltage divider circuit at the last MCP detector. The signal from the MCP is used as a ''start'' signal to perform a time-correlated single photon counting experiment. The analog signal representing the position, and hence wavelength, is digitized concomitantly with the start/stop time difference and stored in the two-dimensional histogramming memory of a multiparameter analyzer

Highly selective ratiometric fluorescent detection of Fe{sup 3+} with a polyphenyl derivative

Energy Technology Data Exchange (ETDEWEB)

Li, Zhan-Xian, E-mail: lizx@zzu.edu.cn [The College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou 450001 (China); Zhou, Wan; Zhang, Li-Feng; Yuan, Rui-Li; Liu, Xing-Jiang; Wei, Liu-He [The College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou 450001 (China); Yu, Ming-Ming, E-mail: yumm@zzu.edu.cn [The College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou 450001 (China)

2013-04-15

Compared with other fluorescent probes, ratiometric fluorescence responses are more attractive because the ratio between the two emission intensities can be used to measure the analyte concentration and provide a built-in correction for environmental effects. A highly selective and sensitive ratiometric fluorescent probe for Fe{sup 3+} was synthesized, which exhibits an enhanced fluorescence with a large red-shift in emission from 361 to 455 nm upon addition of Fe{sup 3+}. The red-shift of the emission peak can be ascribed to the reformed orbital, and the increase of emission intensity may be ascribed to the inhibition of the rotation of C–C bonds between each two aromatic rings. -- Graphical abstract: A highly selective and sensitive ratiometric fluorescent probe for Fe{sup 3+} was synthesized, which exhibits an enhanced fluorescence with a large red-shift in emission from 361 to 455 nm upon addition of Fe{sup 3+}. Highlights: ► A ratiometric fluorescent probe for Fe{sup 3+} was synthesized. ► The probe exhibits an enhanced fluorescence with a red-shift upon addition of Fe{sup 3+}. ► Inhibition of the rotation of C–C bonds was possible detection mechanism for Fe{sup 3+}.

Scanning fluorescence detector for high-throughput DNA genotyping

Science.gov (United States)

Rusch, Terry L.; Petsinger, Jeremy; Christensen, Carl; Vaske, David A.; Brumley, Robert L., Jr.; Luckey, John A.; Weber, James L.

1996-04-01

A new scanning fluorescence detector (SCAFUD) was developed for high-throughput genotyping of short tandem repeat polymorphisms (STRPs). Fluorescent dyes are incorporated into relatively short DNA fragments via polymerase chain reaction (PCR) and are separated by electrophoresis in short, wide polyacrylamide gels (144 lanes with well to read distances of 14 cm). Excitation light from an argon laser with primary lines at 488 and 514 nm is introduced into the gel through a fiber optic cable, dichroic mirror, and 40X microscope objective. Emitted fluorescent light is collected confocally through a second fiber. The confocal head is translated across the bottom of the gel at 0.5 Hz. The detection unit utilizes dichroic mirrors and band pass filters to direct light with 10 - 20 nm bandwidths to four photomultiplier tubes (PMTs). PMT signals are independently amplified with variable gain and then sampled at a rate of 2500 points per scan using a computer based A/D board. LabView software (National Instruments) is used for instrument operation. Currently, three fluorescent dyes (Fam, Hex and Rox) are simultaneously detected with peak detection wavelengths of 543, 567, and 613 nm, respectively. The detection limit for fluorescein-labeled primers is about 100 attomoles. Planned SCAFUD upgrades include rearrangement of laser head geometry, use of additional excitation lasers for simultaneous detection of more dyes, and the use of detector arrays instead of individual PMTs. Extensive software has been written for automatic analysis of SCAFUD images. The software enables background subtraction, band identification, multiple- dye signal resolution, lane finding, band sizing and allele calling. Whole genome screens are currently underway to search for loci influencing such complex diseases as diabetes, asthma, and hypertension. Seven production SCAFUDs are currently in operation. Genotyping output for the coming year is projected to be about one million total genotypes (DNA

A novel fluorescent biosensor for Adenosine Triphosphate detection based on the polydopamine nanospheres integrating with enzymatic recycling amplification.

Science.gov (United States)

Ji, Xiaoting; Yi, Bingqing; Xu, Yujuan; Zhao, Yanan; Zhong, Hua; Ding, Caifeng

2017-07-01

Based on the protective performance of polydopamine nanospheres (PDANSs) for DNA against nuclease digestion and the specific recognition characteristic of aptamer, we have developed an enzymatic recycling signal amplification method for highly sensitive and selective detection of adenosine triphosphate (ATP). Fluorescence measurements were carried out to verify the DNA polymerase and exonuclease III (Exo III) assisted target recycling process and fluorescence signal amplification. In the absence of the ATP, initially, the signal DNA-PDANSs complex was in the "off" state due to the efficient fluorescence quenching of 6-carboxyfluorescein (FAM) adjacent to the surface of PDANSs. Due to the binding of the aptamer by ATP, it trigger DNA polymerase and Exo III assisted target recycling process by the product of release, the complex would change into the "on" state as a result of the dissociation of the FAM from the surface of PDANSs, thus providing greatly enhanced fluorescence emission intensity. The method allows quantitative detection of ATP in the range of 20-600nM with a detection limit of 8.32nM. This biosensor requires no complex operations, and is a new high efficiency method for ATP detection. Copyright © 2017 Elsevier B.V. All rights reserved.

Modulation of a fluorescence switch based on photochromic spirooxazine in composite organic nanoparticles

International Nuclear Information System (INIS)

Sheng Xiaohai; Peng Aidong; Fu Hongbing; Liu Yuanyuan; Zhao Yongsheng; Ma Ying; Yao Jiannian

2007-01-01

We describe a versatile and convenient approach to achieve fluorescence modulation by the preparation of composite nanoparticles (CNPs), based on photochromic 5-methoxy-1,3,3-trimethyl-9'-hydroxyspiroindolinenaphthoxazine (SO), fluorescent 4-(dicyanomethylene)-2-methyl-6-(p-dimethyl-aminostyryl)-4H-pyran (DCM), and emissive-assistant 1,3-bis(pyrene) propane (BPP) molecules, employing doping techniques. The mechanism of the fluorescence switch is the intermolecular energy transfer as supported by both steady-state and time-resolved spectroscopy results. The addition of BPP not only enhances the contrast of the fluorescence signal between the 'ON' and 'OFF' state, but also provides a convenient way to tune the excitation wavelength for reading the fluorescence. High-contrast ON/OFF (20:1) fluorescence switching is successfully implemented in the CNPs and also in a more practical PVA film loaded with the CNPs. This system may represent an alternative to the covalent system in potentially rewritable high-density optical data or image storage utilizing luminescence intensity readout schemes

Three-dimensional in vivo fluorescence diffuse optical tomography of breast cancer in humans

Science.gov (United States)

Corlu, Alper; Choe, Regine; Durduran, Turgut; Rosen, Mark A.; Schweiger, Martin; Arridge, Simon R.; Schnall, Mitchell D.; Yodh, Arjun G.

