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A Simple Fluorescent Biosensor for Theophylline Based on its RNA Aptamer  

Microsoft Academic Search

Theophylline is a potent bronchodilator with a narrow therapeutic index. A simple fluorescent biosensor that detects clinically relevant theophylline concentrations has been developed using the well-characterized theophylline binding RNA aptamer. Hybridization of the RNA aptamer to a fluorescently labeled DNA strand (FL-DNA) yields a fluorescent RNA:DNA hybrid that is sensitive to theophylline. The biosensor retains the remarkable selectivity of the

C. J. Rankin; E. N. Fuller; K. H. Hamor; S. A. Gabarra; T. P. Shields



Simple Adhesive-Tape-Based Sampling of Tomato Surfaces Combined with Rapid Fluorescence In Situ Hybridization for Salmonella Detection?  

PubMed Central

A simple adhesive-tape-based method for sampling of tomato surfaces was combined with fluorescence in situ hybridization for rapid culture-independent detection of Salmonella strains. Tapes could also be placed face-down on selective agar for on-tape enrichment of captured Salmonella cells. Overlay of cell-charged tapes with small volumes of liquid enrichment media enabled subsequent detection of tape-captured Salmonella via flow cytometry.

Bisha, Bledar; Brehm-Stecher, Byron F.



Simple and convenient G-quadruplex-based turn-on fluorescence assay for 3' ? 5' exonuclease activity.  


A selective, oligonucleotide-based, label-free, turn-on fluorescence detection method for 3' ? 5' exonuclease activity has been developed using crystal violet as a G-quadruplex-binding probe. The assay is highly simple and rapid, does not require the use of gel-based equipment or radioisotopic labeling, and is amenable to high-throughput and real-time detection. A proof-of-concept of this assay has been demonstrated for prokaryotic Exonuclease III (ExoIII) and human TREX1. PMID:21114271

Leung, Chung-Hang; Chan, Daniel Shiu-Hin; Man, Bradley Yat-Wah; Wang, Chuan-Jen; Lam, Wing; Cheng, Yung-Chi; Fong, Wang-Fun; Hsiao, Wen-Luan Wendy; Ma, Dik-Lung



A pyrene-based simple but highly selective fluorescence sensor for Cu(2+) ions via a static excimer mechanism.  


A pyrene-based simple fluorosensor has been synthesized by a one step process. It exhibited high selectivity towards Cu(2+) ions via fluorescence enhancement of monomer and excimer emission. The origin of excimer formation was examined and established to be of static in nature from the study of absorption and excitation spectra. The observed monomer and excimer emission in the presence and absence of Cu(2+) ion with varying pH was studied and provided probable justification. The effect of varying portions of water content in solvent on the sensor molecule was also examined. The sensor found its proper application in finding accurate and trace amount of Cu(2+) ions present in drinking water samples from various sources. The detection limit of the current sensor was found to be 4 × 10(-8) M. PMID:24133674

Sarkar, Soma; Roy, Swapnadip; Sikdar, Anindita; Saha, R N; Panja, Sujit S



Simple method based on fluorescent detection for the determination of 4-hydroxycyclophosphamide in plasma.  


Cyclophosphamide is a prodrug used both as a single drug and in combination chemotherapy. Cyclophosphamide is converted to its active metabolite (4-hydroxycyclophosphamide) by the cytochrome P450 enzymes. A liquid chromatography method including liquid-liquid extraction and protein precipitation in one step was developed to measure 4-hydroxycyclophosphamide in plasma. The 4-hydroxycyclophosphamide was stabilized and converted to a fluorescent dansylhydrazone derivative, which was chromatographed on a reverse-phase column and detected using a spectrofluorometric detector at excitation of 350 nm and emission of 550 nm. The limit of quantitation was 60 ng/mL and the between-day accuracy and precision were less than 9%. The method was applied to the analysis of plasma from patients who had received an intravenous infusion of 1 g/m(2) cyclophosphamide. PMID:12021633

Griskevicius, Laimonas; Meurling, Lennart; Hassan, Moustapha



Detection of infective poliovirus by a simple, rapid, and sensitive flow cytometry method based on fluorescence resonance energy transfer technology.  


The rapid and effective detection of virus infection is critical for clinical management and prevention of disease spread during an outbreak. Several methods have been developed for this purpose, of which classical serological and viral nucleic acid detection are the most common. We describe an alternative approach that utilizes engineered cells expressing fluorescent proteins undergoing fluorescence resonance energy transfer (FRET) upon cleavage by the viral 2A protease (2A(pro)) as an indication of infection. Quantification of the infectious-virus titers was resolved by using flow cytometry, and utility was demonstrated for the detection of poliovirus 1 (PV1) infection. Engineered buffalo green monkey kidney (BGMK) cells expressing the cyan fluorescent protein (CFP)-yellow fluorescent protein (YFP) substrate linked by a cleavage recognition site for PV1 2A(pro) were infected with different titers of PV1. After incubation at various time points, cells were harvested, washed, and subjected to flow cytometry analysis. The number of infected cells was determined by counting the number of cells with an increased CFP-to-YFP ratio. As early as 5 h postinfection, a significant number of infected cells (3%) was detected by flow cytometry, and cells infected with only 1 PFU were detected after 12 h postinfection. When applied to an environmental water sample spiked with PV1, the flow cytometry-based assay provided a level of sensitivity similar to that of the plaque assay for detecting and quantifying infectious virus particles. This approach, therefore, is more rapid than plaque assays and can be used to detect other viruses that frequently do not form clear plaques on cell cultures. PMID:19933336

Cantera, Jason L; Chen, Wilfred; Yates, Marylynn V



A simple fluorescent probe for the determination of dissolved oxygen based on the catalytic activation of oxygen by iron(II) chelates  

Microsoft Academic Search

This work aims at establishing a simple fluorescent probe for the determination of dissolved oxygen. It is found that iron(II) ions activate oxygen to produce reactive species being capable of oxidizing non-fluorescent coumarin to fluorescent 7-hydroxycoumarin. However, this process is not effective because the yield of the reactive species is very low in the presence of simple iron(II) salts alone.

Wei Luo; M. E. Abbas; Lihua Zhu; Wenyi Zhou; Kejing Li; Heqing Tang; Shushen Liu; Weiying Li



Simple fluorescent sensors engineered with catalytic DNA 'MgZ' based on a non-classic allosteric design.  


Most NAE (nucleic acid enzyme) sensors are composed of an RNA-cleaving catalytic motif and an aptameric receptor. They operate by activating or repressing the catalytic activity of a relevant NAE through the conformational change in the aptamer upon target binding. To transduce a molecular recognition event to a fluorescence signal, a fluorophore-quencher pair is attached to opposite ends of the RNA substrate such that when the NAE cleaves the substrate, an increased level of fluorescence can be generated. However, almost all NAE sensors to date harbor either NAEs that cannot accommodate a fluorophore-quencher pair near the cleavage site or those that can accept such a modification but require divalent transition metal ions for catalysis. Therefore, the signaling magnitude and the versatility of current NAE sensors might not suffice for analytical and biological applications. Here we report an RNA-cleaving DNA enzyme, termed 'MgZ', which depends on Mg(2+) for its activity and can accommodate bulky dye moieties next to the cleavage site. MgZ was created by in vitro selection. The selection scheme entailed acidic buffering and ethanol-based reaction stoppage to remove selfish DNAs. Characterization of MgZ revealed a three-way junction structure, a cleavage rate of 1 min(-1), and 26-fold fluorescence enhancement. Two ligand-responsive NAE sensors were rationally designed by linking an aptamer sequence to the substrate of MgZ. In the absence of the target, the aptamer-linked substrate is locked into a conformation that prohibits MgZ from accessing the substrate. In the presence of the target, the aptamer releases the substrate, which induces MgZ-mediated RNA cleavage. The discovery of MgZ and the introduction of the above NAE sensor design strategy should facilitate future efforts in sensor engineering. PMID:18030352

Chiuman, William; Li, Yingfu



Simple Fluorescent Sensors Engineered with Catalytic DNA 'MgZ' Based on a Non-Classic Allosteric Design  

PubMed Central

Most NAE (nucleic acid enzyme) sensors are composed of an RNA-cleaving catalytic motif and an aptameric receptor. They operate by activating or repressing the catalytic activity of a relevant NAE through the conformational change in the aptamer upon target binding. To transduce a molecular recognition event to a fluorescence signal, a fluorophore-quencher pair is attached to opposite ends of the RNA substrate such that when the NAE cleaves the substrate, an increased level of fluorescence can be generated. However, almost all NAE sensors to date harbor either NAEs that cannot accommodate a fluorophore-quencher pair near the cleavage site or those that can accept such a modification but require divalent transition metal ions for catalysis. Therefore, the signaling magnitude and the versatility of current NAE sensors might not suffice for analytical and biological applications. Here we report an RNA-cleaving DNA enzyme, termed ‘MgZ’, which depends on Mg2+ for its activity and can accommodate bulky dye moieties next to the cleavage site. MgZ was created by in vitro selection. The selection scheme entailed acidic buffering and ethanol-based reaction stoppage to remove selfish DNAs. Characterization of MgZ revealed a three-way junction structure, a cleavage rate of 1 min?1, and 26-fold fluorescence enhancement. Two ligand-responsive NAE sensors were rationally designed by linking an aptamer sequence to the substrate of MgZ. In the absence of the target, the aptamer-linked substrate is locked into a conformation that prohibits MgZ from accessing the substrate. In the presence of the target, the aptamer releases the substrate, which induces MgZ-mediated RNA cleavage. The discovery of MgZ and the introduction of the above NAE sensor design strategy should facilitate future efforts in sensor engineering.

Chiuman, William; Li, Yingfu



Fluorescence-based biosensors.  


The field of optical sensors has been a growing research area over the last three decades. A wide range of books and review articles has been published by experts in the field who have highlighted the advantages of optical sensing over other transduction methods. Fluorescence is by far the method most often applied and comes in a variety of schemes. Nowadays, one of the most common approaches in the field of optical biosensors is to combine the high sensitivity of fluorescence detection in combination with the high selectivity provided by ligand-binding proteins. In this chapter we deal with reviewing our recent results on the implementation of fluorescence-based sensors for monitoring environmentally hazardous gas molecules (e.g. nitric oxide, hydrogen sulfide). Reflectivity-based sensors, fluorescence correlation spectroscopy-based (FCS) systems, and sensors relying on the enhanced fluorescence emission on silver island films (SIFs) coupled to the total internal reflection fluorescence mode (TIRF) for the detection of gliadin and other prolamines considered toxic for celiac patients are also discussed herein. PMID:22573441

Strianese, Maria; Staiano, Maria; Ruggiero, Giuseppe; Labella, Tullio; Pellecchia, Claudio; D'Auria, Sabato



Fluorescence Based Sensor Arrays  

Microsoft Academic Search

\\u000a Fluorescence-based cross reactive sensor arrays have experienced significant development in the last decade because of the\\u000a advantages that they can offer with respect to other transduction mechanisms, in terms of the usual performance parameters\\u000a such as sensitivity, selectivity and so on. From this point of view, a great impulse to this development has been due to the\\u000a realization of novel

Roberto Paolesse; Donato Monti; Francesca Dini; Corrado Di Natale


A simple and accurate method to determine nitrite and nitrate in serum based on high-performance liquid chromatography with fluorescence detection.  


A simple method for accurate determination of nitrite and nitrate in serum was proposed to avoid the variation of nitrate reduction. For nitrite determination, serum samples were directly precipitated with methanol pre-nitrate conversion, and then the supernatant reacted with 2,3-diaminonaphthalene (DAN) to form 2,3-naphthotriazole (NAT), which was quantitatively analyzed by high-performance liquid chromatography coupled with fluorescence detection (HPLC-FL). For nitrate determination, samples were firstly heated at 70°C for 10 min to inactivate endogenous reductase-inhibiting proteins, then nitrate in the samples was quantitatively reduced to nitrite by reductase added experimentally. The difference in total nitrite concentrations between pre- and post-nitrate conversion was used to calculate the amount of nitrate in the samples. In addition to good specificity, high sensitivity, satisfactory accuracy and reproducibility, our method is simple and suitable for the quantitative determination of nanomolar level of nitrite and nitrate in a large number of serum samples. Copyright © 2013 John Wiley & Sons, Ltd. PMID:23760922

Shu-Yu, Zhan; Qing, Shao; Li, Liu; Xiao-Hui, Fan



A Simple Method for Comparative Study on the Thermal Performance of Light Emitting Diodes (LED) and Fluorescent Lamps  

Microsoft Academic Search

A simple method is proposed to measure the heat dissipation of LEDs and fluorescent lamps in an open system which allows light energy to escape. Based on this method, a comparative study on the thermal and luminous performance of high-brightness LEDs and fluorescent lamps is presented. At rated power, T5 & T8 fluorescent lamps generate about 73% - 77% of

Y. X. Qin; D. Y. Lin; S. Y. R. Hui



Fluorescence-Based Sensors  

NASA Astrophysics Data System (ADS)

The natural luminescent phenomena (from the Latin words "lumen" and "essentia", i.e., "made of light") such as northern lights (aurora borealis), marine brightness, glow-worms, shining putrid fish scales, "bluish"- appearing water when contained in certain wooden cups (quinine fluorescence), some stones heated at high temperatures with reducing agents (BaS phosphorescence), or light emitted while crushing sugar (triboluminescence) already fascinated our ancestors. Nowadays we understand that ultraviolet and visible emission of light originates from a competitive deactivation pathway of the lowest electronic excited state of atoms and molecules that produces the so called luminescence (the sub-terms fluorescence and phosphorescence just designate whether the return of the excited to the ground state is an "allowed" or "forbidden" process, namely it is fast or slow, the loosely-defined border between them being a 1-?s-1 rate constant). Actually, luminescence is the only method to generate light in the known Universe regardless it is powered by the nuclear reactions in the stars, the ohmical heating in bulbs, an electric discharge, the absorption of light or a (bio)chemical reaction (chemiluminescence).

Orellana, Guillermo


Aptamer-Based Fluorescent Biosensors  

PubMed Central

Selected from random pools of DNA or RNA molecules through systematic evolution of ligands by exponential enrichment (SELEX), aptamers can bind to target molecules with high affinity and specificity, which makes them ideal recognition elements in the development of biosensors. To date, aptamer-based biosensors have used a wide variety of detection techniques, which are briefly summarized in this article. The focus of this review is on the development of aptamer-based fluorescent biosensors, with emphasis on their design as well as properties such as sensitivity and specificity. These biosensors can be broadly divided into two categories: those using fluorescently-labeled aptamers and others that employ label-free aptamers. Within each category, they can be further divided into “signal-on” and “signal-off” sensors. A number of these aptamer-based fluorescent biosensors have shown promising results in biological samples such as urine and serum, suggesting their potential applications in biomedical research and disease diagnostics.

Wang, Rongsheng E.; Zhang, Yin; Cai, Jianfeng; Cai, Weibo; Gao, Ting



Multiphoton laser-induced-fluorescence studies of simple species  

SciTech Connect

Recent studies have demonstrated multiple-photon excitation of atomic species. Bischel and coworkers have provided a detailed description of two-photon excitation fluorescence in the detection of atoms generated in a low pressure discharge and its possible application as a diagnostic tool in flame and plasmas. It is also believed that such techniques can be useful in detecting molecular transients which are difficult to detect otherwise as demonstrated in two-photon laser-induced fluorescence (LIF) detection of NO. In this paper, we discuss our recent two-photon LIF studies on I and Br atoms, which are produced via laser photolysis of molecular precursors. The two-photon LIF study of HS and DS radicals is presented as a test case for the detection of other important radical species such as C/sub 2/H and CH/sub 3/, which are currently being investigated in our laboratory. In addition, excitation of three-photon resonances of I/sub 2/, N/sub 2/, and H/sub 2/ is discussed.

Tiee, J.J.; Ferris, M.J.; Loge, G.W.; Wampler, F.B.



The toolbox of fluorescence standards: flexible calibration tools for the standardization of fluorescence-based measurements  

Microsoft Academic Search

To improve the reliability of fluorescence data in the life and material sciences and to enable accreditation of fluorescence techniques, standardization concepts are required that guarantee and improve the comparability of fluorescence measurements. At the core of such concepts are simple and evaluated fluorescence standards for the consideration of instrument-specific spectral and intensity distortions of measured signals and for instrument

Ute Resch-Genger; K. Hoffmann; C. Würth; T. Behnke; A. Hoffmann; D. Pfeifer; A. Engel



The Calibration Kit Spectral Fluorescence Standards— A Simple and Certified Tool for the Standardization of the Spectral Characteristics of Fluorescence Instruments  

Microsoft Academic Search

With the Calibration Kit Spectral Fluorescence Standards BAM-F001–BAM-F005, we developed a simple tool for the characterization of the relative spectral responsivity and the long-term stability of the emission channel of fluorescence instruments under routine measurement conditions thereby providing the basis for an improved comparability of fluorescence measurements and eventually standardization. This first set of traceable fluorescence standards, which links fluorescence

D. Pfeifer; K. Hoffmann; A. Hoffmann; C. Monte; U. Resch-Genger



Simple-english for data base communication  

Microsoft Academic Search

Three classes of the so-called natural languages for communication with data bases are defined:English-like, pseudo-English, andsimple-English. It is argued that English-like and pseudo-English languages are normally more difficult to learn and use than artificial programming languages with no overt claim to English likeness. Simple-English is presented as a family of languages in which many restrictions (which hamper learning) are removed

J. A. Moyne



Implantable fluorescence-based glucose sensor development  

NASA Astrophysics Data System (ADS)

An implantable sensor is being created that allows measurement of blood glucose through fluorescent detection of an embedded chemical assay. The sensor is based on the competitive binding reaction between the protein Concanavalin A and various saccharide molecules, specifically a glycodendrimer and glucose. Previous studies have shown the ability of an embedded chemical assay using Con A and dextran with shorter wavelength dyes to both sense changes in glucose and generate sufficient fluorescent emission to pass through the dermal tissue. However, due to the chemical constituents of the assay, multivalent binding was evident resulting in poor spectral change due to glucose within the biological range. Use of a glycodendrimer and longer wavelength dyes has improved the sensor"s spectral change due to glucose and the overall signal to noise ratio of the sensor. In this work, a description of this sensor and the results obtained from it will be presented showing a large dynamic range of fluorescence with glucose.

Ibey, Bennett L.; Yadavalli, Vamsi K.; Thomas, Hope R.; Rounds, Rebecca M.; Pishko, Michael V.; Cote, Gerard L.



Simple interface of high-performance liquid chromatography–atomic fluorescence spectrometry hyphenated system for speciation of mercury based on photo-induced chemical vapour generation with formic acid in mobile phase as reaction reagent  

Microsoft Academic Search

Photo-induced chemical vapour generation (CVG) with formic acid in mobile phase as reaction reagent was developed as interface to on-line couple HPLC with atomic fluorescence spectrometry for the separation and determination of inorganic mercury, methylmercury (MeHg), ethylmercury (EtHg) and phenylmercury (PhHg). In the developed procedure, formic acid in mobile phase was used to decompose organomercuries and reduce Hg2+ to mercury

Yongguang Yin; Jingfu Liu; Bin He; Jianbo Shi; Guibin Jiang



A simple chlorophyll fluorescence parameter that correlates with the rate coefficient of photoinactivation of Photosystem II  

Microsoft Academic Search

A method of partitioning the energy in a mixed population of active and photoinactivated Photosystem II (PS II) complexes based on chlorophyll fluorescence measurements is presented. There are four energy fluxes, each with its quantum efficiency: a flux associated with photochemical electron flow in active PS II reaction centres (JPSII), thermal dissipation in photoinactivated, non-functional PS IIs (JNF), light-regulated thermal

Luke Hendrickson; Britta Förster; Barry J. Pogson; Wah Soon Chow



Simple triac dimmable Compact Fluorescent Lamp ballast and Light Emitting Diode driver  

Microsoft Academic Search

Compact Fluorescent Lamp (CFL) and Light Emitting Diode (LED) are energy efficient light sources. However, they have a disadvantage that they cannot be directly dimmed with a triac dimmer. This article briefly explains the incompatibility issues, and suggests a CFL ballast circuit to overcome this disadvantage. The ballast is based on resonant mode topology, with additional circuitry for triac interface

Andre Tjokrorahardjo



A rapid and simple assay for human blood malignancy engraftment, homing and chemotherapy treatment using fluorescent imaging of avian embryos.  


Detection of grafted human cells in mice using fluorescence is a rapid and simple technique whose use is continually expanding. Robust engraftment of human hematological malignancy (HHM) lines and patient cells into the naturally immunodeficient turkey embryo has recently been demonstrated by polymerase chain reaction (PCR), fluorescence activated cell sorting (FACS) and histology. We demonstrate here that fluorescence imaging is a rapid and simple technique for detecting engraftment and homing of cells derived from HHM in turkey embryos. Raji lymphoma cells expressing a far-red fluorescent protein were injected intravascularly into turkey embryos and fluorescence was detected 8 days later in their limbs and skulls. Much stronger signals were obtained after removal of the bones from the limbs. Unlabeled Raji cells did not give a fluorescent signal. Treatment with doxorubicin dramatically reduced the fluorescent signal. Intravenously injected HL-60 leukemia cells labeled with infrared-fluorescing dye were detected in the bone marrow after 16 h. Homing was active, although some non-specific fluorescence was present. Use of fluorescence imaging of HHM in turkey embryos is therefore feasible and reduces the time, effort and expense for detecting engraftment. This technique has potential to become a high-throughput xenograft system for hematological chemotherapy development and testing, and for study of hematological cell homing. PMID:21895546

Grinberg, Igor; Dukhovny, Anna; Goldstein, Ronald S



Fluorescent DNA base analogs: preparation, incorporation into oligonucleotides, and time-resolved fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

The synthesis of fluorescent DNA base analogs that can replace the natural bases adenine, thymine and cytosine and their incorporation into synthetic oligodeoxynucleotides is described. The effect on the stability of such modified nucleotides like 2-aminopurine and some pteridine derivatives, is studied by thermal melting studies and comparison with the corresponding unaltered oligonucleotides. The fluorescence spectroscopic properties and several applications of these new fluorescent DNA probes are described in greater detail. Structural information on the conformation of special oligonucleotides like hairpins, junctions and bulged duplexes can be obtained from fluorescence lifetime and fluorescence depolarization data. For example the fluorescence lifetime pattern of 2-aminopurine is a sensitive indicator of DNA base pairing. As examples the structure of the oligonucleotide-linker junction in a synthetically linked oligonucleotide hairpin and the base-pairing of the first 'deoxyribozyme' are discussed. A third application uses doubly, i.e., donor and acceptor, labeled oligonucleotides to measure distances by fluorescence resonance energy transfer.

Hochstrasser, Remo A.



Fluorescent silica nanoparticle-based probe for the detection of ozone via fluorescence resonance energy transfer.  


A fluorescence resonance energy transfer (FRET) platform for the detection of ozone was developed by combining the overlap of the fluorescence spectrum of Ru(bpy)3(2+)-doped silica nanoparticles with the absorption spectrum of indigo carmine at around 600 nm. This FRET system can be used to detect ozone simply within 10 min. Simple qualitative ozone detection methods using cotton swabs or paper were also developed. PMID:24049767

Qi, Wenjing; Wu, Di; Zhao, Jianming; Liu, Zhongyuan; Xu, Min; Anjum, Saima; Xu, Guobao



Development of a Novel Efficient Fluorescence-Based Plaque Reduction Microneutralization Assay for Measles Virus Immunity  

Microsoft Academic Search

The measurement of functional measles virus-specific neutralizing antibodies is of considerable interest for vaccine-related research. In this study, we developed and standardized a simple, rapid, highly sensitive, and reproducible fluorescence-based plaque reduction microneutralization (PRMN) assay with visual and auto- mated readout, using a recombinant measles virus engineered to express enhanced green fluorescent protein. The assay is performed in micro format,

Iana H. Haralambieva; Inna G. Ovsyannikova; Robert A. Vierkant; Gregory A. Poland



Teaching about photosynthesis with simple equipment: analysis of light-induced changes in fluorescence and reflectance of plant leaves.  


Solar energy absorbed by plants results in either reflection or absorption. The latter results in photosynthesis, fluorescence, or heat. Measurements of fluorescence changes have been used for monitoring processes associated with photosynthesis. A simple method to follow changes in leaf fluorescence and leaf reflectance associated with nonphotochemical quenching and light acclimation of leaves is described. The main equipment needed consists of a green-light emitting laser pointer, a digital camera, and a personal computer equipped with the camera acquisition software and the programs ImageJ and Excel. Otherwise, only commonly available cheap materials are required. PMID:23728512

Björn, Lars Olof; Li, Shaoshan



Fluorescence-based active site probes for profiling deubiquitinating enzymes.  


Novel ubiquitin-based active site probes including a fluorescent tag have been developed and evaluated. A new, functionalizable electrophilic trap is utilized allowing for late stage diversification of the probe. Attachment of fluorescent dyes allowed direct detection of endogenous deubiquitinating enzyme (DUB) activities in cell extracts by in-gel fluorescence imaging. PMID:22453277

McGouran, Joanna F; Kramer, Holger B; Mackeen, Mukram M; di Gleria, Katalin; Altun, Mikael; Kessler, Benedikt M



A simple fluorescence labeling method for studies of protein oxidation, protein modification, and proteolysis.  


Proteins are sensitive to oxidation, and oxidized proteins are excellent substrates for degradation by proteolytic enzymes such as the proteasome and the mitochondrial Lon protease. Protein labeling is required for studies of protein turnover. Unfortunately, most labeling techniques involve (3)H or (14)C methylation, which is expensive, exposes researchers to radioactivity, generates large amounts of radioactive waste, and allows only single-point assays because samples require acid precipitation. Alternative labeling methods have largely proven unsuitable, either because the probe itself is modified by the oxidant(s) being studied or because the alternative labeling techniques are too complex or too costly for routine use. What is needed is a simple, quick, and cheap labeling technique that uses a non-radioactive marker, binds strongly to proteins, is resistant to oxidative modification, and emits a strong signal. We have devised a new reductive method for labeling free carboxyl groups of proteins with the small fluorophore 7-amino-4-methycoumarin (AMC). When bound to target proteins, AMC fluoresces very weakly but when AMC is released by proteinases, proteases, or peptidases, it fluoresces strongly. Thus, without acid precipitation, the proteolysis of any target protein can be studied continuously, in multiwell plates. In direct comparisons, (3)H-labeled proteins and AMC-labeled proteins exhibited essentially identical degradation patterns during incubation with trypsin, cell extracts, and purified proteasome. AMC-labeled proteins are well suited to studying increased proteolytic susceptibility after protein modification, because the AMC-protein bond is resistant to oxidizing agents such as hydrogen peroxide and peroxynitrite and is stable over time and to extremes of pH, temperature (even boiling), freeze-thaw, mercaptoethanol, and methanol. PMID:21988844

Pickering, Andrew M; Davies, Kelvin J A



Fluorescence-lifetime-based sensing: applications to clinical chemistry and cellular imaging  

NASA Astrophysics Data System (ADS)

Measurements of fluorescence lifetimes, rather than intensity or intensity ratios, offer many advantages in clinical chemistry and imaging. However, measurements of time-resolved fluorescence are normally associated with complex laser light sources and instrumentation. In this lecture, we show how emerging technology is enabling the design and use of simple instrumentation for time-resolved fluorescence. In particular, it is now possible to imagine lifetime-based measurements of blood gases and blood glucose, and lifetime imaging of calcium and other ions in microscopic samples.

Lakowicz, Joseph R.; Szmacinski, Henryk; Thompson, Richard B.



A label-free G-quadruplex-based switch-on fluorescence assay for the selective detection of ATP.  


A G-quadruplex-based, label-free, switch-on fluorescence detection method has been developed for the selective detection of ATP in aqueous solution using crystal violet as a G-quadruplex-selective probe. The assay is highly simple and rapid, and does not require the use of fluorescent labeling. PMID:22343772

He, Hong-Zhang; Ma, Victor Pui-Yan; Leung, Ka-Ho; Chan, Daniel Shiu-Hin; Yang, Hui; Cheng, Zhen; Leung, Chung-Hang; Ma, Dik-Lung



Simple, Rapid and Inexpensive Quantitative Fluorescent PCR Method for Detection of Microdeletion and Microduplication Syndromes  

PubMed Central

Because of economic limitations, the cost-effective diagnosis of patients affected with rare microdeletion or microduplication syndromes is a challenge in developing countries. Here we report a sensitive, rapid, and affordable detection method that we have called Microdeletion/Microduplication Quantitative Fluorescent PCR (MQF-PCR). Our procedure is based on the finding of genomic regions with high homology to segments of the critical microdeletion/microduplication region. PCR amplification of both using the same primer pair, establishes competitive kinetics and relative quantification of amplicons, as happens in microsatellite-based Quantitative Fluorescence PCR. We used patients with two common microdeletion syndromes, the Williams-Beuren syndrome (7q11.23 microdeletion) and the 22q11.2 microdeletion syndromes and discovered that MQF-PCR could detect both with 100% sensitivity and 100% specificity. Additionally, we demonstrated that the same principle could be reliably used for detection of microduplication syndromes, by using patients with the Lubs (MECP2 duplication) syndrome and the 17q11.2 microduplication involving the NF1 gene. We propose that MQF-PCR is a useful procedure for laboratory confirmation of the clinical diagnosis of microdeletion/microduplication syndromes, ideally suited for use in developing countries, but having general applicability as well.

Stofanko, Martin; Goncalves-Dornelas, Higgor; Cunha, Pricila Silva; Pena, Heloisa B.; Vianna-Morgante, Angela M.; Pena, Sergio Danilo Junho



Simple non-fluorescent polarity labeling of microtubules for molecular motor assays.  


Transport of intracellular organelles along the microtubule cytoskeleton occurs in a bidirectional manner due to opposing activity of microtubule-associated motor proteins of the kinesin and dynein families. Regulation of this opposing activity and the resultant motion is believed to generate a polarized distribution of many organelles within the cell. The bidirectional motion can be reconstituted on in vitro assembled microtubules using organelles extracted from cells. This provides an opportunity to understand the regulation of intracellular transport through quantitative analysis of the motion of organelles in a controlled environment. Such analysis requires the use of polarity-labeled microtubules to resolve the plus and minus components of bidirectional motion. However, existing methods of in vitro microtubule polarity labeling are unsuitable for high-resolution recording of motion. Here we present a simple and reliable method that uses avidin-coated magnetic beads to prepare microtubules labeled at the minus end. The microtubule polarity can be identified without any need for fluorescence excitation. We demonstrate video-rate high-resolution imaging of single cellular organelles moving along plus and minus directions on labeled microtubules. Quantitative analysis of this motion indicates that these organelles are likely to be driven by multiple dynein motors in vivo. PMID:19594454

Soppina, Virupakshi; Rai, Arpan; Mallik, Roop



Hollow waveguide optimization for fluorescence based detection  

Microsoft Academic Search

Previously, we created antiresonant reflecting optical waveguides (ARROWs) with hollow cores that guide light through gas and liquid media. We have demonstrated that these ARROWs can be used in sensing applications with single particle sensitivity using fluorescence correlation spectroscopy. To increase sensitivity for single molecule sensing, we have improved our initial designs and fabrication methods to decrease ARROW background fluorescence

Evan J. Lunt; Brian S. Phillips; Cory J. Jones; Aaron R. Hawkins; Philip Measor; Sergei Kuehn; Holger Schmidt



Nanoparticle-based energy transfer for rapid and simple detection of protein glycosylation  

Microsoft Academic Search

Glycan moiety of glycoproteins plays an essential role in its biological activity in vivo, and the analysis of glycosylation is of great importance in the development of protein therapeutics. In this study, we report a rapid and simple detection of protein glycosylation based on the fluorescence resonance energy transfer (FRET) between concanavalin A-conjugated gold nanoparticles (ConA-AuNPs) and dextran-conjugated quantum dots

Eunkeu Oh; Dohoon Lee; Young-Pil Kim; Seung YOUP Cha; Doo-Beyong Oh; Jungbae Kim; Hyun AH Kang; Hak-Sung Kim



A Simple Sequence Repeat-Based Linkage Map of Barley  

Microsoft Academic Search

A total of 568 new simple sequence repeat (SSR)-based markers for barley have been developed from a combination of database sequences and small insert genomic libraries enriched for a range of short simple sequence repeats. Analysis of the SSRs on 16 barley cultivars revealed variable levels of informativeness but no obvious correlation was found with SSR repeat length, motif type,

L. Ramsay; M. Macaulay; K. MacLean; L. Cardle; J. Fuller; K. J. Edwards; S. Tuvesson; M. Morgante; A. Massari; E. Maestri; N. Marmiroli; T. Sjakste; M. Ganal; W. Powell; R. Waugh


A spectrofluorometer to measure difference in fluorescence spectra: a simple method for improving sensitivity.  


To accurately measure small changes in fluorescence spectra a difference spectrofluorometer was designed and constructed. The instrument simultaneously measures fluorescence from two samples. Routinely, two identical samples are used; one serving as the reference while the other is subjected to experimentation. This procedure minimizes variations in fluorescence arising from instabilities in either the sensitivity of the instrument or from the sample with time. PMID:3734325

Brody, S S; Feliccia, V L



A simple fluorescent technique for screening cervical cells prior to nuclear analysis.  


Monolayer spreads of cervical cells were prepared and stained with haematoxylin and rhodamine-alpha-N-agmatine, a fluorescent marker for a cell surface protease. Mature epithelial cells from normal cervices lacked this cell surface enzyme and did not fluoresce. The abnormal cells possessed the cell surface enzyme, bound the probe and were quickly detected by fluorescence microscopy. The degree of abnormality of these fluorescent cells was determined by examination of their nuclear details, with the result that mild, moderate and severe dyskaryotic cells could be defined. PMID:1295461

Steven, F S; Johnson, J; Eason, P


Phytoplankton photocompensation from space-based fluorescence measurements  

NASA Astrophysics Data System (ADS)

Recent satellite-derived observations linked global scale phytoplankton fluorescence variability with iron stress and hinted at photophysiological responses associated with changing light levels. These photocompensation reactions, the sum of photoacclimation and photoadaptation, were examined with climatological data for the Gulf of Maine. Significant seasonal variability was observed in the fluorescence quantum yield that was unrelated to patterns of biomass. Up to 89% of the variability in the fluorescence quantum yield was explained by a physiology-based photocompensation model. Spatial variability in seasonal patterns was associated with differing hydrodynamic regimes. This variability in the quantum yield demonstrates that satellite-based fluorescence is inappropriate for phytoplankton biomass determinations. More importantly, the work presented here provides the modeling foundation for fluorescence-based investigations of temporal and spatial variability in phytoplankton physiology associated with growth irradiance. These space-based physiological observations have the potential to decrease uncertainties in future ocean color derived primary productivity estimates.

Morrison, J. Ruairidh; Goodwin, Deborah S.



Disposable nitrate-selective optical sensor based on fluorescent dye  

Technology Transfer Automated Retrieval System (TEKTRAN)

A simple, disposable thin-film optical nitrate sensor was developed. The sensor was fabricated by applying a nitrate-selective polymer membrane on the surface of a thin polyester film. The membrane was composed of polyvinylchloride (PVC), plasticizer, fluorescent dye, and nitrate-selective ionophore...


A simple fluorescence technique to stain the plasma membrane of human neutrophils  

Microsoft Academic Search

Three different fluorochromes were tested for their ability to label the plasma membrane proteins of neutrophils without labelling intracellular structures. A fluorescence quenching technique was used to differentiate between extra- and intracellularly localized fluorescence. Fluorescamin and fluoresceinisothiocyanate were shown to stain intracellular structures as well as the plasma membranes of the cells. Another fluorochrome, Evans Blue, is proposed since this

J. Hedl; C. Dahlgren; I. Rundquist



A highly selective fluorescent probe for Al3+ based on 4-aminoantipyrine  

NASA Astrophysics Data System (ADS)

A novel and simple Schiff base based on 2-pyridine formaldehyde and 4-aminoantipyrine was synthesized and characterized as a fluorescent probe. In the presence of Al3+, the fluorescent intensity has a dramatic enhancement over other examined metal ions in aqueous solution. The method of Job's plot indicated the formation of 1:1 complex between probe and Al3+, and the possible binding mode of the system was also proposed. Moreover, other examined metal ions had no effect on the detection of Al3+.

Zhou, Yanmei; Zhang, Junli; Zhou, Hua; Hu, Xiaoyi; Zhang, Lin; Zhang, Min



A highly selective fluorescent probe for Al3+ based on 4-aminoantipyrine.  


A novel and simple Schiff base based on 2-pyridine formaldehyde and 4-aminoantipyrine was synthesized and characterized as a fluorescent probe. In the presence of Al(3+), the fluorescent intensity has a dramatic enhancement over other examined metal ions in aqueous solution. The method of Job's plot indicated the formation of 1:1 complex between probe and Al(3+), and the possible binding mode of the system was also proposed. Moreover, other examined metal ions had no effect on the detection of Al(3+). PMID:23376261

Zhou, Yanmei; Zhang, Junli; Zhou, Hua; Hu, Xiaoyi; Zhang, Lin; Zhang, Min



Characterization of Flavin-Based Fluorescent Proteins: An Emerging Class of Fluorescent Reporters  

PubMed Central

Fluorescent reporter proteins based on flavin-binding photosensors were recently developed as a new class of genetically encoded probes characterized by small size and oxygen-independent maturation of fluorescence. Flavin-based fluorescent proteins (FbFPs) address two major limitations associated with existing fluorescent reporters derived from the green fluorescent protein (GFP)–namely, the overall large size and oxygen-dependent maturation of fluorescence of GFP. However, FbFPs are at a nascent stage of development and have been utilized in only a handful of biological studies. Importantly, a full understanding of the performance and properties of FbFPs as a practical set of biological probes is lacking. In this work, we extensively characterize three FbFPs isolated from Pseudomonas putida, Bacillus subtilis, and Arabidopsis thaliana, using in vitro studies to assess probe brightness, oligomeric state, maturation time, fraction of fluorescent holoprotein, pH tolerance, redox sensitivity, and thermal stability. Furthermore, we validate FbFPs as stable molecular tags using in vivo studies by constructing a series of FbFP-based transcriptional constructs to probe promoter activity in Escherichia coli. Overall, FbFPs show key advantages as broad-spectrum biological reporters including robust pH tolerance (4–11), thermal stability (up to 60°C), and rapid maturation of fluorescence (<3 min.). In addition, the FbFP derived from Arabidopsis thaliana (iLOV) emerged as a stable and nonperturbative reporter of promoter activity in Escherichia coli. Our results demonstrate that FbFP-based reporters have the potential to address key limitations associated with the use of GFP, such as pH-sensitive fluorescence and slow kinetics of fluorescence maturation (10–40 minutes for half maximal fluorescence recovery). From this view, FbFPs represent a useful new addition to the fluorescent reporter protein palette, and our results constitute an important framework to enable researchers to implement and further engineer improved FbFP-based reporters with enhanced brightness and tighter flavin binding, which will maximize their potential benefits.

Mukherjee, Arnab; Schroeder, Charles M.



Toward quantitatively fluorescent carbon-based ``quantum'' dots  

Microsoft Academic Search

Carbon-based ``quantum'' dots (or ``carbon dots'') are generally defined as surface-passivated small carbon nanoparticles that are brightly fluorescent. Apparently, the carbon particle surface passivation in carbon dots is critical to their fluorescence performance. An effective way to improve the surface passivation is to dope the surface of the precursor carbon nanoparticles with an inorganic salt, followed by the typical functionalization

Parambath Anilkumar; Xin Wang; Li Cao; Sushant Sahu; Jia-Hui Liu; Ping Wang; Katerina Korch; Kenneth N. Tackett II; Alexander Parenzan; Ya-Ping Sun



Multistage Spatial Property Based Segmentation for Quantification of Fluorescence Distribution in Cells  

NASA Astrophysics Data System (ADS)

The interpretation of the distribution of fluorescence in cells is often by simple visualization of microscope-derived images for qualitative studies. In other cases, however, it is desirable to be able to quantify the distribution of fluorescence using digital image processing techniques. In this paper, the challenges of fluorescence segmentation due to the noise present in the data are addressed. We report that intensity measurements alone do not allow separation of overlapping data between target and background. Consequently, spatial properties derived from neighborhood profile were included. Mathematical Morphological operations were implemented for cell boundary extraction and a window based contrast measure was developed for fluorescence puncta identification. All of these operations were applied in the proposed multistage processing scheme. The testing results show that the spatial measures effectively enhance the target separability.

Zhang, Guangyun; Jia, Xiuping; Pham, Tuan D.; Crane, Denis I.



Fluorescence-based resource for semi-automated genomic analyses  

SciTech Connect

To facilitate the practical application of highly efficient semi-automated methods for general application in genomic analyses, we have developed a fluorescence-based marker resource. Ninety highly polymorphic simple tandem repeat markers were combined to provide a rapid, accurate, and highly efficient initial genome-wide screening system. These markers are spaced on average every 33 recombination units, with a mean heterozygosity of 81% (range 65-94%), covering 22 autosomes and the X and Y chromosomes. Less than 3% of the genome lies beyond 30 cM of the nearest marker. Markers were placed in a vertical ladder that we have termed a SET according to the size of the PCR fragments they produce during electrophoresis. Each SET was designed to avoid overlap between loci during gel separations to assure accuracy when scoring genotypes. We have constructed 15 SETS of markers. Three SETS, each labelled with one of three fluors, were combined into what we have termed a GROUP, which is co-electrophoresed with internal size standards that are labelled with a fourth flour. Five GROUPS of markers were assembled that contain a total of 15 SETS of markers. Each GROUP cover 18 regions of the genome that can be detected simultaneously, since this genomic analysis system is fully compatible with automated fragment analyzers using simultaneous four-color fluorescence-based detection systems. This allows for multiplex detection and a throughput of 1,944 genotypes daily per instrument. This system will be highly beneficial in a number of clinical and research applications including: linkage, cancer genetics, forensics, and cytogenetics.

Kiser, M.B.; Dragwa, C.; Jedlicka, A.E. [Johns Hopkins Medical Institutions, Baltimore, MD (United States)] [and others



Hollow waveguide optimization for fluorescence based detection  

NASA Astrophysics Data System (ADS)

Previously, we created antiresonant reflecting optical waveguides (ARROWs) with hollow cores that guide light through gas and liquid media. We have demonstrated that these ARROWs can be used in sensing applications with single particle sensitivity using fluorescence correlation spectroscopy. To increase sensitivity for single molecule sensing, we have improved our initial designs and fabrication methods to decrease ARROW background fluorescence and improve transitions between solid and hollow waveguides. Photoluminescence of ARROW layers creates background fluorescence that masks the desired fluorescence signals. To improve sensitivity, we have optimized the PECVD ARROW layers to minimize the photoluminescence of each layer. Sensing applications require that hollow waveguides interface with solid waveguides on the substrate to direct light into and out of test media. Our previous ARROW designs required light at these interfaces to pass through the anti-resonant layers. Although in theory, high transmission through ARROW layers can be achieved, in practice, passing through these layers has limited transmission efficiencies. A new design coats the top and sides of the hollow core with only silicon dioxide, allowing light at interfaces to pass directly from silicon dioxide into the hollow core. This new design exhibits good mode confinement in the hollow core.

Lunt, Evan J.; Phillips, Brian S.; Jones, Cory J.; Hawkins, Aaron R.; Measor, Philip; Kuehn, Sergei; Schmidt, Holger



Fluorescence-based optochemical sensor on flexible foils  

NASA Astrophysics Data System (ADS)

This paper describes the implementation of a low-cost technology platform for fluorescence-based optochemical sensors made up of arrays of multimode waveguides and coupling structures integrated onto a flexible substrate. Such a configuration is ideal for multi-analyte detection owing to a possibility of future integration of different dyes in each waveguides. The presence of light sources, fluorescent sensing elements and photodetectors in a foil platform makes it a compact optochemical sensor, which has wide-range of applications in medical, biochemical, and environmental diagnostics. Flexible lightguides fabricated using soft-lithography based replication techniques, are used in combination with 45° micromirror coupling structures, having a loss of 0.5dB. Fluorescent dyes are incorporated with the lightguides enabling a detection of shift in fluorescence-peaks in contact with gases, which are read-out at the detection. Initial measurements yielded promising results of the waveguides mixed with fluorescent dyes showing response to toluene.

Kalathimekkad, Sandeep; Missinne, Jeroen; Arias Espinoza, Juan Diego; Van Hoe, Bram; Bosman, Erwin; Smits, Edsger; Mandamparambil, Rajesh; Van Steenberge, Geert; Vanfleteren, Jan



Spectral properties of a simple azine Schiff base and its sensing ability towards protic environment through hydrogen bonding interaction  

NASA Astrophysics Data System (ADS)

A simple azine linkage containing Schiff base p-N,N-diethylaminobenzaldazine (PDEAB) has been synthesized and its spectroscopic properties have been investigated using steady state absorption and fluorescence measurement. Both the absorption and emission studies indicate that the compound PDEAB forms intermolecular hydrogen bond with protic solvents. The formation of intermolecular hydrogen bond between PDEAB and protic solvents is further verified by Quantum chemical calculation using Density Functional Theory (DFT) (B3LYP/6-31++G(d,p)) and Natural Bond Orbital (NBO) analysis. The non-fluorescent nature (fluorescence off) of PDEAB in aprotic environment can be switched over to a fluorescent system (fluorescence on) in presence of protic solvents and hence this molecule can be used as highly sensitive fluorosensor for protic solvent in aprotic medium like ACN or DOX.

Ray, Debarati; Dalapati, Sasanka; Guchhait, Nikhil



Spectral properties of a simple azine Schiff base and its sensing ability towards protic environment through hydrogen bonding interaction.  


A simple azine linkage containing Schiff base p-N,N-diethylaminobenzaldazine (PDEAB) has been synthesized and its spectroscopic properties have been investigated using steady state absorption and fluorescence measurement. Both the absorption and emission studies indicate that the compound PDEAB forms intermolecular hydrogen bond with protic solvents. The formation of intermolecular hydrogen bond between PDEAB and protic solvents is further verified by Quantum chemical calculation using Density Functional Theory (DFT) (B3LYP/6-31++G(d,p)) and Natural Bond Orbital (NBO) analysis. The non-fluorescent nature (fluorescence off) of PDEAB in aprotic environment can be switched over to a fluorescent system (fluorescence on) in presence of protic solvents and hence this molecule can be used as highly sensitive fluorosensor for protic solvent in aprotic medium like ACN or DOX. PMID:23835054

Ray, Debarati; Dalapati, Sasanka; Guchhait, Nikhil



Fluorescence enhancement upon gelation and thermally-driven fluorescence switches based on tetraphenylsilole-based organic gelators  

Microsoft Academic Search

Two new organic gelators 1 and 2 based on the silole (silacyclopentadiene) framework were designed with the end to develop switchable fluorescent organogels, by making use of the aggregation-induced emission (AIE) feature of silole derivatives. As for other silole derivatives, compounds 1 and 2 exhibited AIE behavior as indicated by the significant fluorescence enhancement by introducing water to the THF

Ming Wang; Deqing Zhang; Guanxin Zhang; Daoben Zhu



10 CFR 429.35 - Bare or covered (no reflector) medium base compact fluorescent lamps.  

Code of Federal Regulations, 2013 CFR

...false Bare or covered (no reflector) medium base compact fluorescent lamps. 429...35 Bare or covered (no reflector) medium base compact fluorescent lamps. ...applicable to bare or covered (no reflector) medium base compact fluorescent lamps;...



Knowledge-Based Vision and Simple Visual Machines  

Microsoft Academic Search

The vast majority of work in machine vision emphasizes the representation of perceived objects and events: it is these internal representations that incorporate the 'knowledge' in knowledge-based vision or form the 'models' in model-based vision. In this paper, we discuss simple machine vision systems developed by artificial evolution rather than traditional engineering design techniques, and note that the task of

Dave Cliff; Jason Noble



Simple Rule-Based Part of Speech Tagger.  

National Technical Information Service (NTIS)

Automatic part of speech tagging is an area of natural language processing where statistical techniques have been more successful than rule-based methods. In this paper, we present a simple rule-based part of speech tagger which automatically acquires its...

E. Brill



Reversible "off-on" fluorescent chemosensor for Hg 2+ based on rhodamine derivative  

NASA Astrophysics Data System (ADS)

A novel and simple fluorescent chemosensor based on rhodamine was designed and synthesized to detect Hg 2+ with high selectivity. The structure of chemosensor 1 was characterized by IR, 1H NMR, and HRMS spectroscopies. Chemosensor 1 exhibited distinct fluorescent and colorimetric changes toward Hg 2+ in an ethanol/water (80/20, v/v) solution, which resulted in the formation of 1/Hg 2+ complex with the Hg 2+-induced ring opening of the spirolactam ring in rhodamine. The reversibility of chemosensor 1 was verified through its spectral response toward Hg 2+ ions and TBAI (tetrabutylammonium iodide) titration experiments.

Liu, Weimin; Chen, Jianhong; Xu, Liwei; Wu, Jiasheng; Xu, Haitao; Zhang, Hongyan; Wang, Pengfei



Fluorescence tomography of biological tissue based on ultrasound tagging technique  

NASA Astrophysics Data System (ADS)

We report a study for the development of tomographic imaging technique of fluorescence in biological tissue for assays of biological function. Ultrasonic modulation of light based on the acousto-optic effect (so called ultrasound 'tagging') is applied for imaging of fluorescence distribution in the light-scattering media. Sound-field characteristics that affect the light by modulating its amplitude through variation of the refractive index in the medium were determined. With using focused ultrasound, selectively modulated fluorescence on a depth-axis of the medium can be detected. Ultrasound tagging technique applied measuring the optical absorption in light scattering media is well known, and it is principally based on the modulation of speckle pattern. On the contrary, in the case of fluorescence, displacement of scattering particles and variation of the refractive index that is induced by density distribution in a sound field might produce the intensity modulation of scattered light. We have experimentally shown that ultrasound tagging technique is also available for fluorescence measurement. In this paper, we demonstrate the result of tomographic images of fluorescence in dense scattering media using porcine muscle as a biological tissue, and bovine adipose. Tissue samples had the dimension of 40 x 40 mm in section and fluorophore which had the 3mm size was embedded in the center of the tissue. The localized image of the fluorophore was determined with the spatial resolution of focus size of the ultrasound, suggesting the applicability of this technique for visualization of fluorescent probes in deep portion of living body.

Kobayashi, Masaki; Mizumoto, Takashi; Duc, Trinh Quang; Takeda, Motohiro



A rapid, simple measurement of human albumin in whole blood using a fluorescence immunoassay (I)  

Microsoft Academic Search

Background: Human serum albumin (HSA) is the most abundant plasma protein and plays key a role in metabolism. The variation in albumin concentration provides valuable information related to metabolic diseases and diagnostic application. Methods: We constructed two assay systems to quantify the albumin concentration. The immunoassay used a fluorescence (FL) dye to detect albumin in samples and employed the conventional

Sunga Choi; Eui Yul Choi; Dong Joon Kim; Jae Hoon Kim; Tai Sun Kim; Sang Wook Oh



Fluorescence quenching of Rhodamine B base by two amines  

NASA Astrophysics Data System (ADS)

Fluorescence quenching of Rhodamine B base (RhB) in DMF solution has been studied at different concentrations of the amine Triethyl amine (TEA) and n-butyl amine (NBA) at room temperature. It has been observed that the fluorescence intensity of RhB decrease with increase in the concentration of the TEA and NBA. It has been observed that the quenching due to amines proceeds via dynamic quenching process. The rate constants for the quenching process have been calculated using Stern-Volmer equation. Time resolved fluorescence study and 1H NMR spectral study have also been carried out and discussed.

Bakkialakshmi, S.; Selvarani, P.; Chenthamarai, S.



A green-fluorescent-protein-based assay for the characterization of G-protein-coupled receptors  

Microsoft Academic Search

A simple and sensitive system to monitor activation and pharmacology of G-protein-coupled receptors in mammalian cells is described. It is based on cAMP-responsible-element-regulated expression of green fluorescent protein (GFP). Cotransfection with appropriate G-protein-coupled receptors and subsequent activation with agonists induces expression of GFP in a dose-dependent manner. This system is suited for the analysis of most G-protein-coupled receptors, including those

Thomas Roeder; Derk Görich; Dörte Heyden; Michael Gewecke



Rhodamine-based ratiometric fluorescent ion-selective bulk optodes  

Microsoft Academic Search

We developed the highly sensitive bulk optodes with new synthesized rhodamine type of fluorescent chromoionophore. The fluorescent turn-on through spirolactam ring-opening of rhodamine dye inside the hydrophobic polymer phase can only be obtained by incorporating cation exchanger into the sensing membrane. The rhodamine-based optode containing copper ionophore exhibited nanomolar detection limit to copper ion at pH 5.5. By introducing an

Xiaojiang Xie; Xue Li; Yifan Ge; Yu Qin; Hong-Yuan Chen



Ultrasensitive Fluorescence-Based Detection of Nascent Proteins in Gels  

Microsoft Academic Search

The most common method of analysis of proteins synthesized in a cell-free translation system (e.g., nascent proteins) involves the use of radioactive amino acids such as [35S]methionine or [14C]leucine. We report a sensitive, nonisotopic, fluorescence-based method for the detection of nascent proteins directly in polyacrylamide gels. A fluorescent reporter group is incorporated at the N-terminus of nascent proteins using an

Sadanand Gite; Sergey Mamaev; Jerzy Olejnik; Kenneth Rothschild



A simple RTD-based circuit for Boolean CNN cells  

Microsoft Academic Search

We investigate the use of resonant tunneling diodes (RTDs) in circuits for Boolean cellular neural networks (CNNs). RTDs excel in their electronic properties, their size, and switching speed, and can readily be integrated together with GaAs FETs. A simple RTD-based circuit is proposed and shown to be capable of realizing linearly separable Boolean functions. To implement the network parameters, the

M. Hanggi; L. O. Chua; R. Dogaru



Confinement effects in the adsorption of simple bases by zeolites  

Microsoft Academic Search

An analysis of the heats of adsorption of ammonia and several simple amines on zeolites MOR and MFI, based on recent literature data, shows that these adsorption heats are determined by both chemical (heat of protonation) and physical (confinement) factors. Confinement effects which arise from van der Waals interactions cannot be ignored as they may represent up to 40% of

Eric G Derouane; Clarence D Chang



[Simple quantitation of arsenic by energy dispersive fluorescence X-ray spectrometer using Reinsch's test].  


We examined the clinical usefulness of the Reinsch's test for the detection of the small amounts of the heavy metals such as arsenic and mercury using the fluorescence X-ray spectrometry. We tried t o measure various kinds of biological samples, including serum, urine, and gastric contents using this method. 0.4 ml or 1 ml of hydrochloric acid were added to 2 ml of serum or 6 ml urine and gastric content, respectively, and a copper plate (5.0x 0.8 cm) was immersed into this solution. The mixture heated at 90 degrees C by a heating block for 30 minutes. After heating, the copper plate was washed with water and dried. The copper arsenide that stuck to the copper plate due to Reinsch's test dissolved by methanol/ammonia (8:2) solution at 60 degrees C for 15 minutes. A drop gave the solution to a filter paper fluorescence X-ray analysis and completely dried the filter paper, and applied to the fluorescence X-ray spectrometer. As a result, this method showed about 20 times high sensitivity in comparison with the measurement with condition of solution. The minimal detectable limits of rsenic was 0.4 ppm in serum and were 0.2 ppm in rine and gastric content. The calibration curve could be made for 0.5 to 50 ppm. It will take about 90 min for the measurement using this method for the detection of arsenic in biological samples. We showed the usefulness of the Reinsch's test using the fluorescence X-ray spectrometry in the clinical toxicology. PMID:15678930

Ozo, Yumiko; Yoshizawa, Mie; Murata, Atsuo; Shimazaki, Syuji; Kajiwara, Masahiro; Takagi, Tetsuya; Sato, Yoshinobu



Wide field-of-view Talbot grid-based microscopy for multicolor fluorescence imaging.  


The capability to perform multicolor, wide field-of-view (FOV) fluorescence microscopy imaging is important in screening and pathology applications. We developed a microscopic slide-imaging system that can achieve multicolor, wide FOV, fluorescence imaging based on the Talbot effect. In this system, a light-spot grid generated by the Talbot effect illuminates the sample. By tilting the excitation beam, the Talbot-focused spot scans across the sample. The images are reconstructed by collecting the fluorescence emissions that correspond to each focused spot with a relay optics arrangement. The prototype system achieved an FOV of 12 × 10 mm(2) at an acquisition time as fast as 23 s for one fluorescence channel. The resolution is fundamentally limited by spot size, with a demonstrated full-width at half-maximum spot diameter of 1.2 ?m. The prototype was used to nimage green fluorescent beads, double-stained human breast cancer SK-BR-3 cells, Giardia lamblia cysts, and the Cryptosporidium parvum oocysts. This imaging method is scalable and simple for implementation of high-speed wide FOV fluorescence microscopy. PMID:23787643

Pang, Shuo; Han, Chao; Erath, Jessey; Rodriguez, Ana; Yang, Changhuei



Probing subtle fluorescence dynamics in cellular proteins by streak camera based fluorescence lifetime imaging microscopy  

NASA Astrophysics Data System (ADS)

We report the cell biological applications of a recently developed multiphoton fluorescence lifetime imaging microscopy system using a streak camera (StreakFLIM). The system was calibrated with standard fluorophore specimens and was shown to have high accuracy and reproducibility. We demonstrate the applicability of this instrument in living cells for measuring the effects of protein targeting and point mutations in the protein sequence, which are not obtainable by conventional intensity-based fluorescence microscopy methods. We discuss the relevance of such time-resolved information in quantitative energy transfer microscopy and in measurement of the parameters that characterize the intracellular physiology.

Krishnan, R. V.; Biener, Eva; Zhang, Jian-Hua; Heckel, Robert; Herman, Brian



A quick and simple FISH protocol with hybridization-sensitive fluorescent linear oligodeoxynucleotide probes  

PubMed Central

Fluorescence in situ hybridization (FISH) is a powerful tool used in karyotyping, cytogenotyping, cancer diagnosis, species specification, and gene-expression analysis. Although widely used, conventional FISH protocols are cumbersome and time consuming. We have now developed a FISH method using exciton-controlled hybridization-sensitive fluorescent oligodeoxynucleotide (ECHO) probes. ECHO–FISH uses a 25-min protocol from fixation to mounting that includes no stringency washing steps. We use ECHO–FISH to detect both specific DNA and RNA sequences with multicolor probes. ECHO–FISH is highly reproducible, stringent, and compatible with other fluorescent cellular labeling techniques. The resolution allows detection of intranuclear speckles of poly(A) RNA in HeLa cells and dissociated hippocampal primary cultures, and mRNAs in the distal dendrites of hippocampal neurons. We also demonstrate detection of telomeric and centromeric DNA on metaphase mouse chromosomes. The simplicity of the ECHO–FISH method will likely accelerate cytogenetic and gene-expression analysis with high resolution.

Wang, Dan Ohtan; Matsuno, Hitomi; Ikeda, Shuji; Nakamura, Akiko; Yanagisawa, Hiroyuki; Hayashi, Yasunori; Okamoto, Akimitsu



CdSe/ZnS quantum dots based fluorescence quenching method for determination of paeonol.  


Aqueous polymethylmethacrylate (PMMA)-capped CdSe/ZnS quantum dots were used as fluorescence probes for paeonol determination. Based on the fluorescence quenching of aqueous CdSe/ZnS quantum dots caused by paeonol, a simple, sensitive and rapid method was developed. Under the optimal conditions, with excitation and emission wavelengths at 350 nm and 620 nm, respectively, the calibration plot of F0-F with concentration of paeonol was linear in the range of 25.04-175.2 mg L(-1) with correlation coefficient of 0.9986. The limit of detection was 0.017 mg L(-1). The concentration of paeonol in paeonol ointment was determined by the proposed method and the result agreed with the claimed value. Furthermore, the possible fluorescence quenching mechanism was discussed. PMID:21147020

Dong, Wei; Shen, Huai-Bin; Liu, Xiu-Hua; Li, Ming-Jing; Li, Lin-Song



Doped semiconductor nanocrystal based fluorescent cellular imaging probes.  


Doped semiconductor nanocrystals such as Mn doped ZnS, Mn doped ZnSe and Cu doped InZnS, are considered as new classes of fluorescent biological probes with low toxicity. Although the synthesis in high quality of such nanomaterials is now well established, transforming them into functional fluorescent probes remains a challenge. Here we report a fluorescent cellular imaging probe made of high quality doped semiconductor nanocrystals. We have identified two different coating approaches suitable for transforming the as synthesized hydrophobic doped semiconductor nanocrystals into water-soluble functional nanoparticles. Following these approaches we have synthesized TAT-peptide- and folate-functionalized nanoparticles of 10-80 nm hydrodynamic diameter and used them as a fluorescent cell label. The results shows that doped semiconductor nanocrystals can be an attractive alternative for conventional cadmium based quantum dots with low toxicity. PMID:23674276

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



Combination of DNA ligase reaction and gold nanoparticle-quenched fluorescent oligonucleotides: a simple and efficient approach for fluorescent assaying of single-nucleotide polymorphisms.  


A new fluorescent sensing approach for detection of single-nucleotide polymorphisms (SNPs) is proposed based on the ligase reaction and gold nanoparticle (AuNPs)-quenched fluorescent oligonucleotides. The design exploits the strong fluorescence quenching of AuNPs for organic dyes and the difference in noncovalent interactions of the nanoparticles with single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA), where ssDNA can be adsorbed onto the surface of AuNPs while dsDNA cannot be. In the assay, two half primer DNA probes, one being labeled with a dye and the other being phosphorylated, were first incubated with a target DNA template. In the presence of DNA ligase, the two captured ssDNAs are linked for the perfectly matched DNA target to form a stable duplex, but the duplex could not be formed by the single-base mismatched DNA template. After addition of AuNPs, the fluorescence of dye-tagged DNA probe will be efficiently quenched unless the perfectly matched DNA target is present. To demonstrate the feasibility of this design, the performance of SNP detection using two different DNA ligases, T4 DNA ligase and Escherichia coli DNA ligase, were investigated. In the case of T4 DNA ligase, the signal enhancement of the dye-tagged DNA for perfectly matched DNA target is 4.6-fold higher than that for the single-base mismatched DNA. While in the presence of E. coli DNA ligase, the value raises to be 30.2, suggesting excellent capability for SNP discrimination. PMID:20726510

Wang, Hao; Li, Jishan; Wang, Yongxiang; Jin, Jiangyu; Yang, Ronghua; Wang, Kemin; Tan, Weihong



Luminol as a fluorescent acid-base indicator.  


The acid and base dissociation constants of luminol are determined at various ionic strengths. The transition interval occurs at pH 7.7-9.0, therefore luminol is a fluorescent indicator for the titration of strong and weak acids and strong bases. Its value as an indicator is established by titrating milk, red wine and cherry juice. PMID:18959899

Erdey, L; Buzás, I; Vigh, K



mb-FLIM: model-based fluorescence lifetime imaging  

NASA Astrophysics Data System (ADS)

We have developed a model-based, parallel procedure to estimate fluorescence lifetimes. Multiple frequencies are present in the excitation signal. Modeling the entire fluorescence and measurement process produces an analytical ratio of polynomials in the lifetime variable ?. A non-linear model-fitting procedure is then used to estimate ?. We have analyzed this model-based approach by simulating a 10 ?M fluorescein solution (? = 4 ns) and all relevant noise sources. We have used real LED data to drive the simulation. Using 240 ?s of data, we estimate ? = 3.99 ns. Preliminary experiments on real fluorescent images taken from fluorescein solutions (measured ? = 4.1 ns), green plastic test slides (measured ? = 3.0 ns), and GFP in U2OS (osteosarcoma) cells (measured ? = 2.1 ns) demonstrate that this model-based measurement technique works.

Zhao, Qiaole; Young, Ian Ted; Schouten, Raymond; Stallinga, Sjoerd; Jalink, Kees; de Jong, Sander



A Simple Visualization of Double Bond Properties: Chemical Reactivity and UV Fluorescence  

ERIC Educational Resources Information Center

|A simple, easily visualized thin-layer chromatography (TLC) staining experiment is presented that highlights the difference in reactivity between aromatic double bonds and nonaromatic double bonds. Although the stability of aromatic systems is a major theme in organic chemistry, the concept is rarely reinforced "visually" in the undergraduate…

Grayson, Scott M.



Fluorescence-based planar waveguide biosensors  

Microsoft Academic Search

In this paper, different luminescence-based sensing configurations and examples of applications in bioaffinity assays relevant for the key life science areas, demonstrating the achieved system performance, will be presented and compared with literature results from refraction-based techniques.

Gert L. Duveneck



A simple rate-equation-based thermal VCSEL model  

Microsoft Academic Search

Motivated by the potentially large number of devices and simulations involved in optoelectronic system design, and the associated need for compact optoelectronic device models, we present a simple thermal model of vertical-cavity surface-emitting laser (VCSEL) light-current (LI) characteristics based on the laser rate equations and a thermal offset current. The model was implemented in conventional SPICE-like circuit simulators, including HSPICE,

P. V. Mena; J. J. Morikuni; S.-M. Kang; A. V. Harton; K. W. Wyatt



Direct detection of sulfide ions [S2-] in aqueous media based on fluorescence quenching of functionalized CdS QDs at trace levels: analytical applications to environmental analysis.  


A novel, simple but highly selective fluorescent probe is developed for the direct detection of sulfide ions [S(2-)] based on the fluorescence quenching of the functionalized CdS QDs in aqueous solution at trace levels and successfully applied for quantitation of S(2-) from water samples in a complex matrix exclusive of pretreatment by standard addition method. PMID:23334151

Gore, Anil H; Vatre, Sandip B; Anbhule, Prashant V; Han, Sung-Hwan; Patil, Shivajirao R; Kolekar, Govind B



Graphene oxide based fluorescent aptasensor for adenosine deaminase detection using adenosine as the substrate.  


We present a novel fluorescent aptasensor for simple and accurate detection of adenosine deaminase (ADA) activity and inhibition on the basis of graphene oxide (GO) using adenosine (AD) as the substrate. This aptasensor consists of a dye-labeled single-stranded AD specific aptamer, GO and AD. The fluorescence intensity of the dye-labeled AD specific aptamer is quenched very efficiently by GO as a result of strong ?-? stacking interaction and excellent electronic transference of GO. In the presence of AD, the fluorescence of the GO-based probe is recovered since the competitive binding of AD and GO with the dye-labeled aptamer prevents the adsorption of dye-labeled aptamer on GO. When ADA was introduced to this GO-based probe solution, the fluorescence of the probe was quenched owing to ADA can convert AD into inosine which has no affinity to the dye-labeled aptamer, thus allowing quantitative investigation of ADA activity. The as-proposed sensor is highly selective and sensitive for the assay of ADA activity with a detection limit of 0.0129U/mL in clean buffer, which is more than one order of magnitude lower than the previous reports. Meanwhile, a good linear relationship with the correlation coefficient of R=0.9922 was obtained by testing 5% human serum containing a series of concentrations of ADA. Additionally, the inhibition effect of erythro-9-(2-hydroxy-3-nonyl) adenine on ADA activity was investigated in this design. The GO-based fluorescence aptasensor not only provides a simple, cost-effective and sensitive platform for the detection of ADA and its inhibitor but also shows great potential in the diagnosis of ADA-relevant diseases and drug development. PMID:22613226

Xing, Xiao-Jing; Liu, Xue-Guo; Yue-He; Luo, Qing-Ying; Tang, Hong-Wu; Pang, Dai-Wen



Analysis of plasma catecholamines by high-performance liquid chromatography with fluorescence detection: simple sample preparation for pre-column fluorescence derivatization.  


Analysis of plasma catecholamines (norepinephrine, epinephrine and dopamine) by high-performance liquid chromatography using 1,2-diphenylethylenediamine as a fluorescent reagent is described. We have developed an automatic catecholamine analyser, based on pre-column fluorescence derivatization and column switching. The analysis time for one assay was 15 min. The correlation coefficients of the linear regression equations were greater than 0.9996 in the range 10-10,000 pg/ml. The detection limit, at a signal-to-noise ratio of 3, was 2 pg/ml for dopamine. A new method of sample preparation for the pre-column fluorescence derivatization of plasma catecholamines was used. In order to protect the catecholamines from decomposition, an ion-pair complex between boric acid and the diol group in the catecholamine was formed at a weakly alkaline pH. The stabilities of plasma catecholamines were evaluated at several temperatures. After complex formation, the catecholamines were very stable at 17 degrees C for 8 h, and the coefficients of variation for norepinephrine, epinephrine and dopamine were 1.2, 4.2 and 9.3%, respectively. PMID:1939468

Kamahori, M; Taki, M; Watanabe, Y; Miura, J



A Rapid Fluorescence-based Assay for Soluble Methane Monooxgyenase  

SciTech Connect

A fluorescence-based assay was developed to estimate soluble methane monooxygenase (sMMO) activity in solution. Whole cells of Methylosinus trichosporium OB3b expressing sMMO were used to oxidize various compounds to screen for fluorescent products. Of the 12 compounds tested, only coumarin yielded a fluorescent product. The UV absorbance spectrum of the product matches that of 7-hydroxycoumarin, and this identification was confirmed by 13C-NMR spectroscopy. The dependence of the fluorescent reaction on sMMO activity was investigated by pre-incubation with acetylene, a known inhibitor of sMMO activity. Apparent kinetic parameters for whole cells were determined to be Km(app)=262 µM and Vmax(app)=821 nmol 7-hydroxycoumarin min–1 mg protein–1. The rate of coumarin oxidation by sMMO correlates well with those of trichloroethylene degradation and naphthalene oxidation. Advantages of the fluorescence-based coumarin oxidation assay over the naphthalene oxidation assay include a more stable product, direct detection of the product without additional reagents, and greater speed and convenience.

Miller, Amber Reese; Keener, William Kelvin; Roberto, Francisco Figueroa; Watwood, Maribeth E.



A microcontroller-based emergency ballast for fluorescent lamps  

Microsoft Academic Search

This paper presents a new emergency ballast for fluorescent lamps. The fundamental block is the microcontroller-based control circuit, which performs the supervision and control function. High-frequency electronics techniques are proposed for the high power factor battery charger and the lamp driver, which provides high luminous efficacy. In this way, size and weight have been minimized for the whole system. With

J. Marcos Alonso; Pedro J. Villegas; J. Diaz; C. Blanco; M. Rico



Fluorescent lamp model based on the equivalent resistance variation  

Microsoft Academic Search

A circuit model simulating the electrical characteristics of a fluorescent lamp operating at high frequency is proposed. The model is based on exponential approximation that represents the equivalent resistance variation as function of power, constructed by experimental results for several power levels. Simulations and experimental results are presented to verify the feasibility of the model and, moreover, an electronic ballast

Murilo Cervi; A. R. Seidel; F. E. Bisogno; R. N. do Prado



Electrospun sol-gel fibers for fluorescence-based sensing  

Microsoft Academic Search

Fluorescence based biosensors have the ability to provide reliable pathogen detection. However, the performance could be improved by enhancing the effective surface area of the biosensor. We report on a new nanofibrous fluorescencebased biosensor, whereas a sol-gel platform mesh was constructed by utilizing electrospinning techniques. Furthermore, incorporating cetyltrimethylammonium bromide (CTAB) and conducting pore-forming techniques resulted in a high surface area

Jasenka Memisevic; Lela Riley; Sheila A. Grant



Dendrimer based fluorescent glucose sensor for diabetic monitoring  

NASA Astrophysics Data System (ADS)

Fluorescent glucose assays based on the affinity reaction between Concanavalin A and dextran have been extensively studied. However, advancements in polymer science have allowed for new macromolecules capable of replacing dextran which may improve the performance of this well-known assay. Dendrimer macromolecules, being highly ordered and spherical, allow for the binding of specific residues to the terminal (peripheral) binding sites, enabling researchers to customize the molecule. In this research, glycosylated dendrimers have been engineered to replace dextran to allow for more controlled chemical and fluorescent responses (eliminate multivalent binding and improve reversibility). This new assay has been shown to form small aggregate particles containing many Con A and glycosylated dendrimers resulting in a substantial loss in fluorescent intensity. Overall, this assay shows promise for use as part of an implantable glucose monitoring device, but more research needs to be done to increase sensor stability and optimize the sensor response to glucose.

Ibey, Bennett L.; Beier, Hope T.; Rounds, Rebecca M.; Pishko, Michael V.; Coté, Gerard L.



Polyacrylamide based ICG nanocarriers for enhanced fluorescence and photoacoustic imaging  

NASA Astrophysics Data System (ADS)

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

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



A new approach for fluorescence correlation spectroscopy (FCS) based immunoassays.  


Fluorescence correlation spectroscopy (FCS) is a powerful technique for measuring physicochemical properties, such as concentration and diffusion constant, of bio-molecules in complex mixtures. Although, as such, FCS is well suited for development of homogeneous immunoassays, a major obstacle lies in the relatively high molecular weight of antibodies. This is because in FCS discrimination between unbound fluorescently-labelled antibodies and the same antibodies bound to immune complexes is based on the difference of their respective diffusion coefficients. To overcome this limitation we here propose to use a fluorescently-labelled tag which has two crucial properties: (a) its molecular weight is significantly lower than that of an antibody and (b) it is capable to discriminate between free antibodies and immune complexes. We have evaluated the feasibility of this approach in a model system consisting of mouse monoclonal IgG directed against the Lewis X antigen, and Protein A as a low molecular weight tag. PMID:14711501

Mayboroda, Oleg A; van Remoortere, Alexandra; Tanke, Hans J; Hokke, Cornelis H; Deelder, André M



Simple super-resolution live-cell imaging based on diffusion-assisted F?rster resonance energy transfer  

PubMed Central

Despite the recent development of several super-resolution fluorescence microscopic techniques, there are still few techniques that can be readily employed in conventional imaging systems. We present a very simple, rapid, general and cost-efficient super-resolution imaging method, which can be directly employed in a simple fluorescent imaging system with general fluorophores. Based on diffusion-assisted Förster resonance energy transfer (FRET), fluorescent donor molecules that label specific target structures can be stochastically quenched by diffusing acceptor molecules, thereby temporally separating otherwise spatially overlapped fluorescence signals and allowing super-resolution imaging. The proposed method provides two- to three-fold-enhancement in spatial resolution, a significant optical sectioning property, and favorable temporal resolution in live-cell imaging. We demonstrate super-resolution live-cell dynamic imaging using general fluorophores in a standard epi-fluorescence microscope with light-emitting diode (LED) illumination. Due to the simplicity of this approach, we expect that the proposed method will prove an attractive option for super-resolution imaging.

Cho, Sangyeon; Jang, Jaeduck; Song, Chaeyeon; Lee, Heeyoung; Ganesan, Prabhakar; Yoon, Tae-Young; Kim, Mahn Won; Choi, Myung Chul; Ihee, Hyotcherl; Heo, Won Do; Park, YongKeun



Simple HPLC evaluation of lipoamidase activity in tissue using a newly synthesized fluorescent substrate, dansyl-?-lipoyllysine.  


?-Lipoic acid (LA) is a naturally occurring disulfide-containing compound used as an antioxidant supplement which also has been used as a medicine for diabetic neuropathy in Europe. Physiologically LA acts as a coenzyme of mitochondrial multienzyme complex in its protein bound form but it is not yet clear how the externally administrated LA is incorporated into other proteins in the same protein-bound form or why the bound form is active as an antioxidant. The binding and cleavage of LA to or from the protein is mediated by lipoamidase and thus determines LA distribution in tissues. We have developed a simple sensitive assay for lipoamidase using a fluorescent substrate, dansyl-?-lipoyllysine (DLL). Lipoamidase in tissues cleaves the amide bond between LA and the ?-amino-lysine moiety to release dansylated lysine (DL). A HPLC comparison of the fluorescence intensity between DLL and DL was used to quantify the enzyme activity. The hydrolytic reaction did not occur when the tissue was heat-treated before incubation with DLL and was inhibited by free LA, especially by the R-enantiomer of LA (physiologically active form). N(?)-Acetyl-L-lysine did not compete with DLL in the cleavage reaction. The method was applied for the determination of lipoamidase activity levels in various rat tissues. It was revealed the spleen had the highest activity followed by the kidney, heart, lung and liver. The activity in the brain was below the detection limit of the assay. PMID:22293216

Motafakkerazad, Rouhollah; Wang, Man-Yuan; Wada, Naoki; Matsugo, Seiichi; Konishi, Tetsuya



Gaseous ammonia fluorescence probe based on cellulose acetate modified microstructured optical fiber  

NASA Astrophysics Data System (ADS)

In this article, we report a novel fluorescent ammonia gas probe based on microstructured optical fiber (MOF) which is modified with eosin-doped cellulose acetate film. This probe was fabricated by liquid fluxion coating process. Polymer solution doped with eosin was directly inhaled into 18 array holes of MOF and then formed matrix film in them. The sensing properties of the optical fiber sensor to gaseous ammonia at room temperature were investigated. The sensing probe showed different fluorescence intensity at 576 nm to different concentrations of trace ammonia in carrier gas of nitrogen. The response range was 50-400 ppm, with short response time within 500 ms. Furthermore, the response range could be tailored through CTAB co-entrapment process in the sensing film. These test results demonstrated that low cost, simple structured fiber optic sensors for detecting ammonia gas samples could be developed based on MOF.

Peng, Lirong; Yang, Xinghua; Yuan, Libo; Wang, Lili; Zhao, Enming; Tian, Fengjun; Liu, Yanxin



Fluorescence sensing of adenosine deaminase based on adenosine induced self-assembly of aptamer structures.  


A new approach is proposed for simple detection of adenosine deaminase (ADA) based on adenosine induced self-assembly of two pieces of single-stranded DNA (ssDNA). These ssDNA are two fragments of the aptamer that has a strong affinity for adenosine and are labeled with carboxyfluorescein and black hole quencher-1, respectively. The complementarities of the bases in the two pieces of ssDNA are insufficient to form a stable structure. In the presence of adenosine, however, the ssDNA can be assembled into the intact aptamer tertiary structure, which results in fluorescence quenching of the carboxyfluorescein-labeled aptamer fragment. As a result, the adenosine-ssDNA complex shows a low background signal, which is rather desired for achieving sensitive detection. Reaction of the complex with ADA causes a great fluorescence enhancement by converting adenosine into inosine that has no affinity for the aptamer. This behaviour leads to the development of a simple and sensitive fluorescent method for assaying ADA activity, with a detection limit of 0.05 U mL(-1), which is more sensitive than most of the existing approaches. Furthermore, the applicability of the method has been demonstrated by detecting ADA in mouse serum samples. PMID:23462984

Feng, Tingting; Ma, Huimin



Highly selective and sensitive detection of mercuric ion based on a visual fluorescence method.  


The instant and on-site detection of trace aqueous mercuric ion still remains a challenge for environmental monitoring and protection. This work demonstrates a new analytical method and its utility for visual detection of aqueous Hg(2+) on the basis of a novel water-soluble CdSe-ZnS quantum dots (QDs) functionalized with a bidentate ligand of 2-hydroxyethyldithiocarbamate (HDTC). The fluorescence of the aqueous HDTC modified QDs (HDTC-QDs) could be selectively and efficiently quenched by Hg(2+) through a surface chelating reaction between HDTC and Hg(2+), and the detection limit was measured to be 1 ppb. Most interestingly, the orange fluorescence of the HDTC-QDs gradually changes to red upon the increasing amount of Hg(2+) added besides the decreasing of the fluorescence intensity. By taking advantage of this optical phenomenon, a paper-based sensor for aqueous Hg(2+) detection has been developed by immobilizing the HDTC-QDs on cellulose acetate paper which has low background fluorescence in the wavelength range. The paper-based sensor showed high sensitivity and selectivity for Hg(2+) visual detection. When Hg(2+) was dropped onto the paper-sensor, an obviously distinguishable fluorescence color evolution (from orange to red) could be clearly observed depending on the concentration of Hg(2+). The limit of detection of the visual method for aqueous Hg(2+) detection was as low as 0.2 ppm. The very simple and effective strategy reported here should facilitate the development of portable and reliable fluorescence chemosensors for mercuric pollution control. PMID:23121315

Yuan, Chao; Zhang, Kui; Zhang, Zhongping; Wang, Suhua



A rapid and simple method of evaluating the dimeric tendency of fluorescent proteins in living cells using a truncated protein of importin ? as fusion tag.  


Enhanced green fluorescent protein (EGFP) and its yellow variant (Venus) are weakly dimeric under physiological conditions. We designed a simple method to evaluate the dimeric tendency of fluorescent proteins in living mammalian cells. A novel single mutation, A206L, interfering with the hydrophobic interactions of the dimer interface in Venus, contributed to its monomerization, and was as effective as the A206K mutation in this assay. PMID:22313767

Nakagawa, Chika; Nishimura, Shigenori; Senda-Murata, Kaori; Sugimoto, Kenji



A simple and sensitive CE method for the simultaneous determination of catecholamines in urine with in-column optical fiber light-emitting diode-induced fluorescence detection  

Microsoft Academic Search

A simple and sensitive method has been developed for simultaneous analysis of three catecholamines: dopamine (DA), epinephrine (EP) and norepinephrine (NE) in urine by capillary electrophoresis (CE) coupled with in-column fiber-optic light-emitting diode-induced fluorescence detection (ICFO-LED-IFD). Fluorescein isothiocyanate was used as the fluorescence tagged reagent for derivatization of DA, EP and NE. The CE conditions for separation of these catecholamines

Peiyu Diao; Hongyan Yuan; Feng Huo; Lifen Chen; Dan Xiao; Man Chin Paau; Martin M. F. Choi


A simple, rapid method to isolate salt glands for three-dimensional visualization, fluorescence imaging and cytological studies  

PubMed Central

Background Some plants inhabiting saline environment remove salts via the salt glands embedded in the epidermal tissues. Cytological studies of salt glands will provide valuable information to our understanding of the secretory process. Previous studies on salt gland histology relied mainly on two-dimensional microscopic observations of microtome sections. Optical sectioning properties of confocal laser scanning microscope offer alternative approach for obtaining three-dimensional structural information of salt glands. Difficulty in light penetration through intact leaves and interference from neighbouring leaf cells, however, impede the acquiring of good optical salt gland sections and limit its applications in salt gland imaging. Freeing the glands from adjacent leaf tissues will allow better manipulations for three-dimensional imaging through confocal laser scanning microscopy. Results Here, we present a simple and fast method for the isolation of individual salt glands released from the interference of neighbouring cells. About 100-200 salt glands could be isolated from just one cm2 of Avicennia officinalis leaf within hours and microscopic visualization of isolated salt glands was made possible within a day. Using these isolated glands, confocal laser scanning microscopic techniques could be applied and better resolution salt gland images could be achieved. By making use of their intrinsic fluorescent properties, optical sections of the gland cells could be acquired without the use of fluorescent probes and the corresponding three-dimensional images constructed. Useful cytological information of the salt gland cells could also be obtained through the applications of fluorescent dyes (e.g., LysoTracker® Red, FM®4-64, Texas Red®). Conclusions The study of salt glands directly at the glandular level are made possible with the successful isolation of these specialized structures. Preparation of materials for subsequent microscopic observations of salt glands could be achieved within a day. Potential applications of confocal fluorescence microscopic techniques could also be performed using these isolated glands. Experiments designed and targeted directly at the salt glands were explored and cytological information obtained herein could be further incorporated towards the understanding of the mechanism underlying secretion in plant salt glands.



A simple and sensitive fluorescence quenching method for the determination of H(2)O(2) using Rhodamine B and FE(3)O(4) nanocatalyst.  


In pH 1.99 sodium acetate-HCl buffer solutions at 60 °C, Rhodamine B exhibited a strong fluorescence peak at 584 nm using an excitation wavelength of 548 nm. The fluorescence quenching occurred when Fe(3)O(4) nanoparticles catalyzed H(2)O(2) oxidation of Rhodamine B. Under the chosen conditions, the fluorescence intensity at 584 nm decreased when the concentration of H(2)O(2) increased. The fluorescence quenching intensity is linear with the concentration of H(2)O(2) in the range of 10-200 nmol/L. Thus, a new and simple and sensitive nanocatalytic fluorescence method was proposed for the determination of H(2)O(2) in synthetic sample, with satisfactory results. PMID:21617998

Jiang, Zhiliang; Kun, Li; Ouyang, Huixiang; Liang, Aihui; Jiang, Hesheng



Immunosensor systems with the Langmuir-film-based fluorescence detection  

SciTech Connect

A method is developed for detecting protein antigens for fluorescent immunoassay using a model system based on the technique for preparation of Langmuir films. Fluorescein isothiocyanate and donor-acceptor energy-transfer pairs of markers (the Yb complex of tetraphenyl porphyrin - benzoyl trifluoroacetoneisothiocyanate and derivatives of tetra(carboxyphenyl) porphyrin - cyanine dye containing a five-membered polyene chain), which were nor studied earlier, were used as markers for detecting the binding of an antigen on the surface of Langmuir films of antibodies. Fluorescence was detected in the near-IR region (for the first pair) and in the visible spectral range (for the second pair). To reduce the nonspecific sorption of a protein (antigen), a method was proposed for the preparation of a nonpolar surface by applying an even number of layers of stearic acid as a substrate for the Langmuir - Blodgett film. A high sensitivity of model systems to a protein antigen in solution was achieved ({approx}10{sup -11} M), the assay time being 6 - 8 min. The model system with the first donor - acceptor pair was tested in analysis of the blood plasma. The fluorescence of the Dy{sup 3+}, Tm{sup 3+}, and Yb{sup 3+} complexes of tetraphenyl porphyrin sensitised by diketonate complexes of lanthanides was studied for the first time and the enhancement of the IR fluorescence of these complexes in a Langmuir film was demonstrated. (papers devoted to the memory of academician a m prokhorov)

Chudinova, G K; Nagovitsyn, I A; Savranskii, V V [Natural Science Center, A.M. Prokhorov General Physics Institute, Russian Academy of Sciences, Moscow (Russian Federation); Karpov, R E [Photochemistry Center, Russian Academy of Sciences, Moscow (Russian Federation)



Electrospun sol-gel fibers for fluorescence-based sensing  

NASA Astrophysics Data System (ADS)

Fluorescence based biosensors have the ability to provide reliable pathogen detection. However, the performance could be improved by enhancing the effective surface area of the biosensor. We report on a new nanofibrous fluorescencebased biosensor, whereas a sol-gel platform mesh was constructed by utilizing electrospinning techniques. Furthermore, incorporating cetyltrimethylammonium bromide (CTAB) and conducting pore-forming techniques resulted in a high surface area material suitable for biosensor immobilization. The biosensor was designed to detect Helicobacter hepaticus bacterium by sandwiching the pathogen between two antibodies, one labeled with Alexa Fluor 546 fluorescent dye and the other with 20nm Au nanoparticles. In the presence of pathogen, the close proximity of Au nanoparticles quenched the Alexa Fluor fluorescence, suggesting that the electrospun fiber platforms are suitable for sensing H. Hepaticus. Additionally, sol-gel fibers used as biosensor platform have the added benefit of increased immobilization, as fluorescence intensity from immobilized biosensors is 8.5x106 cps higher on fibers than on a flat, non-porous substrate.