2007-05-01

We present three-dimensional (3D) in vivo images of human breast cancer based on fluorescence diffuse optical tomography (FDOT). To our knowledge, this work represents the first reported 3D fluorescence tomography of human breast cancer in vivo. In our protocol, the fluorophore Indocyanine Green (ICG) is injected intravenously. Fluorescence excitation and detection are accomplished in the soft-compression, parallel-plane, transmission geometry using laser sources at 786 nm and spectrally filtered CCD detection. Phantom and in vivo studies confirm the signals are due to ICG fluorescence, rather than tissue autofluorescence and excitation light leakage. Fluorescence images of breast tumors were in good agreement with those of MRI, and with DOT based on endogenous contrast. Tumorto- normal tissue contrast based on ICG fluorescence was two-to-four-fold higher than contrast based on hemoglobin and scattering parameters. In total the measurements demonstrate that FDOT of breast cancer is feasible and promising.

Super-resolution fluorescence microscopy by stepwise optical saturation

Science.gov (United States)

Zhang, Yide; Nallathamby, Prakash D.; Vigil, Genevieve D.; Khan, Aamir A.; Mason, Devon E.; Boerckel, Joel D.; Roeder, Ryan K.; Howard, Scott S.

2018-01-01

Super-resolution fluorescence microscopy is an important tool in biomedical research for its ability to discern features smaller than the diffraction limit. However, due to its difficult implementation and high cost, the super-resolution microscopy is not feasible in many applications. In this paper, we propose and demonstrate a saturation-based super-resolution fluorescence microscopy technique that can be easily implemented and requires neither additional hardware nor complex post-processing. The method is based on the principle of stepwise optical saturation (SOS), where M steps of raw fluorescence images are linearly combined to generate an image with a M-fold increase in resolution compared with conventional diffraction-limited images. For example, linearly combining (scaling and subtracting) two images obtained at regular powers extends the resolution by a factor of 1.4 beyond the diffraction limit. The resolution improvement in SOS microscopy is theoretically infinite but practically is limited by the signal-to-noise ratio. We perform simulations and experimentally demonstrate super-resolution microscopy with both one-photon (confocal) and multiphoton excitation fluorescence. We show that with the multiphoton modality, the SOS microscopy can provide super-resolution imaging deep in scattering samples. PMID:29675306

Probing GFP-actin diffusion in living cells using fluorescence correlation spectroscopy

International Nuclear Information System (INIS)

Engelke, Hanna; Heinrich, Doris; Rädler, Joachim O.

2010-01-01

The cytoskeleton of eukaryotic cells is continuously remodeled by polymerization and depolymerization of actin. Consequently, the relative content of polymerized filamentous actin (F-actin) and monomeric globular actin (G-actin) is subject to temporal and spatial fluctuations. Since fluorescence correlation spectroscopy (FCS) can measure the diffusion of fluorescently labeled actin it seems likely that FCS allows us to determine the dynamics and hence indirectly the structural properties of the cytoskeleton components with high spatial resolution. To this end we investigate the FCS signal of GFP-actin in living Dictyostelium discoideum cells and explore the inherent spatial and temporal signatures of the actin cytoskeleton. Using the free green fluorescent protein (GFP) as a reference, we find that actin diffusion inside cells is dominated by G-actin and slower than diffusion in diluted cell extract. The FCS signal in the dense cortical F-actin network near the cell membrane is probed using the cytoskeleton protein LIM and is found to be slower than cytosolic G-actin diffusion. Furthermore, we show that polymerization of the cytoskeleton induced by Jasplakinolide leads to a substantial decrease of G-actin diffusion. Pronounced fluctuations in the distribution of the FCS correlation curves can be induced by latrunculin, which is known to induce actin waves. Our work suggests that the FCS signal of GFP-actin in combination with scanning or spatial correlation techniques yield valuable information about the local dynamics and concomitant cytoskeletal properties

Electron detachment dissociation of fluorescently labeled sialylated oligosaccharides.

Science.gov (United States)

Zhou, Wen; Håkansson, Kristina

2011-12-01

We explored the application of electron detachment dissociation (EDD) and infrared multiphoton dissociation (IRMPD) tandem mass spectrometry to fluorescently labeled sialylated oligosaccharides. Standard sialylated oligosaccharides and a sialylated N-linked glycan released from human transferrin were investigated. EDD yielded extensive glycosidic cleavages and cross-ring cleavages in all cases studied, consistently providing complementary structural information compared with infrared multiphoton dissociation. Neutral losses and satellite ions such as C-2H ions were also observed following EDD. In addition, we examined the influence of different fluorescent labels. The acidic label 2-aminobenzoic acid (2-AA) enhanced signal abundance in negative-ion mode. However, few cross-ring fragments were observed for 2-AA-labeled oligosaccharides. The neutral label 2-aminobenzamide (2-AB) resulted in more cross-ring cleavages compared with 2-AA-labeled species, but not as extensive fragmentation as for native oligosaccharides, likely resulting from altered negative charge locations from introduction of the fluorescent tag. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The TUS space fluorescence detector for study of UHECR and other phenomena of variable fluorescence light in the atmosphere

Science.gov (United States)

Abrashkin, V.; Alexandrov, V.; Arakcheev, Y.; Bitkin, E.; Cordero, A.; Eremin, S.; Finger, M.; Garipov, G.; Grebenyuk, V.; Kalmykov, N.; Khrenov, B.; Koval, V.; Martinez, O.; Matyushkin, A.; Moreno, E.; Naumov, D.; Olshevsky, A.; Panasyuk, M.; Park, I.; Robledo, C.; Rubinstein, I.; Sharakin, S.; Silaev, A.; Tkatchev, L.; Tulupov, V.; Tyukaev, R.; Sabirov, B.; Salazar, H.; Saprykin, O.; Syromyatnikov, V.; Urmantsev, F.; Villasenor, L.; Yashin, I.; Zaikin, N.; Zepeda, A.

The Tracking Ultraviolet Set Up (TUS) instrument has been designed to observe from space the fluorescence light in the atmosphere when Extensive Air Shower (EAS) or other phenomena such as meteors or dust grains traverse it. The TUS design concepts will allow us to construct the next generation of fluorescence detectors with increasing light collection power and higher resolution. The KLYPVE instrument with collection power 5 times larger of the TUS will be the next space detector. Light collection is obtained with the help of segmented “low frequency Fresnel type” mirrors. Photo receiver retina in the focal consists of modules of PM tubes. For stable performance in conditions of variable light noise and variable temperature the tube type with a multi-alcali cathode was chosen. Voltage supplies for PMT in one module were designed for keeping the performance of photo receiver retina uniform when the tube gain change. From every tube the signal amplitude is recorded in time bins of 400 ns. The digital data are kept and analyzed in the module FPGA connected to the central FPGA controlling all data. The RAM memory is large, capable to record events with different duration of the light signal (up to several seconds). The preliminary event data are analyzed in the triggering system of the central FPGA. The trigger criteria have several options for events of different origin (different pixel signal duration). The trigger integration time is controlled from the space mission center. The performances of the detector were simulated and zenith angle dependent trigger efficiencies were calculated. The TUS detector will be efficient in recording “horizontal” EAS (zenith angles more than 60°), developed to their maximum above the cloud cover. The EAS Cherenkov light, back scattered from the cloud cover, will be recorded and will improve data on the EAS direction and position of maximum. For better accuracy in physical parameters of the events and for the experimental

The Effect of a Fluorophore Photo-Physics on the Lipid Vesicle Diffusion Coefficient Studied by Fluorescence Correlation Spectroscopy.