Memisevic, Jasenka; Riley, Lela; Grant, Sheila A.



Neurotransmitter imaging in living cells based on native fluorescence detection  

SciTech Connect

A UV laser-based optical microscope and CCD detection system with high sensitivity has been developed to image neurotransmitters in living cells. We demonstrate the detection of serotonin that has been taken up into individual living glial cells (astrocytes) based on its native fluorescence. We found that the fluorescence intensity of astrocytes increased by up to 10 times after serotonin uptake. The temporal resolution of this detection system at 10{sup -4} M serotonin is as fast as 50 ms, and the spatial resolution is diffraction limited. This UV laser microscope imaging system shows promise for studies of spatial-temporal dynamics of neurotransmitter levels in living neurons and glia. 19 refs., 5 figs., 1 tab.

Tan, W.; Yeung, E.S. [Ames Lab., IA (United States)]|[Iowa State Univ., Ames, IA (United States); Parpura, V.; Haydon, P.G. [Iowa State Univ., Ames, IA (United States)



Fluorescence-based biosensing of zinc using carbonic anhydrase  

Microsoft Academic Search

Measurement of free zinc levels and imaging of zinc fluxes remains technically difficult due to low levels and the presence of interfering cations such as Mg and Ca. We have developed a series of fluorescent zinc indicators based on the superb sensitivity and selectivity of a protein, human apo-carbonic anhydrase II, for Zn(II). These indicators transduce the level of free zinc

Carol A. Fierke; Richard B. Thompson



Development of fluorescence-based LIDAR technology for biological sensing  

Microsoft Academic Search

Results of our on-going development of biological warfare agents (BWA) detection systems based on spectral detection of ultraviolet (UV) laser induced fluorescence (LIF) are presented. A compact optical parametric oscillator (OPO) with intracavity sum-frequency mixing (SFM) to generate 293 nm UV laser irradiation was developed. The OPO\\/SFM device was pumped by a diode-pumped Nd:YAG laser (1064 nm), including subsequent second-harmonic

Per Jonsson; Fredrik Kullander; Mikael Tiihonen; Melker Nordstrand; Gøran Olofsson; Mikael Lindgren


High efficient solar tracker based on a simple shutter structure  

NASA Astrophysics Data System (ADS)

In many photovoltaic (PV) or sunlight-illumination systems, solar trackers are always essential to obtain high energy/flux concentration efficiency, and that would lead to increase cost and extra power consumption due to the complex structure and heavy weight of the trackers. To decrease the cost while without sacrificing efficiency, a Fresnellens concentrator incorporated with a simple and cheap shutter, which consists of high reflective mirrors instead of conventional trackers, is proposed in this paper to provide solar tracking during the daytime. Thus, the time-variant and slant-incident sunlight rays can be redirected to vertically incident upon the surface of the Fresnel lens by appropriately arranging mirrors and swinging them to the proper slant angles with respect to the orientation of sunlight. The computer simulation results show that power concentration efficiency over 90%, as compared with the efficiency of directly normal incident sunlight, can be achieved with the mirror reflectance of 0.97 and for any solar incident angle within +/-75 degrees to the normal of the Fresnel lens. To verify the feasibility and performance of the concentrator with the proposed shutter, a sunlight illumination system based on this novel structure is demonstrated. Both computer simulation and practical measurement results for the prototype of the sunlight illumination system are also given to compare with. The results prove the simple and high efficient shutter applicable to general PV or sunlight-illumination systems for solar tracking.

Chen, Jin-Jia; Liu, Te-Shu; Huang, Kuang-Lung; Lin, Po-Chih



A schiff-based colorimetric fluorescent sensor with the potential for detection of fluoride ions.  


A simple Schiff-based colorimetric fluorescent receptor 1 was prepared. It exhibits a "turn-on-type" mode with high sensitivity in the presence of F(-). The change in color is very easily observed by the naked eye in the presence of F(-), whereas other anions do not induce such a change. Job plot indicated a 1:2 complexation stoichiometry between receptor 1 and F(-). The association constant for 1-F(-) in CH3CN was determined as 1.32*10(5) M(-2) by a Hill plot. PMID:23888325

Huang, Cheng-Yin; Wan, Chin-Feng; Chir, Jiun-Ly; Wu, An-Tai



Nanoparticle-based energy transfer for rapid and simple detection of protein glycosylation  

SciTech Connect

Glycan moiety of glycoproteins plays an essential role in its biological activity in vivo, and the analysis of glycosylation is of great importance in the development of protein therapeutics. In this study, we report a rapid and simple detection of protein glycosylation based on the fluorescence resonance energy transfer (FRET) between concanavalin A-conjugated gold nanoparticles (ConA-AuNPs) and dextran-conjugated quantum dots (Dex-QDs). The increased photoluminescence (PL) signals of Dex-QDs due to the competitive inhibition of glycoproteins were well correlated with the glycosylation chain length of glucose oxidases as well as the mannosylation degree of bovine serum albumin (BSA). The parallel analysis of the diversely mannosylated BSAs using an image analyzer further demonstrated the potential of this new technique in high-throughput screening of glycoprotein and carbohydrate therapeutics.

Oh, Eunkeu; Lee, Dohoon; Kim, Young-Pil; Cha, Seung YOUP; Oh, Doo BEYONG; Kim, Jungbae; Kang, Hyun AH; Kim, Hak SUNG



A label-free, fluorescence based assay for microarray  

NASA Astrophysics Data System (ADS)

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 intensity of excitation light. The fluorescence contrast is used to quantify the amount of probe-target hybridization. A mathematical model that considers multiple reflections and scattering is developed to explain the mechanism of the fluorescence contrast which depends on the thickness of the PS film. Scattering is the dominant factor that contributes to the contrast. The potential of this assay to detect single nucleotide polymorphism is also tested.

Niu, Sanjun


Wide and scalable field-of-view Talbot-grid-based fluorescence microscopy  

PubMed Central

Here we report a low-cost and simple wide field-of-view (FOV) on-chip fluorescence-imaging platform, termed fluorescence Talbot microscopy (FTM), which utilizes the Talbot self-imaging effect to enable efficient fluorescence imaging over a large and directly scalable FOV. The FTM prototype has a resolution of 1.2 ?m and an FOV of 3.9 mm × 3.5 mm. We demonstrate the imaging capability of FTM on fluorescently labeled breast cancer cells (SK-BR-3) and human embryonic kidney 293 (HEK) cells expressing green fluorescent protein.

Pang, Shuo; Han, Chao; Kato, Mihoko; Sternberg, Paul W.; Yang, Changhuei



Wide and scalable field-of-view Talbot-grid-based fluorescence microscopy.  


Here we report a low-cost and simple wide field-of-view (FOV) on-chip fluorescence-imaging platform, termed fluorescence Talbot microscopy (FTM), which utilizes the Talbot self-imaging effect to enable efficient fluorescence imaging over a large and directly scalable FOV. The FTM prototype has a resolution of 1.2 ?m and an FOV of 3.9 mm × 3.5 mm. We demonstrate the imaging capability of FTM on fluorescently labeled breast cancer cells (SK-BR-3) and human embryonic kidney 293 (HEK) cells expressing green fluorescent protein. PMID:23202123

Pang, Shuo; Han, Chao; Kato, Mihoko; Sternberg, Paul W; Yang, Changhuei



Note: A portable, light-emitting diode-based ruby fluorescence spectrometer for high-pressure calibration  

NASA Astrophysics Data System (ADS)

Ruby (Al2O3, with ~0.5 wt. % Cr doping) is one of the most widely used manometers at the giga-Pascal scale. Traditionally, its fluorescence is excited with intense laser sources. Here, I present a simple, robust, and portable design that employs light-emitting diodes (LEDs) instead. This LED-based system is safer in comparison with laser-based ones.

Feng, Yejun



Note: A portable, light-emitting diode-based ruby fluorescence spectrometer for high-pressure calibration  

SciTech Connect

Ruby (Al{sub 2}O{sub 3}, with {approx}0.5 wt. % Cr doping) is one of the most widely used manometers at the giga-Pascal scale. Traditionally, its fluorescence is excited with intense laser sources. Here, I present a simple, robust, and portable design that employs light-emitting diodes (LEDs) instead. This LED-based system is safer in comparison with laser-based ones.

Feng Yejun [Advanced Photon Source, Argonne National Laboratory, Argonne, Illinois 60439 (United States)



Study plasma disintegration based on the fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

It is known that plasma is very important in the diagnosis and therapy of disease so that more and more scientific workers attach importance to the plasma storage life or storage environment. We research plasma disintegration with the increases of storage time based on the fluorescence spectroscopy. Based on the experimental researches and theoretical analysis, we find that their plasma fluorescence intensity is increasing within about 10 hours, and but is decreasing gradually and is nearly a straight line after this. It is indicated that plasma proteins have begun to disintegrate so as to make the fluorescence quenching after storage time beyond 10 hours. And the disintegration speed of plasma in the case of different concentration is different, the concentration is higher and the speed is lower in them, and but they are almost same after about 35 hours. Therefore, we think that plasma under higher concentration is deposited easier. These research consequences may order a theoretical and experimental reference to know the changes of plasma in structure in different disintegration time. It may make sense for understand the plasma disintegrative mechanism and distinguishing the fine plasma with faulty.

Gao, Shumei; Li, Rongqing; Chen, Guoqing; Lu, Jun



X-ray fluorescence analysis based on Kumakhov optics  

NASA Astrophysics Data System (ADS)

The use of Kumakhov optics in x-ray fluorescence analysis is considered. Thanks to high efficiency of a polycapillary lens the concentration of x-ray detector in a close proximity to the sample decreases sufficiently the time of exposure. It is shown experimentally that in the case of use of a small x-ray source with power of 2W the minimal detection limit may be of the order of 0.1 pg. A new portable x-ray fluorescence analyzer based on Kumakhov optics is described. Minimal detection limit may achieve 10(superscript -16) g if 100 W microfocus source and the lens with approximately 10 micrometers focal spot are used.

Nikitina, Svetlana V.; Ibraimov, Nariman S.; Stcherbakov, Alexander S.



Reaction-based genetically encoded fluorescent hydrogen sulfide sensors.  


The detection of hydrogen sulfide (H(2)S), a toxic gas and an important biological signaling molecule, has been a long-time challenge. Here we report genetically encoded fluorescent protein (FP)-based probes that can selectively detect H(2)S. By expanding the genetic codes of E. coli and mammalian cells, FP chromophores were modified with the sulfide-reactive azide functional group. These structurally modified chromophores were selectively reduced by H(2)S, resulting in sensitive fluorescence enhancement detectable by spectroscopic and microscopic techniques. Exploration of a circularly permuted FP led to an improved sensor with faster responses, and the feasibility of using such a genetically encoded probe to monitor H(2)S in living mammalian cells has also been demonstrated. PMID:22642566

Chen, Si; Chen, Zhi-jie; Ren, Wei; Ai, Hui-wang



A simple, low-cost, remote fiber-optic micro volume fluorescence flowcell for capillary flow-injection analysis  

Microsoft Academic Search

A small volume flowcell for fluorescence detection in capillary flow injection (CFI) analysis has been created by using a low cost, commercially available fluidic device. Fluorescence detection is achieved using an optical fiber to deliver excitation light to the sample flowing through the device and another optical fiber to collect fluorescence emission. The flowcell is a standard fluidic cross with

Sean J. Hart; Renee D. Jiji



A simple method for high-throughput quantification of genome-wide DNA methylation by fluorescence polarization.  


To rapidly determine DNA methylation levels from a large number of biological or clinical samples, we have developed an accurate and sensitive method for high-throughput quantification of global methylation of 5'-Cm ( 5) CGG-3' sites in the genome, visualized by fluorescence polarization (FP) based measurement of DNA methylation (FPDM). In FPDM, the methyl-sensitive HpaII and methyl-insensitive MspI restriction enzymes were employed to achieve DNA cleavage, followed by incorporation of fluorescent dCMP into the enzyme-cleavage products through polymerase chain extension, yielding an FP-ratio between the HpaII- and MspI-restricted preparations as a measure of methylation. FPDM provided stable estimates of methylation level of submicrograms of lambda or human DNA, and of a 255-bp DNA segment containing a single HpaII/MspI restriction site in accord with, and more accurate than, determination by gel electrophoresis. FPDM was also applied to measure dose-dependent DNA hypomethylation in human embryonic kidney 293T cells treated with the DNA-methyltransferase inhibitor 5-aza-dC. PMID:22419070

Zhao, Cunyou; Xue, Hong



Fluorescence resonance energy transfer from sulfonated graphene to riboflavin: a simple way to detect vitamin b2.  


We have prepared sulfonated graphene (SG) by diazonium coupling technique and it has been characterized by UV-vis absorption spectroscopy, Raman spectroscopy, electron microscopy, energy-dispersive spectroscopy (EDS), EDS elemental mapping, X-ray photoelectron spectroscopy (XPS), and FTIR spectroscopy. The photoluminescence (PL) property of SG at different pH (pH 4, 7, and 9.2) has been investigated and SG shows highest PL-intensity and quantum yield at pH 4 compared to those at higher pH and that of GO at pH 4. Due to the strong overlap between the emission spectrum of SG and absorption spectrum of riboflavin (RF, vitamin B2) at pH 4, it has been tactfully used as donor for the fluorescence resonance energy transfer (FRET) process. However, graphene oxide (GO) does not exhibit any FRET with RF at an identical condition due to its much lower quantum yield. We have demonstrated a selective detection of vitamin B2 in presence of nucleic acid (DNA, RNA), protein (BSA), amino acid (Lysine) and other water-soluble vitamins (Becosules, Zevit capsules) based on the spontaneous FRET from PL-active SG (donor) to RF (acceptor). The calibration curve indicates excellent affirmation to detect vitamin B2 using FRET and it is superior to the ordinary fluorescence method of detecting RF in presence of different biomolecules. PMID:23838272

Kundu, Aniruddha; Nandi, Sudipta; Layek, Rama K; Nandi, Arun K



Simple and sensitive high-performance liquid chromatography-fluorescence method for the determination of citrinin application to the analysis of fungal cultures and cheese extracts  

Microsoft Academic Search

A new and highly sensitive method for the detection of the important mycotoxin, citrinin, has been developed. Spectroscopic studies demonstrate that the fluorescence of this metabolite is influenced by the pH of the environment. This fact was exploited in the chromatographic determination of citrinin with fluorescence detection. The proposed method, based on the addition of 1 M hydrochloric acid as

C. M. Franco; C. A. Fente; B. Vazquez; A. Cepeda; L. Lallaoui; P. Prognon; G. Mahuzier



Frog melanophores cultured on fluorescent microbeads: biomimic-based biosensing.  


Melanophores are pigmented cells in lower vertebrates capable of quick color changes and thereby suitable as whole cell biosensors. In the frog dermis skin layer, the large and dark pigmented melanophore surrounds a core of other pigmented cells. Upon hormonal stimulation the black-brown pigment organelles will redistribute within the melanophore, and thereby cover or uncover the core, making complex color changes possible in the dermis. Previously, melanophores have only been cultured on flat surfaces. Here we mimic the three dimensional biological geometry in the frog dermis by culturing melanophores on fluorescent plastic microbeads. To demonstrate biosensing we use the hormones melatonin and alpha-melanocyte stimulating hormone (alpha-MSH) as lightening or darkening stimuli, respectively. Cellular responses were successfully demonstrated on single cell level by fluorescence microscopy, and in cell suspension by a fluorescence microplate reader and a previously demonstrated computer screen photo-assisted technique. The demonstrated principle is the first step towards "single well/multiple read-out" biosensor arrays based on suspensions of different selective-responding melanophores, each cultured on microbeads with distinctive spectral characteristics. By applying small amount of a clinical sample, or a candidate substance in early drug screening, to a single well containing combinations of melanophores on beads, multiple parameter read-outs will be possible. PMID:15967358

Andersson, Tony P M; Filippini, Daniel; Suska, Anke; Johansson, Therese L; Svensson, Samuel P S; Lundström, Ingemar



Novel fiber optic glucose biosensor based on fluorescence quenching  

NASA Astrophysics Data System (ADS)

A new fiber optical glucose biosensor based on oxygen fluorescence quenching using lock-in technology has been presented. Ruthenium complex, Ru(bpy)3Cl2 were used as the fluorescence indicator and cellulose acetate (CA) were used as the matrix membrane to immobilize the indicator and GOD, and the optimal conditions for the preparation of CA membrane has been studied. For the immobilization of GOD to CA membrane, albumin of bovine serum (BSA) and glutaraldehyde (GA) were used through covalence - cross bonding process. The relationship between the concentration of glucose and the phase delay (Phi) has been studied, the results show that (Phi) has good relationship within the range of 50mg/dl 500mg/dl and the response time is less than 30 sec. Several factors to influence the sensor such as pH of the medium, the additional membrane layers, the properties of CA membrane have also been studied. The results indicate that its best pH range is between pH 6.0 and 7.0, and the thickness of the additional membrane layers on the sensor head will influence the response time greatly, that is, the response time is controlled by the speed of dispersion of the dissolved oxygen into the fluorescence dye layer.

Jiang, Desheng; Liu, Er; Huang, Jun



Immunospot assay based on fluorescent nanoparticles for Dengue fever detection.  


Dengue fever is one of the most neglected tropical diseases and of highest international public health importance, with 50 million cases worldwide every year. Early detection can decrease mortality rates from more than 20% to less than 1% and the relevant early diagnosis analyte is the viral non-structural glycoprotein, NS1. Currently, enzyme linked immunosorbent assay (ELISA) is the method of choice to detect NS1. However, this is a time consuming method, requiring 3-5h, and it is the bottleneck for routine of clinical analysis laboratory in epidemic periods, when hundreds of samples should be tested. Here we describe an easy method combining principles of fluorophore linked immunosorbent assay (FLISA) and enzyme linked immunospotting (ELISPOT). For detection, we used mouse anti-NS1 IgG labeled with fluorescent nanoparticles. The presented procedure needs only 4 ?L of serum samples and requires 45-60 min. The detection limit, 5.2 ng/mL, is comparable to ELISA tests. The comparison of 83 samples with a commercial ELISA revealed a sensitivity of 81% and specificity of 88%. The use of fluorescent nanoparticles provides a higher sensitivity than an assay using usual fluorescent dye molecules, besides avoiding bleaching effects. Based on the results, the proposed method provides fast, specific and sensitive results, and proves to be a suitable method for Dengue NS1 detection in impoverished regions or epidemic areas. PMID:22981010

Linares, Elisângela M; Pannuti, Claudio S; Kubota, Lauro T; Thalhammer, Stefan



Data storage based on photochromic and photoconvertible fluorescent proteins.  


The recent discovery of photoconvertible and photoswitchable fluorescent proteins (PCFPs and RSFPs, respectively) that can undergo photoinduced changes of their absorption/emission spectra opened new research possibilities in subdiffraction microscopy and optical data storage. Here we demonstrate the proof-of-principle for read only and rewritable data storage both in 2D and 3D, using PCFPs and RSFPs. The irreversible burning of information was achieved by photoconverting from green to red defined areas in a layer of the PCFP Kaede. Data were also written and erased several times in layers of the photochromic fluorescent protein Dronpa. Using IrisFP, which combines the properties of PCFPs and RSFPs, we performed the first encoding of data in four colours using only one type of fluorescent protein. Finally, three-dimensional optical data storage was demonstrated using three mutants of EosFP (d1EosFP, mEosFP and IrisFP) in their crystalline form. Two-photon excitation allowed the precise addressing of regions of interest (ROIs) within the three-dimensional crystalline matrix without excitation of out-of-focus optical planes. Hence, this contribution highlights several data storage schemes based on the remarkable properties of PCFPs/RSFPs. PMID:20416344

Adam, Virgile; Mizuno, Hideaki; Grichine, Alexei; Hotta, Jun-ichi; Yamagata, Yutaka; Moeyaert, Benjamien; Nienhaus, G Ulrich; Miyawaki, Atsushi; Bourgeois, Dominique; Hofkens, Johan



Flow imaging of dilute colloidal suspension in PDMS-based microfluidic chip using fluorescence microscopy  

Microsoft Academic Search

A slit-like flow channel was designed to allow for fluorescent microscope visualization in the microfluidic chip fabricated with glass substrate and polydimethylsiloxane (PDMS). For the exposure time, moving fluorescent polystyrene latexes of 1?m radius result in image streaks, where the latex concentration is sufficiently dilute underlying the condition of simple fluid. Applying the data processing method for particle streak imaging

Myung-Suk Chun; Sangwoo Lee



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


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

Mayer-Cumblidge, M. Uljana; Cao, Haishi



A simple enzyme based biosensor on flexible plastic substrate  

NASA Astrophysics Data System (ADS)

An enzyme based biosensor was fabricated by employing a simple, inexpensive and rapid xurography fabrication process. The electrodes and channel were made from the conducting polymer poly(3,4-ethyelenedioxythiphene) poly(styrene sulfonate) (PEDOT:PSS). PEDOT:PSS was selectively deposited using a polyimide tape mask. The tape mask was peeled off from the substrate after annealing the polymer in vacuum. Polymer wells of defined dimensions were made and were attached to the device to accommodate the solutions. This sensor utilizes the change in current as a parameter to measure different analyte concentrations. Initial experiments were done by using the sensor for glucose detection. The sensor is able to detect the glucose concentrations approximately from 1 ?M to 10 mM range covering glucose in human saliva (8-210 ?M). The glucose oxidase activity was independently measured using colorimetric method and the results indicate that the sensor retains the enzyme activity and can be used as a biosensor to detect various analytes. The analyte of interest can be measured by preloading the corresponding enzyme into the wells.

Kanakamedala, Senaka K.; Alshakhouri, Haidar T.; Agarwal, Mangilal; Fang, Ji; Decoster, Mark A.



“Off-On” based fluorescent chemosensor for Cu 2+ in aqueous media and living cells  

Microsoft Academic Search

A novel Cu2+-specific “off-on” fluorescent chemosensor of naphthalimide modified rhodamine B (naphthalimide modified rhodamine B chemosensor, NRC) was designed and synthesized, based on the equilibrium between the spirolactam (non-fluorescence) and the ring-opened amide (fluorescence). The chemosensor NRC showed high Cu2+-selective fluorescence enhancement over commonly coexistent metal ions or anions in neutral aqueous media. The limit of detection (LOD) based on

Chunwei Yu; Lingxin Chen; Jun Zhang; Jinhua Li; Ping Liu; Wenhai Wang; Bing Yan


Sensitive and selective detection of nitrite ion based on fluorescence superquenching of conjugated polyelectrolyte.  


In recent years, conjugated polyelectrolytes (CPEs) that feature good water-solubility have drawn great attention as optical transducers in high sensitive bio- and chemo-sensors due to their predominant optical/electronic properties and remarkable signal amplification. Herein, a sensitive and selective assay for nitrite ion has successfully been developed based on the fluorescence superquenching of an anionic CPE, PPESO(3). With the sensor format composed of PPESO(3) and H(+), Fe(2+) can easily be oxidized into Fe(3+) in the presence of NO(2)(-), and the later dramatically quenches the fluorescence of PPESO(3). Indeed, the inclusion of conjugated polyelectrolyte into the sensory scheme can give rise to a notable enhancement of fluorescence response, which endows the newly proposed NO(2)(-) probe with high sensitivity. Thus, nitrite ion within a relatively wide concentration range (0-70 microM) can be determined in a rather simple and sensitive manner with a detection limit of 0.62 microM (approximately 28 ppb). Additionally, most other anions such as halogen ions, acetate, sulfate, carbonate, phosphate and even nitrate, show minor interference on the NO(2)(-) detection. PMID:20188893

Zhang, Tao; Fan, Hongliang; Jin, Qinhan



Folding Coupled with Assembly in Split Green Fluorescent Proteins Studied by Structure-based Molecular Simulations.  


Split green fluorescent protein (GFP) is a powerful tool for imaging of protein-protein interactions in living cells, but molecular mechanisms of the folding and the assembly of split GFPs are poorly understood. Here, using a simple Go model that is based on the energy landscape theory, we performed comprehensive folding simulations of six split GFPs with different split points. Of the six, the fluorescence recovery was reported in four but not in the other two. In the simulations, we found that when the complete folding and assembly were observed, the N-terminal fragment always folded earlier than the C-terminal fragment. The in silico folding rates of the split GFPs were larger for the four split GFPs that the fluorescence recovery was reported in literature. The stability of standalone N-terminal fragments were well-correlated with the folding rates of split GFPs. These suggest that the efficient folding and assembly of split GFPs are realized when the N-terminal fragment folds spontaneously with the central ?-helix as a nucleation core and that the C-terminal fragment folding is coupled to the assembly to the preformed N-terminal fragment. PMID:23679014

Ito, Mashiho; Ozawa, Takeaki; Takada, Shoji



"Fitting" makes "sensing" simple: label-free detection strategies based on nucleic acid aptamers.  


Nucleic acid aptamers are small sequences of DNA made via in vitro selection techniques to bind targets with high affinity and specificity. The term aptamer derives from the Latin, aptus, meaning "to fit", emphasizing the lock-and-key relationship between aptamers and their binding targets. In 2004, aptamers began to attract researchers' attention as new binding elements for biosensors (i.e. aptasensors). Their advantages over other sensors include a diverse range of possible target molecules, high target affinity, simple synthesis, and ability to form Watson-Crick base pairs. These attributes create an enormous array of possible sensing applications and target molecules, spanning nearly all detection methods and readout techniques. In particular, aptamers provide an opportunity for designing "label-free" sensors, meaning sensors that do not require covalently labeling a signal probe to either the analyte or the recognition element (here, the aptamer). "Label-free" systems previously could only analyze large molecules using a few readout techniques, such as when employing the other recognition elements like antibodies. "Label-free" methods are one of the most effective and promising strategies for faster, simpler, and more convenient detection, since they avoid the expensive and tedious labeling process and challenging labeling reactions, while retaining the highest degree of activity and affinity for the recognition element. "Label-free" sensors are one of the most promising future biosensors. In this Account, we describe our efforts exploring and constructing such label-free sensing strategies based on aptamers. Our methods have included using various readout techniques, employing novel nanomaterials, importing lab-on-a-chip platforms, and improving logical recognition. The resulting sensors demonstrate that aptamers are ideal tools for "label-free" sensors. We divide this Account into three main parts describing three strategies for designing "label-free" sensors: (1) Label-free, separation-free strategies. These include colorimetric sensors based on G-quadruplex-hemin complex, and fluorescent sensors based on fluorescent small molecules, novel conjugated polymers, and metal ion clusters. (2) Label-free, separation-required strategies. In this part, electrochemical sensors are introduced, including sensors with different subtechniques using an electrode array. (3) Logic sensors. Some logic recognition systems are introduced. We emphasize that label-free aptasensors are not merely simple. We hope our introduction illustrates the powerful, flexible, and smart functions of aptamers in carrying out various detection tasks or playing various recognition games. Our work is only a start. We believe this field will bring additional knowledge on general designs, anti-interference, multianalysis, minimization, and auto-operation of aptamer biosensors. PMID:23214491

Du, Yan; Li, Bingling; Wang, Erkang



Miniature fiber optic sensor based on fluorescence energy transfer  

NASA Astrophysics Data System (ADS)

Optical fiber biosensors based on fluorescence assays have several distinct advantages when measuring biological analytes such as metabolites, cofactors, toxins, etc. Not only are optical signals immune to electronic interferences, but the polychromatic nature of most fluorochemical assays provides more potentially useful data about the system being studied. One of the most common difficulties normally encountered with optical biosensors is the inability to routinely recalibrate the optical and electronic components of the system throughout the life of the sensor. With this in mind, we present an optical fiber assay system for glucose based on a homogeneous singlet/singlet energy transfer assay along with the electronic instrumentation built to support the sensor system. In the sensor probe, glucose concentrations are indirectly measured from the level of fluorescence quenching caused by the homogeneous competition assay between TRITC labeled concanavalin A (receptor) and FITC labeled Dextran (ligand). The FITC signal is used to indicate glucose concentrations and the TRITC signal is used for internal calibration. Data is also presented on a protein derivatization procedure that was used to prevent aggregation of the receptor protein in solution. Also, a molecular model is described for the singlet/singlet energy transfer interactions that can occur in a model system composed of a monovalent ligand (FITC labeled papain) and a monovalent receptor (TRITC labeled concanavalin A).

Meadows, David L.; Schultz, Jerome S.



BODIPY-conjugated thermoresponsive copolymer as a fluorescent thermometer based on polymer microviscosity.  


A simple copolymer, poly(NIPAM-co-BODIPY), consisting of N-isopropylacrylamide (NIPAM) and boradiazaindacene (BODIPY) units, behaves as a fluorescent thermometer in water. The copolymer exhibits weak fluorescence at <23 degrees C, but the intensity increases with a rise in temperature up to 35 degrees C, enabling an accurate indication of the solution temperature at 23-35 degrees C. The heat-induced fluorescence enhancement is driven by an increase in the polymer microviscosity, associated with a phase transition of the polymer from the coil to globule state. The viscous domain formed inside the globule-state polymer suppresses the rotation of the meso-pyridinium group of the excited-state BODIPY units, resulting in heat-induced fluorescence enhancement. The polymer shows reversible fluorescence enhancement/quenching regardless of the heating/cooling process and displays high reusability with a simple recovery process. PMID:19821567

Wang, Dongping; Miyamoto, Ryo; Shiraishi, Yasuhiro; Hirai, Takayuki



Monitoring water supplies for weaponized bacteria and bacterial toxins using rapid fluorescence-based viability and affinity assays  

NASA Astrophysics Data System (ADS)

The rapid detection of weaponized bacteria and toxins is a major problem during a biological attack. Although sensitive detection formats exist for many biowarfare agents, they often require advanced training and complex procedures. Luna has developed simple, rapid means for determining the presence of pathogens and bacterial toxins in water supplies using fluorescence-based assays that can be adapted for field use. The batteries of rapid assays are designed for i) determining cell viability and bacterial loads by exploiting metabolic markers (e.g., acid-production, redox potentials, etc) and ii) detecting bacterial toxins using fluorescent, polymerized affinity liposomes (fluorosomes). The viability assays were characterized using E. coli, S. aureus and the anthrax simulant, B. globigii. The viability assays detected bacterial loads of ~ 104 CFU/ml and with simple filtration ~ 100CFU/ml could be detected. The affinity fluorosomes were characterized using cholera toxin (CT). Affinity liposomes displaying GM1 and anti-CT antibodies could detect CT at fluorescence system, Luna characterized the binding of affinity fluorosomes to respective targets and determined the responses of bacterial loads in the fluorescent viability assays. Using this two-tiered approach, Luna demonstrated that water susceptible to sabotage could be easily monitored and confirmed for specific agents using simple, general and specific fluorescence-based detection schemes based on metabolism and ligand-target interactions.

Van Tassell, Roger L.; Evans, Mishell


Poly(N-isopropylacrylamide)-Based Thermoresponsive Behavior of Fluorescent Organic Nanocrystals  

NASA Astrophysics Data System (ADS)

In this article, we describe the poly(N-isopropylacrylamide)-based thermoresponsive behaviors of fluorescent organic nanocrystals. Aqueous dispersions of thermoresponsive fluorescent nanocrystals were prepared by the reprecipitation method. Fluorescent dyes used for these nanocrystals were perylene, quinacridone, and zinc phthalocyanine. The perylene nanocrystals in the aqueous system using poly(N-isopropylacrylamide) exhibited a significantly increased fluorescence intensity assigned to the emission of the perylene monomer above the cloud point (˜35 °C). Significantly increased fluorescence intensities of quinacridone nanocrystals and zinc phthalocyanine nanocrystals were also observed above the cloud point (˜35 °C). These unique fluorescence behaviors were only observed in the aqueous system using poly(N-isopropylacrylamide).

Baba, Koichi; Kasai, Hitoshi; Nishida, Kohji; Nakanishi, Hachiro



Fluorescent indicators for Ca2+based on green fluorescent proteins and calmodulin  

Microsoft Academic Search

Important Ca2+ signals in the cytosol and organelles are often extremely localized and hard to measure. To overcome this problem we have constructed new fluorescent indicators for Ca2+ that are genetically encoded without cofactors and are targetable to specific intracellular locations. We have dubbed these fluorescent indicators `cameleons'. They consist of tandem fusions of a blue- or cyan-emitting mutant of

Atsushi Miyawaki; Juan Llopis; Roger Heim; J. Michael McCaffery; Joseph A. Adams; Mitsuhiko Ikura; Roger Y. Tsien



Facile fluorescence-based detection of PAD4-mediated citrullination.  


The post-translational modifications of histone proteins are highly diverse and dynamic processes. It is becoming increasingly evident that modifying histone proteins can have a direct influence on both cellular homeostasis and disease states. Protein arginine deiminase 4 (PAD4) is an enzyme that converts peptidyl-arginine to citrulline. The overexpression of PAD4 has been found in numerous types of human cancer and autoimmune diseases. We report a new, facile, fluorescence-based assay for the detection of PAD4 activity that exploits the substrate specificity of trypsin to monitor the citrullination reaction carried out by PAD4 based on the fact that, upon citrullination, the positively charged arginine side chain is converted to the neutral citrulline. We show that the assay can be performed rapidly with readily available reagents and that it responds accordingly to a known PAD4 inhibitor. PMID:23640867

Wildeman, Erin; Pires, Marcos M



Beer's-Law-Based, Simple Spectral Model for Direct Normal and Diffuse Horizontal Irradiance.  

National Technical Information Service (NTIS)

A spectral model for cloudless days that uses simple mathematical expressions and tabulated look-up tables to generate direct normal and diffuse horizontal irradiance is presented. The model is based on modifications to previously published simple models ...

R. E. Bird



A fluorescence lifetime-based assay for serine and threonine kinases that is suitable for high-throughput screening.  


We describe the development of a novel method for the assay of serine/threonine protein kinases based on fluorescence lifetime. The assay consists of three generic peptides (which have been used by others in the assay of >140 protein kinases in various assay formats) labeled with a long lifetime fluorescent dye (14 or 17 ns) that act as substrates for protein kinases and an iron(III) chelate that modulates the fluorescence lifetime of the peptide only when it is phosphorylated. The decrease in average fluorescence lifetime as measured in a recently developed fluorescence lifetime plate reader (Edinburgh Instruments) is a measure of the degree of phosphorylation of the peptide. We present data showing that the assay performs as well as, and in some cases better than, the "gold standard" radiometric kinase assays with respect to Z' values, demonstrating its utility in high-throughput screening applications. We also show that the assay gives nearly identical results in trial screening to those obtained by radiometric assays and that it is less prone to interference than simple fluorescence intensity measurements. PMID:20230774

Paterson, Michael J; Dunsmore, Colin J; Hurteaux, Reynald; Maltman, Beatrice A; Cotton, Graham J; Gray, Alexander



A universal and label-free aptasensor for fluorescent detection of ATP and thrombin based on SYBR Green I dye.  


A facile and universal aptamer-based label-free approach for selective and sensitive fluorescence detection of proteins and small biomolecules by using the SYBR Green I (SGI) dye is developed. This robust versatile biosensing strategy relies on fluorescence turn-off changes of SGI, resulting from target-induced structure switching of aptamers. Upon binding with the targets, the aptamers dissociate from the respective cDNA/aptamer duplexes, leading to the release of the dsDNA-intercalated SGI into solution and the quenching of the corresponding fluorescence intensities. Such target-induced conformational changes and release of aptamers from the DNA duplexes essentially lead to the change in the fluorescence signal of the SGI and thus constitute the mechanism of our aptamer-based label-free fluorescence biosensor for specific target analyses. Under optimized conditions, our method exhibits high sensitivity and selectivity for the quantification of ATP and thrombin with low detection limits (23.4 nM and 1.1 nM, respectively). Compared with previous reported methods for aptamer-based detection of ATP and thrombin, this label-free approach is selective, simple, convenient and cost-efficient without any chemical labeling of the probe or the target. Therefore, the present strategy could be easily applicable to biosensors that target a wide range of biomolecules. PMID:23202351

Kong, Ling; Xu, Jin; Xu, Yunying; Xiang, Yun; Yuan, Ruo; Chai, Yaqin



Three-dimensional nanographene based on triptycene: synthesis and its application in fluorescence imaging.  


A novel kind of three-dimensional (3D) nanographene based on a triptycene structure bearing three hexa-peri-hexabenzocoronene (HBC) moieties was synthesized efficiently from triiodotriptycene. With the characteristic of intrinsic fluorescence, the 3D nanographene was used as a fluorescent agent for in vitro and in vivo fluorescence imaging with good antiphotobleaching ability and little toxicity. PMID:23163295

Zhang, Chun; Liu, Ying; Xiong, Xiao-Qin; Peng, Lian-Hui; Gan, Lu; Chen, Chuan-Feng; Xu, Hui-Bi



Rhodamine-based 'turn-on' fluorescent probe for Cu(II) and its fluorescence imaging in living cells.  


A novel rhodamine spirolactam derivative 3',6'-Bis(diethylamino)-2-(2-hydroxyethylamino) spiro[isoindoline-1,9'-xanthen]-3-one (RO1) was synthesized, and characterized by high-resolution mass spectrometry (HRMS), X-ray crystallography, Infrared spectroscopy (IR), and (1)H NMR and (13)C NMR spectroscopy. RO1 exhibited highly sensitive and exclusively selective fluorescence response toward Cu(2+) over other metal ions with a detection limit of 0.56ppb in mixed aqueous solution. The fluorescence was pH-independent in the wide range pH 3.1-11.6. The turn-on fluorescence enhancement of the probe is based on Cu(2+) induced ring-opening mechanism of the rhodamine spirolactam. Moreover, by means of fluorescence microscopy experiments, it was demonstrated that RO1 could monitor trace Cu(2+) changes by live cell imaging. PMID:23570786

Tian, Mao-Zhong; Hu, Ming-Ming; Fan, Jiang-Li; Peng, Xiao-Jun; Wang, Jing-Yun; Sun, Shi-Guo; Zhang, Rong



Coordination polymer based on cyano: Synthesis, crystal structure, and fluorescence  

SciTech Connect

One novel 2-D polythreading framework named as [Cu{sub 3}(CN){sub 3}(NH{sub 3})] (1), was obtained through the self-assembling of CuCN under hydrothermal reaction. It is remarkable that there is a 26-membered [Cu{sub 10}(CN){sub 8}] decanuclear metallamacrocycle with the effective size of ca. 16.8x6.83 A{sup 2} along the a-axis. These 2-D layers stack in an ..ABAB...staggered fashion, with the lateral {l_brace}(CN)Cu{sub 3}(NH{sub 3}){r_brace} moieties of each layer inserting into the voids of the decanuclear metallamacrocycles from two adjacent layers. Optical diffuse reflectance spectrum and the result of DFT calculation reveal that 1 is potential direct semiconducting material. In the solid state at room temperature, 1 shows bright yellow fluorescence under ultraviolet light illumination. Its emissive excited state is primarily attributed to the LMCT, LLCT and {sup 3}[MMLCT] excited state, based on the result of DFPT calculation. - Graphical abstract: One novel 2-D polythreading framework [Cu{sub 3}(CN){sub 3}(NH{sub 3})] (1) obtained through the self-assembling of CuCN under hydrothermal reaction shows bright yellow fluorescence at room temperature.

Fang Zhenlan; He Jiangang; Ju Qiang; Wu Xiaoyuan [State Key Laboratory of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China); Lu Canzhong, E-mail: [State Key Laboratory of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China)



Optical biopsy fiber-based fluorescence spectroscopy instrumentation  

NASA Astrophysics Data System (ADS)

Native fluorescence spectroscopy of biomolecules has emerged as a new modality to the medical community in characterizing the various physiological conditions of tissues. In the past several years, many groups have been working to introduce the spectroscopic methods to diagnose cancer. Researchers have successfully used native fluorescence to distinguish cancerous from normal tissue samples in rat and human tissue. We have developed three generations of instruments, called the CD-scan, CD-ratiometer and CD-map, to allow the medical community to use optics for diagnosing tissue. Using ultraviolet excitation and emission spectral measurements on both normal and cancerous tissue of the breast, gynecology, colon, and aerodigestive tract can be separated. For example, from emission intensities at 340 nm to 440 nm (300 nm excitation), a statistically consistent difference between malignant tissue and normal or benign tissue is observed. In order to utilize optical biopsy techniques in a clinical setting, the CD-scan instrument was developed, which allows for rapid and reliable in-vitro and in-vivo florescence measurements of the aerodigestive tract with high accuracy. The instrumentation employs high sensitivity detection techniques which allows for lamp excitation, small diameter optical fiber probes; the higher spatial resolution afforded by the small diameter probes can increase the ability to detect smaller tumors. The fiber optic probes allow for usage in the aerodigestive tract, cervix and colon. Needle based fiber probes have been developed for in-vivo detection of breast cancer.

Katz, Al; Ganesan, S.; Yang, Yuan-Long; Tang, Gui C.; Budansky, Y.; Celmer, Edward J.; Savage, Howard E.; Schantz, Stimson P.; Alfano, Robert R.



Synthesis of fluorescent carbon dots via simple acid hydrolysis of bovine serum albumin and its potential as sensitive sensing probe for lead (II) ions.  


Carbon dots have great potential to be utilised as an optical sensing probe due to its unique photoluminescence and less toxic properties. This work reports a simple and novel synthesis method of carbon dots via direct acid hydrolysis of bovine serum albumin protein in a one-pot approach. Optimisation of the important synthetic parameters has been performed which consists of temperature effect, acid to protein ratio and kinetics of reaction. Higher temperature has promoted better yield with shorter reaction time. The carbon dots obtained shows a strong emission at the wavelength of 400nm with an optimum excitation of 305nm. The potential of the carbon dots as optical sensing probe has been investigated on with different cations that are of environmental and health concern. The fluorescence of the carbon dots was significantly quenched particularly by lead (II) ions in a selective manner. Further analytical study has been performed to leverage the performance of the carbon dots for lead (II) ions sensing using the standard Stern-Volmer relationship. The sensing probe has a dynamic linear range up to 6.0mM with a Stern-Volmer constant of 605.99M(-1) and a limit of detection (LOD) of 5.05?M. The probe performance was highly repeatable with a standard deviation below 3.0%. The probe suggested in this study demonstrates the potential of a more economical and greener approach that uses protein based carbon dots for sensing of heavy metal ions. PMID:24148375

Wee, Shui Shui; Ng, Yann Huey; Ng, Sing Muk



Simple resazurin-based microplate assay for measuring Chlamydia infections.  


The conventional method for quantification of Chlamydia infection using fluorescence microscopy typically involves time- and labor-intensive manual enumeration, which is not applicable for a large-scale analysis required for an inhibitory compound screen. In this study, an alamarBlue (resazurin) assay was adopted to measure Chlamydia infection by measuring the redox capability of infected host cells in a 96-well format. The assay provided measurements comparable to those of the conventional microscopy method while drastically reducing the time required for analysis. PMID:23507273

Osaka, Ichie; Hefty, P Scott



Development of an image processing support system based on fluorescent dye to prevent elderly people with dementia from wandering.  


The wandering of elderly people with dementia is a significant behavioral problem and is a heavy burden on caregivers in residential and nursing homes. Thus, warning systems have been developed to prevent elderly people with dementia from leaving the premises. Some of these systems use radio waves. However, systems based on radio waves present several practical problems. For instance, the transmitter must be carried and may become lost; in addition, the battery of the transmitter must be changed. To solve these problems, we developed a support system that prevents elderly people with dementia from wandering. The system employs image processing technology based on fluorescent dye. The composition of the support system can be described as follows: fluorescent dye is painted in a simple shape on the clothes of an elderly person. The fluorescent color becomes visible by irradiation with a long wavelength of ultraviolet light. In the present paper, the relationship between the color of the dye and the cloth was investigated. A 3D video camera was used to acquire a 3D image and detect the simple shape. As a preliminary experiment, 3 colors (red, green and blue) of fluorescent dye were applied to cloths of 9 different colors. All fluorescent colors were detected on 6 of the cloths, but red and blue dye could not be detected on the other 3 cloths. In contrast, green dye was detectable on all 9 of the cloths. Additionally, we determined whether green dye could be detected in an actual environment. A rectangular shaped patch of green fluorescent dye was painted on the shoulder area of a subject, from the scapula to the clavicle. As a result, the green dye was detected on all 9 different colored cloths. PMID:24111431

Nishigaki, Yutaka; Tanaka, Kentaro; Kim, Juhyon; Nakajima, Kazuki



Studies of bio-mimetic medium of ionic and non-ionic micelles by a simple charge transfer fluorescence probe N,N-dimethylaminonapthyl-(acrylo)-nitrile.  


In this report we have studied micellization process of anionic, cationic and non-ionic surfactants using N,N-dimethylaminonapthyl-(acrylo)-nitrile (DMANAN) as an external fluorescence probe. Micropolarity, microviscosity, critical micellar concentration of these micelles based on steady state absorption and fluorescence and time resolved emission spectroscopy of the probe DMANAN show that the molecule resides in the micelle-water interface for ionic micelles and in the core for the non-ionic micelle. The effect of variation of pH of the micellar solution as well as fluorescence quenching measurements of DMANAN provide further support for the location of the probe in the micelles. PMID:21393054

Samanta, Anuva; Paul, Bijan Kumar; Guchhait, N



Frequency-domain flow cytometry: fluorescence-lifetime-based sensing technology for analyzing cells and chromosomes labeled with fluorescent probes  

NASA Astrophysics Data System (ADS)

A flow cytometer has been developed that combines flow cytometry (FCM) and fluorescence lifetime spectroscopy measurement principles to provide unique capabilities for making frequency-domain, excited-state lifetime measurements on cells/chromosomes labeled with fluorescent probes, while preserving conventional FCM capabilities. Cells are analyzed as they intersect a high-frequency, intensity-modulated (sine-wave) laser excitation beam. Fluorescence signals are processed by (1) low-pass filtering to obtain conventional FCM dc-excited signals and (2) phase-sensitive detection electronics to resolve heterogeneous fluorescence based on differences in lifetimes expressed as phase-shifts and to quantify fluorescence lifetimes in real time. Processed signals are displayed as frequency distribution histograms and bivariate contour diagrams. Recent examples of biological applications include: (1) lifetime histograms recorded on autofluorescent human lung fibroblasts, murine thymus cells labeled with antibodies conjugated to fluorophores for studying fluorescence quenching as a function of antibody dilution and F/P ratio, and on cultured cells, nuclei, and chromosomes stained with DNA-binding fluorochromes and (2) phase-resolved, fluorescence signal- intensity histograms recorded on autofluorescent HLFs labeled with immunofluorescence markers and on murine thymus cells labeled with Red 613-antiThy 1.2 and propidium iodide (PI positive `dead' cells) to demonstrate the resolution of signals from highly overlapping emission spectra. This technology will increase the number of fluorescent markers usable in multilabeling studies and lifetimes can be used as spectroscopic probes to study the interaction of markers with their targets, each other, and the surrounding microenvironment.

Steinkamp, John A.; Crissman, Harry A.; Lehnert, Bruce E.; Lehnert, Nancy M.; Deka, Chiranjit



A simple, low-cost, remote fiber-optic micro volume fluorescence flowcell for capillary flow-injection analysis.  


A small volume flowcell for fluorescence detection in capillary flow injection (CFI) analysis has been created by using a low cost, commercially available fluidic device. Fluorescence detection is achieved using an optical fiber to deliver excitation light to the sample flowing through the device and another optical fiber to collect fluorescence emission. The flowcell is a standard fluidic cross with a swept volume of 721 nL. Optical fibers were oriented at right angles using standard sleeves and ferrules to set their position near the cross intersection. Multiple excitation sources were used including a low power UV laser and blue and UV light emitting diodes (LED). The full emission spectrum detection limits, using the laser, for fluorescein and bovine serum albumin (BSA) were 0.30 ppb and 2.1 x 10(-4)% (w/w), respectively. Two fluidic crosses were used in series for multi-wavelength fluorescence excitation using fiber-optically coupled LED. PMID:12373382

Hart, Sean J; Jiji, Renee D



A SIMPLE based discontinuous Galerkin solver for steady incompressible flows  

NASA Astrophysics Data System (ADS)

In this paper we present how the well-known SIMPLE algorithm can be extended to solve the steady incompressible Navier-Stokes equations discretized by the discontinuous Galerkin method. The convective part is discretized by the local Lax-Friedrichs fluxes and the viscous part by the symmetric interior penalty method. Within the SIMPLE algorithm, the equations are solved in an iterative process. The discretized equations are linearized and an equation for the pressure is derived on the discrete level. The equations obtained for each velocity component and the pressure are decoupled and therefore can be solved sequentially, leading to an efficient solution procedure. The extension of the proposed scheme to the unsteady case is straightforward, where fully implicit time schemes can be used.Various test cases are carried out: the Poiseuille flow, the channel flow with constant transpiration, the Kovasznay flow, the flow into a corner and the backward-facing step flow. Using a mixed-order formulation, i.e. order k for the velocity and order k-1 for the pressure, the scheme is numerically stable for all test cases. Convergence rates of k+1 and k in the L2-norm are observed for velocity and pressure, respectively. A study of the convergence behavior of the SIMPLE algorithm shows that no under-relaxation for the pressure is needed, which is in strong contrast to the application of the SIMPLE algorithm in the context of the finite volume method or the continuous finite element method. We conclude that the proposed scheme is efficient to solve the steady incompressible Navier-Stokes equations in the context of the discontinuous Galerkin method comprising hp-accuracy.

Klein, Benedikt; Kummer, Florian; Oberlack, Martin



Simple, effective estimation of frequency based on Prony's method  

Microsoft Academic Search

We address the design of hardware-efficient digital implementations for the estimation of sinusoidal frequency from noisy data. We are motivated by the design of digital microwave receivers with high sample rates, which thus require simple processing to be cost-effective. By a controlled introduction of potential ambiguities and by use of formulas for the noise-free Prony (1795), or linear prediction coefficients,

Donald W. Tufts; Paul D. Fiore



Movement-based compatibility in simple response tasks  

Microsoft Academic Search

Previous studies reported that movement observation affected movement execution. Using one and the same set of responses (i.e., lifting or tapping the finger), correspondence effects were observed for simple responses when the go-signals were similar to the responses (i.e., movies of finger movements) but not when they were dissimilar (i.e., moving squares). The difference was attributed to a higher degree

Simone Bosbach; Wolfgang Prinz; Dirk Kerzel



Fluorescent detection of protein kinase based on zirconium ions-immobilized magnetic nanoparticles.  


We report here an affinity separation-based fluorometric method for monitoring the activity and inhibition of protein kinase. In this assay, when the fluorescein isothiocyanate (FITC) labeled substrate peptides (S-peptide) are phosphorylated by kinase, the product peptides (P-peptide) will be adsorbed and concentrated onto the surface of Zr(4+)-immobilized nitrilotriacetic acid-coated magnetic nanoparticles (Zr-NTA MNPs) through the chelation of Zr(4+) and phosphate groups. After magnetic separation, the fluorescence intensity of the homogeneous solution changes dramatically. Hence the fluorescence response allows this MNPs-based method to easily probe kinase activity by a spectrometer. The feasibility of the method has been demonstrated by sensitive measurement of the activity of cAMP-dependent protein kinase (PKA) with a low detection limit (0.5 mU ?L(-1)). Moreover, the system is successfully applied to estimate the IC50 value of PKA inhibitor H-89 and detect the Forskolin/3-isobutyl-1-methylxanthine (IBMX) stimulated activation of PKA in cell lysate. Additionally, Zr-NTA MNPs are reusable by stripping Zr(4+) ions from NTA-coated MNPs and rechelating again. This method, which relies on the surface-functionalized MNPs, presents a promising candidate for simple and cost-effective assay of kinase activity and inhibitor screening. PMID:23680555

Tan, Penglong; Lei, Chunyang; Liu, Xin; Qing, Meng; Nie, Zhou; Guo, Manli; Huang, Yan; Yao, Shouzhuo



Development of a novel efficient fluorescence-based plaque reduction microneutralization assay for measles virus immunity.  


The measurement of functional measles virus-specific neutralizing antibodies is of considerable interest for vaccine-related research. In this study, we developed and standardized a simple, rapid, highly sensitive, and reproducible fluorescence-based plaque reduction microneutralization (PRMN) assay with visual and automated readout, using a recombinant measles virus engineered to express enhanced green fluorescent protein. The assay is performed in micro format, requires less time to complete (2 versus 4 to 7 days), and is less labor-intensive and less costly than the classical plaque reduction neutralization (PRN) test, widely accepted as the "gold standard" in measles serology. Two available WHO international anti-measles virus standards and one in-house reference serum were used to develop and standardize the new assay. The mean PRMN values from repeated assays were found to be similar to those reported in the literature or assigned to the WHO standards by the classical PRN assay. For validation, we used three groups of low, moderate, and high measles virus vaccine responders' sera with moderate values of correlation in antibody levels (mIU/ml) between PRMN and the Dade Behring immunoglobulin G enzyme immunoassay (EIA). The PRMN assay was more sensitive at low antibody levels and more informative in terms of protection than this commercial EIA. In conclusion, we have developed and validated a sensitive and high-throughput measles virus-specific PRMN that can be readily used in large population-based measles studies. PMID:18463223

Haralambieva, Iana H; Ovsyannikova, Inna G; Vierkant, Robert A; Poland, Gregory A



Ultrathin and nanostructured ZnO-based films for fluorescence biosensing applications.  


The fluorescence-based sensing capability of ultrathin ZnO-SiO(2) nanoplatforms, deposited by an integrated approach of colloidal lithography and metal organic chemical vapor deposition, has been investigated upon adsorption of fluorescein-labeled albumin, used as model analyte biomolecule. The protein immobilization process after spontaneous adsorption/desorption significantly enhances the green emission of the different ZnO-based films, as evidenced by scanning confocal microscopy, corresponding to a comparable protein coverage detected by X-ray photoelectron spectroscopy. Moreover, experiments of fluorescence recovery after photobleaching evidence that the protein lateral diffusion at the biointerface is affected by the chemical and/or topographical patterning of hybrid ZnO-SiO(2) surfaces. The used approach is very promising for biomolecular detection applications of these ZnO-SiO(2) nanoplatforms, by simple sizing of the 2D vs. 3D patterning design, which in turn is accomplished by the fine tuning of the integrated colloidal lithography-chemical vapor deposition processes. PMID:21978402

Satriano, Cristina; Fragalà, Maria Elena; Aleeva, Yana



Label-free fluorescent detection of thrombin activity based on a recombinant enhanced green fluorescence protein and nickel ions immobilized nitrilotriacetic acid-coated magnetic nanoparticles.  


Herein, a novel label-free fluorescent assay has been developed to detect the activity of thrombin and its inhibitor, based on a recombinant enhanced green fluorescence protein (EGFP) and Ni(2+) ions immobilized nitrilotriacetic acid-coated magnetic nanoparticles (Ni(2+)-NTA MNPs). The EGFP, containing a thrombin cleavage site and a hexahistidine sequence (His-tag) at its N-terminal, was adsorbed onto Ni(2+)-NTA MNPs through Ni(2+)-hexahistidine interaction, and dragged out of the solution by magnetic separation. Thrombin can selectively digest EGFP accompanied by His-tag peptide sequence leaving, and the resulting EGFP cannot be captured by Ni(2+)-NTA MNPs and kept in supernatant. Hence the fluorescence change of supernatant can clearly represent the activity of thrombin. Under optimized conditions, such assay showed a relatively low detection limit (3.0×10(-4)UmL(-1)), and was also used to detect the thrombin inhibitor, Hirudin, and further applied to detect thrombin activity in serum. Combined with the satisfactory reusability of Ni(2+)-NTA MNPs, our method presents a promising candidate for simple, sensitive, and cost-saving protease activity detecting and inhibitor screening. PMID:24148431

Wang, Ming; Lei, Chunyang; Nie, Zhou; Guo, Manli; Huang, Yan; Yao, Shouzhuo



Gold Nanoparticle Based Surface Enhanced Fluorescence For Detection of Organophosphorus Agents  

PubMed Central

Organophosphorus agents (OPA) represent a serious concern to public safety as nerve agents and pesticides. Here we report the development of gold nanoparticle based surface enhanced fluorescence (NSEF) spectroscopy for rapid and sensitive screening of organophosphorus agents. Fluorescent from Eu3+ ions that are bound within the electromagnetic field of gold nanoparticles exhibit a strong enhancement. In the presence of OPA, Eu3+ ions are released from the gold nanoparticle surface and thus a very distinct fluorescence signal change was observed. We discussed the mechanism of fluorescence enhancement and the role of OPA for fluorescence intensity change in the presence of gold nanoparticles.

Dasary, Samuel S. R.; Rai, Uma S.; Yu, Hongtao; Anjaneyulu, Yerramilli; Dubey, Madan




Microsoft Academic Search

Guanosine 3?,5?-cyclic monophosphate (cGMP) plays a role as a second messenger in many different biological systems. Given the ubiquitous nature of cGMP, a simple method of detecting cGMP is of interest. To that end a fluorescent polymer with recognition sites for cGMP has been prepared. Its selectivity and sensitivity were investigated and a dose-dependant decrease in fluorescence of the polymer

Nguyen Thi Kim Thanh; Daniel L. Rathbone; David C. Billington; Nicholas A. Hartell



Fluorescent quenching-based quantitative detection of specific DNA\\/RNA using a BODIPY(R) FL-labeled probe or primer  

Microsoft Academic Search

We have developed a simple method for the quantitative detection of specific DNA or RNA molecules based on the finding that BODIPY® FL fluorescence was quenched by its interaction with a uniquely positioned guanine. This approach makes use of an oligonucleo- tide probe or primer containing a BODIPY® FL-modified cytosine at its 5'-end. When such a probe was hybrid- ized

Shinya Kurata; Takahiro Kanagawa; Kazutaka Yamada; Masaki Torimura; Toyokazu Yokomaku; Yoichi Kamagata; Ryuichiro Kurane



A label-free fluorescence DNA probe based on ligation reaction with quadruplex formation for highly sensitive and selective detection of nicotinamide adenine dinucleotide.  


A simple label-free fluorescent sensing scheme for sensitive and selective detection of nicotinamide adenine dinucleotide (NAD(+)) has been developed based on DNA ligation reaction with ligand-responsive quadruplex formation. This approach can detect 0.5 nM NAD(+) with high selectivity against other NAD(+) analogs. PMID:22456321

Zhao, Jingjin; Zhang, Liangliang; Jiang, Jianhui; Shen, Guoli; Yu, Ruqin



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

Microsoft Academic Search

Fluorescence lifetime-based chemical sensors have recently been described for applications in medicine, environmental monitoring, and bioprocess control. These sensors transduce the level of the analyte as a change in the apparent fluorescence lifetime of an indicator phase. We have previously developed a wavelength-ratiometric fluorescence biosensor for zinc based on binding of zinc and dansylamide to apo-carbonic anhydrase which exhibited high

Richard B. Thompson; Marcia W. Patchan



The first bifluoride sensor based on fluorescent enhancement.  


The first fluorescent sensor for HF2(-) anion, N(1), N(3)-di(naphthalene-1-yl)isophthalamide (L) has been derived from ?-Napthylamine and isopthaloyl chloride. In 1:1 (v/v) DMSO:H2O, L exhibits high selectivity towards HF2(-) anion with a 4-fold enhancement in fluorescent intensity. Very little enhancement in fluorescence intensity is observed for F(-), Cl(-), Br(-), I(-), SCN(-), PO4(3-), SO4(2-), and CH3COO(-) anions. The stoichiometry interaction between L and HF2 (-) is found to be 1:1 from fluorescence and UV/Visible spectral data. DFT calculation shows that binding between HF2(-) and L is 1:1 and increases the relative planarity between the two naphthyl rings causing fluorescence enhancement. A shift of 0.080 V in oxidation potential of L is observed on interaction with HF2(-) by cyclic voltammetry and square wave voltammetry. PMID:23525971

Dutta, Kaku; Deka, Ramesh Ch; Das, Diganta Kumar



Fluorescent detection of single nucleotide polymorphism utilizing a hairpin DNA containing a nucleotide base analog pyrrolo-deoxycytidine as a fluorescent probe  

Microsoft Academic Search

A novel fluorescent method for the detection of single nucleotide polymorphism (SNP) was developed using a hairpin DNA containing nucleotide base analog pyrrolo-deoxycytidine (P-dC) as a fluorescent probe. This fluorescent probe was designed by incorporating a fluorescent P-dC into a stem of the hairpin DNA, whose sequence of the loop moiety complemented the target single strand DNA (ss-DNA). In the

Hongge Zhang; Minjuan Wang; Qiang Gao; Honglan Qi; Chengxiao Zhang



A fluorescent chemosensor for Hg2+ based on naphthalimide derivative by fluorescence enhancement in aqueous solution.  


Naphthalimide derivative (compound 1) containing hydrophilic hexanoic acid group was synthesized and used to recognize Hg(2+) in aqueous solution. The fluorescence enhancement of 1 is attributed to the formation of a complex between 1 and Hg(2+) by 1:1 complex ratio (K=2.08×10(5)), which has been utilized as the basis of fabrication of the Hg(2+)-sensitive fluorescent chemosensor. The comparison of this method with some other fluorescence methods for the determination of Hg(2+) indicated that the method can be applied in aqueous solution rather than organic solution. The analytical performance characteristics of the proposed Hg(2+)-sensitive chemosensor were investigated. The chemosensor can be applied to the quantification of Hg(2+) with a linear range covering from 2.57×10(-7) to 9.27×10(-5) M and a detection limit of 4.93×10(-8) M. The experiment results show that the response behavior of 1 toward Hg(2+) is pH independent in medium condition (pH 4.0-8.0). Most importantly, the fluorescence changes of the chemosensor are remarkably specific for Hg(2+) in the presence of other metal ions, which meet the selective requirements for practical application. Moreover, the response of the chemosensor toward Hg(2+) is fast (response time less than 1 min). In addition, the chemosensor has been used for determination of Hg(2+) in hair samples with satisfactory results, which further demonstrates its value of practical applications. PMID:22304823

Li, Chun-Yan; Xu, Fen; Li, Yong-Fei; Zhou, Kai; Zhou, Yu



Enhanced fluorescence sensing of melamine based on thioglycolic acid-capped CdS quantum dots.  


A sensitive and simple method for the determination of melamine (MA) was developed based on the fluorescence enhancement effect of MA for thioglycolic acid-capped (TGA-capped) CdS quantum dots (QDs). Under optimum conditions, a good linear relationship was obtained from 2.0 × 10(-9) to 5.0 × 10(-5)M. The detection limit was 1.0 × 10(-9)M, which was much lower than the safety limit (2.5 ppm in USA and the UK; 1 ppm for infant formula in China). The solution pH, the adding sequence of the buffer solution and MA and surface modifiers of CdS QDs greatly influenced the enhancement extent of MA for CdS QDs. The fluorescence enhancement was attributed to the surface passivation of the surface states of QDs by amine group of MA. The method was applied to detect MA in raw milk with satisfactory results. The proposed method manifested several advantages such as high sensitivity, short analysis time, low cost and ease of operation. PMID:22483928

Wang, Guang-Li; Jiao, Huan-Jun; Zhu, Xiao-Ying; Dong, Yu-Ming; Li, Zai-Jun



Fluorescence resonance energy transfer-based near-infrared fluorescence sensor for glucose monitoring.  


A novel near-infrared (NIR) fluorescence affinity sensor for continuous glucose monitoring was developed and characterized. The sensor operates by fluorescence resonance energy transfer between a NIR chromophore linked to concanavalin A (ConA) and an NIR fluorophore linked to free dextran. The binding of dextran with ConA in the absence of glucose results in low fluorescence due to quenching; however, the quenching is reversed by competitive displacement of dextran from ConA by glucose. In order to increase thermodynamic stability and the lifetime of the sensor, ConA was immobilized within a macroporous bead matrix. The sensor was contained within a sealed hollow dialysis fiber (o.d. 215 microm, wall thickness 20 microm), preventing the macromolecules from leaking out and enabling glucose to rapidly enter the fiber lumen. A glucose-insensitive reference fluorophore was also incorporated to allow for ratiometric measurements, resulting in a robust sensor output that is independent of positional and/or light intensity changes. The response of the fluorescence affinity sensor to glucose was tested continuously in an automated test chamber at 37 degrees C. The sensor showed good dynamic range within physiologically relevant glucose concentration range (15% change over 2.5-30 mM, no hysteresis), fast response time (2-4 min), and a remarkable long-term stability (6 months). We interpret the improved longevity of this sensor to be the result of an optimized photo exposure regime and immobilization of ConA to the matrix. Its small size, ratiometric output, and NIR fluorescence make this sensor well suited for dermal implantation and continuous transdermal monitoring. PMID:15117585

Ballerstadt, Ralph; Gowda, Ashok; McNichols, Roger



Self-tuned Power System Stabilizer Based on a Simple Fuzzy Logic Controller  

Microsoft Academic Search

A self-tuned fuzzy logic power system stabilizer to enhance the damping of power system oscillations is presented. The design of the self-tuned fuzzy logic power system stabilizer is based on a simple fuzzy logic controller that possesses a significantly reduced rule base, a small number of tuning parameters, and is implemented through a simple control algorithm and architecture. The control

Miguel Ramirez-Gonzalez; O. P. Malik



A simple machine [based on the SK-combinator reduction mechanism  

Microsoft Academic Search

A simple machine architecture based on an extension to the combinator notation is described. Code travels along a unidirectional stream and is executed in parallel by simple finite state machine based cells separated by sections of a first-in, first-out (FIFO) buffer. The resulting regularity and structural simplicity ensure a good match to VLSI implementation. It is verified that the extended

Alex Dickinson; Michael T. Pope



Fluorescence lifetime-based sensing in tissues: a computational study.  

PubMed Central

We have numerically solved the photon diffusion equation to predict the distribution of light in a tissue model system with a uniform concentration of fluorophore. Our results show that time-dependent measurements of light propagation can be used to monitor the fluorescent lifetimes of a uniformly distributed fluorophore in tissues. With proper referencing, frequency-domain measurements of phase-shift, theta, may allow quantitation of fluorescent lifetimes, tau, independent of changes in the local absorption and scattering properties. These results point to a new approach for noninvasive diagnostic monitoring through quantitation of fluorescent lifetime, tau, when the lifetime of the fluorophore is comparable with photon migration times. Images FIGURE 1

Hutchinson, C L; Lakowicz, J R; Sevick-Muraca, E M



Neoplasm diagnostics based on fluorescence of polymethine dyes  

NASA Astrophysics Data System (ADS)

Investigated polymethine dye TICS has near IR bands of fluorescence and absorption within the transparency region of biological tissues. It can be detected up to 1.5 cm from the surface of the skin. The intensity of a fluorescence signal of TICS is linear for doses up to 2 mg/kg in both tumor and muscle tissue. The ratio of an intensity of light induced fluorescence in tumor tissue to one in muscle tissue is up to 3.6 for rapidly growing tumors. The retention time of TICS is 7 days in all tissues. TICS can be used in the detection of tumor boundaries and tumor internal structure.

Samtsov, Michael P.; Voropay, Eugene S.; Chalov, Vadim N.; Zhavrid, Edvard A.



Translation on demand by a simple RNA-based thermosensor  

PubMed Central

Structured RNA regions are important gene control elements in prokaryotes and eukaryotes. Here, we show that the mRNA of a cyanobacterial heat shock gene contains a built-in thermosensor critical for photosynthetic activity under stress conditions. The exceptionally short 5?-untranslated region is comprised of a single hairpin with an internal asymmetric loop. It inhibits translation of the Synechocystis hsp17 transcript at normal growth conditions, permits translation initiation under stress conditions and shuts down Hsp17 production in the recovery phase. Point mutations that stabilized or destabilized the RNA structure deregulated reporter gene expression in vivo and ribosome binding in vitro. Introduction of such point mutations into the Synechocystis genome produced severe phenotypic defects. Reversible formation of the open and closed structure was beneficial for viability, integrity of the photosystem and oxygen evolution. Continuous production of Hsp17 was detrimental when the stress declined indicating that shutting-off heat shock protein production is an important, previously unrecognized function of RNA thermometers. We discovered a simple biosensor that strictly adjusts the cellular level of a molecular chaperone to the physiological need.

Kortmann, Jens; Sczodrok, Simon; Rinnenthal, Jorg; Schwalbe, Harald; Narberhaus, Franz



Synthesis and properties of novel base-discriminating fluorescent (BDF) nucleosides.  


We designed a new type of pyrene-labeled base-discrimination fluorescent (BDF) nucleosides (Py)U, (Py)C, (8Py)A and (MePy)dA, which emitted strong fluorescence only when the bases opposite the BDF base are A, G, T and C, respectively. The DNA probes containing four different BDF bases enable us to distinguish single base alterations by simply mixing with a sample solution of target DNA. PMID:17150679

Saito, Yoshio; Hanawa, Kazuo; Hayashi, Keigo; Motegi, Kaori; Okaoto, Akimitsu; Saito, Isao



ALA-based fluorescent diagnosis of malignant oral lesions in the presence of bacterial porphyrin formation  

NASA Astrophysics Data System (ADS)

The aminolevulinic acid (5-ALA) -based fluorescence diagnosis has been found to be promising for an early detection and demarcation of superficial oral squamous cell carcinomas (OSCC). This method has previously demonstrated high sensitivity, however this clinical trial showed a specificity of approximately 62 %. This specificity was mainly restricted by tumor detection in the oral cavity in the presence of bacteria. After topical ALA application in the mouth of patients with previously diagnosed OSSC, red fluorescent areas were observed which did not correlate to confirm histological findings. Swabs and plaque samples were taken from 44 patients and cultivated microbiologically. Fluorescence was investigated (OMA-system) from 32 different bacteria strains found naturally in the oral cavity. After ALA incubation, 30 of 32 strains were found to synthesize fluorescent porphyrins, mainly Protoporphyrin IX. Also multiple fluorescent spectra were obtained having peak wavelengths of 636 nm and around 618 nm - 620 nm indicating synthesis of different porphyrins, such as the lipophylic Protoporphyrin IX (PpIX) and hydrophylic porphyrins (water soluble porphyrins, wsp). Of the 32 fluorescent bacterial strains, 18 produced wsp, often in combination with PpIX, and 5 produced solely wsp. These results clarify that ALA-based fluorescence diagnosis without consideration or suppression of bacteria fluorescence may lead to false-positive findings. It is necessary to suppress bacteria fluorescence with suitable antiseptics before starting the procedure. In this study, when specific antiseptic pre-treatment was performed bacterial associated fluorescence was significantly reduced.

Schleier, P.; Berndt, A.; Zinner, K.; Zenk, W.; Dietel, W.; Pfister, W.



A new boronic acid based fluorescent reporter for catechol.  


Catechol skeleton widely exists in natural products and bioactive substances. Fluorescent reporters which could recognize catechol are very promising for the construction of chemosensors to detect catechol and its derivatives in biological environment. Herein, we reported a novel catechol reporter, 2-(4-boronophenyl)quinoline-4-carboxylic acid, which exhibits significant fluorescent property changes upon binding catechol containing molecules in an aqueous solution. PMID:23079526

Wu, Zhongyu; Li, Minyong; Fang, Hao; Wang, Binghe



A coumarin based fluorescent photoinduced electron transfer cation sensor  

Microsoft Academic Search

We wish to report 4-[(Bis(pyridin-2-ylmethyl)amino)methyl]-7-methoxychromen-2-one, 1, as a fluorescent photoinduced electron transfer cation sensor that is capable of indicating the presence of Zn2+, Cd2+ and Pb2+ ions via a fluorescence signal. The log binding constants, ?, for these metal ions have been measured, and were found to be 6.10±0.4, 6.37±0.3, and 5.67±0.3, respectively.

Chandrika P. Kulatilleke; Saliya A. de Silva; Yair Eliav



Development of fluorescent FeIII sensor based on chalcone  

Microsoft Academic Search

In this paper, 4-dimethylamino 2,5-dihydroxy chalcone (DMADHC), which exhibits excited state intramolecular charge transfer (ICT) characteristics, was synthesized and characterized. A sensitive optochemical sensor for Fe3+ ion was developed using DMADHC as fluorescence receptor. The fluorescence of DMADHC was gradually quenched with the addition of Fe3+ ion, which attributed to the formation of 1:1 complex between DMADHC and Fe3+ ion.

Yanli Wei; Guojie Qin; Wenyan Wang; Wei Bian; Shaomin Shuang; Chuan Dong



Quantum dots and fluorescent protein FRET-based biosensors.  


There has been considerable recent interest in the creation of nanoparticle-biomolecule hybrid materials for uses such as in vitro and in vivo biosensing, biological imaging, and drug -delivery. Nanoparticles have a high surface to volume ratio, making them capable of being decorated with -various biomolecules on their surface which retain their biological activity. Techniques to bind these biomolecules to nanoparticle surfaces are also advancing rapidly. Here we demonstrate hybrid materials assembled around CdSe/ZnS core/shell semiconductor quantum dots (QDs). These intrinsically fluorescent materials are conjugated to the fluorescent proteins YFP, mCherry and the light harvesting complex b-phycoerythrin (b-PE). QDs have fluorescent properties that make them ideal as donor fluorophores for Förster resonance energy transfer (FRET) while the fluorescent proteins are able to act as FRET acceptors displaying many advantages over organic dyes. We examine FRET interactions between QDs and all three fluorescent proteins. Furthermore, we show QD-mCherry hybrid materials can be utilized for in vitro biosensing of caspase-3 enzymatic activity. We further show that QDs and fluorescent proteins can be conjugated together intracellularly with strong potential for live-cell imaging and biosensing applications. PMID:22101713

Boeneman, Kelly; Delehanty, James B; Susumu, Kimihiro; Stewart, Michael H; Deschamps, Jeffrey R; Medintz, Igor L



Photobleaching-Based Quantitative Analysis of Fluorescence Resonance Energy Transfer inside Single Living Cell  

Microsoft Academic Search

The current advances of fluorescence microscopy and new fluorescent probes make fluorescence resonance energy transfer (FRET)\\u000a a powerful technique for studying protein-protein interactions inside living cells. It is very hard to quantitatively analyze\\u000a FRET efficiency using intensity-based FRET imaging microscopy due to the presence of autofluorescence and spectral crosstalks.\\u000a In this study, we for the first time developed a novel

Longxiang Wang; Tongsheng Chen; Junle Qu; Xunbin Wei



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


The experimentally obtained time-resolved fluorescence spectra of photosystem II (PS II) core complexes, purified from a thermophilic cyanobacterium Thermosynechococcus vulcanus, at 5-180 K are compared with simulations. Dynamic localization effects of excitons are treated implicitly by introducing exciton domains of strongly coupled pigments. Exciton relaxations within a domain and exciton transfers between domains are treated on the basis of Redfield theory and generalized Förster theory, respectively. The excitonic couplings between the pigments are calculated by a quantum chemical/electrostatic method (Poisson-TrEsp). Starting with previously published values, a refined set of site energies of the pigments is obtained through optimization cycles of the fits of stationary optical spectra of PS II. Satisfactorily agreement between the experimental and simulated spectra is obtained for the absorption spectrum including its temperature dependence and the linear dichroism spectrum of PS II core complexes (PS II-CC). Furthermore, the refined site energies well reproduce the temperature dependence of the time-resolved fluorescence spectrum of PS II-CC, which is characterized by the emergence of a 695 nm fluorescence peak upon cooling down to 77 K and the decrease of its relative intensity upon further cooling below 77 K. The blue shift of the fluorescence band upon cooling below 77 K is explained by the existence of two red-shifted chlorophyll pools emitting at around 685 and 695 nm. The former pool is assigned to Chl45 or Chl43 in CP43 (Chl numbering according to the nomenclature of Loll et al. Nature2005, 438, 1040) while the latter is assigned to Chl29 in CP47. The 695 nm emitting chlorophyll is suggested to attract excitations from the peripheral light-harvesting complexes and might also be involved in photoprotection. PMID:23537277

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



Ionic Calcium Determination in Skim Milk with Molecular Probes and Front-Face Fluorescence Spectroscopy: Simple Linear Regression  

Microsoft Academic Search

The purpose of this study was to determine if the ionic calcium content of skim milk could be determined using molecular probes and front-face fluorescence spectroscopy. Current methods for determining ionic calciumarenotsensitive,overestimateioniccalcium,or require complex procedures. Molecular probes designed specifically for measuring ionic calcium could poten- tially be used to determine the ionic calcium content of skim milk. The goal of

R. R. Gangidi; L. E. Metzger



A simple robot paths planning based on Quadtree  

Microsoft Academic Search

Based on traditional methods, according with practical application conditions, and the special structure of Quadtree, this paper gives a practical method of paths planning for the further expansion of Quadtree in the field of paths planning, except for segmentation obstacles. The experimental and simulation results demonstrate that the algorithm described in the text could be able to find all the

Fule Wangl; Changle Zhou; H. de Garis



Reliability-Based Optimum Design of Simple Plastic Frames.  

National Technical Information Service (NTIS)

Reliability analysis and optimum reliability-based design are discussed and illustrated in this report. A new iterative optimization scheme is exploited where at each stage of the design process both upper and lower bounds on the true optimum value of the...

E. H. Vanmarcke J. Diaz-Padilla D. A. Roth



A simple raster-based model for flood inundation simulation  

Microsoft Academic Search

In this paper the development of a new model for simulating flood inundation is outlined. The model is designed to operate with high-resolution raster Digital Elevation Models, which are becoming increasingly available for many lowland floodplain rivers and is based on what we hypothesise to be the simplest possible process representation capable of simulating dynamic flood inundation. This consists of

P. D Bates; A. P. J De Roo



Quantification of fluorescence intensity of labeled human mesenchymal stem cells and cell counting of unlabeled cells in phase-contrast imaging: an open-source-based algorithm.  


Assessment of cell fate is indispensable to evaluate cell-based therapies in regenerative medicine. Therefore, a widely used technique is fluorescence labeling. A major problem still is the standardized, noninvasive, and reliable quantification of fluorescence intensity of adherent cell populations on single-cell level, since total fluorescence intensity must be correlated to the cell number. Consequently, the aim of the present study was to produce and validate an open-source-based algorithm, capable of measuring the total fluorescence intensity of cell populations and assessing the total cell number in phase-contrast images. To verify the algorithms' capacity to assess fluorescence intensity, human mesenchymal stem cells were transduced to stably express enhanced green fluorescent protein and results produced by the algorithm were compared to flow cytometry analysis. No significant differences could be observed at any time (p ? 0.443). For validation of the algorithm for cell counting in phase-contrast images, adherent human mesenchymal stem cells were manually counted and compared to results produced by the algorithm (correlation coefficient [CC] r = 0.975), nuclei staining (CC r = 0.997), and hemocytometer (CC r = 0.629). We conclude that applying the developed algorithm in routine practice allows robust, fast, and reproducible assessment of fluorescence intensity and cell numbers in simple large-scale microscopy. The method is easy to perform and open source based. PMID:20218817

Polzer, Hans; Haasters, Florian; Prall, Wolf Christian; Saller, Maximilian Michael; Volkmer, Elias; Drosse, Inga; Mutschler, Wolf; Schieker, Matthias



Piezoelectric immunosensor based on magnetic nanoparticles with simple immobilization procedures  

Microsoft Academic Search

A novel method for immobilizing antibodies (antigens) based on magnetic nanoparticles has been proposed for piezoelectric immunoassay. The goat-anti-IgG antibody (IgGAb) as the model analyte was first covalently immobilized to magnetic nanoparticles, which were surface modified with amino-groups. The magnetic bio-nanoparticles (MBN-s) formed were attached to the surfaces of quartz crystal with the help of a permanent magnet. The detection

Jishan Li; Xiaoxiao He; Zhaoyang Wu; Kemin Wang; Guoli Shen; Ruqin Yu



Simultaneous detection of SERS and fluorescence using a single excitation for microbead-based analysis.  


We demonstrate simultaneous detection of surface-enhanced Raman scattering (SERS) and fluorescence signals from a silver microbead. For the dual signal generation, silver microbeads with a diameter of 15 microm were functionalized with benzenethiol (BT) as a Raman tag and a cardiac troponin I (cTnI) antibody on their surface. SERS and fluorescence signals were obtained using a single argon laser source with 488 nm wavelength. The SERS signals from Raman tag can be used as identification indices for decoding a particular microbead, while the fluorescence signals provide the information about molecular interactions with a specific biomarker. With simultaneous detection of SERS and fluorescence signals using single excitation on the functionalized microbeads, we successfully showed the possibility of a simple barcoding strategy for multiplex analysis using suspension arrays. PMID:23909138

Lee, Sa Ram; Jeon, Chang Su; Hwang, Inseong; Chung, Taek Dong; Kim, Hee Chan



Designation of rapid detection system for chlorophyll fluorescence parameters based on LED irradiation  

NASA Astrophysics Data System (ADS)

Adopting high-power light-emitting diode (LED) as excitation light source, the study designed a rapid detection system for fluorescence parameters based on MINIPAM. The system uses a microcomputer as the core of the programmable power supply to provide constant current drive of the LED array, and the LED array as a fluorescence excitation light source produces light photochemical system needed. It also uses MINIPAM to detect the fluorescence, analyzing the fluorescence parameters of the mathematical model, studying the plant photosystem& light response curve. The System is of great significance in the evaluation of chlorophyll photosynthesis ability and the plant physiological stress response and the appropriate mechanism.

Li, Zhengming; Ji, Jianwei; Xu, Minghu



Fluorescent quenching-based quantitative detection of specific DNA/RNA using a BODIPY® FL-labeled probe or primer  

PubMed Central

We have developed a simple method for the quantitative detection of specific DNA or RNA molecules based on the finding that BODIPY® FL fluorescence was quenched by its interaction with a uniquely positioned guanine. This approach makes use of an oligonucleotide probe or primer containing a BODIPY® FL-modified cytosine at its 5?-end. When such a probe was hybridized with a target DNA, its fluorescence was quenched by the guanine in the target, complementary to the modified cytosine, and the quench rate was proportional to the amount of target DNA. This widely applicable technique will be used directly with larger samples or in conjunction with the polymerase chain reaction to quantify small DNA samples.

Kurata, Shinya; Kanagawa, Takahiro; Yamada, Kazutaka; Torimura, Masaki; Yokomaku, Toyokazu; Kamagata, Yoichi; Kurane, Ryuichiro



An improved respiratory syncytial virus neutralization assay based on the detection of green fluorescent protein expression and automated plaque counting  

PubMed Central

Background Virus neutralizing antibodies against respiratory syncytial virus (RSV) are considered important correlates of protection for vaccine evaluation. The established plaque reduction assay is time consuming, labor intensive and highly variable. Methods Here, a neutralization assay based on a modified RSV strain expressing the green fluorescent protein in combination with automated detection and quantification of plaques is described. Results The fluorescence plaque reduction assay in microplate format requires only two days to complete and is simple and reproducible. A good correlation between visual and automated counting methods to determine RSV neutralizing serum antibody titers was observed. Conclusions The developed virus neutralization assay is suitable for high-throughput testing and can be used for both animal studies and (large scale) vaccine clinical trials.



Development of a fluorescence-based point detector for biological sensing  

NASA Astrophysics Data System (ADS)

This paper presents the status of an ongoing development of a point detector for biological warfare agent sensing based on ultraviolet laser-induced fluorescence from single particles in air. The detector will measure the fluorescence spectra of single particles in a sheath flow air beam. The spectral detection part of the system consists of a grating and a photomultiplier tube array with 32 channels, which measure fluorescence spectra in the wavelength band from 300 nm to 650 nm. The detector is designed to measure laser induced fluorescence from single laser pulses and has been tested by measuring fluorescence from simulants of biological warfare agents in aqueous solution. The solutions were excited with laser pulses at the wavelengths of 293 nm and 337 nm. The paper also presents preliminary results on the sheath flow particle injector and time-resolved measurements of fluorescence from biological warfare agent simulants in solution.

Jonsson, Per; Kullander, Fredrik; Nordstrand, Melker; Tjarnhage, Torbjorn; Wasterby, Par; Lindgren, Mikael



Simple and sensitive detection method for Cobalt(II) in water using CePO4:Tb3+ nanocrystals as fluorescent probes.  


A simple and sensitive method for detecting cobalt by synchronous fluorescence spectrometry technique with a novel fluorescence probe CePO4:Tb(3+) has been developed. CePO4:Tb(3+) nanocrystals were synthesized in aqueous solutions and characterized by transmission electron microscopy, electron diffraction pattern spectroscopy and spectrofluorometry. When ??=210 nm, the selected synchronous fluorescence is produced at 284 nm. CePO4:Tb(3+) nanocrystals were negatively charged under weakly basic conditions (pH=8.2), which can interact with Co(2+) via electrostatic interaction. Moreover, there is the spectrum overlap between the emission wavelength of CePO4:Tb(3+) NCs and the absorbance of Co(2+). So the energy transfer would occur, leading to the quenching phenomenon. The quenching equation of the system was agreed with the Stern-Volmer equation. The linear range and detection limit of Co(2+) were 5-1.8 ?M and 3.5 nM, respectively. The method is successfully applied to the quantification of Co(2+) in water samples. PMID:23416919

Chen, Hongqi; Yuan, Fei; Xu, Juan; Zhang, Yiyang; Wu, Yong; Wang, Lun



Three-flat test solutions based on simple mirror symmetry  

SciTech Connect

In interferometric surface and wavefront metrology, three-flat tests are the archetypes of measurement procedures to separate errors in the interferometer reference wavefront from errors due to the test part surface, so-called absolute tests. What is believed to be a new class of solutions of the three-flat problem for circular flats is described in terms of functions that are symmetric or antisymmetric with respect to reflections at a single line passing through the center of the flat surfaces. The new solutions are simpler and easier to calculate than the known solutions based on twofold mirror symmetry or rotation symmetry.Strategies for effective azimuthal averaging and a method for determining the averaging error are also discussed.

Griesmann, Ulf



Simple communication using a SSVEP-based BCI  

NASA Astrophysics Data System (ADS)

Majority of Brain-Computer Interface (BCI) for communication purposes are speller, i.e., the user has to select letter by letter. In this work, is proposed a different approach where the user can select words from a word set designed in order to answer a wide range of questions. The word selection process is commanded by a Steady-state visual evoked potential (SSVEP) based-BCI that allows selecting a word in an average time of 26 s with accuracies of 92% on average. This BCI is focus in the first stages on rehabilitation or even in first moments of some diseases (such as stroke), when the person is eager to communicate with family and doctors.