Science.gov (United States)

Drabik, Dominik; Przybyło, Magda; Sikorski, Aleksander; Langner, Marek

2016-03-01

Fluorescence Correlation Spectroscopy (FCS) is a technique, which allows determination of the diffusion coefficient and concentration of fluorescent objects suspended in the solution. The measured parameter is the fluctuation of the fluorescence signal emitted by diffusing molecules. When 100 nm DOPC vesicles labeled with various fluorescent dyes (Fluorescein-PE, NBD-PE, Atto488 DOPE or βBodipy FL) were measured, different values of diffusion coefficients have been obtained. These diffusion coefficients were different from the expected values measured using the dynamic light scattering method (DLS). The FCS was initially developed for solutions containing small fluorescent molecules therefore the observed inconsistency may result from the nature of vesicle suspension itself. The duration of the fluorescence signal may depend on the following factors: the exposure time of the labeled object to the excitation beam, the photo-physical properties (e.g., stability) of a fluorophore, the theoretical model used for the calculations of the diffusion coefficient and optical properties of the vesicle suspension. The diffusion coefficients determined for differently labeled liposomes show that its dependence on vesicle size and quantity of fluorescent probed used for labeling was significant demonstrating that the fluorescence properties of the fluorophore itself (bleaching and/or blinking) were critical factors for a correct outcome of FCS experiment. The new, based on combined FCS and DLS measurements, method for the determination of the focal volume prove itself to be useful for the evaluation of a fluorescence dye with respect to its applicability for FCS experiment.

Time-Gated Raman Spectroscopy for Quantitative Determination of Solid-State Forms of Fluorescent Pharmaceuticals.

Science.gov (United States)

Lipiäinen, Tiina; Pessi, Jenni; Movahedi, Parisa; Koivistoinen, Juha; Kurki, Lauri; Tenhunen, Mari; Yliruusi, Jouko; Juppo, Anne M; Heikkonen, Jukka; Pahikkala, Tapio; Strachan, Clare J

2018-04-03

Raman spectroscopy is widely used for quantitative pharmaceutical analysis, but a common obstacle to its use is sample fluorescence masking the Raman signal. Time-gating provides an instrument-based method for rejecting fluorescence through temporal resolution of the spectral signal and allows Raman spectra of fluorescent materials to be obtained. An additional practical advantage is that analysis is possible in ambient lighting. This study assesses the efficacy of time-gated Raman spectroscopy for the quantitative measurement of fluorescent pharmaceuticals. Time-gated Raman spectroscopy with a 128 × (2) × 4 CMOS SPAD detector was applied for quantitative analysis of ternary mixtures of solid-state forms of the model drug, piroxicam (PRX). Partial least-squares (PLS) regression allowed quantification, with Raman-active time domain selection (based on visual inspection) improving performance. Model performance was further improved by using kernel-based regularized least-squares (RLS) regression with greedy feature selection in which the data use in both the Raman shift and time dimensions was statistically optimized. Overall, time-gated Raman spectroscopy, especially with optimized data analysis in both the spectral and time dimensions, shows potential for sensitive and relatively routine quantitative analysis of photoluminescent pharmaceuticals during drug development and manufacturing.

Multicolor fluorescent in situ hybridization to define abutting and overlapping gene expression in the embryonic zebrafish brain

Directory of Open Access Journals (Sweden)

Hauptmann Giselbert

2011-04-01

Full Text Available Abstract Background In recent years, mapping of overlapping and abutting regulatory gene expression domains by chromogenic two-color in situ hybridization has helped define molecular subdivisions of the developing vertebrate brain and shed light on its basic organization. Despite the benefits of this technique, visualization of overlapping transcript distributions by differently colored precipitates remains difficult because of masking of lighter signals by darker color precipitates and lack of three-dimensional visualization properties. Fluorescent detection of transcript distributions may be able to solve these issues. However, despite the use of signal amplification systems for increasing sensitivity, fluorescent detection in whole-mounts suffers from rapid quenching of peroxidase (POD activity compared to alkaline phosphatase chromogenic reactions. Thus, less strongly expressed genes cannot be efficiently detected. Results We developed an optimized procedure for fluorescent detection of transcript distribution in whole-mount zebrafish embryos using tyramide signal amplification (TSA. Conditions for hybridization and POD-TSA reaction were optimized by the application of the viscosity-increasing polymer dextran sulfate and the use of the substituted phenol compounds 4-iodophenol and vanillin as enhancers of POD activity. In combination with highly effective bench-made tyramide substrates, these improvements resulted in dramatically increased signal-to-noise ratios. The strongly enhanced signal intensities permitted fluorescent visualization of less abundant transcripts of tissue-specific regulatory genes. When performing multicolor fluorescent in situ hybridization (FISH experiments, the highly sensitive POD reaction conditions required effective POD inactivation after each detection cycle by glycine-hydrochloric acid treatment. This optimized FISH procedure permitted the simultaneous fluorescent visualization of up to three unique transcripts

Chromophore photophysics and dynamics in fluorescent proteins of the GFP family

International Nuclear Information System (INIS)

Nienhaus, Karin; Nienhaus, G Ulrich

2016-01-01

Proteins of the green fluorescent protein (GFP) family are indispensable for fluorescence imaging experiments in the life sciences, particularly of living specimens. Their essential role as genetically encoded fluorescence markers has motivated many researchers over the last 20 years to further advance and optimize these proteins by using protein engineering. Amino acids can be exchanged by site-specific mutagenesis, starting with naturally occurring proteins as templates. Optical properties of the fluorescent chromophore are strongly tuned by the surrounding protein environment, and a targeted modification of chromophore-protein interactions requires a profound knowledge of the underlying photophysics and photochemistry, which has by now been well established from a large number of structural and spectroscopic experiments and molecular-mechanical and quantum-mechanical computations on many variants of fluorescent proteins. Nevertheless, such rational engineering often does not meet with success and thus is complemented by random mutagenesis and selection based on the optical properties. In this topical review, we present an overview of the key structural and spectroscopic properties of fluorescent proteins. We address protein-chromophore interactions that govern ground state optical properties as well as processes occurring in the electronically excited state. Special emphasis is placed on photoactivation of fluorescent proteins. These light-induced reactions result in large structural changes that drastically alter the fluorescence properties of the protein, which enables some of the most exciting applications, including single particle tracking, pulse chase imaging and super-resolution imaging. We also present a few examples of fluorescent protein application in live-cell imaging experiments. (topical review)