Sanchez, Guillermo; Diez, Pablo F.; Avila, Enrique; Laciar Leber, Eric



Simple DNA transformation in Pseudomonas based on the Yoshida effect.  


Current protocols of recombinant DNA research, including gene cloning and complementation, quantification of gene expression and tagging with reporter proteins, are usually limited by the availability of effective bacteria transformation tools different from Escherichia coli. This is particularly relevant with respect to the Pseudomonas species due to their biotechnological and sanitary importance. Here, we describe an optimized and efficient plasmid transference protocol based on the Yoshida effect, a method that relies on DNA uptake mediated by friction forces. The main advantages of this method are: (i) no competent cell preparation is needed, (ii) cells in any physiological state can be used, (iii) the procedure is performed directly on agar plates and (iv) the protocol, which is neither time-consuming nor labor-intensive, offers good efficiency. This approach promises to become the gold standard for day to day genetic manipulation in Pseudomonas. PMID:22405834

Rodríguez-Beltrán, Jerónimo; Elabed, Hamouda; Gaddour, Kamel; Blázquez, Jesús; Rodríguez-Rojas, Alexandro



Donor-Acceptor Compound Based on Rhodanineacetic Acid-Pyrene Derivative: Red-Light Emitting Fluorescent Organic Nanoparticles  

Microsoft Academic Search

A donor-acceptor compound based on Rhodanineacetic acid-pyrene derivative (RAAP), which emits weak yellow-green fluorescence\\u000a in the methanol solution, was investigated. RAAP nanoparticles with a mean diameter of 50–60 nm were prepared by a simple\\u000a reprecipitation method without surfactants. The observation of RAAP nanoparticles were undertaken through SEM and TEM method.\\u000a The emission spectra of RAAP nanoparticles are red-shifted (? ?em?=?86 nm) to

Bo Zhang; Wei Diao; Chun Bi; Jian Sun; Guoxia Han; Yi Shi; Li Sheng; Gui Yin; Lin Pu


Folate-based near-infrared fluorescent theranostic gemcitabine delivery.  


A series of heptamethine cyanine (1-3) derivatives bearing a carbamate ethyl disulfide group and gemcitabine, an anticancer drug, have been newly synthesized. Their disulfide bonds are readily cleaved by various thiols including glutathione, to result in a subsequent decomposition of the carbamate into amine followed by release of the active gemcitabine, which can be monitored by the fluorescence changes. In the biological experiment, prodrug 1 is preferentially up-taken by folate-positive KB cells over folate-negative A549 cells via receptor-mediated endocytosis to release gemcitabine causing cell death and to emit fluorescence in endoplasmic reticulum. Moreover, it is selectively accumulated in the KB cells which were treated to mice by dorsal subcutaneous injection. This drug delivery system is a new theranostic agent, wherein both therapeutic effect and drug uptake can be easily monitored at the subcellular level, by in vivo and in vitro fluorescence imaging. PMID:23865715

Yang, Zhigang; Lee, Jae Hong; Jeon, Hyun Mi; Han, Ji Hye; Park, Nayoung; He, Yanxia; Lee, Hyunseung; Hong, Kwan Soo; Kang, Chulhun; Kim, Jong Seung



A simple model for thermal conductivity of carbon nanotube-based composites  

Microsoft Academic Search

A quite simple formula for the thermal conductivity enhancement in carbon nanotube composites is presented based on a conventional model. This simple formula predicts much higher thermal conductivity enhancement even in the dilute case of the carbon nanotubes, due to ultrahigh thermal conductivity and aspect ratio of the carbon nanotubes. By applying this model to nanotube suspensions recently reported in

C.-W. Nan; Z. Shi; Y. Lin



Comparing simple role based access control models and access control lists  

Microsoft Academic Search

The RBAC metaphor is powerful in its ability to ex- press access control policy in terms of the way in which administrators view organizations. The functionality of simple Role Based Access Control (RBAC) models are compared to access control lists (ACL). A very simple RBAC model is shown to be no different from a group ACL mechanism from the point

John F. Barkley



Beer's-law-based, simple spectral model for direct normal and diffuse horizontal irradiance  

Microsoft Academic Search

A spectral model for cloudless days that uses simple mathematical expressions and tabulated lock up tables to generate direct normal and diffuse horizontal irradiance is presented. The model is based on modifications to previously published simple models and comparisons with rigorous radiative transfer codes. This model is expected to be more accurate and to be applicable to a broader range

R. E. Bird



Gold nanoparticle based surface enhanced fluorescence for detection of organophosphorus agents  

Microsoft Academic Search

Organophosphorus agents (OPA) represent a serious concern to public safety as nerve agents and pesticides. Here we report the development of gold nanoparticle based surface enhanced fluorescence (NSEF) spectroscopy for rapid and sensitive screening of organophosphorus agents. Fluorescent from Eu3+ ions that are bound within the electromagnetic field of gold nanoparticles exhibit a strong enhancement. In the presence of OPA,

Samuel S. R. Dasary; Uma S. Rai; Hongtao Yu; Yerramilli Anjaneyulu; Madan Dubey; Paresh Chandra Ray



Iron complex-based fluorescent probes for intracellular hydrogen peroxide detection.  


A metal-based fluorescent probe for H2O2, named MBFh2, releases a highly fluorescent resorufin in the seconds time scale even in the presence of 5 ?M H2O2. The use of MBFh2 enabled the visualization of intracellular H2O2 that was generated after stimulation of the epidermal growth factor. PMID:24000350

Hitomi, Yutaka; Takeyasu, Toshiyuki; Kodera, Masahito



Polyoxometalate-based inorganic-organic hybrid film structure with reversible electroswitchable fluorescence property.  


A novel inorganic-organic hybrid film structure based on polyoxometalate and conventional organic dye has been fabricated, whose fluorescence can be reversibly switched using the electrochromic component to activate or suppress the related fluorescence quenching mechanism upon applying reduction or oxidation potentials of polyoxometalates. PMID:22245959

Jin, Lihua; Fang, Youxing; Hu, Peng; Zhai, Yanling; Wang, Erkang; Dong, Shaojun



Rational Design of a Fluorescent Hydrogen Peroxide Probe Based on the Umbelliferone Fluorophore  

PubMed Central

In this study, we report a novel water-soluble umbelliferone-based fluorescent probe for hydrogen peroxide. This probe shows very large increases (up to 100 fold) in fluorescent intensity upon reaction with hydrogen peroxide, and good selectivity over other reactive oxygen species (ROS).

Du, Lupei; Li, Minyong; Zheng, Shilong; Wang, Binghe



Micro-fluidic-based optical detection platform for characterizing fluorescing objects with integrated wavelength detection  

Microsoft Academic Search

This presentation will give a brief overview on on-the-flow analyte detection based on native fluorescence spectroscopy. This is a very promising approach that does not require specific binding or tagging of the analyte. However, the variety of cells is large compared to the number of basic molecular building blocks. Therefore, the fluorescence spectra of different species are often very similar,

Peter Kiesel; Markus Beck; Michael Bassler; Noble Johnson



Ruthenium(II) complex-based fluorescent sensor for peroxynitrite.  


We have developed a new ruthenium complex, Ru(bpy)(2)[4-(2,2'-bipyridin-4-yloxy)phenol]Cl(2) (RuL), as a fluorescent sensor to detect peroxynitrite (ONOO(-)). The results showed that the addition of ONOO(-) to the aqueous solution of RuL would result in distinct fluorescence quenching at 600nm. RuL exhibits a good selectivity for ONOO(-) over other reactive oxygen species (ROS) and reactive nitrite species (RNS), and the reaction time is less than 1.5s. The sensing mechanism is proposed as the oxidative O-dealkylation reaction. PMID:22554897

Ma, Jingjin; Wu, Jiasheng; Liu, Weimin; Wang, Pengfei; Fan, Zhiyuan



Fluorescence-based microendoscopes for breast cancer ductoscopy  

NASA Astrophysics Data System (ADS)

Recently microendoscopes are being developed as a tool to detection cancer or pre-cancerous lesions in the milk ducts of the human breast. The microendoscope can be inserted into the duct through the nipple. Integration of fluorescence spectroscopy into microendoscopy can provide an improved platform for real-time cancer detection followed by immediate intervention. Typically, the optical fibers employed by existing microendoscope systems transmit in the 450 to 900 nm range. A prototype system combining fluorescence spectroscopy with visible imaging by microendoscopy is described and preliminary measurements on ex vivo human breast tissues are presented. Image resolution and distortion are discussed.

Zeylikovich, Iosif; Tang, Guichen C.; Katz, A.; Budansky, Yury; Alfano, R. R.



A fluorescence polarization based assay for glucose sensing  

NASA Astrophysics Data System (ADS)

A fluorescence polarization (FP) assay was developed to determine concentrations of glucose using concanavalin A (ConA) and fluorescently-labeled dextran. Predictive FP responses to glucose were elicited for different assay configurations using mathematical modeling and displayed herein. Using 4 kDa FITC-dextran, we predicted a change of 0.120 P units from 0 mg/dL glucose to 500 mg/dL. This shows the potential that a homogenous, reproducible FP assay can be engineered to measure glucose concentrations using tetrameric ConA and 4k kDa FITC-dextran.

Cummins, Brian M.; Coté, Gerard L.



SIL-based confocal fluorescence microscope for investigating individual nanostructures  

NASA Astrophysics Data System (ADS)

We developed a fluorescence confocal microscope equipped with a hemispherical solid immersion lens (SIL) and apply it to study the optical properties of light-harvesting complexes. We demonstrate that the collection efficiency of the SIL-equipped microscope is significantly improved, as is the spatial resolution, which reaches 600 nm. This experimental setup is suitable for detailed studies of physical phenomena in hybrid nanostructures. In particular, we compare the results of fluorescence intensity measurements for a light-harvesting peridinin-chlorophyll-protein (PCP) complex with and without the SIL.

Krajnik, Bartosz; Schulte, Tim; Pi?tkowski, Dawid; Czechowski, Nikodem; Hofmann, Eckhard; Mackowski, Sebastian



An improved bimolecular fluorescence complementation tool based on superfolder green fluorescent protein.  


Bimolecular fluorescence complementation (BiFC) has been widely used in the analysis of protein-protein interactions (PPIs) in recent years. There are many notable advantages of BiFC such as convenience and direct visualization of PPI in cells. However, BiFC has one common limitation: the separated non-fluorescent fragments can be spontaneously self-assembled into an intact protein, which leads to false-positive results. In this study, a pair of complementary fragments (sfGFPN and sfGFPC) was constructed by splitting superfolder GFP (sfGFP) between the 214 and 215 amino acid residue, and sfGFPC was mutated by site-directed gene mutagenesis to decrease the signal of negative control. Our results showed that mutations in sfGFPC (sfGFPC(m12)) can effectively decrease the signal of negative control. Thus, we provide an improved BiFC tool for the analysis of PPI. Further, since the self-assembly problem is a common shortcoming for application of BiFC, our research provides a feasible strategy for other BiFC candidate proteins with the same problem. PMID:21273204

Zhou, Jun; Lin, Jian; Zhou, Cuihong; Deng, Xiaoyan; Xia, Bin



Neural bases of categorization of simple speech and nonspeech sounds.  


Categorization is fundamental to our perception and understanding of the environment. However, little is known about the neural bases underlying the categorization of sounds. Using human functional magnetic resonance imaging (fMRI) we compared the brain responses to a category discrimination task with an auditory discrimination task using identical sets of sounds. Our stimuli differed along two dimensions: a speech-nonspeech dimension and a fast-slow temporal dynamics dimension. All stimuli activated regions in the primary and nonprimary auditory cortices in the temporal cortex and in the parietal and frontal cortices for the two tasks. When comparing the activation patterns for the category discrimination task to those for the auditory discrimination task, the results show that a core group of regions beyond the auditory cortices, including inferior and middle frontal gyri, dorsomedial frontal gyrus, and intraparietal sulcus, were preferentially activated for familiar speech categories and for novel nonspeech categories. These regions have been shown to play a role in working memory tasks by a number of studies. Additionally, the categorization of nonspeech sounds activated left middle frontal gyrus and right parietal cortex to a greater extent than did the categorization of speech sounds. Processing the temporal aspects of the stimuli had a greater impact on the left lateralization of the categorization network than did other factors, particularly in the inferior frontal gyrus, suggesting that there is no inherent left hemisphere advantage in the categorical processing of speech stimuli, or for the categorization task itself. PMID:16281285

Husain, Fatima T; Fromm, Stephen J; Pursley, Randall H; Hosey, Lara A; Braun, Allen R; Horwitz, Barry



Cyclam-based "clickates": homogeneous and heterogeneous fluorescent sensors for Zn(II).  


In an effort to improve upon the recently reported cyclam based zinc sensor 1, the "click"-generated 1,8-disubstituted analogue 2 has been prepared. The ligand shows a 2-fold increase in its fluorescence emission compared to 1 exclusively in the presence of Zn(II) that is typical of switch-on PET fluorescent sensors. Single crystal X-ray diffraction of complexes of model ligand 10 reveals that the configuration adopted by the macrocyclic framework is extremely sensitive to the metal ion to which it coordinates. For Zn(II), Mg(II), and Li(I) the metal ions adopt an octahedral geometry with a trans III configuration of the cyclam ring. In contrast for Ni(II) the ligand adopts the rare cis V configuration, while for Cu(II) a clear preference for five-coordinate geometry is displayed with a trans I configuration of the macrocyclic ring being observed in two essentially isostructural compounds prepared via different routes. The ligand displays an increased selectivity for Zn(II) compared to 1 in the majority of cases with excellent selectivity upheld over Na(I), Mg(II), Ca(II), Mn(II), Ni(II), Co(II), and Fe(III). In contrast for Cu(II) and Hg(II) little improvement was observed for 2 compared to 1 and for Cd(II) the selectivity of the new ligand was inferior. In the light of these findings and the slower response times for ligand 2, our original "click"-generated cyclam sensor system 1 was employed in a proof of concept study to prepare a heterogeneous sol-gel based material which retains its PET response to Zn(II). The versatile nature of the sol-gel process importantly allows the simple preparation of a variety of nanostructured materials displaying high surface area-volume ratio using fabrication methods such as soft lithography, electrospinning, and nanopipetting. PMID:20297799

Tamanini, Emiliano; Flavin, Kevin; Motevalli, Majid; Piperno, Silvia; Gheber, Levi A; Todd, Matthew H; Watkinson, Michael



A new Schiff base fluorescent probe for imaging Cu2+ in living cells.  


A novel probe based on ferrocenyl-1,3,4-thiadiazol-containing Schiff base was synthesized by the reaction of 5-ferrocenyl-1,3,4-thiadiazol-2-amine and 4-(diethylamino)salicylaldehyde, and characterized by IR, NMR, HRMS and X-ray analysis. UV-vis spectral and fluorescence property of the probe were investigated. The probe can be used to colorimetric sensitive and selective fluorescent recognition of Cu(2+) in buffer solution. Moreover, the probe can detect Cu(2+) by electrochemical method. Additionally, the Schiff base was successfully used as a selective and sensitive fluorescent probe for monitoring Cu(2+) ions in living cells. PMID:23666347

Ye, Hui; Ge, Fei; Zhou, Yi-Ming; Liu, Jin-Ting; Zhao, Bao-Xiang



A new Schiff base fluorescent probe for imaging Cu2+ in living cells  

NASA Astrophysics Data System (ADS)

A novel probe based on ferrocenyl-1,3,4-thiadiazol-containing Schiff base was synthesized by the reaction of 5-ferrocenyl-1,3,4-thiadiazol-2-amine and 4-(diethylamino)salicylaldehyde, and characterized by IR, NMR, HRMS and X-ray analysis. UV-vis spectral and fluorescence property of the probe were investigated. The probe can be used to colorimetric sensitive and selective fluorescent recognition of Cu2+ in buffer solution. Moreover, the probe can detect Cu2+ by electrochemical method. Additionally, the Schiff base was successfully used as a selective and sensitive fluorescent probe for monitoring Cu2+ ions in living cells.

Ye, Hui; Ge, Fei; Zhou, Yi-Ming; Liu, Jin-Ting; Zhao, Bao-Xiang



Selective recognition of acetate ion based on fluorescence enhancement chemosensor.  


Fluorescence study of the complexation between uranyl salophen (L) and some common anions in acetonitrile-water (90:10, v/v) solution showed a tendency of L toward acetate ion (AcO-). The fluorescence enhancement of L is attributed to a 1:1 complex formation between L and acetate ion which was utilized as the basis for the selective detection of AcO-. The association constant of the 1:1 complex formation of L-AcO- was calculated as 6.60?×?10(6) . The linear response range of the fluorescent chemosensor covers a AcO- concentration range of 1.6?×?10(-7) to 2.5?×?10(-5) ?mol/L, with a detection limit of 2.5?×?10(-8) ?mol/L. L showed a selective and sensitive fluorescence enhancement response toward acetate ion over I3- , NO3-, CN-, CO3 (2-), Br-, Cl-, F-, H2 PO4- and SO4 (2-) , which was attributed to the higher stability of inorganic complex between acetate and L. PMID:22371380

Hosseini, Morteza; Ganjali, Mohammad Reza; Veismohammadi, Bahareh; Faridbod, Farnoush; Abkenar, Shiva Dehghan; Salavati-Niasari, Masoud



Enhanced fluorescence of isophorone derivatives in DNA based materials  

NASA Astrophysics Data System (ADS)

Photophysical properties studies of three push–pull isophorone derivatives are reported.Two matrices were used: DNA–CTMA complex and PMMA and results compared.Weaker interaction of chromophores with DNA–CTMA is observed.Fluorescence quantum yield is larger in DNA–CTMA matrix.

Massin, Julien; Parola, Stéphane; Andraud, Chantal; Kajzar, François; Rau, Ileana



Development of a Green Fluorescent Protein-Based Laboratory Curriculum  

ERIC Educational Resources Information Center

|A laboratory curriculum has been designed for an undergraduate biochemistry course that focuses on the investigation of the green fluorescent protein (GFP). The sequence of procedures extends from analysis of the DNA sequence through PCR amplification, recombinant plasmid DNA synthesis, bacterial transformation, expression, isolation, and…

Larkin, Patrick D.; Hartberg, Yasha



Caged-fluorescent-dye-based studies of turbulent scalar mixing  

Microsoft Academic Search

The initialization of a flow field with distinct and spatially segregated scalar components represents a significant experimental difficulty. Many theoretical modeling efforts in turbulent mixing, however, seek to describe the temporal evolution of a scalar concentration field that begins with this type of idealized initial conditions experimentally. This technique uses photoactivatable (caged) fluorescence dyes dissolved in the flow medium. Caged

James Guilkey; Kyle Gee; Joseph Klewicki; Patrick R. McMurtry



Simple fiber-optic-based interface to facilitate spectroscopic measurements in supercritical solvents  

SciTech Connect

A new fiber-optic-based interface is described to facilitate fluorescence measurements in supercritical solvents. Steady-state and time-resolved measurements are possible, and assembly/disassembly is significantly easier compared to that required for traditional high-pressure optical cell designs. 9 refs., 6 figs., 1 tab.

Zagrobelny, J.; Ming Li; Run Wang; Betts, T.A.; Bright, F.V. (State Univ. of New York, Buffalo (United States))



A novel, simple and efficient dye laser with low amplified spontaneous emission background for analytical fluorescence and ionization spectroscopy  

SciTech Connect

A new, simple, compact and efficient, grazing- incidence type of dye laser is suggested which has a low level of Amplified Spontaneous Emission. By using a Coumarin dye (LD 5000) pumped with a 20 mJ XeCl excimer laser, and a diffraction grating with 3000 grooves/mm, an efficiency of 11%, a spectral bandwidth of 0.6 cm{sup -1} and a tuning range from 458 to 517 nm have been obtained.

Matveev, Oleg I.; Omenetto, Nicolo' [EC, Joint Research Centre, Environment Institute, 21020 Ispra, Varese (Italy)



Tunable fluorescent pH sensor based on water-soluble perylene tetracarboxylic acid/Fe(3+).  


A novel fluorescent pH sensor with tunable response range was designed based on highly fluorescent 3,4:9,10-perylene tetracarboxylic ammonium, which could coordinate the paramagnetic Fe(3+) ions to turn off its fluorescence and could also release Fe(3+) to turn on the fluorescence again at higher pH. The fluorescent pH sensor was tunable in the presence of different ligands in aqueous solution. PMID:21984374

Xu, Ping; Pan, Cuicui; Zhao, Yingjie; Kong, Xiangxue; Sun, Juanjuan; Xu, Maoyou; Shi, Zhiqiang



Fluorescence resonance energy transfer-based molecular logic circuit using a DNA scaffold  

NASA Astrophysics Data System (ADS)

This paper presents a method of information processing using biomolecular input signals and fluorescence resonance energy transfer (FRET) signaling constructed on a DNA scaffold. Logic operations are achieved by encoding molecular inputs into an arrangement of fluorescence dyes using simple DNA reactions and by evaluating a logic expression using local photonic signaling that is much faster than DNA reactions. Experimental results verify the operation of a complete set of Boolean logic functions (AND, OR, NOT) and combinational logic operations using a FRET-signal cascade.

Nishimura, Takahiro; Ogura, Yusuke; Tanida, Jun



Time-domain imaging with quench-based fluorescent contrast agents  

NASA Astrophysics Data System (ADS)

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

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



A Rapid Fluorescence-Based Assay for Classification of iNKT Cell Activating Glycolipids  

PubMed Central

Structural variants of ?-galactosylceramide (?GC) that activate invariant natural killer T cells (iNKT cells) are being developed as potential immunomodulatory agents for a variety of applications. Identification of specific forms of these glycolipids that bias responses to favor production of proinflammatory vs anti-inflammatory cytokines is central to current efforts, but this goal has been hampered by the lack of in vitro screening assays that reliably predict the in vivo biological activity of these compounds. Here we describe a fluorescence-based assay to identify functionally distinct ?GC analogues. Our assay is based on recent findings showing that presentation of glycolipid antigens by CD1d molecules localized to plasma membrane detergent-resistant microdomains (lipid rafts) is correlated with induction of interferon-? secretion and Th1-biased cytokine responses. Using an assay that measures lipid raft residency of CD1d molecules loaded with ?GC, we screened a library of ?200 synthetic ?GC analogues and identified 19 agonists with potential Th1-biasing activity. Analysis of a subset of these novel candidate Th1 type agonists in vivo in mice confirmed their ability to induce systemic cytokine responses consistent with a Th1 type bias. These results demonstrate the predictive value of this novel in vitro assay for assessing the in vivo functionality of glycolipid agonists and provide the basis for a relatively simple high-throughput assay for identification and functional classification of iNKT cell activating glycolipids.



A seminaphthofluorescein-based fluorescent chemodosimeter for the highly selective detection of cysteine†  

PubMed Central

A fluorescent chemodosimeter for cysteine detection was developed based on a tandem conjugate addition and intramolecular cyclization reaction. The method exhibited an excellent selectivity for cysteine over other biothiols such as homocysteine and glutathione.

Guo, Yixing



Fluorescence-based video profile beam diagnostics: Theory and experience  

SciTech Connect

Inelastic collisions between accelerated particles and residual gas in the accelerator vessel can cause the residual gas to fluoresce. The gas fluorescence intensity is proportional to the current density of the particle beam. This process provides the foundation for a video diagnostic system to measure the profile and position of accelerated particle beams. This, in fact, has proven to be a useful diagnostic at several installations. This paper describes the light production process resulting from beam -- residual gas interactions and gives formulas for estimating the beam radiance for various conditions. Ground Test Accelerator (GTA) radiance calculations will be used as an example. In addition, measurement experiences with the GTA video diagnostics system will be discussed.

Sandoval, D.; Gilpatrick, D.; Shinas, M.; Garcia, R.; Yuan, V.; Zander, M.



Fluorescence-based video profile beam diagnostics: Theory and experience  

SciTech Connect

Inelastic collisions between accelerated particles and residual gas in the accelerator vessel can cause the residual gas to fluoresce. The gas fluorescence intensity is proportional to the current density of the particle beam. This process provides the foundation for a video diagnostics system to measure the profile and position of accelerated particle beams. This, in fact, has proven to be a useful diagnostic at several installations. This paper describes the light production process resulting from beam-residual gas interactions and gives formulas for estimating the beam radiance for various conditions. Ground Test Accelerator (GTA) radiance calculations will be used as an example. In addition, measurement experiences with the GTA video diagnostics system will be discussed.

Sandoval, D.P.; Garcia, R.C.; Gilpatrick, J.D.; Shinas, M.A.; Wright, R.; Yuan, V.; Zander, M.E. (Los Alamos National Laboratory, Los Alamos, New Mexico 87545 (United States))



Quantum dots (QDs) based fluorescent sensor for the selective determination of nimesulide.  


Fluorescent PET (Photoinduced Electron Transfer) has been of particular growth in recent times. A novel PET based fluorescent sensor using unmodified CdSe quantum dots (QDs) has been developed for the trace determination of Nimesulide (NIM). The sensor is based on the selective fluorescence quenching of quantum dots by NIM in presence of other NSAIDs and is found that intensity of quenching is linearly related to NIM concentration in the range 8.2 × 10(-7) - 4.01 × 10(-5) M. The mechanism of interaction is discussed. Finally, the potential application of the proposed method for the trace determination of NIM in pharmaceutical formulation is demonstrated. PMID:23397489

Thomas, Divya; Lonappan, Laina; Rajith, Leena; Cyriac, Soumya T; Kumar, Krishnapillai Girish



Fluorescent-based chemical sensor for organophosphate detection  

Microsoft Academic Search

We present a new optical sensor for the detection of organophosphates by incorporating fluorescent indicator dye into sol-gel material. We used different configurations of immobilization matrices such as thin film and spherical nanoparticles. The sensor thin films were prepared by using acid-catalyzed sol-gel process and the spherical nanoparticles by modified Stöber method. The effects of configuration matrices on the sensor's

Spela Korent Urek; Aleksandra Lobnik



MEMS scanner based handheld fluorescent hyperspectral imaging system  

Microsoft Academic Search

We demonstrate a hand-held hyperspectral fluorescence imaging microsystem, where multiple narrow-band spectral images can be captured simultaneously across the area under examination. In addition to the advantages of combined functional imaging and spectroscopy, the system demonstrates the fast scanning over large field-of-view (FOV) provided by a CMOS compatible 2-axis microelectromechanical system (MEMS) scanning mirror in the probe. Spectral information from

Sheldon Bish; Youmin Wang; James W. Tunnell; Xiaojing Zhang



Submicrometer fluorescence microprobe based on Bragg-Fresnel optics  

Microsoft Academic Search

An x-ray fluorescence microprobe with linear Bragg-Fresnel lens was tested at the European Synchrotron Radiation Facility Microfocus beamline. A focal line with a width of 0.8 ?m was observed using the knife-edge technique at a wavelength of 0.85 A? (14.6 eV). Wiener filtering with the Fast Fourier Transformation was applied to solve a convolution equation to evaluate the size of

S. M. Kuznetsov; I. I. Snigireva; A. A. Snigirev; P. Engström; C. Riekel



Quantitative and qualitative 5-aminolevulinic acid-induced protoporphyrin IX fluorescence in skull base meningiomas  

PubMed Central

Object Complete resection of skull base meningiomas provides patients with the best chance for a cure; however, surgery is frequently difficult given the proximity of lesions to vital structures, such as cranial nerves, major vessels, and venous sinuses. Accurate discrimination between tumor and normal tissue is crucial for optimal tumor resection. Qualitative assessment of protoporphyrin IX (PpIX) fluorescence following the exogenous administration of 5-aminolevulinic acid (ALA) has demonstrated utility in malignant glioma resection but limited use in meningiomas. Here the authors demonstrate the use of ALA-induced PpIX fluorescence guidance in resecting a skull base meningioma and elaborate on the advantages and disadvantages provided by both quantitative and qualitative fluorescence methodologies in skull base meningioma resection. Methods A 52-year-old patient with a sphenoid wing WHO Grade I meningioma underwent tumor resection as part of an institutional review board–approved prospective study of fluorescence-guided resection. A surgical microscope modified for fluorescence imaging was used for the qualitative assessment of visible fluorescence, and an intraoperative probe for in situ fluorescence detection was utilized for quantitative measurements of PpIX. The authors assessed the detection capabilities of both the qualitative and quantitative fluorescence approaches. Results The patient harboring a sphenoid wing meningioma with intraorbital extension underwent radical resection of the tumor with both visibly and nonvisibly fluorescent regions. The patient underwent a complete resection without any complications. Some areas of the tumor demonstrated visible fluorescence. The quantitative probe detected neoplastic tissue better than the qualitative modified surgical microscope. The intraoperative probe was particularly useful in areas that did not reveal visible fluorescence, and tissue from these areas was confirmed as tumor following histopathological analysis. Conclusions Fluorescence-guided resection may be a useful adjunct in the resection of skull base meningiomas. The use of a quantitative intraoperative probe to detect PpIX concentration allows more accurate determination of neoplastic tissue in meningiomas than visible fluorescence and is readily applicable in areas, such as the skull base, where complete resection is critical but difficult because of the vital structures surrounding the pathology.

Bekelis, Kimon; Valdes, Pablo A.; Erkmen, Kadir; Leblond, Frederic; Kim, Anthony; Wilson, Brian C.; Harris, Brent T.; Paulsen, Keith D.; Roberts, David W.



Analysis of total aerobic viable counts in samples of raw meat using fluorescence-based probe and oxygen consumption assay  

Microsoft Academic Search

A simple test for determining total aerobic viable counts in raw meat is presented. Homogenates of meat samples are prepared in full PBW medium, dispensed in the wells of 96-well plate together with the oxygen-sensing probe, Redlight, covered with oil and monitored on a fluorescent reader at 30°C. The probe produces characteristic sigmoidal profiles of fluorescence reflecting depletion of sample

Fiach O’Mahony; Rebecca A. Green; Chris Baylis; Richard Fernandes; Dmitri B. Papkovsky



Specific fluorescent bands on chromosomes produced by acridine orange after prestaining with base specific non-fluorescent DNA ligands  

Microsoft Academic Search

Metaphase chromosomes stained with acridine orange exhibit uniform yellow-green fluorescence. Chromosome preparations treated with the non-fluorescent A-T specific antibiotic distamycin A prior to acridine orange staining exhibit longitudinal fluorescent banding patterns similar to those produced by a number of fluorescent R-band techniques. Similarly, chromosome preparations treated with the non-fluorescent G-C specific antibiotic actinomycin D followed by acridine orange staining exhibit

C. C. Lin; K. F. Jorgenson; J. H. van de Sande



Molecular dynamics study of the structure and performance of simple and double bases propellants.  


To investigate the structure and performance of simple and double bases propellants, the nitrocellulose (NC), nitroglycerin (NG), and double mixed system (NC+NG) have been simulated by using the molecular dynamics (MD) method with the COMPASS force field. The interactions between NC and NG have been analyzed by means of pair correlation functions. The mechanical properties of the three model systems, i.e. elastic coefficients, modulus, Cauchy pressure, and Poisson's ratio, etc., have been obtained. It is found that the rigidity, ductibility, and tenacity of the double bases propellants (NC+NG) are stronger than those of simple base propellants (NC), which attributes to the effect of NG and the strong interactions between NC and NG. The detonation properties of the three systems have also been calculated and the results show that compared with the simple base propellant (NC), the detonation heat and detonation velocity of the double base propellants (NC+NG) are increased. PMID:18243539

Ma, Xiufang; Zhu, Weihua; Xiao, Jijun; Xiao, Heming



Droplet temperature measurement based on 2-color laser-induced exciplex fluorescence  

NASA Astrophysics Data System (ADS)

Measurements of liquid phase temperature distributions in liquid-vapor co-existing conditions (such as in evaporating sprays) are important to understand the physics of droplet evaporation. The techniques based on laser-induced fluorescence are not suitable for evaporating case since both liquid and vapor phases emit fluorescence with the same wavelength. In this study, the fluorescence from liquid and vapor phases was separated by use of laser-induced exciplex fluorescence (LIEF) technique. Two fluorescence bands from the liquid phase fluorescence spectra were detected simultaneously, and their intensity ratio was correlated to the liquid phase temperature. For the LIEF imaging system, FB-DEMA- n-hexane was selected as it was a typical LIEF system for the vapor concentration diagnostic, and thus easily to be extended to a simultaneous diagnostic on the vapor concentration and the droplet temperature. The fluorescence spectra were obtained in the temperature range from 303 to 423 K. The effects of liquid temperature, liquid pressure, dopant concentration and laser energy on the temperature measurement were investigated. The results show a good linear relationship between the fluorescence ratio and the temperature function. Increasing the dopant concentration can raise the signal-to-noise ratio but deteriorate temperature sensitivity. The optimal range of the dopant concentration was found between 0.1 % and 0.5 %. After calibration, the technique was applied to a monosized droplet stream, and the measurement results demonstrated excellent measurement accuracy with error below 1 % in the range of 303-423 K.

Zhang, Yuyin; Zhang, Gaoming; Xu, Min; Wang, Jianxin



Fabrication, optimization and application of a dip-probe fluorescence spectrometer based on white-light excitation fluorescence  

NASA Astrophysics Data System (ADS)

A dip-probe adapter was designed for easy analysis of complex multifluorophoric systems. The newly fabricated fiber optic compatible spectrometer design with dip-probe adapter could reach and analyze fluorophores kept at a distance from the analyzer. The new design and its parameters were optimized using standard fluorophores to get an optimal spectral response. This white-light excitation fluorescence (WLEF) based spectrometer could analyze any fluorophoric system rapidly without changing the instrument configuration. Fuel-oil blends such as petrol-2T oil and petrol-4T oil were precisely quantified with high accuracy (0.3% error) using a WLEF-multivariate analysis combination.

Prakash, John; Mishra, A. K.



Determination of L-Argininamide Based on Water-Soluble Fluorescent Conjugated Polymer-Aptamer  

PubMed Central

Water-soluble fluorescent conjugated polymer is a promising material which could be used as an optical platform in highly sensitive molecular sensors. In this paper, a simple label-free DNA sensor, which consisted of a poly(3-alkoxy-4-methylthiophene) and an aptamer, was used to detect L-argininamide (L-Arm). Due to the specific binding reaction between L-Arm and its aptamer, the proposed method can easily determinate the L-Arm through the recovery of fluorescence without any modification. Other ions or similar molecules had little effect on the detection. Moreover, there was a linear relationship between fluorescence intensity and the concentration of L-Arm. The detection limit of L-Arm was as low as 4.7?nM.

Guan, Hongliang; He, Zhike



Detection of enzyme activity in orthotopic murine breast cancer by fluorescence lifetime imaging using a fluorescence resonance energy transfer-based molecular probe  

NASA Astrophysics Data System (ADS)

Cancer-related enzyme activity can be detected noninvasively using activatable fluorescent molecular probes. In contrast to ``always-on'' fluorescent molecular probes, activatable probes are relatively nonfluorescent at the time of administration due to intramolecular fluorescence resonance energy transfer (FRET). Enzyme-mediated hydrolysis of peptide linkers results in reduced FRET and increase of fluorescence yield. Separation of signal from active and inactive probe can be difficult with conventional intensity-based fluorescence imaging. Fluorescence lifetime (FLT) measurement is an alternative method to detect changes in FRET. Thus, we investigate FLT imaging for in vivo detection of FRET-based molecular probe activation in an orthotopic breast cancer model. Indeed, the measured FLT of the enzyme-activatable molecular probe increases from 0.62 ns just after injection to 0.78 ns in tumor tissue after 4 h. A significant increase in FLT is not observed for an always-on targeted molecular probe with the same fluorescent reporter. These results show that FLT contrast is a powerful addition to preclinical imaging because it can report molecular activity in vivo due to changes in FRET. Fluorescence lifetime imaging exploits unique characteristics of fluorescent molecular probes that can be further translated into clinical applications, including noninvasive detection of cancer-related enzyme activity.

Solomon, Metasebya; Guo, Kevin; Sudlow, Gail P.; Berezin, Mikhail Y.; Edwards, W. Barry; Achilefu, Samuel; Akers, Walter J.



Autoregressive-model-based fluorescence-lifetime measurements by phase-modulation fluorometry using a pulsed-excitation light source and a high-gain photomultiplier tube.  


We propose a novel method for measuring fluorescence lifetimes by use of a pulsed-excitation light source and an ordinary or a high-gain photomultiplier tube (PMT) with a high-load resistor. In order to obtain the values of fluorescence lifetimes, we adopt a normal data-processing procedure used in phase-modulation fluorometry. We apply an autoregressive (AR)-model-based data-analysis technique to fluorescence- and reference-response time-series data obtained from the PMT in order to derive plural values of phase differences at a repetition frequency of the pulsed-excitation light source and its harmonic ones. The connection of the high-load resistor enhances sensitivity in signal detection in a certain condition. Introduction of the AR-model-based data-analysis technique improves precision in estimating the values of fluorescence lifetimes. Depending on the value of the load resistor and that of the repetition frequency, plural values of fluorescence lifetimes are obtained at one time by utilizing the phase information of harmonic frequencies. Because the proposed measurement system is simple to construct, it might be effective when we need to know approximate values of fluorescence lifetimes readily, such as in the field of biochemistry for a screening purpose. PMID:19891834

Iwata, Tetsuo; Ito, Ritsuki; Mizutani, Yasuhiro; Araki, Tsutomu



A new Schiff base based on vanillin and naphthalimide as a fluorescent probe for Ag+ in aqueous solution  

NASA Astrophysics Data System (ADS)

A new Schiff base based on vanillin and naphthalimide was designed and synthesized as fluorescent probe. The probe showed high selectivity for Ag+ over other metal ions such as Pb2+, Na+, K+, Cd2+, Ba2+, Cr3+, Zn2+, Cu2+, Ni2+, Ca2+, Al3+ and Mg2+ in aqueous solution. A new fluorescence emission was observed at 682 nm in the presence of Ag+ ion. The fluorescence intensity quenched with increasing the concentration of Ag+ at 682 nm. The method of job's plot confirmed the 1:2 complex between Ag+ and probe, and the mechanism was proposed.

Zhou, Yanmei; Zhou, Hua; Ma, Tongsen; Zhang, Junli; Niu, Jingyang



A fluorescence-based assay for fatty acid amide hydrolase compatible with high-throughput screening  

Microsoft Academic Search

A novel fluorescent assay to continuously monitor fatty acid amide hydrolase (FAAH) activity that is simple, sensitive, and amenable to high-throughput screening (HTS) of compound libraries is described in this article. Stable Chinese hamster ovary (CHO) cell lines expressing either human FAAH or an inactive mutant, FAAH-S241A, were established. Arachidonyl 7-amino, 4-methyl coumarin amide (AAMCA), a novel fluorogenic substrate for

Manjunath K. Ramarao; Elizabeth A. Murphy; Marina W. H. Shen; Yuren Wang; Kristen N. Bushell; Nelson Huang; Ning Pan; Cara Williams; James D. Clark



Enantioselective Fluorescent Recognition of Chiral Acids by Cyclohexane-1,2-diamine-Based Bisbinaphthyl Molecules  

PubMed Central

The cyclohexane-1,2-diamine-based bisbinaphthyl macrocycles (S)-/(R)-5 and their cyclic and acyclic analogs are synthesized. The interactions of these compounds with various chiral acids are studied. Compounds (S)-/(R)-5 exhibit highly enantioselective fluorescent responses and high fluorescent sensitivity toward ?-hydroxycarboxylic acids and N-protected amino acids. Among these interactions, (S)-mandelic acid (10?3 M) led to over 20 fold fluorescence enhancement of (S)-5 (1.0 × 10?5 M in benzene/0.05% DME) at the monomer emission and (S)-hexahydromandelic acid (10?3 M) led to over 80 fold fluorescence enhancement. These results demonstrate that (S)-5 is useful as an enantioselective fluorescent sensor for the recognition of the chiral acids. On the basis of the study of the structures of (S)-5 and the previously reported 1,2-diphenylethylenediamine-based bisbinaphthyl macrocycle (S)-4, the large fluorescence enhancement of (S)-5 with achirality-matched ?-hydroxycarboxylic acid is attributed to the formation of a structurally rigidified host-guest complex and the further interaction of this complex with the acid to suppress the photo-induced electron transfer fluorescent quenching caused by the nitrogens in (S)-5.

Li, Zi-Bo; Lin, Jing; Sabat, Michal; Hyacinth, Marilise; Pu, Lin



Prospects for fluorescence based imaging/visualization of hydrodynamic systems on the National Ignition Facility  

SciTech Connect

The next generation of large, high power lasers, such as the National Ignition Facility (NIF) [1] in the United States, Laser Mega Joule [2] in France or Helen Successor [3] in the United Kingdom offer the prospect of x-ray fluorescence based diagnosis of hydrodynamic experiments The x-ray fluorescence could be pumped by at least two techniques One technique is to use a sizable fraction of these facilities` high power to efficiently make multi-kilovolt x-rays which, in turn, causes dopants placed in experimental packages to fluoresce We call this ``externally pumped x-ray fluorescence`` The second technique is to use the sizable multi-kilovolt photon background that we expect to be present in many hohlraum based experiments, while the driving laser is on, to pump x-ray fluorescence The fluorescing medium could be a dopant in an experimental package or, possibly, a relatively thick slab of material in the hohlraum wall which could serve as a backlighter We call this ``hohlraum hot-corona pumped fluorescence``.

Suter, L. J., LLNL



An in-situ fluorescence-based optical extensometry system for imaging mechanically loaded bone  

PubMed Central

The application and quantification of well-controlled tissue strains is required for quantitative investigations into mechanisms of tissue adaptation within the musculoskeletal system. Although there are many commercial and custom extensometry systems for large biological samples, integrated loading/strain measurement for small samples are not as readily available. Advanced imaging modules such as laser scanning microscopy provide in situ, minimally invasive tools to probe cellular and molecular processes with high spatiotemporal resolution. Currently, there is a need to devise loading/strain measurement systems that can be integrated with such advanced imaging modules. In this paper we describe the development and validation of a fluorescence-based, optical extensometry system directly integrated within a confocal microscopy platform. This system will allow in-situ measurement of surface strain that is compatible with the direct imaging of cellular processes within small bone samples. This optical extensometry system is capable of accurately and reproducibly measuring physiologically relevant surface strains (200-3000 micro-strain) in beams machined from various well-characterized materials, including bovine femoral cortex, and in intact murine tibia. This simple optical extensometry system provides a powerful tool to further our investigation of the relationships between mechanical loading, fluid and solute transport, and mechanosensation within the musculoskeletal system.

Price, Christopher; Li, Wen; Novotny, John E.; Wang, Liyun



An in-situ fluorescence-based optical extensometry system for imaging mechanically loaded bone.  


The application and quantification of well-controlled tissue strains is required for investigations into mechanisms of tissue adaptation within the musculoskeletal system. Although many commercial and custom extensometry systems exist for large biological samples, integrated loading/strain measurement for small samples is not as readily available. Advanced imaging modules such as laser scanning microscopy provide in situ, minimally invasive tools to probe cellular and molecular processes with high spatiotemporal resolution. Currently, a need exists to devise loading/strain measurement systems that can be integrated with such advanced imaging modules. We describe the development and validation of a fluorescence-based, optical extensometry system directly integrated within a confocal microscopy platform. This system allows in situ measurement of surface strain and is compatible with the direct imaging of cellular processes within small bone samples. This optical extensometry system can accurately and reproducibly measure physiologically relevant surface strains (200 to 3000 microstrain) in beams machined from various well-characterized materials, including bovine femoral cortex, and in intact murine tibia. This simple system provides a powerful tool to further our investigation of the relationships between mechanical loading, fluid and solute transport, and mechanosensation within the musculoskeletal system. PMID:20041487

Price, Christopher; Li, Wen; Novotny, John E; Wang, Liyun



Cell trafficking of carbon nanotubes based on fluorescence detection.  