Chromophore photophysics and dynamics in fluorescent proteins of the GFP family

Science.gov (United States)

Nienhaus, Karin; Nienhaus, G. Ulrich

2016-11-01

Proteins of the green fluorescent protein (GFP) family are indispensable for fluorescence imaging experiments in the life sciences, particularly of living specimens. Their essential role as genetically encoded fluorescence markers has motivated many researchers over the last 20 years to further advance and optimize these proteins by using protein engineering. Amino acids can be exchanged by site-specific mutagenesis, starting with naturally occurring proteins as templates. Optical properties of the fluorescent chromophore are strongly tuned by the surrounding protein environment, and a targeted modification of chromophore-protein interactions requires a profound knowledge of the underlying photophysics and photochemistry, which has by now been well established from a large number of structural and spectroscopic experiments and molecular-mechanical and quantum-mechanical computations on many variants of fluorescent proteins. Nevertheless, such rational engineering often does not meet with success and thus is complemented by random mutagenesis and selection based on the optical properties. In this topical review, we present an overview of the key structural and spectroscopic properties of fluorescent proteins. We address protein-chromophore interactions that govern ground state optical properties as well as processes occurring in the electronically excited state. Special emphasis is placed on photoactivation of fluorescent proteins. These light-induced reactions result in large structural changes that drastically alter the fluorescence properties of the protein, which enables some of the most exciting applications, including single particle tracking, pulse chase imaging and super-resolution imaging. We also present a few examples of fluorescent protein application in live-cell imaging experiments.

Using fluorescence measurement of zinc ions liberated from ZnS nanoparticle labels in bioassay for Escherichia coli O157:H7

International Nuclear Information System (INIS)

Cowles, Chad L.; Zhu Xiaoshan; Pai, Chi-Yun

2011-01-01

In this study, an alternative approach using ZnS nanoparticle biolabels as fluorescence signal transducers is reported for the immunoassay of E. coli O157:H7 in tap water samples. Instead of measuring the fluorescence of ZnS nanoparticles in the assay, the fluorescence signal is generated through the binding of zinc ions released from nanoparticle labels with zinc-ion sensitive fluorescence indicator Fluozin-3. In the assay, ZnS nanoparticles around 50 nm in diameter were synthesized, bioconjugated, and applied for the detection of E. coli O157:H7. The assay shows a detection range over two orders of magnitude and a detection limit around 1000 colony-forming units (cfu) of E. coli O157:H7.

Nano-Gap Embedded Plasmonic Gratings for Surface Plasmon Enhanced Fluorescence

Science.gov (United States)

Bhatnagar, Kunal; Bok, Sangho; Korampally, Venumadhav; Gangopadhyay, Shubhra

2012-02-01

Plasmonic nanostructures have been extensively used in the past few decades for applications in sub-wavelength optics, data storage, optoelectronic circuits, microscopy and bio-photonics. The enhanced electromagnetic field produced at the metal/dielectric interface by the excitation of surface plasmons via incident radiation can be used for signal enhancement in fluorescence and surface enhanced Raman scattering studies. Novel plasmonic structures on the sub wavelength scale have been shown to provide very efficient and extreme light concentration at the nano-scale. The enhanced electric field produced within a few hundred nanometers of these structures can be used to excite fluorophores in the surrounding environment. Fluorescence based bio-detection and bio-imaging are two of the most important tools in the life sciences. Improving the qualities and capabilities of fluorescence based detectors and imaging equipment has been a big challenge to the industry manufacturers. We report the novel fabrication of nano-gap embedded periodic grating substrates on the nanoscale using micro-contact printing and polymethylsilsesquioxane (PMSSQ) polymer. Fluorescence enhancement of up to 118 times was observed with these silver nanostructures in conjugation with Rhodamine-590 fluorescent dye. These substrates are ideal candidates for low-level fluorescence detection and single molecule imaging.

In Vivo Imaging of Far-red Fluorescent Proteins after DNA Electrotransfer to Muscle Tissue

DEFF Research Database (Denmark)

Hojman, Pernille; Eriksen, Jens; Gehl, Julie

2009-01-01

DNA electrotransfer to muscle tissue yields long-term, high levels of gene expression; showing great promise for future gene therapy. We want to characterize the novel far-red fluorescent protein Katushka as a marker for gene expression using time domain fluorescence in vivo imaging. Highly...... weeks. Depth and 3D analysis proved that the expression was located in the target muscle. In vivo bio-imaging using the novel Katushka fluorescent protein enables excellent evaluation of the transfection efficacy, and spatial distribution, but lacks long-term stability....... efficient transgenic expression was observed after DNA electrotransfer with 100-fold increase in fluorescent intensity. The fluorescent signal peaked 1 week after transfection and returned to background level within 4 weeks. Katushka expression was not as stable as GFP expression, which was detectable for 8...

SignalPlant: an open signal processing software platform.

Science.gov (United States)

Plesinger, F; Jurco, J; Halamek, J; Jurak, P

2016-07-01

The growing technical standard of acquisition systems allows the acquisition of large records, often reaching gigabytes or more in size as is the case with whole-day electroencephalograph (EEG) recordings, for example. Although current 64-bit software for signal processing is able to process (e.g. filter, analyze, etc) such data, visual inspection and labeling will probably suffer from rather long latency during the rendering of large portions of recorded signals. For this reason, we have developed SignalPlant-a stand-alone application for signal inspection, labeling and processing. The main motivation was to supply investigators with a tool allowing fast and interactive work with large multichannel records produced by EEG, electrocardiograph and similar devices. The rendering latency was compared with EEGLAB and proves significantly faster when displaying an image from a large number of samples (e.g. 163-times faster for 75  ×  10(6) samples). The presented SignalPlant software is available free and does not depend on any other computation software. Furthermore, it can be extended with plugins by third parties ensuring its adaptability to future research tasks and new data formats.

Chlorophyll fluorescence, photochemical reflective index and normalized difference vegetative index during plant senescence.

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Cordon, Gabriela; Lagorio, M Gabriela; Paruelo, José M

2016-07-20

The relationship between the Photochemical Reflectance Index (PRI), Normalized Difference Vegetation Index (NDVI) and chlorophyll fluorescence along senescence was investigated in this work. Reflectance and radiance measurements were performed at canopy level in grass species presenting different photosynthetic metabolism: Avena sativa (C3) and Setaria italica (C4), at different stages of the natural senescence process. Sun induced-chlorophyll fluorescence at 760nm (SIF 760 ) and the apparent fluorescence yield (SIF 760 /a, with a=irradiance at time of measurement) were extracted from the radiance spectra of canopies using the Fraunhofer Line Discrimination-method. The photosynthetic parameters derived from Kautsky kinetics and pigment content were also calculated at leaf level. Whilst stand level NDVI patterns were related to changes in the structure of canopies and not in pigment content, stand level PRI patterns suggested changes both in terms of canopy and of pigment content in leaves. Both SIF 760 /a and Φ PSII decreased progressively along senescence in both species. A strong increment in NPQ was evident in A. sativa while in S. italica NPQ values were lower. Our most important finding was that two chlorophyll fluorescence signals, Φ PSII and SIF 760 /a, correlated with the canopy PRI values in the two grasses assessed, even when tissues at different ontogenic stages were present. Even though significant changes occurred in the Total Chlr/Car ratio along senescence in both studied species, significant correlations between PRI and chlorophyll fluorescence signals might indicate the usefulness of this reflectance index as a proxy of photosynthetic RUE, at least under the conditions of this study. The relationships between stand level PRI and the fluorescence estimators (Φ PSII and SIF 760 /a) were positive in both cases. Therefore, an increase in PRI values as in the fluorescence parameters would indicate higher RUE. Copyright © 2016 Elsevier GmbH. All

Lipidots: competitive organic alternative to quantum dots for in vivo fluorescence imaging

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Gravier, Julien; Navarro, Fabrice P.; Delmas, Thomas; Mittler, Frédérique; Couffin, Anne-Claude; Vinet, Françoise; Texier, Isabelle

2011-09-01

The use of fluorescent nanostructures can bring several benefits on the signal to background ratio for in vitro microscopy, in vivo small animal imaging, and image-guided surgery. Fluorescent quantum dots (QDs) display outstanding optical properties, with high brightness and low photobleaching rate. However, because of their toxic element core composition and their potential long term retention in reticulo-endothelial organs such as liver, their in vivo human applications seem compromised. The development of new dye-loaded (DiO, DiI, DiD, DiR, and Indocyanine Green (ICG)) lipid nanoparticles for fluorescence imaging (lipidots) is described here. Lipidot optical properties quantitatively compete with those of commercial QDs (QTracker®705). Multichannel in vivo imaging of lymph nodes in mice is demonstrated for doses as low as 2 pmols of particles. Along with their optical properties, fluorescent lipidots display very low cytotoxicity (IC50 > 75 nM), which make them suitable tools for in vitro, and especially in vivo, fluorescence imaging applications.

Fiber optic-based fluorescence detection system for in vivo studies of exogenous chromophore pharmacokinetics

Science.gov (United States)

Doiron, Daniel R.; Dunn, J. B.; Mitchell, W. L.; Dalton, Brian K.; Garbo, Greta M.; Warner, Jon A.

1995-05-01

The detection and quantification of the concentration of exogenous chromophores in-vivo by their fluorescence is complicated by many physical and geometrical parameters. Measurement of such signals is advantageous in determining the pharmacokinetics of photosensitizers such as those used in photodynamic therapy (PDT) or to assist in the diagnosis of tissue histological state. To overcome these difficulties a ratio based fiber optic contact fluorometer has been developed. This fluorescence detection system (FDS) uses the ratio of the fluorescence emission peak of the exogenous chromophore to that of endogenous chromophores, i.e. autofluorescence, to correct for a variety of parameters affecting the magnitude of the measured signals. By doing so it also minimizes the range of baseline measurements prior to exogenous drug injection, for various tissue types. Design of the FDS and results of its testing in animals and patients using the second generation photosensitizer Tin ethyletiopurpurin (SnET2) are presented. These results support the feasibility and usefulness of the Ratio FDS system.

A new s-adenosylhomocysteine hydrolase-linked method for adenosine detection based on DNA-templated fluorescent Cu/Ag nanoclusters.

Science.gov (United States)

Ahn, Jun Ki; Kim, Hyo Yong; Baek, Songyi; Park, Hyun Gyu

2017-07-15

We herein describe a novel fluorescent method for the rapid and selective detection of adenosine by utilizing DNA-templated Cu/Ag nanoclusters (NCs) and employing s-adenosylhomocysteine hydrolase (SAHH). SAHH is allowed to promote hydrolysis reaction of s-adenosylhomocysteine (SAH) and consequently produces homocysteine, which would quench the fluorescence signal from DNA-templated Cu/Ag nanoclusters employed as a signaling probe in this study. On the other hand, adenosine significantly inhibits the hydrolysis reaction and prevent the formation of homocysteine. Consequently, highly enhanced fluorescence signal from DNA-Cu/Ag NCs is retained, which could be used to identify the presence of adenosine. By employing this design principle, adenosine was sensitively detected down to 19nM with high specificity over other adenosine analogs such as AMP, ADP, ATP, cAMP, guanosine, cytidine, and urine. Finally, the diagnostic capability of this method was successfully verified by reliably detecting adenosine present in a real human serum sample. Copyright © 2016 Elsevier B.V. All rights reserved.

Sensitive detection of mercury and copper ions by fluorescent DNA/Ag nanoclusters in guanine-rich DNA hybridization.

Science.gov (United States)

Peng, Jun; Ling, Jian; Zhang, Xiu-Qing; Bai, Hui-Ping; Zheng, Liyan; Cao, Qiu-E; Ding, Zhong-Tao

2015-02-25

In this work, we designed a new fluorescent oligonucleotides-stabilized silver nanoclusters (DNA/AgNCs) probe for sensitive detection of mercury and copper ions. This probe contains two tailored DNA sequence. One is a signal probe contains a cytosine-rich sequence template for AgNCs synthesis and link sequence at both ends. The other is a guanine-rich sequence for signal enhancement and link sequence complementary to the link sequence of the signal probe. After hybridization, the fluorescence of hybridized double-strand DNA/AgNCs is 200-fold enhanced based on the fluorescence enhancement effect of DNA/AgNCs in proximity of guanine-rich DNA sequence. The double-strand DNA/AgNCs probe is brighter and stable than that of single-strand DNA/AgNCs, and more importantly, can be used as novel fluorescent probes for detecting mercury and copper ions. Mercury and copper ions in the range of 6.0-160.0 and 6-240 nM, can be linearly detected with the detection limits of 2.1 and 3.4 nM, respectively. Our results indicated that the analytical parameters of the method for mercury and copper ions detection are much better than which using a single-strand DNA/AgNCs. Copyright © 2014 Elsevier B.V. All rights reserved.

Predicting fluorescence quantum yield for anisole at elevated temperatures and pressures

Science.gov (United States)

Wang, Q.; Tran, K. H.; Morin, C.; Bonnety, J.; Legros, G.; Guibert, P.