Cell trafficking of carbon nanotubes (CNTs) is an area of scientific inquiry that has great implications in medicine, biosensing, and environmental science and engineering. The essence of this inquiry resides in the interaction of carbon nanostructures and cell membranes, regulated by the laws of molecular cell biology and the physiochemical properties of the nanostructures. Of equal importance to this inquiry is a description of cellular responses to the integration of man-made materials; yet, how cellular responses may invoke whole-organism level reaction remains unclear. In this chapter, we show three experimental studies, which may be beneficial to obtaining such an understanding. Among the reservoir of methodologies, which have proved of merit, we focus our attention on fluorescence microscopy, one of the most powerful and yet least invasive means of probing nanoparticles in biological systems. Especially, we present the method of fluorescence energy transfer induced between a lysophospholipid molecule and a single-walled CNT upon cellular uptake, and describe coating nanotubes with RNA and suspending fullerenes with phenolic acids for facilitating their translocation across cell membranes and shuttling between cell organelles. Finally, we comment on the perspective of using molecular simulations for facilitating and guiding such experiments. PMID:20422387

Lamm, Monica H; Ke, Pu Chun



Boronate-Based Fluorescent Probes for Imaging Cellular Hydrogen Peroxide  

PubMed Central

The syntheses, properties, and biological applications of the Peroxysensor family, a new class of fluorescent probes for hydrogen peroxide, are presented. These reagents utilize a boronate deprotection mechanism to provide high selectivity and optical dynamic range for detecting H2O2 in aqueous solution over similar reactive oxygen species (ROS) including superoxide, nitric oxide, tert-butyl hydroperoxide, hypochlorite, singlet oxygen, ozone, and hydroxyl radical. Peroxyresorufin-1 (PR1), Peroxyfluor-1 (PF1), and Peroxyxanthone-1 (PX1) are first-generation probes that respond to H2O2 by an increase in red, green, and blue fluorescence, respectively. The boronate dyes are cell-permeable and can detect micromolar changes in H2O2 concentrations in living cells, including hippocampal neurons, using confocal microscopy and two-photon microscopy. The unique combination of ROS selectivity, membrane permeability, and a range of available excitation/emission colors establishes the potential value of PR1, PF1, PX1, and related probes for interrogating the physiology and pathology of cellular H2O2.

Miller, Evan W.; Albers, Aaron E.; Chang, Christopher J.; Pralle, Arnd; Isacoff, Ehud Y.



Computed tomography-based spectral imaging for fluorescence microscopy.  

PubMed Central

The computed tomography imaging spectrometer (CTIS) is a non-scanning instrument capable of simultaneously acquiring full spectral information (450-750 nm) from every position element within its field of view (75 microm x 75 microm). The current spatial and spectral sampling intervals of the spectrometer are 1.0 microm and 10 nm, respectively. This level of resolution is adequate to resolve signal responses from multiple fluorescence probes located within individual cells or different locations within the same cell. Spectral imaging results are presented from the CTIS combined with a commercial inverted fluorescence microscope. Results demonstrate the capability of the CTIS to monitor the spatiotemporal evolution of pH in rat insulinoma cells loaded with SNARF-1. The ability to analyze full spectral information for two-dimensional (x, y) images allows precise evaluation of heterogeneous physiological responses within cell populations. Due to low signal levels, integration times up to 2 s were required. However, reasonable modifications to the instrument design will provide higher system transmission efficiency with increased temporal and spatial resolution. Specifically, a custom optical design including the use of a larger format detector array is under development for a second-generation system.

Ford, B K; Volin, C E; Murphy, S M; Lynch, R M; Descour, M R



Temperature-modulated fluorescence tomography based on both concentration and lifetime contrast  

PubMed Central

Abstract. It is challenging to image fluorescence objects with high spatial resolution in a highly scattering medium. Recently reported temperature-sensitive indocyanine green-loaded pluronic nanocapsules can potentially alleviate this problem. Here we demonstrate a frequency-domain temperature-modulated fluorescence tomography system that could acquire images at high intensity-focused ultrasound resolution with use of these nanocapsules. The system is experimentally verified with a phantom study, where a 3-mm fluorescence object embedded 2 cm deep in a turbid medium is successfully recovered based on both intensity and lifetime contrast.

Lin, Yuting; Kwong, Tiffany C.; Bolisay, Linden; Gulsen, Gultekin



A bio-aerosol detection technique based on tryptophan intrinsic fluorescence measurement  

NASA Astrophysics Data System (ADS)

Based on the measurement of intrinsic fluorescence, a set of bio-aerosol including virus aerosols detection instrument is developed, with which a method of calibration is proposed using tryptophan as the target. The experimental results show a good linear relationship between the fluorescence voltage of the instrument and the concentration of the tryptophan aerosol. An excellent correlation (R2>=0.99) with the sensitivity of 4000PPL is obtained. The research demonstrates the reliability of the bio-aerosol detection by measuring the content of tryptophan. Further more the feasibility of prejudgment to the species of bio-aerosol particles with the multi-channel fluorescence detection technology is discussed.

Cai, Shuyao; Zhang, Pei; Zhu, Linglin; Zhao, Yongkai; Huang, Huijie



Quinoline-Based Fluorescent Probe for Ratiometric Detection of Lysosomal pH.  


A new pH-responsive fluorescent probe has been reported based on protonation-activable resonance charge transfer. In aqueous solution, probe PQ-Lyso exhibits ratiometric detection of pH changes with a large hypsochromic shift of 76 nm and remarkable changes in the fluorescence intensity ratio (R = F494 nm/F570 nm, R/R0 = 105). Furthermore, PQ-Lyso, which is localized to lysosomes in living cells, can calibrate lysosomal pH using fluorescence ratiometry. PMID:24040756

Li, Guoping; Zhu, Dongjian; Xue, Lin; Jiang, Hua



A simple agent-based social impact theory model of student STEM selection  

Microsoft Academic Search

There is a growing body of knowledge describing the economic and social challenge faced by the United States because of the small (14%) and decreasing number of students pursuing Science, Technology, Engineering, and Mathematics (STEM) majors. We propose a simple two-period, agent-based simulation based on social impact theory to predict the % yield of STEM majors. The model indicates that

Theodore T. Allen; Nixon Davis



Simple 4Bit Processor Based On Quantum-Dot Cellular Automata (QCA)  

Microsoft Academic Search

We describe the design and layout of a simple 4-bit processor based on quantum dot cellular automata (QCA) using the QCADesigner design tool. The processor design is based on an accumulator architecture which reduces the required hardware complexity and allows for reasonable simulation times. Our aim is to provide evidence that QCA has potential applications in future computers provided that

Konrad Walus; Mike Mazur; Gabriel Schulhof; Graham A. Jullien



Development of label-free molecular beacons based on abasic site-binding fluorescence molecules.  


Here we report on a class of label-free molecular beacons (MBs) based on a non-covalent interaction with abasic site (AP site)-binding fluorescence molecules. In contrast to conventional MBs that require the chemical labelling with fluorophores and quenchers, our MB simply contains the AP site in the stem moiety, so that a small molecule specifically binds to the AP site. This binding event is accompanied by a significant quenching of its fluorescence, and thus a closed state of the AP site-containing MB (APMB) shows almost no fluorescence. Upon hybridization with a complementary DNA, APMB undergoes a conformational change to take an open state, resulting in an effective fluorescence enhancement due to a release of the molecule from the AP site. These sensing functions of APMB are discussed with a view towards further development of gene detection chemistry based on DNA-binding small molecules. PMID:18776283

Sato, Yusuke; Nishizawa, Seiichi; Teramae, Norio



[Subcutaneous microdialysis: a simple technique for monitoring the extracellular biochemical environment. Combination with capillary electrophoresis and laser-induced fluorescence detection].  


Microdialysis is a simple technique that allows monitoring endogenous or exogenous substances in any extracellular compartment. It has many useful experimental and clinical applications. The sampling of the extracellular fluid of the subcutaneous compartment is especially useful for metabolic evaluation in critically ill patients, pharmacokinetic studies and blood glucose monitoring. We built a subcutaneous microdialysis probe, with a cellulose hollow fiber (13,000 molecular weight cut off, 200 microns outside diameter) glued to stainless steel tubing (26 ga. outside diameter). It was implanted in the subcutaneous tissue of a critically ill child or anesthetized mice to obtain amino acids patterns by means of capillary electrophoresis with laser induced fluorescence detection (CE-LIFD). The probe was also implanted in ambulatory volunteers to monitor glucose. The results confirmed that subcutaneous microdialysis is a very simple, inexpensive and not aggressive method with advantages over repeated venipuncture sampling and endovenous microdialysis sampling. The present report shows that subcutaneous microdialysis with the proper analytical technique can be used to monitor the chemical composition of the interstitial compartment in very different preclinical or clinical conditions. PMID:14552061

Páez, Ximena; Mazzei-Dávila, Carmen Amalia; Hernández, Luis



Fluorescence of Dendrons based on Donors and Accepter with Different Linkages  

NASA Astrophysics Data System (ADS)

Earlier indirect studies utilizing wavelength and bias spectra of photocurrent in simple photovoltaic cells demonstrated charge transfer (CT) in 1st generation dendritic macromolecules prepared using two different donor (tetraphenylporphyrin) groups bound to an accepter (naphthalenediimide) group. We report here fluorescence for solid-state films and solutions of these donor and dendrons. Using 460nm excitation, fluorescence (660nm, 715nm) in solution samples can be observed for both donor and dendron but fluorescence in the solid state can be observable only in donor sample due to fluorescence quenching within the dendron. This demonstrates intermolecular CT from donor to accepter. Fluorescence lifetime measurements (460nm 1.5nsec FWHM pulse excitation) of donor and dendron solutions show that it depends on length of the linkage between donor and accepter. This shows a direct relaxation path from donor to accepter (intramolecular CT). The separation of the exciton to separate electron and on the donor and acceptor portions of the dendron would open the potential for its use in photovoltaic application. Supported in part by DOE #DE-FG02-01ER45931

Park, J. H.; Wu, Y.; Modarelli, D. A.; Parquette, J. R.; Epstein, A. J.



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


Cell surface glycans are a class of sophisticated biomolecules related to cancer development and progression, and their analysis is of great significance for early cancer diagnosis and treatment. In this paper, we proposed a fluorescence assay to evaluate glycan expression on living cancer cells based on a competitive strategy coupled with dual-functionalized nanobiocomposites. The competitive assay was conducted between living cancer cells and thiomannosyl derivatives using concanavalin A (Con A)-modified electrode as the interaction platform. To impart fluorescence signaling ability to competitive derivatives, quantum dots (QDs) were anchored on BSA-protected Au nanoparticles, and thiomannosyl derivatives were further immobilized on the nanoparticle surface through Au-S binding. Due to the spacing between QDs and Au nanoparticles by BSA, the {QDs-Au-BSA-mannose} nanobiocomposites maintained the fluorescence of QDs and showed binding ability with the Con A-modified electrode. Au nanorods (AuNRs)-modified electrode was used as an effective substrate to immobilize Con A. This assay was successfully applied to the analysis of two cancer cells lines (A549 and QGY-7701). The method is simple and shows promise for the study of glycan expression on living cancer cells. PMID:24098881

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



A fluorescence resonance energy transfer-based fluorometer assay for screening anti-coxsackievirus B3 compounds.  


In view of the need to develop a simple and rapid method to screen for antiviral therapeutic agents, a fluorescence resonance energy transfer (FRET)-based reporter system consisting of engineered mammalian cells expressing a cyan fluorescent protein-yellow fluorescent protein (CFP-YFP) pair linked by a short peptide containing the cleavage site of viral protease 2A (2A(pro)) was developed. By detecting the 2A(pro) produced early during the virus infection cycle, the CFP-YFP pair effectively identifies infectious coxsackievirus B3 (CVB3), a picornavirus that causes viral myocarditis in humans. The reporter system was used to screen a library of 2000 drugs and natural products for potential antiviral compounds. The reporter cells were treated with the test compounds, challenged with CVB3, and then examined using a fluorometer at 24h post-infection. Sixty-four compounds, mostly therapeutic drugs, antimicrobial compounds and compounds with unknown functions, caused at least 50% inhibition of 2A(pro) activity. Three known antiviral compounds, cosmosiin, ribavirin and baicalein, were also identified in the screening. The developed method is an effective strategy for rapid screening, and identifies compounds that inhibit CVB3 2A(pro). This method should be a valuable aid in the antiviral drug discovery effort. PMID:21029747

Cantera, Jason L; Chen, Wilfred; Yates, Marylynn V



Cyclohexyl "base pairs" stabilize duplexes and intensify pyrene fluorescence by shielding it from natural base pairs.  


In this study, we investigated the stability and structure of artificial base pairs that contain cyclohexyl rings. The introduction of a single pair of isopropylcyclohexanes into the middle of DNA slightly destabilized the duplex. Interestingly, as the number of the "base pairs" increased, the duplex was remarkably stabilized. A duplex with six base pairs was even more stable than one containing six A-T pairs. Thermodynamic analysis revealed that changes in entropy and not enthalpy contributed to duplex stability, demonstrating that hydrophobic interactions between isopropyl groups facilitated the base pairing, and thus stabilized the duplex. NOESY of a duplex containing an isopropylcyclohexane-methylcyclohexane pair unambiguously demonstrated its "pairing" in the duplex because distinct NOEs between the protons of cyclohexyl moieties and imino protons of both of the neighboring natural base pairs were observed. CD spectra of duplexes tethering cyclohexyl moieties also showed a positive-negative couplet that is characteristic of the B-form DNA duplex. Taken together, these results showed that cyclohexyl moieties formed base pairs in the DNA duplex without severely disturbing the helical structure of natural DNA. Next, we introduced cyclohexyl base pairs between pyrene and nucleobases as an "insulator" that suppresses electron transfer between them. We found a massive increase in the quantum yield of pyrene due to the efficient shielding of pyrene from nucleobases. The cyclohexyl base pairs reported here have the potential to prepare highly fluorescent labeling agents by multiplying fluorophores and insulators alternately into DNA duplexes. PMID:22068299

Kashida, Hiromu; Sekiguchi, Koji; Higashiyama, Naofumi; Kato, Tomohiro; Asanuma, Hiroyuki



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

Microsoft Academic Search

A new fluorescent host-guest system based on the inclusion of the fluorophore 2,3-diazabicyclo[2.2.2]oct-2-ene (DBO) into the cavity of the molecular container compound cucurbit[7]uril (CB7) has been designed which possesses an exceedingly long-lived emission (690 ns in aerated water). The large binding constant of (4±1)×105 M-1 along with the resistance of the CB7·DBO complex toward external fluorescence quenchers allow the use

Cesar Marquez; Fang Huang; Werner M. Nau



High-speed front end for LED-Photodiode based fluorescence lifetime measurement system  

Microsoft Academic Search

An optoelectronic front end for a high-speed solid-state based time-domain fluorescence measurement system is investigated. It consists of a source light emitting diode (LED) and photodetector (PD) pair connected to an original transimpedance amplifier. The amount of light reaching the detector from the fluorescence and the effect of different noise sources on the proposed system are evaluated. The simulation results

Clement Joseph; Mounir Boukadoum; Joe Charlson; David Starikov; Abdelhak Bensaoula



Sensitive fluorescent vesicles based on the supramolecular inclusion of ?-cyclodextrins withN-alkylamino-l-anthraquinone  

Microsoft Academic Search

Self-assembly fluorescent vesicles were designed and prepared based on the supramolecular interaction of cyclodextrins and N-alkylamino-l-anthraquinone (n-AQ). As the guest molecules, n-AQs with alkyl lengths ranging from C0 to C18 were synthesised by the direct reaction of alkylamine with l-nitroanthraquinone in N,N-dimethylformamide. Transmission electron microscopy (TEM), scanning electron microscopy, dynamic light scattering and epi fluorescence microscopy were employed to study

Tao Sun; Huacheng Zhang; Hui Yan; Jianye Li; Guanghui Cheng; Aiyou Hao; Hongwei Qiao; Feifei Xin



Multiplex immunoassays of equine virus based on fluorescent encoded magnetic composite nanoparticles  

Microsoft Academic Search

A new detection format for multiplexed analysis based on fluorescent encoded magnetic composite nanoparticles is presented.\\u000a Two kinds of virus were analyzed by this new method: equine influenza virus (EIV) and equine infectious anemia virus (EIAV).\\u000a Firstly, EIV antigen and EIAV antigen were conjugated to two kinds of fluorescent encoded magnetic composite nanoparticles,\\u000a while the green-emitting CdTe quantum dots (QDs)

Guannan Wang; Yuan Gao; Hui Huang; Xingguang Su



Gold Nanoparticle-Based Fluorescence Resonance Energy Transfer Aptasensor for Ochratoxin a Detection  

Microsoft Academic Search

In this paper, a sensitive and specific fluorescence resonance energy transfer (FRET) aptasensor for the detection of Ochratoxin A (OTA) is developed based on a dye-tagged ssDNA hybridized with aptamer-conjugated Au nanoparticles (Au NPs). The binding between the aptamer-Au NPs conjugate and the dye-labeled ssDNA leads to the fluorescence quenching of FAM due to its close proximity. The addition of

Nuo Duan; Shijia Wu; Xiaoyuan Ma; Xiujuan Chen; Yukun Huang; Zhouping Wang



A flash-lamp based device for fluorescence detection and identification of individual pollen grains  

NASA Astrophysics Data System (ADS)

We present a novel optical aerosol particle detector based on Xe flash lamp excitation and spectrally resolved fluorescence acquisition. We demonstrate its performances on three natural pollens acquiring in real-time scattering intensity at two wavelengths, sub-microsecond time-resolved scattering traces of the particles' passage in the focus, and UV-excited fluorescence spectra. We show that the device gives access to a rather specific detection of the bioaerosol particles.

Kiselev, Denis; Bonacina, Luigi; Wolf, Jean-Pierre



Fluorescent-based chemical sensor for organophosphate detection  

NASA Astrophysics Data System (ADS)

We present a new optical sensor for the detection of organophosphates by incorporating fluorescent indicator dye into sol-gel material. We used different configurations of immobilization matrices such as thin film and spherical nanoparticles. The sensor thin films were prepared by using acid-catalyzed sol-gel process and the spherical nanoparticles by modified Stöber method. The effects of configuration matrices on the sensor's characteristic were studied. The use of dye-doped nanoparticles improved the detection limit from 0.69 ?M to 17 nM, response time from 600 s to 12 s, precision and sensitivity, but reduced the sensor's working rage from 6.9×10-7 M - 6.9×10-3 M to 1.75×10-8M - 2.3×10-7 M.

Urek, Špela Korent; Lobnik, Aleksandra



A sensitive method based on fluorescence-detected circular dichroism for protein local structure analysis.  


We report an improved fluorescence-detected circular dichroism (FDCD)-based analytical method that is useful for probing protein three-dimensional structures. The method uses a novel FDCD device with an ellipsoidal mirror that functions on a standard circular dichroism (CD) spectrometer and eliminates all artifacts. Our experiments demonstrated three important findings. First, the method is applicable to any proteins either by using intrinsic fluorescence derived from tryptophan residues or by introducing a fluorescent label onto nonfluorescent proteins. Second, by using intrinsic fluorescence, FDCD spectroscopy can detect a structural change in the tertiary structure of metmyoglobin due to stepwise denaturation on a change in pH. Such changes could not be detected by conventional CD spectroscopy. Third, based on the typical advantages of fluorescence-based analyses, FDCD measurements enable observation of only the target proteins in a solution even in the presence of other peptides. Using our ellipsoidal mirror FDCD device, we could observe structural changes of fluorescently labeled calmodulin on binding with Ca(2+) and/or interacting with binding peptides. Because FDCD appears to reflect the protein's local structure around the fluorophore, it may provide a useful means for "pinpoint analysis" of protein structures. PMID:22940648

Nehira, Tatsuo; Ishihara, Kaoru; Matsuo, Koichi; Izumi, Shunsuke; Yamazaki, Takeshi; Ishida, Atsuhiko



A fluorescence-based screen for ribosome binding antibiotics.  


The development of new antibacterial agents has become necessary to treat the large number of emerging bacterial strains resistant to current antibiotics. Despite the different methods of resistance developed by these new strains, the A-site of the bacterial ribosome remains an attractive target for new antibiotics. To develop new drugs that target the ribosomal A-site, a high-throughput screen is necessary to identify compounds that bind to the target with high affinity. To this end, we present an assay that uses a novel fluorescein-conjugated neomycin (F-neo) molecule as a binding probe to determine the relative binding affinity of a drug library. We show here that the binding of F-neo to a model Escherichia coli ribosomal A-site results in a large decrease in the fluorescence of the molecule. Furthermore, we have determined that the change in fluorescence is due to the relative change in the pK(a) of the probe resulting from the change in the electrostatic environment that occurs when the probe is taken from the solvent and localized into the negative potential of the A-site major groove. Finally, we demonstrate that F-neo can be used in a robust, highly reproducible assay, determined by a Z'-factor greater than 0.80 for 3 consecutive days. The assay is capable of rapidly determining the relative binding affinity of a compound library in a 96-well plate format using a single channel electronic pipette. The current assay format will be easily adaptable to a high-throughput format with the use of a liquid handling robot for large drug libraries currently available and under development. PMID:23262284

Watkins, Derrick; Norris, F A; Kumar, Sunil; Arya, Dev P



Rapid, photoactivatable turn-on fluorescent probes based on an intramolecular photoclick reaction.  


Photoactivatable fluorescent probes are invaluable tools for the study of biological processes with high resolution in space and time. Numerous strategies have been developed in generating photoactivatable fluorescent probes, most of which rely on the photo-"uncaging" and photoisomerization reactions. To broaden photoactivation modalities, here we report a new strategy in which the fluorophore is generated in situ through an intramolecular tetrazole-alkene cycloaddition reaction ("photoclick chemistry"). By conjugating a specific microtubule-binding taxoid core to the tetrazole/alkene prefluorophores, robust photoactivatable fluorescent probes were obtained with fast photoactivation (?1 min) and high fluorescence turn-on ratio (up to 112-fold) in acetonitrile/PBS (1:1). Highly efficient photoactivation of the taxoid-tetrazoles inside the mammalian cells was also observed under a confocal fluorescence microscope when the treated cells were exposed to either a metal halide lamp light passing through a 300/395 filter or a 405 nm laser beam. Furthermore, a spatially controlled fluorescent labeling of microtubules in live CHO cells was demonstrated with a long-wavelength photoactivatable taxoid-tetrazole probe. Because of its modular design and tunability of the photoactivation efficiency and photophysical properties, this intramolecular photoclick reaction based approach should provide a versatile platform for designing photoactivatable fluorescent probes for various biological processes. PMID:21736329

Yu, Zhipeng; Ho, Lok Yin; Lin, Qing



Flavin Mononucleotide-Based Fluorescent Proteins Function in Mammalian Cells without Oxygen Requirement  

PubMed Central

Usage of the enhanced green fluorescent protein (eGFP) in living mammalian cells is limited to aerobic conditions due to requirement of oxygen during chromophore formation. Since many diseases or disease models are associated with acute or chronic hypoxia, eGFP-labeling of structures of interest in experimental studies might be unreliable leading to biased results. Thus, a chromophore yielding a stable fluorescence under hypoxic conditions is desirable. The fluorescence of flavin mononucleotide (FMN)-based fluorescent proteins (FbFPs) does not require molecular oxygen. Recently, the advantages of FbFPs for several bacterial strains and yeasts were described, specifically, their usage as a real time fluorescence marker in bacterial expression studies and their ability of chromophore formation under anaerobic conditions. Our objective was to verify if FbFPs also function in mammalian cells in order to potentially broaden the repertoire of chromophores with ones that can reliably be used in mammalian studies under hypoxic conditions. In the present study, we demonstrate for the first time, that FbFPs can be expressed in different mammalian cells, among them murine neural stem cells during proliferative and differentiated stages. Fluorescence intensities were comparable to eGFP. In contrast to eGFP, the FbFP fluorescence did not decrease when cells were exposed to defined hypoxic conditions neither in proliferating nor in differentiated cells. Thus, FbFPs can be regarded as an alternative to eGFP in studies that target cellular structures which are exposed to hypoxic conditions.

Walter, Janine; Hausmann, Sascha; Drepper, Thomas; Puls, Michael; Eggert, Thorsten; Dihne, Marcel



Comparison of fluorescence-based semi-automated genotyping of multiple microsatellite loci with autoradiographic techniques  

SciTech Connect

The practical application of highly efficient fluorescence-based methods for the semi-automated genotyping of polymerase chain reaction-based microsatellite markers will depend on the development of robust protocols that provide accurate and reproducible data. In the present report the authors compare the accuracy of a fluorescence-based protocol with a benchmark radiolabeling method that depends on a known sequence ladder or amplified DNA from reference individuals for sizing by autoradiography. Three microsatellite markers, IGF (mfd1), D4S174 (mfd 59), and D5S211 (mfd 154), with products overlapping in size were each labeled with a different fluorophore and run simultaneously with an internal size standard in a single electrophoretic lane. The size of each allele was compared for these markers by using both techniques for five larger CEPH families (884, 1331, 1333, and 1362). Of 462 possible alleles, four discrepancies (0.8%) were identified when the two approaches were compared. The authors conclude that the fluorescence-based protocol is at least as accurate as the standard radiolabeling technique since none of the sizing errors arose as a result of the fluorescence-based technique. They describe the adaptation of the fluorescence-based protocol to the simultaneous analysis of up to 24 microsatellite loci per electrophoretic lane. These highly accurate and efficient semi-automated techniques will be useful in high-resolution genomic analyses. 18 refs., 4 tabs., 3 figs.

Schwengel, D.A.; Jedlicka, A.E.; Levitt, R.C. [Johns Hopkins Medical Institutions, Baltimore, MD (United States)] [and others



A method for fluorescence sensing of adenosine and alkaline phosphatase based on the inhibition of S-adenosylhomocysteine hydrolase activity.  


This study presents a simple fluorescent method for the sensitive and selective detection of adenosine, based on adenosine inhibiting S-adenosylhomocysteine hydrolase (SAHH)-catalyzed hydrolysis of S-adenosylhomocysteine (SAH). Because of homocysteine (HCys) belonging to the thiol and amino groups, 2,3-naphthalenedicarboxaldehyde (NDA) can selectively react with HCys to form a 6-membered ring without the addition of nucleophiles. Electrospray ionization mass spectrometry was used to obtain the molecular mass of the resulting products, which is helpful in proposing the possible reaction mechanism between NDA and HCys. When SAHH catalyzed the cleavage of SAH, the generated HCys reacted with NDA to form highly fluorescent products with a quantum yield of 34%. The addition of adenosine to an SAH solution resulted in the inhibition of SAHH activity. Consequently, HCys production decreased with an increase in adenosine concentration. Under optimal NDA derivatization conditions, the SAHH-based probe showed a limit of detection (at a signal-to-noise ratio of 3) for adenosine of 0.3 ?M. Selectivity of the SAHH-based probe is more than 100-fold for adenosine over any adenosine analog. We validated the applicability of this probe by determining adenosine concentration in urine samples. The SAHH-based probe was also used to evaluate the activity and inhibition of alkaline phosphatase, which can convert adenosine monophosphate to adenosine. PMID:23040372

Lin, Jia-Hui; Tseng, Wei-Lung



A novel fluorescent assay for oxytetracycline hydrochloride based on fluorescence quenching of water-soluble CdTe nanocrystals.  


A novel assay for oxytetracycline hydrochloride (OTC) based on fluorescence quenching was developed from the interaction between functionalized cadmium telluride quantum dots (CdTe QDs) and OTC. Optimum conditions for the detection of OTC were found after investigating all factors. Under optimum conditions, luminescence of CdTe nanocrystals (? ex = 365 nm, ? em = 562 nm) was quenched by OTC in a concentration-dependent manner best described by a modified Stern-Volmer type equation. Good linearity was obtained with a regression coefficient of 0.9999 in the range of 1.34 ~ 13.4 x 10(-5) mol/L and a limit of detection of 3.08 x 10(-7) mol/L. In addition, the quenching mechanism was also established. The results imply that the close proximity of OTC-CdTe was driven by electrostatic attraction and the resulting effective electron transfer from OTC to QDs could be responsible for fluorescence quenching of CdTe-QDs. PMID:22715152

Gao, Chao; Liu, Zhen; Chen, Jianqiu; Yan, Zhengyu



Characterization of photophysical and base-mimicking properties of a novel fluorescent adenine analogue in DNA  

PubMed Central

To increase the diversity of fluorescent base analogues with improved properties, we here present the straightforward click-chemistry-based synthesis of a novel fluorescent adenine-analogue triazole adenine (AT) and its photophysical characterization inside DNA. AT shows promising properties compared to the widely used adenine analogue 2-aminopurine. Quantum yields reach >20% and >5% in single- and double-stranded DNA, respectively, and show dependence on neighbouring bases. Moreover, AT shows only a minor destabilization of DNA duplexes, comparable to 2-aminopurine, and circular dichroism investigations suggest that AT only causes minimal structural perturbations to normal B-DNA. Furthermore, we find that AT shows favourable base-pairing properties with thymine and more surprisingly also with normal adenine. In conclusion, AT shows strong potential as a new fluorescent adenine analogue for monitoring changes within its microenvironment in DNA.

Dierckx, Anke; Diner, Peter; El-Sagheer, Afaf H.; Kumar, Joshi Dhruval; Brown, Tom; Gr?tli, Morten; Wilhelmsson, L. Marcus



Characterization and use of an unprecedentedly bright and structurally non-perturbing fluorescent DNA base analogue  

PubMed Central

This article presents the first evidence that the DNA base analogue 1,3-diaza-2-oxophenoxazine, tCO, is highly fluorescent, both as free nucleoside and incorporated in an arbitrary DNA structure. tCO is thoroughly characterized with respect to its photophysical properties and structural performance in single- and double-stranded oligonucleotides. The lowest energy absorption band at 360 nm (? = 9000 M?1 cm?1) is dominated by a single in-plane polarized electronic transition and the fluorescence, centred at 465 nm, has a quantum yield of 0.3. When incorporated into double-stranded DNA, tCO shows only minor variations in fluorescence intensity and lifetime with neighbouring bases, and the average quantum yield is 0.22. These features make tCO, on average, the brightest DNA-incorporated base analogue so far reported. Furthermore, it base pairs exclusively with guanine and causes minimal perturbations to the native structure of DNA. These properties make tCO a promising base analogue that is perfectly suited for e.g. photophysical studies of DNA interacting with macromolecules (proteins) or for determining size and shape of DNA tertiary structures using techniques such as fluorescence anisotropy and fluorescence resonance energy transfer (FRET).

Sandin, Peter; Borjesson, Karl; Li, Hong; Martensson, Jerker; Brown, Tom; Wilhelmsson, L. Marcus; Albinsson, Bo



Simultaneous quantitative analysis of three compounds using three-dimensional fluorescence spectra based on digital image techniques.  


Digital image processing has been applied on various fields such as classification and qualitative analysis. In this work, a very simple quantitative approach was proposed for the first time. Based on the digital grayscale images of three-dimensional fluorescence spectra, several wavelet moment invariants were calculated, and used to establish the linear models for the quantitative analysis. This approach was applied to the quantitative analysis of Tryptophan, Tyrosine and Phenylalanine in mixture samples, and the correlation coefficients R(2) of the obtained linear models were more than 0.99, which were supported by the strict statistical parameters as well as leave-one-out and Jackknife cross-validations. Our study indicates that the selected wavelet moment invariants are immune from the noise and background signals, and the quantitative analysis can be performed accurately based on the overlapping peaks of compounds in mixture. This proposed approach provides a novel pathway for the analysis of three-dimensional spectra. PMID:22477062

Zhai, Hong Lin; Shan, Zhi Jie; Li, Rui Na; Yu, E



Highly fluorescent colloids based on rhodamine 6G, modified layered silicate, and organic solvent.  


Synthetic layered silicate saponite was modified with dodecyltrimethylammonium (C12), octadecyltrimethylammonium (C18), and dioctadecyldimethylammonium (2C18) cations for use as sorbents of the laser dye, rhodamine 6G (R6G). Via solvent exchange, transparent colloids in xylene were prepared and investigated using absorption and fluorescence spectroscopies. Molecular aggregation and partial quenching of the fluorescence were observed for the colloids based on 2C18 cations. Maximal fluorescence yields were observed for the colloids with C12 and C18 cations. Transparent gels without an apparent loss of luminescent efficiency could be prepared by concentrating the colloids. These highly fluorescent colloids and gels represent new types of materials with interesting optical properties. PMID:22995248

Bujdák, Juraj; Iyi, Nobuo



Visualizing Hg2+ ions in living cells using a FRET-based fluorescent sensor  

NASA Astrophysics Data System (ADS)

A novel FRET fluorescent sensor for Hg2+ imaging in living cells is rationally designed based on a coumarin-rhodamine platform. RBC1 exhibit high selectivity and excellent sensitivity in both absorbance and fluorescence detection of Hg2+ in aqueous solution. After addition of increasing concentrations of Hg2+, it result in the decrease of coumarin emission at 467 nm and a new emission profile of rhodamine at 590 nm gradually increased. The response time to Hg2+ is less than 2 min, and other metal ions including Fe2+, Mn2+, Ni2+, Co2+, Cu2+, Zn2+, Cd2+, Pb2+, and Cr3+ had no interference. In addition, fluorescent imaging of Hg2+ in A375 cells is also successfully demonstrated. The design strategy of two fluorophores switching in this work would help to extend the development of FRET fluorescent sensors.

Zhou, Yi; Chu, Kaihui; Zhen, Haifu; Fang, Yuan; Yao, Cheng



Fluorescent Nanoparticle-Based Indirect Immunofluorescence Microscopy for Detection of Mycobacterium tuberculosis  

PubMed Central

A method of fluorescent nanoparticle-based indirect immunofluorescence microscopy (FNP-IIFM) was developed for the rapid detection of Mycobacterium tuberculosis. An anti-Mycobacterium tuberculosis antibody was used as primary antibody to recognize Mycobacterium tuberculosis, and then an antibody binding protein (Protein A) labeled with Tris(2,2-bipyridyl)dichlororuthenium(II) hexahydrate (RuBpy)-doped silica nanoparticles was used to generate fluorescent signal for microscopic examination. Prior to the detection, Protein A was immobilized on RuBpy-doped silica nanoparticles with a coverage of ?5.1×102 molecules/nanoparticle. With this method, Mycobacterium tuberculosis in bacterial mixture as well as in spiked sputum was detected. The use of the fluorescent nanoparticles reveals amplified signal intensity and higher photostability than the direct use of conventional fluorescent dye as label. Our preliminary studies have demonstrated the potential application of the FNP-IIFM method for rapid detection of Mycobacterium tuberculosis in clinical samples.

Qin, Dilan; He, Xiaoxiao; Wang, Kemin; Zhao, Xiaojun Julia; Tan, Weihong; Chen, Jiyun



Static hyperspectral fluorescence imaging of viscous materials based on a linear variable filter spectrometer.  


This paper presents a low-cost hyperspectral measurement setup in a new application based on fluorescence detection in the visible (Vis) wavelength range. The aim of the setup is to take hyperspectral fluorescence images of viscous materials. Based on these images, fluorescent and non-fluorescent impurities in the viscous materials can be detected. For the illumination of the measurement object, a narrow-band high-power light-emitting diode (LED) with a center wavelength of 370 nm was used. The low-cost acquisition unit for the imaging consists of a linear variable filter (LVF) and a complementary metal oxide semiconductor (CMOS) 2D sensor array. The translucent wavelength range of the LVF is from 400 nm to 700 nm. For the confirmation of the concept, static measurements of fluorescent viscous materials with a non-fluorescent impurity have been performed and analyzed. With the presented setup, measurement surfaces in the micrometer range can be provided. The measureable minimum particle size of the impurities is in the nanometer range. The recording rate for the measurements depends on the exposure time of the used CMOS 2D sensor array and has been found to be in the microsecond range. PMID:24064604

Murr, Patrik J; Schardt, Michael; Koch, Alexander W



A rhodamine based fluorescent probe for Hg2+ and its application to cellular imaging  

NASA Astrophysics Data System (ADS)

A new rhodamine-based fluorescent probe (Rh-F) for detection of Hg2+ ions was synthesized, which could bind Hg2+ in aqueous ethanol (7:3, v/v) at pH 7.0 with detectable change in color and fluorescence. The response is based on a ring opening reaction and formation of a 1:1 complex, while ring-opening process of spirolactam enables large fluorescent enhancement and colorimetric change upon the addition of Hg2+. The response is reversible and linear in the range between 200 nM and 1000 nM, with a detection limit of 4.2 nM. Selectivity and competition experiments with various other metal ion revealed that Rh-F possesses highly selective fluorescent response to Hg2+. Furthermore, the probe was successfully applied to fluorescent imaging of Hg2+ in L-929 cells confirm that Rh-F can be used as a fluorescent probe for monitoring Hg2+ in living cells.

Yan, Fanyong; Cao, Donglei; Yang, Ning; Wang, Meng; Dai, Linfeng; Li, Chuying; Chen, Li



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


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

Mayer-Cumblidge, M. Uljana (Richland, WA); Cao, Haishi (Richland, WA)



Molecular dynamics study of the structure and performance of simple and double bases propellants  

Microsoft Academic Search

To investigate the structure and performance of simple and double bases propellants, the nitrocellulose (NC), nitroglycerin (NG), and double mixed system (NC+NG) have been simulated by using the molecular dynamics (MD) method with the COMPASS force field. The interactions between NC and NG have been analyzed by means of pair correlation functions. The mechanical properties of the three model systems,

Xiufang Ma; Weihua Zhu; Jijun Xiao; Heming Xiao



A Simple System for Observing Dynamic Phase Equilibrium via an Inquiry-Based Laboratory or Demonstration  

ERIC Educational Resources Information Center

|This article describes an activity that can be used as an inquiry-based laboratory or demonstration for either high school or undergraduate chemistry students to provide a basis for understanding both vapor pressure and the concept of dynamic phase equilibrium. The activity includes a simple setup to create a closed system of only water liquid…

Cloonan, Carrie A.; Andrew, Julie A.; Nichol, Carolyn A.; Hutchinson, John S.



Flagellar Length Control System: Testing a Simple Model Based on Intraflagellar Transport and Turnover  

Microsoft Academic Search

Flagellar length regulation provides a simple model system for addressing the general problem of organelle size control. Based on a systems-level analysis of flagellar dynamics, we have proposed a mechanism for flagellar length control in which length is set by the balance of continuous flagellar assembly and disassembly. The model proposes that the assembly rate is length dependent due to

Wallace F. Marshall; Hongmin Qin; Monica Rodrigo Brenni; Joel L. Rosenbaum



A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences  

Microsoft Academic Search

Summary Some simple formulae were obtained which enable us to estimate evolutionary distances in terms of the number of nucleotide substitutions (and, also, the evolutionary rates when the divergence times are known). In comparing a pair of nucleotide sequences, we distinguish two types of differences; if homologous sites are occupied by different nucleotide bases but both are purines or both

Motoo Kimura



A simple GPU-based approach for 3D Voronoi diagram construction and visualization  

Microsoft Academic Search

In this paper we propose a simple GPU-based approach for discrete incremental approximation of 3D Voronoi diagram. By constructing region maps via GPU. Nearest sites, space clustering, and shortest distance query can be quickly answered by looking up the region map. In addition, we propose another representation of the 3D Voronoi diagram for visualization.

Hsien-hsi Hsieh; Wen-kai Tai



High-throughput retrotransposon-based fluorescent markers: improved information content and allele discrimination  

PubMed Central

Background Dense genetic maps, together with the efficiency and accuracy of their construction, are integral to genetic studies and marker assisted selection for plant breeding. High-throughput multiplex markers that are robust and reproducible can contribute to both efficiency and accuracy. Multiplex markers are often dominant and so have low information content, this coupled with the pressure to find alternatives to radio-labelling, has led us to adapt the SSAP (sequence specific amplified polymorphism) marker method from a 33P labelling procedure to fluorescently tagged markers analysed from an automated ABI 3730 xl platform. This method is illustrated for multiplexed SSAP markers based on retrotransposon insertions of pea and is applicable for the rapid and efficient generation of markers from genomes where repetitive element sequence information is available for primer design. We cross-reference SSAP markers previously generated using the 33P manual PAGE system to fluorescent peaks, and use these high-throughput fluorescent SSAP markers for further genetic studies in Pisum. Results The optimal conditions for the fluorescent-labelling method used a triplex set of primers in the PCR. These included a fluorescently labelled specific primer together with its unlabelled counterpart, plus an adapter-based primer with two bases of selection on the 3' end. The introduction of the unlabelled specific primer helped to optimise the fluorescent signal across the range of fragment sizes expected, and eliminated the need for extensive dilutions of PCR amplicons. The software (GeneMarker Version 1.6) used for the high-throughput data analysis provided an assessment of amplicon size in nucleotides, peak areas and fluorescence intensity in a table format, so providing additional information content for each marker. The method has been tested in a small-scale study with 12 pea accessions resulting in 467 polymorphic fluorescent SSAP markers of which 260 were identified as having been mapped previously using the radio-labelling technique. Heterozygous individuals from pea cultivar crosses were identifiable after peak area data analysis using the fluorescent SSAP method. Conclusion As well as developing a rapid, and high-throughput marker method for genetic studies, the fluorescent SSAP system improved the accuracy of amplicon scoring, increased the available marker number, improved allele discrimination, and was sensitive enough to identify heterozygous loci in F1 and F2 progeny, indicating the potential to develop high-throughput codominant SSAPs.

Knox, Maggie; Moreau, Carol; Lipscombe, James; Baker, David; Ellis, Noel



A novel lophine-based fluorescence probe and its binding to human serum albumin.  


The binding of a lophine-based fluorescence probe, 4-[4-(4-dimethylaminophenyl)-5-phenyl-1H-imidazol-2-yl]benzoic acid methyl ester (DAPIM) with human serum albumin (HSA) was investigated by fluorescence spectroscopy under physiological conditions. While DAPIM shows extreme low fluorescence in aqueous solution, DAPIM binding with HSA emits strong fluorescence at 510nm. The binding constant and binding number determined by Scatchard plot was 3.65×10(6)M(-1) and 1.07, respectively. Competitive binding between DAPIM and other ligands such as warfarin, valproic acid, diazepam and oleic acid, were also studied fluorometrically. The results indicated that the primary binding site of DAPIM to HSA is site II at subdomain IIIA. DAPIM can be a useful fluorescence probe for the characterization of drug-binding sites. In addition to the interaction study, because the fluorescence intensity of DAPIM increased in proportion to HSA concentration, its potential in HSA assay for serum sample was also evaluated. PMID:23680544

Kishikawa, Naoya; Ohyama, Kaname; Saiki, Akane; Matsuo, Aya; Ali, Marwa Fathy Bakr; Wada, Mitsuhiro; Nakashima, Kenichiro; Kuroda, Naotaka



Förster resonance energy transfer-based total internal reflection fluorescence reader for apoptosis  

NASA Astrophysics Data System (ADS)

A fluorescence reader for the detection of Förster resonance energy transfer (FRET) on surfaces of living cells is described. The method is based on multiple total internal reflections (TIR) of an incident laser beam within a glass slide, such that individual samples on top of the glass slide are illuminated simultaneously by an evanescent electromagnetic field. Enhanced cyan fluorescent protein (ECFP) anchored to the inner leaflet of the plasma membrane is optically excited and transfers its excitation energy via the peptide linker Asp-Glu-Val-Asp (DEVD) to an enhanced yellow fluorescent protein. Upon apoptosis, DEVD is cleaved, and energy transfer is disrupted, as proven by an increase of fluorescence intensity as well as of fluorescence lifetime of the donor ECFP. Due to selective excitation of membrane-associated fluorophores, intracellular fluorescence and background luminescence from the surrounding medium are eliminated. Therefore, this test system appears to be a sensitive device for the detection of apoptosis and more generally for drug screening or in vitro diagnosis on a nanometer scale.