2017-07-01

Aromatic molecules are promising candidates for using as a fluorescent tracer for gas-phase scalar parameter diagnostics in a drastic environment like engines. Along with anisole turning out an excellent temperature tracer by Planar Laser-Induced Fluorescence (PLIF) diagnostics in Rapid Compression Machine (RCM), its fluorescence signal evolution versus pressure and temperature variation in a high-pressure and high-temperature cell have been reported in our recent paper on Applied Phys. B by Tran et al. Parallel to this experimental study, a photophysical model to determine anisole Fluorescence Quantum Yield (FQY) is delivered in this paper. The key to development of the model is the identification of pressure, temperature, and ambient gases, where the FQY is dominated by certain processes of the model (quenching effect, vibrational relaxation, etc.). In addition to optimization of the vibrational relaxation energy cascade coefficient and the collision probability with oxygen, the non-radiative pathways are mainly discussed. The common non-radiative rate (intersystem crossing and internal conversion) is simulated in parametric form as a function of excess vibrational energy, derived from the data acquired at different pressures and temperatures from the literature. A new non-radiative rate, namely, the equivalent Intramolecular Vibrational Redistribution or Randomization (IVR) rate, is proposed to characterize anisole deactivated processes. The new model exhibits satisfactory results which are validated against experimental measurements of fluorescence signal induced at a wavelength of 266 nm in a cell with different bath gases (N2, CO2, Ar and O2), a pressure range from 0.2 to 4 MPa, and a temperature range from 473 to 873 K.

Large scale statistical inference of signaling pathways from RNAi and microarray data

Directory of Open Access Journals (Sweden)

Poustka Annemarie

2007-10-01

Full Text Available Abstract Background The advent of RNA interference techniques enables the selective silencing of biologically interesting genes in an efficient way. In combination with DNA microarray technology this enables researchers to gain insights into signaling pathways by observing downstream effects of individual knock-downs on gene expression. These secondary effects can be used to computationally reverse engineer features of the upstream signaling pathway. Results In this paper we address this challenging problem by extending previous work by Markowetz et al., who proposed a statistical framework to score networks hypotheses in a Bayesian manner. Our extensions go in three directions: First, we introduce a way to omit the data discretization step needed in the original framework via a calculation based on p-values instead. Second, we show how prior assumptions on the network structure can be incorporated into the scoring scheme using regularization techniques. Third and most important, we propose methods to scale up the original approach, which is limited to around 5 genes, to large scale networks. Conclusion Comparisons of these methods on artificial data are conducted. Our proposed module network is employed to infer the signaling network between 13 genes in the ER-α pathway in human MCF-7 breast cancer cells. Using a bootstrapping approach this reconstruction can be found with good statistical stability. The code for the module network inference method is available in the latest version of the R-package nem, which can be obtained from the Bioconductor homepage.

Correction of fluorescence for depth-specific optical and vascular properties using reflectance and differential path-length spectroscopy during PDT

Science.gov (United States)

van Zaane, F.; Middelburg, T. A.; de Bruijn, H. S.; van der Ploeg-van den Heuvel, A.; de Haas, E. R. M.; Sterenborg, H. J. C. M.; Neumann, H. A. M.; Robinson, D. J.

2009-06-01

Introduction: The rate of PpIX fluorescence photobleaching is routinely used as a dose metric for ALA-PDT. Diffuse reflection spectroscopy is often used to account for variations in tissue optical properties at the photosensitizer excitation and emission bands. It can be used to quantify changes in vascular parameters, such as blood volume fraction and saturation, and can aid understanding of tissue response to PDT. The volume and(/or) depth over which these signals are acquired are critical. The aim of this study is to use quantitative reflectance spectroscopy (DPS) to correct fluorescence for changes in tissue optical properties and monitor PDT. Materials & Methods: ALA was topically applied to hairless mice skin and the incubated spot was treated with PDT according to fractionated illumination schemes. DPS measurements of vascular parameters and optical properties were performed directly before and after illumination. Both the differential signal, delivery-and-collection-fiber signal and the collection fiber signal, which all probe different measurement volumes, are analyzed. Results & Conclusions: Analysis of DPS measurements shows that at the depth where most fluorescence originates, there is almost no blood present. During PDT vascular parameters at this depth stay constant. In more oxygenated layers of the tissue, the optical properties do change during PDT, suggesting that only a small part of PpIX fluorescence originates from the interesting depths where vascular response occurs. Correcting fluorescence emission spectra for optical changes at specific depths and not for the total of changes in a larger volume, as is usually done now, can improve PpIX photobleaching based treatment monitoring.

Tolerance of a knotted near infrared fluorescent protein to random circular permutation

Science.gov (United States)

Pandey, Naresh; Kuypers, Brianna E.; Nassif, Barbara; Thomas, Emily E.; Alnahhas, Razan N.; Segatori, Laura; Silberg, Jonathan J.

2016-01-01

Bacteriophytochrome photoreceptors (BphP) are knotted proteins that have been developed as near-infrared fluorescent protein (iRFP) reporters of gene expression. To explore how rearrangements in the peptides that interlace into the knot within the BphP photosensory core affect folding, we subjected iRFP to random circular permutation using an improved transposase mutagenesis strategy and screened for variants that fluoresce. We identified twenty seven circularly permuted iRFP that display biliverdin-dependent fluorescence in Escherichia coli. The variants with the brightest whole cell fluorescence initiated translation at residues near the domain linker and knot tails, although fluorescent variants were discovered that initiated translation within the PAS and GAF domains. Circularly permuted iRFP retained sufficient cofactor affinity to fluoresce in tissue culture without the addition of biliverdin, and one variant displayed enhanced fluorescence when expressed in bacteria and tissue culture. This variant displayed a similar quantum yield as iRFP, but exhibited increased resistance to chemical denaturation, suggesting that the observed signal increase arose from more efficient protein maturation. These results show how the contact order of a knotted BphP can be altered without disrupting chromophore binding and fluorescence, an important step towards the creation of near-infrared biosensors with expanded chemical-sensing functions for in vivo imaging. PMID:27304983

Combined X-ray fluorescence and absorption computed tomography using a synchrotron beam

International Nuclear Information System (INIS)

Hall, C

2013-01-01

X-ray computed tomography (CT) and fluorescence X-ray computed tomography (FXCT) using synchrotron sources are both useful tools in biomedical imaging research. Synchrotron CT (SRCT) in its various forms is considered an important technique for biomedical imaging since the phase coherence of SR beams can be exploited to obtain images with high contrast resolution. Using a synchrotron as the source for FXCT ensures a fluorescence signal that is optimally detectable by exploiting the beam monochromaticity and polarisation. The ability to combine these techniques so that SRCT and FXCT images are collected simultaneously, would bring distinct benefits to certain biomedical experiments. Simultaneous image acquisition would alleviate some of the registration difficulties which comes from collecting separate data, and it would provide increased information about the sample: functional X-ray images from the FXCT, with the morphological information from the SRCT. A method is presented for generating simultaneous SRCT and FXCT images. Proof of principle modelling has been used to show that it is possible to recover a fluorescence image of a point-like source from an SRCT apparatus by suitably modulating the illuminating planar X-ray beam. The projection image can be successfully used for reconstruction by removing the static modulation from the sinogram in the normal flat and dark field processing. Detection of the modulated fluorescence signal using an energy resolving detector allows the position of a fluorescent marker to be obtained using inverse reconstruction techniques. A discussion is made of particular reconstruction methods which might be applied by utilising both the CT and FXCT data.