Bruns, Thomas; Angres, Brigitte; Steuer, Heiko; Weber, Petra; Wagner, Michael; Schneckenburger, Herbert



Fluorescence of a photosensitizer based on an indotricarbocyanine dye in photochemotherapy  

NASA Astrophysics Data System (ADS)

We present the results for studies of the spectral luminescence properties of a symmetric indotricarbocyanine dye (PD1) in HeLa tumor cells and animal tissues in vivo during a photochemotherapy session and after the end of the session. We have established that when the dye is exposed to light in tumor tissues, changes occur in the position and half-width of the dye fluorescence spectra, while in a culture of HeLa cells its spectral characteristics are constant. Based on analysis of the effect of overlap between the absorption spectra of endogenous biomolecules and the fluorescence spectra of the dye plus comparison of the experimental data with numerical modeling results, we have concluded that the observed changes in the fluorescence spectra of PD1 in vivo are due to a change in the ratio of the different forms of hemoglobin in the tumor tissue. We have shown that the spectral characteristics of PD1, fluorescing in the near IR range, correlate with the depth of tumor tissue necrosis achieved on exposure to light. We have established that tumor tissue necrosis occurs down to a depth of 2 cm in the case of all strains studied: S-45, SM-1, and W-256, where as a result of exposure to light, we observe an increase in the half-width and a short-wavelength shift of the fluorescence spectrum of the dye PD1, and also the intensity of its fluorescence does not recover.

Samtsov, M. P.; Voropay, E. S.; Liashenka, L. S.; Melnikau, D. G.; Kapleusky, K. N.; Lugovskij, A. P.



Forster resonance energy transfer-based total internal reflection fluorescence reader for apoptosis.  


A fluorescence reader for the detection of Forster resonance energy transfer (FRET) on surfaces of living cells is described. The method is based on multiple total internal reflections (TIR) of an incident laser beam within a glass slide, such that individual samples on top of the glass slide are illuminated simultaneously by an evanescent electromagnetic field. Enhanced cyan fluorescent protein (ECFP) anchored to the inner leaflet of the plasma membrane is optically excited and transfers its excitation energy via the peptide linker Asp-Glu-Val-Asp (DEVD) to an enhanced yellow fluorescent protein. Upon apoptosis, DEVD is cleaved, and energy transfer is disrupted, as proven by an increase of fluorescence intensity as well as of fluorescence lifetime of the donor ECFP. Due to selective excitation of membrane-associated fluorophores, intracellular fluorescence and background luminescence from the surrounding medium are eliminated. Therefore, this test system appears to be a sensitive device for the detection of apoptosis and more generally for drug screening or in vitro diagnosis on a nanometer scale. PMID:19405716

Bruns, Thomas; Angres, Brigitte; Steuer, Heiko; Weber, Petra; Wagner, Michael; Schneckenburger, Herbert


Simple methods for improving speaker-similarity of HMM-based speech synthesis  

Microsoft Academic Search

In this paper we revisit some basic configuration choices of HMM-based speech synthesis, such as waveform sampling rate, auditory frequency warping scale and the logarithmic scaling of F0, with the aim of improving speaker similarity which is an acknowledged weakness of current HMM-based speech synthesisers. All of the techniques investigated are simple but, as we demonstrate using perceptual tests, can

Junichi Yamagishi; Simon King



Simple method of enzyme immobilization for pH-ISFET-based urea biosensors  

Microsoft Academic Search

In this paper, a simple chemical method of urease immobilization on silicon nitride surface is described. As a basic structure to construct urea-biosensor, a pH-sensitive Si3N4-gate ISFET was used. The developed method of chemical immobilization of urease is based on Schiff's base formation. The developed EnFET type urea biosensor are characterized by the following parameters: (1) maximum analytical signal: 120

Dorota Pijanowska; Wladislaw Torbicz



A simple clinical score accurately predicts outcome in a community-based population undergoing stress testing  

Microsoft Academic Search

PurposeScoring systems based on clinical variables are available but not widely applied for evaluating patients with chronic coronary artery disease. The purpose of this study was to validate the prognostic value of a simple clinical scoring system, originally developed in patients referred for a nuclear stress test at a tertiary-care medical center, in a less-selected, community-based population undergoing stress testing

Todd D. Miller; Veronique L. Roger; David O. Hodge; Raymond J. Gibbons



Phthalocyanine-based fluorescent chemosensor for the sensing of Zn (II) in dimethyl sulfoxide-acetonitrile  

Microsoft Academic Search

A tetra-substituted phthalocyanine based on 4-[2-(4-nitrophenoxy)ethoxy]phthalonitrile carrying nitrophenyl group for the\\u000a sensing of Zn2+ has been prepared and characterized by elemental analysis, FT-IR, 1H and 13C NMR, and MS spectral data. The sensing of Zn2+ is based on the fluorescence quenching of Pc. Both absorbance and fluorescence spectra of ZnPc exhibit distinct changes in\\u000a visible region in response to treatment

Yasemin Ça?lar; Nurhan Gümrükçüo?lu; Ece Tu?ba Saka; Miraç Ocak; Halit Kantekin; Ümmühan Ocak


Fluorescent probes for Al(III) and Cr(III) based on a photochromic diarylethene bearing a fluorescent rhodamine unit  

Microsoft Academic Search

A sensitive and selective “turn-on” fluorescent probe (compound 1O) was prepared by combining a photochromic diarylethene with a fluorescent rhodamine unit. This compound displays a favorable\\u000a photochromism on alternating irradiation with UV\\/Vis light. Upon addition of Al(III) or Cr(III) to the probe, its color changes\\u000a from colorless to pink, and its fluorescence is markedly enhanced. The probe displays excellent sensitivity

Weijun Liu; Shouzhi Pu; Duohua Jiang; Shiqiang Cui; Gang Liu; Congbin Fan


Beer's-law-based, simple spectral model for direct normal and diffuse horizontal irradiance  

SciTech Connect

A spectral model for cloudless days that uses simple mathematical expressions and tabulated look-up tables to generate direct normal and diffuse horizontal irradiance is presented. The model is based on modifications to previously published simple models and comparisons with rigorous radiative transfer codes. This model is expected to be more accurate and to be applicable to a broader range of atmospheric conditions than previous simple models. The prime significance of this model is its simplicity, which allows it to be used on small desk-top computers. The spctrum produced by this model is limited to 0.3 to 4.0 wavelength with an approximate resolution of 10 nm.

Bird, R.E.



A Simple High-Throughput Method for Determination of Antiepileptic Analogues of ?-Aminobutyric Acid in Pharmaceutical Dosage Forms Using Microplate Fluorescence Reader.  


Pregabalin (PGB), gabapentin (GBP), and vigabatrin (VGB) are structural analogues of ?-aminobutyric acid used for the treatment of different forms of epilepsy. Their analytical determination is challenging since these molecules have no significant UV or visible absorption. Several derivatization methods have been developed and used for their determination in bulk or pharmaceutical dosage forms. We aimed to develop a high- throughput method using a microplate reader with fluorescence detection and simple derivatization with fluorescamine. Obtained method involves derivatization step of only 5?min at room temperature and simultaneous measurements of 96 samples (?ex 395, ?em 476?nm) thus rendering excellent high-throughput analysis. The method was found to be linear with r(2)>0.998 across investigated analytical ranges of 0.75 to 30.0?µg/mL for PGB, 2.00 to 80.0?µg/mL for GBP, and 1.50 to 60.0?µg/mL for VGB. Intraday and interday precision values did not exceed 4.93%. The accuracy was ranging between 96.6 to 103.5%. The method was also found to be specific since used excipients did not interfere with the method. The robustness study showed that derivatization procedure is more robust than spectrofluorimetric conditions. The developed high-throughput method was successfully applied for determination of drug content and dissolution profiles in pharmaceutical dosage forms of studied antiepileptic drugs. PMID:23856517

Martinc, Boštjan; Vovk, Tomaž



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


A new fluorescent host-guest system based on the inclusion of the fluorophore 2,3-diazabicyclo[2.2.2]oct-2-ene (DBO) into the cavity of the molecular container compound cucurbit[7]uril (CB7) has been designed which possesses an exceedingly long-lived emission (690 ns in aerated water). The large binding constant of (4 +/- 1) x 10(5) M(-1) along with the resistance of the CB7.DBO complex toward external fluorescence quenchers allow the use of CB7 as an enhancer in time-resolved fluorescence-based assays, e.g., to screen enzyme activity or inhibition by using DBO-labeled peptides as substrates. The response of CB7.DBO to different environmental conditions and possible quenchers are described. PMID:15382642

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



Flow-based continuous DNA sequencing via single molecule detection of enzymatically cleaved fluorescent nucleotides  

NASA Astrophysics Data System (ADS)

The development of our rapid, continuous DNA sequencing technology, based on single molecule detection of fluorescently-tagged nucleotides, has proceeded along separate research fronts, each with specific goals: the faithful replication of long sequences of template DNA using one or more fluorescent nucleotide analogues, the incorporation and stable mounting of a single DNA strand into a flow chamber, the enzymatic cleavage of labeled DNA by exonucleases, and the detection of single fluorescent nucleotides in a flow stream by the method of time-gated photon counting. Each individual goal of the sequencing technology has now been realized, and we have begun integrating these efforts in order to demonstrate the feasibility of flow-based sequencing. We are currently detecting photon bursts from TRITC labeled nucleotides which have been cleaved from DNA suspended in our flow cell. The sample size is estimated to be tens of DNA strands.

Schecker, Jay A.; Goodwin, Peter M.; Affleck, Rhett L.; Wu, Ming; Martin, John C.; Jett, James H.; Keller, Richard A.; Harding, John D.



Chemical biology-based approaches on fluorescent labeling of proteins in live cells.  


Recently, significant advances have been made in live cell imaging owing to the rapid development of selective labeling of proteins in vivo. Green fluorescent protein (GFP) was the first example of fluorescent reporters genetically introduced to protein of interest (POI). While GFP and various types of engineered fluorescent proteins (FPs) have been actively used for live cell imaging for many years, the size and the limited windows of fluorescent spectra of GFP and its variants set limits on possible applications. In order to complement FP-based labeling methods, alternative approaches that allow incorporation of synthetic fluorescent probes to target POIs were developed. Synthetic fluorescent probes are smaller than fluorescent proteins, often have improved photochemical properties, and offer a larger variety of colors. These synthetic probes can be introduced to POIs selectively by numerous approaches that can be largely categorized into chemical recognition-based labeling, which utilizes metal-chelating peptide tags and fluorophore-carrying metal complexes, and biological recognition-based labeling, such as (1) specific non-covalent binding between an enzyme tag and its fluorophore-carrying substrate, (2) self-modification of protein tags using substrate variants conjugated to fluorophores, (3) enzymatic reaction to generate a covalent binding between a small molecule substrate and a peptide tag, and (4) split-intein-based C-terminal labeling of target proteins. The chemical recognition-based labeling reaction often suffers from compromised selectivity of metal-ligand interaction in the cytosolic environment, consequently producing high background signals. Use of protein-substrate interactions or enzyme-mediated reactions generally shows improved specificity but each method has its limitations. Some examples are the presence of large linker protein, restriction on the choice of introducible probes due to the substrate specificity of enzymes, and competitive reaction mediated by an endogenous analogue of the introduced protein tag. These limitations have been addressed, in part, by the split-intein-based labeling approach, which introduces fluorescent probes with a minimal size (~4 amino acids) peptide tag. In this review, the advantages and the limitations of each labeling method are discussed. PMID:23318293

Jung, Deokho; Min, Kyoungmi; Jung, Juyeon; Jang, Wonhee; Kwon, Youngeun



An ultra-high sensitive platform for fluorescence detection of micrococcal nuclease based on graphene oxide.  


Micrococcal nuclease (MNase) is the extracellular nuclease of Staphylococcus aureus (S. aureus). It preferentially digests single-stranded nucleic acids. The existence of MNase can be the standard to identify S. aureus and the content of MNase can be used to evaluate the pathogenicity of S. aureus. Herein, an ultra-high sensitive and selective fluorescent sensing platform for MNase is developed based on MNase-induced DNA strand scission and the difference in affinity of graphene oxide (GO) for single-stranded DNA containing different numbers of bases in length. In the absence of MNase, the adsorption of the dye-labeled ssDNA on GO makes the dyes close proximity to GO surface resulting in high efficiency quenching of fluorescence of the dyes. Conversely, and very importantly, in the presence of MNase, it cleaves the dye-labeled ssDNA into small fragments. The introduction of GO into the sensing solution results in weak quenching of the fluorescence of the dyes due to the weak affinity of the short dye-labeled oligonuleotide fragment to GO, and the fluorescence intensity gradually increases with increasing concentration of MNase. MNase can be detected in a range of 8×10?? to 1.6×10?³ unit/mL with a detection limit of 2.7×10?? unit/mL and good selectivity. The detection limit is of two orders of magnitude lower than those reported fluorescence MNase assays. Moreover, when the GO-based biosensor is used in S. aureus sample assays, preeminent fluorescence signals are obtained, thus the platform of the GO-based biosensor can be used to detect MNase in real-world samples. PMID:23238320

He, Yue; Xiong, Ling-Hong; Xing, Xiao-Jing; Tang, Hong-Wu; Pang, Dai-Wen



A novel finite-element-based algorithm for fluorescence molecular tomography of heterogeneous media.  


The knowledge of optical properties distribution of heterogeneous media has significant impact on the reconstructed fluorescence image quality in fluorescence molecular tomography (FMT). In this study, a novel finite-element-based algorithm for FMT of heterogeneous media is proposed. In the algorithm, optical properties are reconstructed using the conjugate gradient method. A modified method based on reverse differential scheme is deduced for calculating the gradient when the detector points are not restricted on boundary nodes. With the recovered optical properties, linear relationship between known surface measurements of emission light and unknown fluorescence yield is then obtained. FMT reconstruction is implemented by combining algebraic reconstruction technique (ART) and Landweber iteration method. With initial value provided by ART, Landweber iteration method improves the quantification smoothly with small step length between neighboring iterations. The algorithm was evaluated using phantoms of different heterogeneity configurations. Results show that the reconstructed fluorescence yield is insensitive to various degrees of heterogeneity for the proposed algorithm. In contrast, when assuming homogeneous optical properties, it shows that more underestimation of optical properties results in more underestimation of the reconstructed fluorescence yield. Fast computation speed of the proposed algorithm is also demonstrated in this study. PMID:19304493

Wang, Daifa; Liu, Xin; Chen, Yanping; Bai, Jing



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

NASA Astrophysics Data System (ADS)

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

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



Remote fluorescence-based sensing of elastic thin-film elastic moduli  

NASA Astrophysics Data System (ADS)

The fluorescence lifetime of dyes embedded within thin polymer films is sensitive to the local mechanical environment. Here we show that this enables direct measurement of elastic properties of thin films and small samples whose investigation by conventional macroscopic mechanical characterization would not be possible. The specific example of poly(ethylene oxide) (PEO) and its nanocomposite thin films is highlighted. The method is also validated by comparison to a family of other polymers of known macroscopic moduli. Being simple, rapid, and reliable, we propose that this analysis can in principle apply generally to a broad class of soft materials and other polymer multilayer films.

Jee, Ah Young; Lee, Minyung



Nucleic acid based fluorescent sensor for mercury detection  


A nucleic acid enzyme comprises an oligonucleotide containing thymine bases. The nucleic acid enzyme is dependent on both Hg.sup.2+and a second ion as cofactors, to produce a product from a substrate. The substrate comprises a ribonucleotide, a deoxyribonucleotide, or both.

Lu, Yi; Liu, Juewen



2-Aminopurine fluorescence studies of base stacking interactions at abasic sites in DNA: metal-ion and base sequence effects.  

PubMed Central

Metal-ion and sequence dependent changes in the stacking interactions of bases surrounding abasic (AB) sites in 10 different DNA duplexes were examined by incorporating the fluorescent nucleotide probe 2-aminopurine (2-AP), opposite to the site (AB-APopp) or adjacent to the site (AB-APadj) on either strand. A detailed study of the fluorescence emission and excitation spectra of these AB duplexes and their corresponding parent duplexes indicates that AB-APoppis significantly less stacked than 2-AP in the corresponding normal duplex. In general, AB-APadjon the AB strand is stacked, but AB-APadjon the opposite strand shows destabilized stacking interactions. The results also indicate that divalent cation binding to the AB duplexes contributes to destabilizaton of the base stacking interactions of AB-APopp, but has little or no effect on the stacking interactions of AB-APadj. Consistent with these results, the fluorescence of AB-APoppis 18-30-fold more sensitive to an externally added quenching agent than the parent normal duplex. When uracil DNA glycosylase binds to AB-APoppin the presence of 2.5 mM MgCl2, a 3-fold decrease in fluorescence is observed ( K d = 400 +/- 90 nM) indicating that the unstacked 2-APoppbecomes more stacked upon binding. On the basis of these fluorescence studies a model for the local base stacking interactions at these AB sites is proposed.

Stivers, J T



A "turn-on" fluorescent Hg2+ chemosensor based on Ferrier carbocyclization.  


A "turn-on" fluorescent chemosensor with excellent selectivity and satisfactory sensitivity on Hg(2+) detection in 100% water media has been established employing a carbohydrate based Ferrier carbocyclization reaction. The probe has also presented satisfactory results for the imaging of Hg(2+) ions in cells and organisms. PMID:22264070

Ma, Xing; Wang, Jing; Shan, Qiuli; Tan, Zhuowei; Wei, Guohua; Wei, Dongbin; Du, Yuguo



Fluorescence Assay Based on Aptamer-Quantum Dot Binding to Bacillus thuringiensis Spores.  

National Technical Information Service (NTIS)

A novel assay was developed for the detection of Bacillus thuringiensis (BT) spores. The assay is based on the fluorescence observed after binding an aptamer-quantum dot conjugate to BT spores. The in vitro selection and amplification technique called SEL...

A. Allman J. G. Bruno M. Ikanovic M. P. Carrillo W. E. Rudzinski



A smart-lighting emergency ballast for fluorescent lamps based on microcontroller  

Microsoft Academic Search

A new emergency ballast for fluorescent lamps is presented. The fundamental block is the control circuit based on a microcontroller that performs the supervision and control function, achieving an increase in the installation security. High frequency electronics techniques are proposed for the battery charger and the lamp driver, with high power factor in the first and high luminous efficacy in

J. M. Alonso; J. Diaz; C. Blanco; M. Rico



An ICT-based fluorescent switch-on probe for hydrogen sulfide in living cells.  


An ICT-based fluorescent turn-on probe for hydrogen sulfide with high selectivity has been designed and synthesized. It exhibits up to 62-fold switch-on response toward H2S at given concentrations and can detect H2S in living cells with high sensitivity. PMID:23949387

Li, Xin; Zhang, Shuai; Cao, Ji; Xie, Nan; Liu, Tao; Yang, Bo; He, Qiaojun; Hu, Yongzhou



Optical detection of pesticides and drugs based on chemiluminescence–fluorescence assays  

Microsoft Academic Search

The paper illustrates two different concepts for monitoring pesticides and drugs. In the first approach double stranded deoxyribonucleic acid (dsDNA) was used as the sensing material, which intercalated drugs with high affinity. The intercalation of drugs such as Chlorpromazine (CP), Clomipramine (CM) and Imipramine (IP) were investigated. The assay was based on competition between the drug molecule and a fluorescent

Bengt Danielsson; Ioana Surugiu; Anatoli Dzgoev; Michael Mecklenburg; Kumaran Ramanathan



Targeted pH-dependent fluorescent activity-based cathepsin probes.  


Bifunctional, pH-activatable BODIPY dyes were developed and incorporated in mannose cluster-containing activity-based probes for cysteine proteases. Mannose receptor-dependent uptake of the probes in dendritic cells, followed by trafficking to acidic cellular compartments resulted in fluorescence as seen by live-cell imaging, and subsequent cathepsin inhibition. PMID:21769329

Hoogendoorn, Sascha; Habets, Kim L; Passemard, Solène; Kuiper, Johan; van der Marel, Gijsbert A; Florea, Bogdan I; Overkleeft, Herman S



Label-free fluorescent assays based on aptamer-target recognition.  


Label-free fluorescent assays were developed based on the competition of intramolecular DNA hybridization and aptamer-target binding. Using small molecule adenosine triphosphate (ATP) and biomacro-molecule thrombin as model targets, our design was proved to be a general method with good sensitivity and high selectivity. PMID:22451893

Tan, Ying; Zhang, Xin; Xie, Yonghua; Zhao, Rui; Tan, Chunyan; Jiang, Yuyang



Fiber optic based fluorescence detection system for in vivo studies of exogenous chromophore pharmacokinetics  

Microsoft Academic Search

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

Daniel R. Doiron; J. B. Dunn; W. L. Mitchell; Brian K. Dalton; Greta M. Garbo; Jon A. Warner



A distance-dependent metal-enhanced fluorescence sensing platform based on molecular beacon design.  


A new metal-enhanced fluorescence (MEF) based platform was developed on the basis of distance-dependent fluorescence quenching-enhancement effect, which combined the easiness of Ag-thiol chemistry with the MEF property of noble-metal structures as well as the molecular beacon design. For the given sized AgNPs, the fluorescence enhancement factor was found to increase with a d(6) dependency in agreement with fluorescence resonance energy transfer mechanism at shorter distance and decrease with a d(-3) dependency in agreement with plasmonic enhancement mechanism at longer distance between the fluorophore and the AgNP surface. As a proof of concept, the platform was demonstrated by a sensitive detection of mercuric ions, using thymine-containing molecular beacon to tune silver nanoparticle (AgNP)-enhanced fluorescence. Mercuric ions were detected via formation of a thymine-mercuric-thymine structure to open the hairpin, facilitating fluorescence recovery and AgNP enhancement to yield a limit of detection of 1nM, which is well below the U.S. Environmental Protection Agency regulation of the Maximum Contaminant Level Goal (10nM) in drinking water. Since the AgNP functioned as not only a quencher to reduce the reagent blank signal but also an enhancement substrate to increase fluorescence of the open hairpin when target mercuric ions were present, the quenching-enhancement strategy can greatly improve the detection sensitivity and can in principle be a universal approach for various targets when combined with molecular beacon design. PMID:24080216

Zhou, Zhenpeng; Huang, Hongduan; Chen, Yang; Liu, Feng; Huang, Cheng Zhi; Li, Na



Ratiometric fluorescent nanosensor based on water soluble carbon nanodots with multiple sensing capacities.  


A construction strategy for ratiometric fluorescent nanosensors based on water soluble C-dots was developed, which could sense temperature (10-82 °C), pH values (lower than 6.0 or higher than 8.6) and Fe(3+) ions (>0.04 ?M) by monitoring the intensity ratios of dual fluorescence bands (Ib/Ig) under 380 nm excitation. Ib/Ig decreased nearly linearly with increasing temperature from 10 to 82 °C. In the pH range from 8.6 to 6.0, the Ib/Ig was nearly constant at 0.75. Ib/Ig gradually decreased from 0.75 to 0.52 in the pH range from 6.0 to 1.9, and increased nearly linearly from 0.52 to 0.75 in the pH range from 1.9 to 1.0. The dual fluorescence behavior was reversible in the pH range from 1.0 to 8.6. As pH increased from 10.6 to 13.0, the green fluorescence band decreased continuously and blue shifted with a nearly linear increase in Ib/Ig from 0.75 to 2.15, while the green fluorescence band cannot be recovered by decreasing the pH value. Ib/Ig was ultrasensitive and selective in presence of Fe(3+) (>0.04 ?M) in neutral aqueous environments. The two fluorescence bands of the C-dots were attributed to different surface states that may produce different fluorescent signal responses to external physical or chemical stimuli. PMID:23673389

Qu, Songnan; Chen, Hong; Zheng, Xuanming; Cao, Junsheng; Liu, Xingyuan



Surface recognition and fluorescence sensing of histone by dansyl-appended cyclophane-based resorcinarene trimer.  


A cyclophane-based resorcinarene trimer (3) bearing a dansyl moiety as an environmentally sensitive fluorophore was prepared by stepwise condensation of a tetraaza[]paracyclophane skeleton with a dansyl moiety and three resorcinarene derivatives having heptacarboxylic acid residues in this sequence. The dansyl-appended cyclophane exhibited the following fluorescence properties regarding solvent polarity dependency and histone surface recognition: With increasing dioxane contents in dioxane/water solvents, the fluorescence intensity originating from the dansyl moiety of 3 increased along with a concomitant blue shift of the fluorescence maximum (lambdaem). The microenvironmentally sensitive fluorescence properties of dansyl fluorophore were maintained, even when the dansyl moiety was covalently attached to a cyclophane. Most interestingly, the cyclophane-based resorcinarene trimer exhibited recognition and fluorescence sensing capabilities toward histone, a small basic protein of eukaryotic chromatins. The fluorescence intensity originating from 3 increased along with a concomitant blue shift of lambdaem upon the addition of histone, reflecting the formation of 3-histone complexes. A relatively large fluorescence polarization (P) value was obtained for the 3-histone complexes (0.15), reflecting highly restricted conformations of 3, and the obtained P value was much larger than that of 3 alone in aqueous medium (0.07). The binding constant (K) of 3 with histone (unit basis) was estimated to be 2.1 x 106 M-1. On the other hand, upon the addition of acetylated histone (Ac-histone) to an aqueous solution containing 3, the extent of change in fluorescence intensity originating from the dansyl group of 3 was almost negligible, indicating that the electrostatic interactions between 3 and Ac-histone were weak. In addition, the fluorescence spectral changes were also small or negligible upon the addition of other proteins such as albumin, ovalbumin, peanut agglutinin, myoglobin, concanavalin A, cytochrome c, and lysozyme, having isoelectric points of 4.7, 4.8, 5.7-6.7, 6.8, 7.1, 9, and 11.0, respectively, to an aqueous solution containing 3. PMID:17929813

Hayashida, Osamu; Ogawa, Naoyuki; Uchiyama, Masaki



A rhodamine-based fluorescent probe for detecting Hg(2+) in a fully aqueous environment.  


A water-soluble fluorescent probe for Hg(2+) based on a rhodamine B derivative was designed and synthesized. The new probe showed reversible colorimetric and fluorescent response to Hg(2+) in a fully aqueous solution. The probe exhibited real-time detection of Hg(2+) with high selectivity in media containing less than 1% organic cosolvent. Furthermore, bioimaging studies indicated that the new probe was cell permeable and suitable for the real-time imaging of Hg(2+) in living cells by confocal microscopy. PMID:23986178

Chen, Xiaoli; Meng, Xiangming; Wang, Shuxing; Cai, Yulei; Wu, Yifan; Feng, Yan; Zhu, Manzhou; Guo, Qingxiang



Turn-on fluorescent chemosensor for hg2+ based on multivalent rhodamine ligands.  


Rhodamine-based fluorescent chemosensors 1 and 2 exhibit selective fluorescence enhancement to Fe3+ and Hg2+ over other metal ions at 580 nm in CH(3)CN/H(2)O (3/1, v/v) solution. Bis(rhodamine) chemosensor 1, under optimized conditions (CH(3)CN/HEPES buffer (0.02 M, pH = 7.0) (95/5, v/v)), shows a high selectivity and sensitivity to Hg2+, with a linear working range of 0-50 ?M, a wide pH span of 4-10, and a detection limit of 0.4 ?M Hg2+. PMID:23222686

Wang, Xuemei; Iqbal, Mudassir; Huskens, Jurriaan; Verboom, Willem



Turn-On Fluorescent Chemosensor for Hg2+ Based on Multivalent Rhodamine Ligands  

PubMed Central

Rhodamine-based fluorescent chemosensors 1 and 2 exhibit selective fluorescence enhancement to Fe3+ and Hg2+ over other metal ions at 580 nm in CH3CN/H2O (3/1, v/v) solution. Bis(rhodamine) chemosensor 1, under optimized conditions (CH3CN/HEPES buffer (0.02 M, pH = 7.0) (95/5, v/v)), shows a high selectivity and sensitivity to Hg2+, with a linear working range of 0–50 ?M, a wide pH span of 4–10, and a detection limit of 0.4 ?M Hg2+.

Wang, Xuemei; Iqbal, Mudassir; Huskens, Jurriaan; Verboom, Willem



Molecular distribution sensing in a fluorescence resonance energy transfer based affinity assay for glucose  

NASA Astrophysics Data System (ADS)

A newly developed method for determining molecular distribution functions is applied to a widely researched glucose affinity sensor. The reduction in fluorescence resonance energy transfer (FRET) to a malachite green (MG)-dextran complex from allophycocyanin (APC) bound to concanavalin A (ConA) due to displacement of the complex by glucose from ConA provides the basis of the assay. The higher sensitivity and specificity of a new approach to fluorescence decay analysis, over the methods based on conventional Förster-type models, is demonstrated and critical parameters in competitive binding FRET sensing derived.

Rolinski, O. J.; Birch, D. J. S.; McCartney, L.; Pickup, J. C.



Solitary and edge-minimal bases for representations of the simple lie algebra G2  

Microsoft Academic Search

We consider two families of weight bases for “one-rowed” irreducible representations of the simple Lie algebra G2 constructed in Donnelly et al [Constructions of representations of o(2n+1,C) that imply Molev and Reiner–Stanton lattices are strongly Sperner, Discrete Math. 263 (2003) 61–79] using two corresponding families of distributive lattices (called “supporting graphs”), here denoted LG2LM(k) and LG2RS(k). We exploit the relationship

Robert G. Donnelly; Scott J. Lewis; Robert Pervine



A simple and cheap end-to-end voice encryption framework over GSM-based networks  

Microsoft Academic Search

This paper presents a framework for voice encryption over GSM-based networks. To evaluate the framework, we have designed and implemented prototypes. The prototypes support full-duplex communication and real-time communication. To make the prototypes cheap and simple, the RC4 algorithm is utilized to protect digital of voice. The Bluetooth technology is utilized to communicate between the voice encryption prototype and a

Prawit Chumchu; Attaphon Phayak; Prakaidao Dokpikul



Simple and sensitive method of microcantilever-based DNA detection using nanoparticles conjugates  

Microsoft Academic Search

As the real sample diagnosis using resonance-based microcantilevers, a simple method is strongly suggested that capture probe immobilized on the cantilever directly hybridize with nanoparticles conjugated detection probe. Two different dimensions of microfabricated piezoelectric cantilevers and two different sizes of nanoparticles were used in this experiment. As a result, the most suitable conditions, the 30times90 mum2 (width times length) of

Byung Hak Cha; Sang-Myung Lee; Kyo Seon Hwang; Sang Kyung Kim; Yoon-Sik Lee; Byeong-Kwon Ju; Tae Song Kim



A simple model for the conformations and structure of polyelectrolytes with acid-base equilibria  

Microsoft Academic Search

We investigate a simple model of polyelectrolytes at non-zero concentration with acid-base equilibria. We assume each monomer contributes electrostatic repulsion Uc and short-range attraction Us, and the interaction between monomers equals f Uc + (1-f) Us, where f is the monomer charge fraction due to the chemical equilibrium. We solve the model using the self-consistent PRISM theory. As the attraction

Chwen-Yang Shew; Arun Yethiraj



Simple inexpensive adaptive optical system for large ground-based telescopes at infrared wavelengths  

Microsoft Academic Search

Simple adaptive optical system are studied that can provide real-time atmospheric turbulence compensation at IR wavelengths for large ground-based astronomical telescopes. A tip-tilt mirror and a 7-element segmented mirror, a long with an iterative adaptive control method, called Far-Field Optimization, provide the turbulence correction. The primary advantage of far-field optimization is that no near-field phase measurements are required, thus eliminating

Naresh C. Mehta



Ka-Band Vector Reflectometer Based on Simple Phase-Shifter Design  

Microsoft Academic Search

An automated handheld simple vector reflectometer design for complex-reflection-coefficient measurements at 35.5 GHz (Ka-band, 26.5-40 GHz) is presented. The proposed reflectometer is based on a standing-wave voltage measuring device and an electronically controlled and custom-designed millimeter-wave phase shifter. The phase shifter consists of p-i-n diode-loaded subresonant slots cut into the wall of a Ka-band waveguide. This paper describes the design

Mohamed Ahmed Abou-Khousa; Mark A. Baumgartner; Sergey Kharkovsky; Reza Zoughi



Simple-Design Low-Noise NLTL-Based Frequency Synthesizers for a CPT Cs Clock  

Microsoft Academic Search

This paper presents simple-architecture low-noise frequency synthesis chains generating a 9-GHz signal. These devices are based on the use of a nonlinear transmission line (NLTL) used as a comb generator. The residual phase noise spectra of the key components are reported. The residual phase noise performance of the chains at 9 GHz is measured to be less than -80 dBrad2\\/Hz

Rodolphe Boudot; Stéphane Guérandel; Emeric de Clercq



Simple spectrophotometric method for determination of zirconium or hafnium in selected molybdenum-base alloys.  


A simple analytical procedure is described for determining zirconium or hafnium in molybdenum-base alloys by formation of the Arsenazo III complex of zirconium or hafnium in 9 M hydrochloric acid medium. The absorbance is measured at 670 nm. Molybdenum (10 mg), titanium (1 mg), and rhenium (10 mg) have no adverse effect. No prior separation is needed. The relative standard deviation is 1.3-2.7%. PMID:18961121

Dupraw, W A



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


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. PMID:22894519

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



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

NASA Astrophysics Data System (ADS)

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.

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



Light sheet-based fluorescence microscopy: more dimensions, more photons, and less photodamage.  


Light-sheet-based fluorescence microscopy (LSFM) is a fluorescence technique that combines optical sectioning, the key capability of confocal and two-photon fluorescence microscopes with multiple-view imaging, which is used in optical tomography. In contrast to conventional wide-field and confocal fluorescence microscopes, a light sheet illuminates only the focal plane of the detection objective lens from the side. Excitation is, thus, restricted to the fluorophores in the volume near the focal plane. This provides optical sectioning and allows the use of regular cameras in the detection process. Compared to confocal fluorescence microscopy, LSFM reduces photo bleaching and photo toxicity by up to three orders of magnitude. In LSFM, the specimen is embedded in a transparent block of hydrogel and positioned relative to the stationary light sheet using precise motorized translation and rotation stages. This feature is used to image any plane in a specimen. Additionally, multiple views obtained along different angles can be combined into a single data set with an improved resolution. LSFMs are very well suited for imaging large live specimens over long periods of time. However, they also perform well with very small specimens such as single yeast cells. This perspective introduces the principles of LSFM, explains the challenges of specimen preparation, and introduces the basics of a microscopy that takes advantage of multiple views. PMID:19404438

Reynaud, Emmanuel G; Krzic, Uros; Greger, Klaus; Stelzer, Ernst H K



Highly sensitive water-soluble fluorescent ph sensors based on the 7-amino-1-methylquinolinium chromophore.  


Highly sensitive water-soluble fluorescent pH sensors have been synthesized employing the 7-amino-1-methylquinolinium (7AMQ) chromophore. These compounds, which contained oligomethylene spacers and substituted amine receptor units, were prepared in high yields and purity by a single reaction from the readily available 7-fluoro-1-methylquinolinium iodide. Density functional theory (DFT) and semiempirical INDO/s calculations have been performed to describe the ground state and the locally excited state of the chromophore. The photophysics and the sensor characteristics have been investigated. Dissociation constants pK(A)* ranging from 5.8 to 9.9 have been obtained. An additional quenching process at pH 11, due to excited state deprotonation at N7, has been observed. Fluorescence quantum yields in the protonated "on-state" between 0.75 and 0.85 and fluorescence enhancements (FE) between 2 and 55 were determined. These values are significantly higher than those of molecules based on other CT-chromophores that contain identical spacer-receptor units. The high fluorescence enhancements may be explained by the low rate of fluorescence ( approximately 6 x 10(7) s(-1)) and the high excited state reduction potential ( approximately 1.6 eV) of the 7AMQ chromophore. PMID:20196564

Jager, Wolter F; Hammink, Tessel S; van den Berg, Otto; Grozema, Ferdinand C



Highly sensitive detection of superoxide dismutase based on an immunoassay with surface-enhanced fluorescence.  


Herein, a novel highly sensitive enhanced-fluorescence immunoassay for detection of superoxide dismutase (SOD) is established by combining surface-enhanced fluorescence (SEF) with immuno-magnetic separation. Based on a sandwich-type immunoassay, analytes in samples are first captured by magnetic beads coated with a monoclonal antibody and then "sandwiched" by another monoclonal antibody on silver nanoparticles labeled with fluorescein-labeled oligonucleotides in the presence of a magnet. Subsequently, the immune complex is enriched by exposure to a magnetic field. Lastly, the fluorescence intensity is measured according to the number of dissociated fluoresceins. The increased fluorescence intensity permits highly sensitive detection of SOD in a linear range of 10-8 × 10(5) pg mL(-1), with a detection limit of 4 pg mL(-1) at a signal-to-noise ratio of 3. Significantly, this method was validated for detection of SOD in human serum, human urine, and cosmetic samples. Moreover, the reliability and accuracy of results obtained by the enhanced-fluorescence method was confirmed by the analysis of high performance liquid chromatography (HPLC). PMID:23615635

Yang, Xiaoming; Dou, Yao; Zhu, Shanshan



Evaluation of fluorescence-based thermal shift assays for hit identification in drug discovery.  


The fluorescence-based thermal shift assay is a general method for identification of inhibitors of target proteins from compound libraries. Using an environmentally sensitive fluorescent dye to monitor protein thermal unfolding, the ligand-binding affinity can be assessed from the shift of the unfolding temperature (Delta Tm) obtained in the presence of ligands relative to that obtained in the absence of ligands. In this article, we report that the thermal shift assay can be conducted in an inexpensive, commercially available device for temperature control and fluorescence detection. The binding affinities obtained from thermal shift assays are compared with the binding affinities measured by isothermal titration calorimetry and with the IC(50) values from enzymatic assays. The potential pitfalls in the data analysis of thermal shift assays are also discussed. PMID:15301960

Lo, Mei-Chu; Aulabaugh, Ann; Jin, Guixian; Cowling, Rebecca; Bard, Jonathan; Malamas, Michael; Ellestad, George



A novel fiber optic biosensor for nitric oxide determination based on vicinal diaminobenzozcridine fluorescent probe  

NASA Astrophysics Data System (ADS)

A novel fiber optic biosensor for the determination of nitric oxide based on vicinal diaminobenzozcridine (VDABA) fluorescent probe was designed and fabricated. The reaction conditions between VDABA and NO, which include concentration of VDABA, temperature and pH, were studied in-depth. The sensitivity of VDABA for NO detection under the optimum conditions and its optical properties were also investigated. The fluorescence responses were concentration-dependent and a good linear relationship (R2=0.9863) was observed over the range 1.8×10-6 to 9×10-6 mol/L NO, the regression equation was F = 3.8889[NO] (mol/L)+217.2. Besides, a complex sensitive film embedding VDABA in cellulose acetate (CA) was prepared, and a fiber optic NO biosensor was fabricated using this film. Then the change of fluorescence phase shift of this biosensor was studied preliminarily by means of the lock-in technology.

Ding, Liyun; Huang, Lanfen; Huang, Jun; Zhong, Yunming; Fan, Dian



Bio-assay based on single molecule fluorescence detection in microfluidic channels.  


A rapid bioassay is described based on the detection of colocalized fluorescent DNA probes bound to DNA targets in a pressure-driven solution flowing through a planar microfluidic channel. By employing total internal reflection excitation of the fluorescent probes and illumination of almost the entire flow channel, single fluorescent molecules can be efficiently detected leading to the rapid analysis of nearly the entire solution flowed through the device. Cross-correlation between images obtained from two spectrally distinct probes is used to determine the target concentration and efficiently reduces the number of false positives. The rapid analysis of DNA targets in the low pM range in less than a minute is demonstrated. PMID:16802123

Hollars, Christopher W; Puls, Jana; Bakajin, Olgica; Olsan, Brad; Talley, Chad E; Lane, Stephen M; Huser, Thomas



Assessment of fluorescein-based fluorescent dyes for tracing Neotyphodium endophytes in planta.  


Fluorescent dyes were assessed for their ability to stain viable hyphae of the fungi Neotyphodium lolii and N. coenophialum, symbiotic endophytes of the Pooideae grasses Lolium perenne and Festuca arundinacea, respectively. The fluorescein-based fluorophores; fluorescein diacetate (FDA), 5(6)-carboxy-fluorescein diacetate (CFDA), 5-chloromethylfluorescein diacetate (CMFDA) and the chitin-binding stain, Calcofluor while M2R, were assessed for staining of endophyte hyphae in vitro from axenic fungal cultures and in planta, including epidermal leaf sheath peels, nodes, ovaries, embryos and meristems. CMFDA produced the greatest intensity of staining of fungal hyphae and gave excellent contrast in planta compared to the plant cells. Compared to the other dyes, CMFDA was also the least affected by photo bleaching and continued to fluoresce up to 2 h after initial excitation. None of the fluorescent dyes stained fungal hyphae in seed. PMID:22802389

Card, Stuart D; Tapper, Brian A; Lloyd-West, Catherine; Wright, Kathryn M



Cell-based fluorescence screen for K+ channels and transporters using an extracellular triazacryptand-based K+ sensor.  


K+ channels and K+-coupled membrane transporters are important targets for drug discovery. We previously developed a triazacryptand (TAC)-based K+ sensor, TAC-Red, and demonstrated its utility to image K+ waves in mouse brain in vivo (Padmawar et al. Nat. Methods. 2005, 2, 825-827). Here, we synthesized a green-fluorescing dextran conjugate of TAC-bodipy ("TAC-Limedex") for use as an extracellular K+ sensor and demonstrated its utility in measuring K+ transport across cell membranes. TAC-Limedex fluorescence increased by 50% with increasing [K+] from 0 to 2 mM and was insensitive to [Na+], [Cl-], or pH. K+ efflux from cells was quantified from increasing extracellular TAC-Limedex fluorescence following cell immersion in K+-free buffer. In HT-29 cells, K+ efflux was 2.0 +/- 0.1 micromol/cm2/s, increasing 8-fold following K+ channel activation by ATP; the increase in K+ efflux was inhibited by a K+ channel blocker or by preventing cytoplasmic calcium elevation. Electroneutral K+/Cl- cotransport was demonstrated in SiHa cells, in which K+ efflux was increased 3-fold by hypotonic challenge; the increase in K+ efflux was fully inhibited by a K+/Cl- transport blocker. K+ efflux measurements were adapted to a commercial fluorescence platereader for automated screening. The fluorescence-based K+ transport assay largely replaces assays requiring radioactive rubidium and is suitable for high-throughput identification of K+ transport modulators. PMID:18512924

Namkung, Wan; Padmawar, Prashant; Mills, Aaron D; Verkman, A S



A fluorescence-based assay for the measurement of S-adenosylhomocysteine hydrolase activity in biological samples.  