In-vivo cancer diagnosis of the esophagus using laser-induced fluorescence

Science.gov (United States)

Vo-Dinh, Tuan; Panjehpour, Masoud; Overholt, Bergein F.; Buckley, Paul F., II; Edwards, Donna H.

1995-04-01

Laser-induced fluorescence (LIF) was used for direct in-vivo cancer diagnosis of the esophagus without requiring biopsy. The methodology was applied to differentiate normal and malignant tumors of the esophagus. Endogenous fluorescence of normal and malignant tissues were measured directly using a fiberoptic probe inserted through an endoscope. The measurements were performed in vivo during routine endoscopy. Detection of the fluorescence signal from the tissue was performed using laser excitation. The results of this LIF approach were compared with histopathology results of the biopsy samples and indicated excellent agreement in the classification of normal and malignant tumors for the samples investigated. The LIF procedure could lead to the development of a rapid and cost-effective technique for cancer diagnosis.

APD detectors for biological fluorescence spectroscopy

International Nuclear Information System (INIS)

Mazeres, S.; Borrel, V.; Magenc, C.; Courrech, J.L.; Bazer-Bachi, R.

2006-01-01

Fluorescence spectroscopy is a very convenient and widely used method for studying the molecular background of biological processes [L. Salome, J.L. Cazeil, A. Lopez, J.F. Tocanne, Eur. Biophys. J. 27 (1998) 391-402]. Chromophores are included in the structure under study and a flash of laser light induces fluorescence (Fluorescence Recovery After Photo-bleaching), the decay of which yields information on the polarity, the speed of rotation, and the speed of diffusion as well as on the temporal and spatial evolution of interactions between molecular species. The method can even be used to study living cells [J.F. Tocanne, L. Cezanne, A. Lopez, Prog. Lipid Res. 33 (1994) 203-237, L. Cezanne, A. Lopez, F. Loste, G. Parnaud, O. Saurel, P. Demange, J.F. Tocanne, Biochemistry 38 (1999) 2779-2786]. This is classically performed with a PM-based system. For biological reasons a decrease of the excitation of the cells is highly desirable. Because the fluorescence response then becomes fainter a significant improvement in detector capability would be welcome. We present here results obtained with an Avalanche Photo Diode (APD)-based system. The small sensitive area of detection allows a very significant improvement in signal/noise ratio, improvement in gain, and the opening-up of a new parameter space. With these new detectors we can begin the study of information transmission between cells through morphine receptors. This work involves both electronics engineers and biophysicists, so results and techniques in both fields will be presented here

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.

Protein A Detection Based on Quantum Dots-Antibody Bioprobe Using Fluorescence Coupled Capillary Electrophoresis

Directory of Open Access Journals (Sweden)

Lin Qiu

2014-01-01

Full Text Available In this report, fluorescence detection coupled capillary electrophoresis (CE-FL was used to detect Protein A. Antibody was first labeled with Cy5 and then mixed with quantum dots (QDs to form QDs-antibody bioprobe. Further, we observed fluorescence resonance energy transfer (FRET from QDs donor to Cy5 acceptor. The bioprobe was formed and brought QDs and Cy5 close enough to allow FRET to occur. After adding protein A, the FRET system was broken and caused the FRET signal to decrease. Thus, a new method for the determination of protein A was proposed based on the FRET signal changes. This study provides a new trail of thought for the detection of protein.

A novel portable fluorescence detection system for microfluidic card

International Nuclear Information System (INIS)

Shen, B; Xie, Y; Irawan, R

2008-01-01

Fluorescence based sensors are widely used in the field of biochemistry and medicine due to their high sensitivity and accuracy. But the cost and time required for each sample to be tested is high. If the diagnostic tools could be miniaturized, made simple to use and much less expensive, and readily available at the point of need such as emergency diagnosis, millions of people would be benefited from it. In this paper, we design a prototype of portable fluorescence detection system based on Fluorescence Filter Block and DAQ card which can emulate signal collection and processing functionalities. After the introduction of system structure and functional modules, we use a resolution approximation method to investigate the system performance. The evaluation shows that our prototype system has the sensitivity of 0.01 mMol/L (333.306 μg/mL) which meets most of the medical requirements.

Image navigation as a means to expand the boundaries of fluorescence-guided surgery.

Science.gov (United States)

Brouwer, Oscar R; Buckle, Tessa; Bunschoten, Anton; Kuil, Joeri; Vahrmeijer, Alexander L; Wendler, Thomas; Valdés-Olmos, Renato A; van der Poel, Henk G; van Leeuwen, Fijs W B

2012-05-21

Hybrid tracers that are both radioactive and fluorescent help extend the use of fluorescence-guided surgery to deeper structures. Such hybrid tracers facilitate preoperative surgical planning using (3D) scintigraphic images and enable synchronous intraoperative radio- and fluorescence guidance. Nevertheless, we previously found that improved orientation during laparoscopic surgery remains desirable. Here we illustrate how intraoperative navigation based on optical tracking of a fluorescence endoscope may help further improve the accuracy of hybrid surgical guidance. After feeding SPECT/CT images with an optical fiducial as a reference target to the navigation system, optical tracking could be used to position the tip of the fluorescence endoscope relative to the preoperative 3D imaging data. This hybrid navigation approach allowed us to accurately identify marker seeds in a phantom setup. The multispectral nature of the fluorescence endoscope enabled stepwise visualization of the two clinically approved fluorescent dyes, fluorescein and indocyanine green. In addition, the approach was used to navigate toward the prostate in a patient undergoing robot-assisted prostatectomy. Navigation of the tracked fluorescence endoscope toward the target identified on SPECT/CT resulted in real-time gradual visualization of the fluorescent signal in the prostate, thus providing an intraoperative confirmation of the navigation accuracy.

X-ray fluorescence holography: A different approach to data collection

International Nuclear Information System (INIS)

Busetto, E.; Kopecky, M.; Lausi, A.; Menk, R.H.; Miculin, M.; Savoia, A.

2000-01-01

The images of nearest neighbors of gallium atoms in a GaAs crystal were obtained by the x-ray fluorescence holography technique. The fluorescence from gallium atoms was selected by means of a thin zinc foil filter that made possible the use of an x-ray silicon photodiode detector without energy resolution. This method makes possible the detection of a much higher signal with respect to all previous experiments, thus reducing drastically measuring times, that is a basic and essential step from contemporary demonstration experiments to possible practical applications of x-ray holography in structure analysis

Quantitative fluorescence microscopy and image deconvolution.

Science.gov (United States)

Swedlow, Jason R

2013-01-01

Quantitative imaging and image deconvolution have become standard techniques for the modern cell biologist because they can form the basis of an increasing number of assays for molecular function in a cellular context. There are two major types of deconvolution approaches--deblurring and restoration algorithms. Deblurring algorithms remove blur but treat a series of optical sections as individual two-dimensional entities and therefore sometimes mishandle blurred light. Restoration algorithms determine an object that, when convolved with the point-spread function of the microscope, could produce the image data. The advantages and disadvantages of these methods are discussed in this chapter. Image deconvolution in fluorescence microscopy has usually been applied to high-resolution imaging to improve contrast and thus detect small, dim objects that might otherwise be obscured. Their proper use demands some consideration of the imaging hardware, the acquisition process, fundamental aspects of photon detection, and image processing. This can prove daunting for some cell biologists, but the power of these techniques has been proven many times in the works cited in the chapter and elsewhere. Their usage is now well defined, so they can be incorporated into the capabilities of most laboratories. A major application of fluorescence microscopy is the quantitative measurement of the localization, dynamics, and interactions of cellular factors. The introduction of green fluorescent protein and its spectral variants has led to a significant increase in the use of fluorescence microscopy as a quantitative assay system. For quantitative imaging assays, it is critical to consider the nature of the image-acquisition system and to validate its response to known standards. Any image-processing algorithms used before quantitative analysis should preserve the relative signal levels in different parts of the image. A very common image-processing algorithm, image deconvolution, is used

Compression of Born ratio for fluorescence molecular tomography/x-ray computed tomography hybrid imaging: methodology and in vivo validation.

Science.gov (United States)

Mohajerani, Pouyan; Ntziachristos, Vasilis

2013-07-01

The 360° rotation geometry of the hybrid fluorescence molecular tomography/x-ray computed tomography modality allows for acquisition of very large datasets, which pose numerical limitations on the reconstruction. We propose a compression method that takes advantage of the correlation of the Born-normalized signal among sources in spatially formed clusters to reduce the size of system model. The proposed method has been validated using an ex vivo study and an in vivo study of a nude mouse with a subcutaneous 4T1 tumor, with and without inclusion of a priori anatomical information. Compression rates of up to two orders of magnitude with minimum distortion of reconstruction have been demonstrated, resulting in large reduction in weight matrix size and reconstruction time.

Absorption and fluorescence spectroscopic characterization of BLUF domain of AppA from Rhodobacter sphaeroides

Energy Technology Data Exchange (ETDEWEB)

Zirak, P. [Institut II - Experimentelle und Angewandte Physik, Universitaet Regensburg, Universitaetstrasse 31, D-93053 Regensburg (Germany); Penzkofer, A. [Institut II - Experimentelle und Angewandte Physik, Universitaet Regensburg, Universitaetstrasse 31, D-93053 Regensburg (Germany)], E-mail: alfons.penzkofer@physik.uni-regensburg.de; Schiereis, T. [Institut fuer Biologie, Experimentelle Biophysik, Humboldt-Universitaet zu Berlin, Invalidenstrasse 42, D-10115 Berlin (Germany); Hegemann, P. [Institut fuer Biologie, Experimentelle Biophysik, Humboldt-Universitaet zu Berlin, Invalidenstrasse 42, D-10115 Berlin (Germany); Jung, A. [Max-Planck-Institut fuer medizinische Forschung, Abteilung Biomolekulare Mechanismen, Jahnstrasse 29, D-69120 Heidelberg (Germany); Schlichting, I. [Max-Planck-Institut fuer medizinische Forschung, Abteilung Biomolekulare Mechanismen, Jahnstrasse 29, D-69120 Heidelberg (Germany)

2005-08-08

The BLUF domain of the transcriptional anti-repressor protein AppA from the non-sulfur anoxyphototrophic purple bacterium Rhodobacter sphaeroides was characterized by absorption and emission spectroscopy. The BLUF domain constructs AppA{sub 148} (consisting of amino-acid residues 1-148) and AppA{sub 126} (amino-acid residues 1-126) are investigated. The cofactor of the investigated domains is found to consist of a mixture of the flavins riboflavin, FMN, and FAD. The dark-adapted domains exist in two different active receptor conformations (receptor states) with different sub-nanosecond fluorescence lifetimes (BLUF{sub r,f} and BLUF{sub r,sl}) and a small non-interacting conformation (BLUF{sub nc}). The active receptor conformations are transformed to putative signalling states (BLUF{sub s,f} and BLUF{sub s,sl}) of low fluorescence efficiency and picosecond fluorescence lifetime by blue-light excitation (light-adapted domains). In the dark at room temperature both signalling states recover back to the initial receptor states with a time constant of about 17 min. A quantum yield of signalling state formation of about 25% was determined by intensity dependent transmission measurements. A photo-cycle scheme is presented including photo-induced charge transfer complex formation, charge recombination, and protein binding pocket reorganisation.

Vertical profiles of atmospheric fluorescent aerosols observed by a mutil-channel lidar spectrometer system

Science.gov (United States)

Huang, Z.; Huang, J.; Zhou, T.; Sugimoto, N.; Bi, J.

2015-12-01

Zhongwei Huang1*, Jianping Huang1, Tian Zhou1, Nobuo Sugimoto2, Jianrong Bi1 and Jinsen Shi11Key Laboratory for Semi-Arid Climate Change of the Ministry of Education, College of Atmospheric Sciences, Lanzhou University, Lanzhou, China. 2Atmospheric Environment Division, National Institutes for Environmental Studies, Tsukuba, Japan Email: huangzhongwei@lzu.edu.cn Abstract Atmospheric aerosols have a significant impact on regional and globe climate. The challenge in quantifying aerosol direct radiative forcing and aerosol-cloud interactions arises from large spatial and temporal heterogeneity of aerosol concentrations, compositions, sizes, shape and optical properties (IPCC, 2007). Lidar offers some remarkable advantages for determining the vertical structure of atmospheric aerosols and their related optical properties. To investigate the characterization of atmospheric aerosols (especially bioaerosols) with high spatial and temporal resolution, we developed a Raman/fluorescence/polarization lidar system employed a multi-channel spectrometer, with capabilities of providing measurements of Raman scattering and laser-induced fluorescence excitation at 355 nm from atmospheric aerosols. Meanwhile, the lidar system operated polarization measurements both at 355nm and 532nm wavelengths, aiming to obtain more information of aerosols. It employs a high power pulsed laser and a received telescope with 350mm diameter. The receiver could simultaneously detect a wide fluorescent spectrum about 178 nm with spectral resolution 5.7 nm, mainly including an F/3.7 Crossed Czerny-Turner spectrograph, a grating (1200 gr/mm) and a PMT array with 32 photocathode elements. Vertical structure of fluorescent aerosols in the atmosphere was observed by the developed lidar system at four sites across northwest China, during 2014 spring field observation that conducted by Lanzhou University. It has been proved that the developed lidar could detect the fluorescent aerosols with high temporal and