The methylation of DNA, RNA, and proteins plays crucial roles in numerous biological processes, including epigenetic control, virus replication, and cell differentiation. In mammals, the rate-limiting step of the S-adenosylmethionine-dependent methylation process is exclusively controlled by S-adenosylhomocysteine (S-AdoHcy) hydrolase (SAHH). SAHH hydrolyzes S-AdoHcy to adenosine and homocysteine (Hcy) and is therefore a potential therapeutic target for various diseases, including cancer, malaria, and viral diseases. However, a simple and highly sensitive assay for the evaluation of SAHH activity, particularly for drug discovery, had not yet been developed. Here we present the development of a fluorescence-based assay for the measurement of SAHH activity in biological samples. We combined the advantages of the detection of fluorescent thiol groups in Hcy by ThioGlo1 with the S-AdoHcy-driven enzyme-coupled reaction. Our results confirmed the reliability of the proposed assay for the measurement of the SAHH activity of purified SAHH and showed the potential of this assay for the measurement of the SAHH activity of biological samples. Therefore, the proposed SAHH activity assay may be utilized in clinical laboratories and in high-throughput screenings for the identification of new SAHH inhibitors with potentially beneficial effects on numerous pathologies. PMID:23079506

Hudec, Roman; Hamada, Kozo; Mikoshiba, Katsuhiko



Fluorescence-based quantitative scratch wound healing assay demonstrating the role of MAPKAPK-2/3 in fibroblast migration.  


The scratch wound healing assay is a sensitive method to characterize cell proliferation and migration, but it is difficult to be quantitatively evaluated. Therefore, we developed an infrared fluorescence detection-based real-time assay for sensitive and accurate quantification of cell migration in vitro. The method offers sensitivity, simplicity, and the potential for integration into automated large-scale screening studies. A live cell staining lipophilic tracer-1,1'-dioctadecyl-3,3,3',3'-tetramethyl indotricarbocyanine iodide (DiR)-is used for accurate imaging of wound closure in a simple 96-well scratch assay. Scratches are made on prestained confluent cell monolayers using a pipette tip and scanned at different time intervals using a fluorescent scanner. Images are analyzed using Image J software and the migration index is calculated. Effect of cell number, time after scratch and software settings are analyzed. The method is validated by showing concentration- and time-dependent effects of cytochalasin-D on fibroblast migration. Using this assay, we quantitatively evaluate the role of the MAPK-activated protein kinases MK2 and MK3 in fibroblast migration. First, the migratory phenotype of MK2-deficient MEFs is analyzed in a retroviral rescue model. In addition, migration of MK2/3-double-deficient cells is determined and the ability of MK3 to rescue cell migration in MK2/3-double-deficient fibroblasts is demonstrated. PMID:19743408

Menon, Manoj B; Ronkina, Natalia; Schwermann, Jessica; Kotlyarov, Alexey; Gaestel, Matthias



Analog Integrated Circuit for Motion Detection with Simple-Shape Recognition Based on Frog Vision System  

NASA Astrophysics Data System (ADS)

We proposed in this research a novel two-dimensional network based on the frog visual system, with a motion detection function and a newly developed simple-shape recognition function, for use in object discrimination by integrated circuits. Specifically, the network mimics the signal processing of the small-field cell in a frog brain, consisting of the tectum and thalamus, which generates signals of the motion and simple shape of an object. The proposed network is constructed from simple analog complementary metal oxide semiconductor (CMOS) circuits; a test chip of the proposed network was fabricated with a 1.2 ?m CMOS process. Measurements on the chip clarified that the proposed network can generate signals of the moving direction, velocity, and simple shape, as well as perform information processing of the small-field cell. Results with the simulation program with integrated circuit emphasis (SPICE) showed that the analog circuits used in the network have low power consumption. Applications of the proposed network are expected to realize advanced vision chips with functions such as object discrimination and target tracking.

Nishio, Kimihiro; Yonezu, Hiroo; Furukawa, Yuzo



Nontemplated Approach to Tuning the Spectral Propertiesof Cyanine-Based Fluorescent NanoGUMBOS  

SciTech Connect

Template-free controlled aggregation and spectral properties in fluorescent organic nanoparticles (FONs) is highly desirable for various applications.Herein, we report a nontemplated method for controlling the aggregation in near-infrared (NIR) cyanine-based nanoparticles derived from a group of uniformmaterials based on organic salts (GUMBOS). Cationic heptamethine cyanine dye 1,10,3,3,30,30-hexamethylindotricarbocyanine (HMT) was coupled with five different anions, viz., [NTf2 -], [BETI-], [TFPB-], [AOT-], and [TFP4B-], by an ion-exchange method to obtain the respective GUMBOS. The nanoGUMBOS obtained via a reprecipitation method were primarily amorphous and spherical (30-100 nm) as suggested by selected area electron diffraction (SAED) and transmission electron microscopy (TEM). The formation of tunable self-assemblies within the nanoGUMBOS was characterized using absorption and fluorescence spectroscopy in conjunction with molecular dynamics simulations. Counterion-controlled spectral properties observed in the nanoGUMBOS were attributed to variations in J/H ratios with different anions. Association with the [AOT-] anion afforded predominant J aggregation enabling the highest fluorescence intensity, whereas [TFP4B-] disabled the fluorescence due to predominantHaggregation in the nanoparticles. Analyses of the stacking angle of the cations based on molecular dynamic simulation results in [HMT][NTf2], [HMT][BETI], and [HMT][AOT] dispersed in water and a visual analysis of the representative simulation snapshots also imply that the type of aggregation was controlled through the counterion associated with the dye cation.

Das, Susmita [Louisiana State University; Bwambok, David [Louisiana State University; El-Zahab, Bilal [Lousianna State University; Monk, Joshua [Louisiana State University; De Rooy, Sergio [Louisiana State University; Challa, Santhosh [Louisiana State University; Li, Min [Lousianna State University; Hung, Francisco [Louisiana State University; Baker, Gary A [ORNL; Warner, Isiah M [ORNL



Ultrathin oligonucleotide layers for fluorescence-based DNA sensors  

NASA Astrophysics Data System (ADS)

Preliminary investigations into the design of an affinity sensor using evanescent wave technology concentrate upon the means of immobilization of the receptor molecules. In this work DNA served as the selective recognition element. The molecular principle of a sequence-selective biosensor for DNA is based on a sandwich-hybridization assay wherein the analyte, a single-stranded (ss)DNA, bound specifically to both an immobilized capture probe and a dye-labeled oligonucleotide in free solution. The efficiency of the capture array depends on the density of highly organized oligonucleotides on the waveguide surface and correlates therefore directly with the specificity and the sensitivity of the sensor. In the present approach using the Langmuir- Blodgett technique cinnamoylbutylether-cellulose monolayers were transferred onto optical fibers or planar waveguides. These films served as matrices for the immobilization of biotinylated oligonucleotides via streptavidin. For the first time streptavidin was immobilized by that manner. The specificity of the streptavidin layer or the following bounded nucleic acid molecules were controlled by an enzyme- linked immunosorbent assay (ELISA). Finally, this application has also shown to be suitable for the detection of Salmonella, which is an important pathogen associated with acute gastroenteritidis and food borne diseases.

Furch, M.; Ueberfeld, J.; Hartmann, Andreas; Bock, Daniel; Seeger, Stefan



Using quantum mechanics to investigate the photophysical properties of the DNA and RNA bases and their fluorescent analogs  

NASA Astrophysics Data System (ADS)

The ability of the nucleic acids to absorb ultraviolet light and remain relatively photostable is a property upon which life depends. The nucleobases, which are the primary chromophores, when irradiated display rapid radiationless decay back to the ground state, in general faster than is needed for photoreaction. Fluorescent analogs of these bases have structures similar to the nucleic acid bases, but display much longer excited state lifetimes. Theoretical investigations using quantum mechanical methods can provide insight into the precise mechanisms of these decay processes, and to the molecular specifics that contribute to them. The results of multi-reference configuration interaction (MRCI) ab initio investigations into these mechanisms are presented, with emphasis on cytosine and its fluorescent analog 5-methyl-2-pyrimidinone (5M2P). A comprehensive picture of the potential energy surfaces of these two bases is given, including stationary points and conical intersections, where radiationless transisitons are promoted, between up to three state surfaces, as well as pathways connecting these points for each base. Cytosine is shown to have two different energetically accessible radiationless decay channels. The fluorescence of 5M2P is also demonstrated theoretically, with mechanism proposed. The potential energy surfaces of the two bases have many close similarities, with the different photophysical properties being attributed to subtle energetic differences between the two bases. Nonadiabatic coupling and the geometric phase effect are analyzed in detail near conical intersections in cytosine, including in a region close to a three-state conical intersection. A substituent effect study on the 2-pyrimidinone ring system shows that the presence, position and orientation of the amino group in cytosine is central to its photophysical properties, particulary its high absorption energy, and can be explained with a simple Frontier Molecular Orbital model. The effects of water solvent on the excitation energies of cytosine and uracil are theoretically investigated using two multi-reference ab initio methods, a quantum mechanical molecular mechanics method using MRCI (MRCI-QM/MM), and the fragment molecular orbital multiconfiguration self-consistent field method (FMO-MCSCF). The solvatochromic shifts calculated from both methods agree well with other more expensive methods and experimental data. The effects of water on the photophysical pathways of cytosine is also investigated using MRCI-QM/MM, including considerations of solvent reorganization. Results show that the overall effect of water on the decay mechanisms is small, with neither decay channel being significantly blocked or favored.

Kistler, Kurt A.


Green fluorescent protein-based monitoring of endoplasmic reticulum redox poise  

PubMed Central

Pathological endoplasmic reticulum (ER) stress is tightly linked to the accumulation of reactive oxidants, which can be both upstream and downstream of ER stress. Accordingly, detrimental intracellular stress signals are amplified through establishment of a vicious cycle. An increasing number of human diseases are characterized by tissue atrophy in response to ER stress and oxidative injury. Experimental monitoring of stress-induced, time-resolved changes in ER reduction-oxidation (redox) states is therefore important. Organelle-specific examination of redox changes has been facilitated by the advent of genetically encoded, fluorescent probes, which can be targeted to different subcellular locations by means of specific amino acid extensions. These probes include redox-sensitive green fluorescent proteins (roGFPs) and the yellow fluorescent protein-based redox biosensor HyPer. In the case of roGFPs, variants with known specificity toward defined redox couples are now available. Here, we review the experimental framework to measure ER redox changes using ER-targeted fluorescent biosensors. Advantages and drawbacks of plate-reader and microscopy-based measurements are discussed, and the power of these techniques demonstrated in the context of selected cell culture models for ER stress.

Birk, Julia; Ramming, Thomas; Odermatt, Alex; Appenzeller-Herzog, Christian



Aptasensor for ampicillin using gold nanoparticle based dual fluorescence-colorimetric methods.  


A gold nanoparticle based dual fluorescence-colorimetric method was developed as an aptasensor to detect ampicillin using its single-stranded DNA (ssDNA) aptamer, which was discovered by a magnetic bead-based SELEX technique. The selected aptamers, AMP4 (5'-CACGGCATGGTGGGCGTCGTG-3'), AMP17 (5'-GCGGGCGGTTGTATAGCGG-3'), and AMP18 (5'-TTAGTTGGGGTTCAGTTGG-3'), were confirmed to have high sensitivity and specificity to ampicillin (K(d), AMP7 = 9.4 nM, AMP17 = 13.4 nM, and AMP18 = 9.8 nM, respectively). The 5'-fluorescein amidite (FAM)-modified aptamer was used as a dual probe for observing fluorescence differences and color changes simultaneously. The lower limits of detection for this dual method were a 2 ng/mL by fluorescence and a 10 ng/mL by colorimetry for ampicillin in the milk as well as in distilled water. Because these detection limits were below the maximum residue limit of ampicillin, this aptasensor was sensitive enough to detect antibiotics in food products, such as milk and animal tissues. In addition, this dual aptasensor will be a more accurate method for antibiotics in food products as it concurrently uses two detection methods: fluorescence and colorimetry. PMID:22222912

Song, Kyung-Mi; Jeong, Euiyoung; Jeon, Weejeong; Cho, Minseon; Ban, Changill



Blood interference in fiber-optical based fluorescence guided resection of glioma using 5-aminolevulinic acid  

NASA Astrophysics Data System (ADS)

Fluorescence guidance in brain tumor resection is performed intra-operatively where bleeding is included. When using fiber-optical probes, the transmission of light to and from the tissue is totally or partially blocked if a small amount of blood appears in front of the probe. Sometimes even after rinsing with saline, the remnant blood cells on the optical probe head, disturb the measurements. In such a case, the corresponding spectrum cannot be reliably quantified and is therefore discarded. The optimal case would be to calculate and take out the blood effect systematically from the collected signals. However, the first step is to study the pattern of blood interference in the fluorescence spectrum. In this study, a fiber-optical based fluorescence spectroscopy system with a laser excitation light of 405 nm (1.4 J/cm2) was used during fluorescence guided brain tumor resection using 5-aminolevulinic acid (5-ALA). The blood interference pattern in the fluorescence spectrum collected from the brain was studied in two patients. The operation situation was modeled in the laboratory by placing blood drops from the finger tip on the skin of forearm and the data was compared to the brain in vivo measurements. Additionally, a theoretical model was developed to simulate the blood interference pattern on the skin autofluorescence. The blood affects the collected fluorescence intensity and leaves traces of oxy and deoxy-hemoglobin absorption peaks. According to the developed theoretical model, the autofluorescence signal is considered to be totally blocked by an approximately 500 ?m thick blood layer.

Haj-Hosseini, Neda; Lowndes, Shannely; Salerud, Göran; Wårdell, Karin



Conditionally fluorescent molecular probes for detecting single base changes in double-stranded DNA  

NASA Astrophysics Data System (ADS)

Small variations in nucleic acid sequences can have far-reaching phenotypic consequences. Reliably distinguishing closely related sequences is therefore important for research and clinical applications. Here, we demonstrate that conditionally fluorescent DNA probes are capable of distinguishing variations of a single base in a stretch of target DNA. These probes use a novel programmable mechanism in which each single nucleotide polymorphism generates two thermodynamically destabilizing mismatch bubbles rather than the single mismatch formed during typical hybridization-based assays. Up to a 12,000-fold excess of a target that contains a single nucleotide polymorphism is required to generate the same fluorescence as one equivalent of the intended target, and detection works reliably over a wide range of conditions. Using these probes we detected point mutations in a 198 base-pair subsequence of the Escherichia coli rpoB gene. That our probes are constructed from multiple oligonucleotides circumvents synthesis limitations and enables long continuous DNA sequences to be probed.

Chen, Sherry Xi; Zhang, David Yu; Seelig, Georg



Rapid High-Throughput Assessment of Aerobic Bacteria in Complex Samples by Fluorescence-Based Oxygen Respirometry  

Microsoft Academic Search

A simple method has been developed for the analysis of aerobic bacteria in complex samples such as broth and food homogenates. It employs commercial phosphorescent oxygen-sensitive probes to monitor oxygen consumption of samples containing bacteria using standard microtiter plates and fluorescence plate readers. As bacteria grow in aqueous medium, at certain points they begin to deplete dissolved oxygen, which is

Fiach C. O'Mahony; Dmitri B. Papkovsky



Fluorescence-based sensing of glucose using engineered glucose/galactose-binding protein: A comparison of fluorescence resonance energy transfer and environmentally sensitive dye labelling strategies  

SciTech Connect

Fluorescence-based glucose sensors using glucose-binding protein (GBP) as the receptor have employed fluorescence resonance energy transfer (FRET) and environmentally sensitive dyes, but with widely varying sensitivity. We therefore compared signal changes in (a) a FRET system constructed by transglutaminase-mediated N-terminal attachment of Alexa Fluor 488/555 as donor and QSY 7 as acceptor at Cys 152 or 182 mutations with (b) GBP labelled with the environmentally sensitive dye badan at C152 or 182. Both FRET systems had a small maximal fluorescence change at saturating glucose (7% and 16%), badan attached at C152 was associated with a 300% maximal fluorescence increase with glucose, though with badan at C182 there was no change. We conclude that glucose sensing based on GBP and FRET does not produce a larger enough signal change for clinical use; both the nature of the environmentally sensitive dye and its site of conjugation seem important for maximum signal change; badan-GBP152C has a large glucose-induced fluorescence change, suitable for development as a glucose sensor.

Khan, Faaizah; Gnudi, Luigi [Metabolic Unit, King's College London School of Medicine, Guy's Hospital, London SE1 9RT (United Kingdom); Pickup, John C. [Metabolic Unit, King's College London School of Medicine, Guy's Hospital, London SE1 9RT (United Kingdom)], E-mail:



Patterning pallet arrays for cell selection based on high-resolution measurements of fluorescent biosensors  

PubMed Central

Pallet arrays enable cells to be separated while they remain adherent to a surface and provide a much greater range of cell selection criteria relative to that of current technologies. However there remains a need to further broaden cell selection criteria to include dynamic intracellular signaling events. To demonstrate the feasibility of measuring cellular protein behavior on the arrays using high resolution microscopy, the surfaces of individual pallets were modified to minimize the impact of scattered light at the pallet edges. The surfaces of the three-dimensional pallets on an array were patterned with a coating such as fibronectin using a customized stamping tool. Micropatterns of varying shape and size were printed in designated regions on the pallets in single or multiple steps to demonstrate the reliability and precision of patterning molecules on the pallet surface. Use of a fibronectin matrix stamped at the center of each pallet permitted the localization of H1299 and mouse embryonic fibroblast (MEF) cells to the pallet centers and away from the edges. Compared to pallet arrays with fibronection coating the entire top surface, arrays with a central fibronectin pattern increased the percentage of cells localized to the pallet center by 3-4 fold. Localization of cells to the pallet center also enabled the physical separation of cells from optical artifacts created by the rough pallet side walls. To demonstrate the measurement of dynamic intracellular signaling on the arrays, fluorescence measurements of high spatial resolution were performed using a RhoA GTPase biosensor. This biosensor utilized fluorescence resonance energy transfer (FRET) between cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP) to measure localized RhoA activity in cellular ruffles at the cell periphery. These results demonstrated the ability to perform spatially resolved measurements of fluorescence-based sensors on the pallet arrays. Thus, the patterned pallet arrays should enable novel cell separations in which cell selection is based on complex cellular signaling properties.

Shadpour, Hamed; Zawistowski, Jon S.; Herman, Annadele; Hahn, Klaus; Allbritton, Nancy L.



Ratiometric fluorescence detection of mercuric ion based on the nanohybrid of fluorescence carbon dots and quantum dots.  


A novel nanohybrid ratiometric fluorescence probe comprised of carbon dots (C-dots) and hydrophilic CdSe@ZnS quantum dots (QDs) has been developed by simply mixing the blue-emission C-dots with red-emission carboxylmethyldithiocarbamate modified CdSe@ZnS QDs (GDTC-QDs). The nanohybrid ratiometric fluorescence probe exhibits dual emissions at 436 nm and 629 nm under a single excitation wavelength. Due to the strong chelating ability of GDTC on the surface of QDs to mercuric ion (Hg(2+)), the fluorescence of the GDTC-QDs in the nanohybrid system could be selectively quenched in the presence of Hg(2+) while the fluorescence of the C-dots remained constant, resulting in an obviously distinguishable fluorescence color evolution (from red to blue) of the nanohybrid system. The detection limit of this method was found to be as low as 0.1 ?M. Furthermore, the recovery result for Hg(2+) in real samples including tap water and lake water by this method was satisfying, suggesting its potential application for Hg(2+) sensing. PMID:23790304

Cao, Benmei; Yuan, Chao; Liu, Bianhua; Jiang, Changlong; Guan, Guijian; Han, Ming-Yong



Structural and dynamical aspects of skin studied by multiphoton excitation fluorescence microscopy-based methods.  


This mini-review reports on applications of particular multiphoton excitation microscopy-based methodologies employed in our laboratory to study skin. These approaches allow in-depth optical sectioning of the tissue, providing spatially resolved information on specific fluorescence probes' parameters. Specifically, by applying these methods, spatially resolved maps of water dipolar relaxation (generalized polarization function using the 6-lauroyl-2-(N,N-dimethylamino)naphthale probe), activity of protons (fluorescence lifetime imaging using a proton sensitive fluorescence probe - 2,7-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein) and diffusion coefficients of distinct fluorescence probes (raster imaging correlation spectroscopy) can be obtained from different regions of the tissue. Comparative studies of different tissue strata, but also between equivalent regions of normal and abnormal excised skin, including applications of fluctuation correlation spectroscopy on transdermal penetration of liposomes are presented and discussed. The data from the different studies reported reveal the intrinsic heterogeneity of skin and also prove these strategies to be powerful noninvasive tools to explore structural and dynamical aspects of the tissue. PMID:23608611

Bloksgaard, Maria; Brewer, Jonathan; Bagatolli, Luis A



Design and validation of a homogeneous time-resolved fluorescence-based leptin receptor binding assay.  


The pleiotropic cytokine hormone leptin, by activating its receptor OB-R, plays a major role in many biological processes, including energy homeostasis, immune function, and cell survival and proliferation. Abnormal leptin action is associated with obesity, autoimmune diseases, and cancer. The pharmacological characterization of OB-R and the development of synthetic OB-R ligands are still in their infancy because currently available binding assays are not compatible with ligand saturation binding experiments and high-throughput screening (HTS) approaches. We have developed here a novel homogeneous time-resolved fluorescence-based binding assay that overcomes these limitations. In this assay, fluorescently labeled leptin or leptin antagonist binds to the SNAP-tagged OB-R covalently labeled with terbium cryptate (Tb). Successful binding is monitored by measuring the energy transfer between the Tb energy donor and the fluorescently labeled leptin energy acceptor. Ligand binding saturation experiments revealed high-affinity dissociation constants in the subnanomolar range with an excellent signal-to-noise ratio. The assay performed in a 384-well format shows high specificity and reproducibility, making it perfectly compatible with HTS applications to identify new OB-R agonists or antagonists. In addition, fluorescently labeled leptin and SNAP-tagged OB-R will be valuable tools for monitoring leptin and OB-R trafficking in cells and tissues. PMID:23333588

Vauthier, Virginie; Derviaux, Carine; Douayry, Najim; Roux, Thomas; Trinquet, Eric; Jockers, Ralf; Dam, Julie



Ultrasound-modulated fluorescence based on a fluorophore-quencher-labeled microbubble system  

NASA Astrophysics Data System (ADS)

Ultrasound-modulated fluorescence from a fluorophore-quencher-labeled microbubble system driven by a single ultrasound pulse was theoretically quantified by solving a modified Herring equation (for bubble oscillation), a two-energy-level rate equation (for fluorophore excitation), and a diffusion equation (for light propagation in tissue). The efficiency of quenching caused by fluorescence resonance energy transfer (FRET) between the fluorophore and the quencher was modulated when the microbubble oscillates in size driven by the ultrasound pulse. Both intensity- and lifetime-based imaging methods are discussed in three different illumination modes of the excitation light: continuous wave (DC), frequency domain (FD), and time domain (TD). Results show that microbubble expansion opens a time period during which the quenching efficiency is dramatically reduced so that the emitted fluorescence strength and fluorophore lifetime are significantly increased. The modulation efficiency may even reach 100%. In addition, an important finding in this study is that in TD illumination mode, the modulated fluorescence photons may be temporally separated from the unmodulated photons, which makes the modulation efficiency limited only by thermal noise of the measurement system.

Yuan, Baohong



Ultrasound-modulated fluorescence based on a fluorophore-quencher-labeled microbubble system.  


Ultrasound-modulated fluorescence from a fluorophore-quencher-labeled microbubble system driven by a single ultrasound pulse was theoretically quantified by solving a modified Herring equation (for bubble oscillation), a two-energy-level rate equation (for fluorophore excitation), and a diffusion equation (for light propagation in tissue). The efficiency of quenching caused by fluorescence resonance energy transfer (FRET) between the fluorophore and the quencher was modulated when the microbubble oscillates in size driven by the ultrasound pulse. Both intensity- and lifetime-based imaging methods are discussed in three different illumination modes of the excitation light: continuous wave (DC), frequency domain (FD), and time domain (TD). Results show that microbubble expansion opens a time period during which the quenching efficiency is dramatically reduced so that the emitted fluorescence strength and fluorophore lifetime are significantly increased. The modulation efficiency may even reach 100%. In addition, an important finding in this study is that in TD illumination mode, the modulated fluorescence photons may be temporally separated from the unmodulated photons, which makes the modulation efficiency limited only by thermal noise of the measurement system. PMID:19405771

Yuan, Baohong


Sensitive signal-on fluorescent sensing for copper ions based on the polyethyleneimine-capped silver nanoclusters-cysteine system.  


In this work, we present a label-free sensor for copper ions. This sensor is composed of silver nanoclusters and cysteine. The fluorescence of the silver nanoclusters was quenched by cysteine, which was recovered in the presence of copper ions. This binding of silver nanoclusters to cysteine promoted agglomeration of silver nanoclusters to yield larger non-fluorescent silver nanoparticles. The presence of copper ions resulted in the oxidation of cysteine to form a disulfide compound, leading to recovery of fluorescence of the silver nanoclusters. The fluorescence of the silver nanoclusters in the presence of cysteine increased with increasing concentration of copper ions in the range of 10-200 nM. The detection limit of this sensor for copper ions was 2.3 nM. The silver nanoclusters-cysteine sensor provides a simple, cost-effective, and sensitive platform for the detection of copper ions. PMID:23890605

Zhang, Na; Qu, Fei; Luo, Hong Qun; Li, Nian Bing



A device for fluorescence temperature measurement based on fast fourier transform  

NASA Astrophysics Data System (ADS)

A sapphire fiber thermal probe with Cr3+ ion-doped end was grown using the laser heated pedestal method. The fluorescence thermal probe offers advantages of compact structure, high performance and the ability to sustain high temperature from the room temperature to 450 °C. Based on the fast fourier transform (FFT), the fluorescence lifetime is obtained from the tangent function of the phase angle of the first non-zeroth item of FFT result. Compared with other traditional fitting methods, our method has advantages such as fast speed, high accuracy and being free from the influence of the base signal. The standard deviation of FFT method is about half of that of the Prony method and close to the one of the Marquardt method. In addition, since the FFT method is immunity to the background noise of the signal, the background noise analysis can be skipped.

Wang, Dong-Sheng; Wang, Gui-Mei; Pan, Wei-Wei; Wang, Yu-Tian



The SlideReactor--a simple hollow fiber based bioreactor suitable for light microscopy.  


Most bioartificial liver support systems are based on hollow fiber capillaries within modified dialysis cartridges or more sophisticated bioreactor constructions. Due to their design microscopic follow-up of reorganization and growth of tissue between the hollow fibers is not possible. The SlideReactor is a simple hollow fiber based bioreactor construction suitable for light microscopy and time-lapse video observation. The SlideReactor offers a cell compartment separated from a medium inflow and outflow compartment. Cell compartment access ports enable easy filling of the cell compartment with cell suspension, as well as fixation of the tissue. For more complex procedures or full access to all the cells, the bioreactor can be opened easily by cutting the silicone seal with a scalpel. Due to its simple design and the utilization of standard materials, it could serve as a suitable, cost-efficient tool to evaluate the behavior of cells cultured between hollow fiber capillaries. The paper describes the production process: similar to open source projects in software engineering, we would like to propose the concept as an open platform to anyone interested in hollow fiber based cell culture. PMID:15725230

Sauer, Igor M; Schwartlander, Ruth; Schmid, Jochen; Efimova, Ekaterina; Vondran, Florian W R; Kehr, Daniel; Pless, Gesine; Spinelli, Antonio; Brandenburg, Börries; Hildt, Eberhardt; Neuhaus, Peter



The Circadian Energy Scale (CIRENS): two simple questions for a reliable chronotype measurement based on energy.  


This study presents the Circadian Energy Scale (CIRENS), a very short and simple chronotype measurement tool based on energy. The CIRENS consists of two introspective questions about the usual energy level (very low, low, moderate, high, or very high, scored 1 to 5) in the morning and in the evening. The difference between energy level scores (-4 to 4) felt by respondents in the evening and morning defines the chronotype score and classification. A concurrent validity analysis of the CIRENS with the widely used Horne and Östberg Morningness-Eveningness Questionnaire (MEQ) was conducted using a sample of 225 college students, and with MSFsc, a sleep-based chronotype assessment tool based on the Munich Chronotype Questionnaire (MCTQ), using a sample of 34,530 subjects (18-83 yrs, 27% males). This large sample was collected in a Web survey for behavioral correlates of the CIRENS with variables previously associated with chronotype differences. The correlation of the CIRENS chronotype score was r?=?-.70 with the MEQ and r?=?.32 with the MSFsc. CIRENS chronotype scores declined with age and were not affected by sex. Both CIRENS and MSFsc chronotype scores were related to differences in tobacco, caffeine, and cola soft-drink consumption (all higher in evening types). The CIRENS provides a simple chronotype index and a measure of absolute energy throughout the day and seems to be a reliable chronotype assessment tool that may be useful both clinically and for large-scale studies. PMID:21452918

Ottoni, Gustavo L; Antoniolli, Eduardo; Lara, Diogo R



Polarization-selective beam splitter based on a highly efficient simple binary diffraction grating.  


A polarization beam splitter (PBS) based on a giant-reflection to zero-order (GIRO) grating is presented. The GIRO grating is a simple binary diffraction grating with parameters chosen such that the excited optical modes in the grating interfere constructively and destructively at the respective interfaces. This interference results in high-zero-order reflection (>99%) with a high polarization-selective extinction ratio (+/-30 dB). The grating shows a low aspect ratio. The GIRO PBS is theoretically and experimentally shown to be an adequate PBS for use as an optical isolator in combination with a quarter-wave plate in a CO2-laser system. PMID:15605556

Delbeke, Danaë; Baets, Roel; Muys, Peter



A simple algorithm for generating streamflow networks for grid-based, macroscale hydrological models  

NASA Astrophysics Data System (ADS)

A simple algorithm for generating streamflow networks for macroscale hydrological models (MHMs) from digital elevation models (DEMs) is presented. Typically these hydrological models are grid based, with the simulated runoff produced within each cell routed through a stream network which connects the centers of cells in the direction of the major streams. Construction of such stream networks is a time consuming task, which has generally been done by hand with the aid of maps. Results indicate that the algorithm works satisfactorily in areas of both high and low relief, and for a wide range of model cell resolutions, although some manual adjustments may be necessary.

O'Donnell, Greg; Nijssen, Bart; Lettenmaier, Dennis P.



Site-specific mutants of carbonic anhydrase for fluorescence energy-transfer-based metal-ion biosensing  

NASA Astrophysics Data System (ADS)

In order to gain wavelength and analyte flexibility, we have recently altered the transduction approach of our fluorescence-based biosensor. Briefly, binding of metal ions such as zinc to the active site of carbonic anhydrase is transduced by metal-dependent binding of a colored inhibitor to a fluorescent derivative of the enzyme; in the absense of metal the inhibitor does not bind and the label fluorescence is unquenched, but at higher metal concentrations the inhibitor binds, energy transfer occurs with moderate efficiency and the fluorescent label exhibits reduced intensity and lifetime. Inasmush as Forster energy transfer is distance dependent the position of the fluorescent label on the surface of the enzyme has some impact on the performance of the sensor. We designed, produced, and expressed site-selective mutants of carbonic anhydrase which could be unambiguously derivatized with suitable fluorescent labels, and which gave much improved responses to zinc ion compared with randomly derivatized wild type enzyme.

Thompson, Richard B.; Ge, Zhengfang; Patchan, Marcia W.; Kiefer, Laura L.; Fierke, Carol A.



A fluorescent nanoprobe based on graphene oxide fluorescence resonance energy transfer for the rapid determination of oncoprotein vascular endothelial growth factor (VEGF).  


Oncoprotein vascular endothelial growth factor (VEGF) is one of the most critical growth factors that regulates tumor growth and division. The vascular endothelial growth factor (VEGF) is also an important biomarker for different diseases and clinical disorders. Herein, we propose a graphene oxide (GO) fluorescence resonance energy transfer (FRET)-based aptasensor for rapid, sensitive, and selective detection of VEGF in homogeneous solution. The fluorescent dye-labeled anti-VEGF aptamer is adsorbed on the surface of GO via ?-? interaction between the flat planar GO sheets and the ring structures in the nucleobases, which results in the fluorescence quenching of the dye due to the highly effective FRET from the dye to GO. Upon recognition and binding with the target VEGF, it specifically forms a VEGF/aptamer complex and then release from the GO surface, leading to the restoration of fluorescence signal of the dye. This GO-based sensing platform exhibits high sensitivity and specificity toward VEGF versus other proteins, with the detection limits corresponding to 2.5×10(-10) M. The sensitivity of this new type of aptamer-based assay is at least one order of magnitude higher than that of conventional homogeneous optical assays. Moreover, the application of this nanosensor for human serum sample analysis is also demonstrated. The GO/aptamer-based assay approach holds great promise as a general platform for detection of a variety of target molecules. PMID:24160878

Wang, Sheng-E; Si, Shihui



Nanosized pi-conjugated molecules based on truxene and porphyrin: synthesis and high fluorescence quantum yields.  


[reaction: see text] A series of novel nanosized pi-conjugated molecules based on both truxene and porphyrin moieties with high fluorescence quantum yields have been prepared via the Suzuki cross-coupling and the Lindsey reactions. The investigation of optical properties demonstrates that various aryl groups as the antenna efficiently enhance the intramolecular and intermolecular energy transfer. These nanosized macromolecules emitting intense red light might be good candidates for photonic and electronic devices. PMID:16146354

Duan, Xiao-Fei; Wang, Jin-Liang; Pei, Jian



X-ray absorption spectroscopy: A fluorescence detection system based on a plastic scintillator  

Microsoft Academic Search

A fluorescence detection system based on a plastic scintillator is presented that can be used for both X-ray absorption near-edge structure (XANES) and extended X-ray absorption fine-structure (EXAFS) spectroscopy. Its counting rate is limited by the pulsation frequency of the synchrotron radiation (3.16×106 counts s-1), and can be theoretically extended to roughly 5×107 counts s-1 if used on a multibunch

G. Tourillon; D. Guay; M. Lemonnier; F. Bartol; M. Badeyan



Indole-based fluorescent sensors for selective detection of Hg2+.  


A series of indole-based fluorescent chemosensors 1-4 were prepared and investigated characteristick features with transition metal ions. Sensors 1 and 2 were selective for Hg(2+) ion among a series of metal ions in H2O-DMSO with association constants of 4.60×10(4) and 5.90×10(4)?M(-1) and detection limits of 140 and 101.6 ?M, respectively. PMID:23479206

Sun, Yao-Lin; Wu, An-Tai



Iterative Behavioral Modeling of Charge-Pump-Based Electronic Ballast–Fluorescent Lamp System  

Microsoft Academic Search

A new behavioral model for analyzing the current-source charge-pump-based ballast with a self-oscillatory gate drive circuit is proposed in this paper. A detailed model derivation is presented and the model is applied to analyze a ballast system of a 16-W compact fluorescent lamp (CFL) as an illustration. The ballast of the CFL consists of a self-oscillatory driven inverter stage and

Pok Wai Tam; Henry Shu-Hung Chung



Mesh-based Monte Carlo code for fluorescence modeling in complex tissues with irregular boundaries  

Microsoft Academic Search

There is a growing need for the development of computational models that can account for complex tissue morphology in simulations of photon propagation. We describe the development and validation of a user-friendly, MATLAB-based Monte Carlo code that uses analytically-defined surface meshes to model heterogeneous tissue geometry. The code can use information from non-linear optical microscopy images to discriminate the fluorescence

Robert H. Wilson; Leng-Chun Chen; William Lloyd; Shiuhyang Kuo; Cynthia Marcelo; Stephen E. Feinberg; Mary-Ann Mycek



Fluorescence detection of single-nucleotide polymorphisms using a thymidine-based molecular beacon  

Microsoft Academic Search

We have developed a universal molecular beacon (T7-MB-T7) for the detection of single-nucleotide polymorphisms (SNPs). The beacon, which contains a 19-mer loop and a stem comprising a pair of seven thymidine (T) bases, forms double-stranded structures with target DNA molecules, leading to increases in the fluorescence of ethidium bromide (EthBr) as a result of intercalation. The interactions of the beacon

Chi-Wei Liu; Yang-Wei Lin; Chih-Ching Huang; Huan-Tsung Chang



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

Microsoft Academic Search

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

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



Stilbene-Based Fluorescent Sensor for Detection of Organophosphorus Warfare Nerve Agents  

Microsoft Academic Search

In this paper, we report the synthesis of stilbene-based fluorophore, 3,4-dihydroxy-4?-aminostilbene (DHAS) for the detection of chemical warfare agents such as organophosphorus nerve gases. DHAS was characterized by various spectroscopic methods and grafted on to electrospun nanofibers. The interaction of DHAS with nerve agents simulant, diethyl chlorophosphate (DCP) was investigated in solution and vapor phase by fluorescence spectroscopy.

Robinson Anandakathir; Umaprasana Ojha; Earl T. Ada; Rudolf Faust; Jayant Kumar



Colorimetric and fluorescent chemosensor for citrate based on a rhodamine and Pb2+ complex in aqueous solution.  


In this paper we unveil a novel rhodamine compound based fluorescent chemosensor (1-Pb(2+)) for colormetric and fluorescent detection of citrate in aqueous solution. This is the first fluorescent chemosensor for citrate based on rhodamine compound. The comparison of this method with some other fluorescence methods for citrate indicates that the method can detect citrate in aqueous solution by both color changes and fluorescent changes with long emission wavelength. In the new developed sensing system, 1-Pb(2+) is fluorescent due to Pb(2+)-induced fluorescence enhancement of 1. However, the addition of citrate may release 1 into the solution with quenching of fluorescence. The chemosensor can be applied to the quantification of citrate with a linear range covering from 1.0×10(-7) to 5.0×10(-5) M and a detection limit of 2.5×10(-8) M. The experiment results show that the response behavior of 1-Pb(2+) towards citrate is pH independent in medium condition (pH 6.0-8.0). Most importantly, the fluorescence changes of the chemosensor are remarkably specific for citrate in the presence of other anions (even those that exist in high concentration), which meet the selective requirements for practical application. Moreover, the response of the chemosensor toward citrate is fast (response time less than 1 min). In addition, the chemosensor has been used for determination of citrate in urine samples with satisfactory results. PMID:23567120

Li, Chun-Yan; Zhou, Yu; Li, Yong-Fei; Kong, Xue-Fei; Zou, Chun-Xiang; Weng, Chao



Detection of HIV1 RNA by Nucleic Acid Sequence-Based Amplification Combined with Fluorescence Correlation Spectroscopy  

Microsoft Academic Search

Nucleic acid sequence-based amplification (NASBA) has proved to be an ultrasensitive method for HIV-1 diagnosis in plasma even in the primary HIV infection stage. This technique was combined with fluorescence correlation spectroscopy (FCS) which enables online detection of the HIV-1 RNA molecules amplified by NASBA. A fluorescently labeled DNA probe at nanomolar concentration was introduced into the NASBA reaction mixture

Frank Oehlenschlager; Petra Schwille; Manfred Eigen



Graphene oxide-based fluorescent biosensor for protein detection via terminal protection of small-molecule-linked DNA.  


A fluorescence method for protein detection is developed based on terminal protection of small-molecule-linked DNA by target protein and a graphene oxide-assisted DNA assay strategy. This design results in fluorescence-enhanced detection that is sensitive and selective for the target protein. PMID:23362224

He, Yue; Xing, Xiaojing; Tang, Hongwu; Pang, Daiwen



Novel ultra-fast deconvolution method for fluorescence lifetime imaging microscopy based on the Laguerre expansion technique  

Microsoft Academic Search

A new deconvolution method for fluorescence lifetime imaging microscopy (FLIM) based on the Laguerre expansion technique is presented. The performance of this method was tested on synthetic FLIM images derived from a multiexponential model and from fluorescence lifetime standards, and then compared to standard algorithms of FLIM analysis. Our results demonstrated significant advantages of the Laguerre method over standard algorithms.

J. A. Jo; Q. Fang; T. Papaioannou; L. Marcu



Assessment and Continued Validation of the Malaria SYBR Green I-Based Fluorescence Assay for Use in Malaria Drug Screening  

Microsoft Academic Search

Several new fluorescence malaria in vitro drug susceptibility microtiter plate assays that detect the presence of malarial DNA in infected erythrocytes have recently been reported, in contrast to traditional isotopic screens that involve radioactive substrate incorporation to measure in vitro malaria growth inhibition. We have assessed and further characterized the malaria SYBR Green I-based fluorescence (MSF) assay for its ability

Jacob D. Johnson; Richard A. Dennull; Lucia Gerena; Miriam Lopez-Sanchez; Norma E. Roncal; Norman C. Waters



Intrinsic Fluorescence-Based Optical Fiber Sensor for Cocaine Using a Molecularly Imprinted Polymer as the Recognition Element  

Microsoft Academic Search

A fiber-optic chemical sensor for the detection of cocaine has been developed, based on a molecularly imprinted polymer (MIP) containing a fluorescein moiety as the signalling group. The fluorescent MIP was formed and covalently attached to the distal end of an optical fiber. The sensor exhibited an increase in fluorescence intensity in response to cocaine in the concentra- tion range

T. Hien Nguyen; Sheila A. Hardwick; Tong Sun; Kenneth T. V. Grattan



High-speed DNA sequencing: An approach based upon fluorescence detection of single molecules  

SciTech Connect

This document discusses the development of a laser based technique for the rapid sequencing of large fragments of DNA based upon the detection of single, fluorescently tagged nucleotides cleaved from a single DNA fragments. Demonstrated is significant progress on several of the important steps of this technique. The projected rate of sequencing is several hundred bases per second which is orders of magnitude faster than existing methods. Once developed, this technology could be utilized by investigators for rapid sequencing of genetic material from virtually any source. 24 refs., 4 figs.

Jett, J.H.; Keller, R.A.; Martin, J.C.; Marrone, B.L.; Moyzis, R.K.; Ratliff, R.L.; Seitzinger, N.K.; Shera, E.B.; Stewart, C.C.



Study on the Fluorescence Characteristics of Alkaline Degradation of Cefadroxil, Cephradine, Cefotaximum Sodium and Amoxicillini  

Microsoft Academic Search

A simple and sensitive fluorescence method for cefadroxil, cephradine, cefotaximum and amoxicillini is described. The method is based on their degradation under alkaline condition producing fluorescence products. The factors affecting the degradation and the determination were studied in detail. Under optimum conditions a linear relationship was obtained between the concentration of the antibiotics and the fluorescence intensity. The linear ranges

Jinghe Yang; Guangjun Zhou; Xihui Cao; Quanli Ma; Jie Dong



Pancreatic tumor detection using hypericin-based fluorescence spectroscopy and cytology  

NASA Astrophysics Data System (ADS)

Hypericin is a novel, highly fluorescent photosensitizer that exhibits selective tumor cell uptake properties and is particularly resistant to photobleaching. In this study, we have characterized hypericin uptake in human pancreatic tumor cells with relation to incubation time, cell number, and drug concentration. Ex vivo hypericin based fluorescence spectroscopy was performed to detect the presence of MIA PaCa-2 pancreatic tumor cells in the peritoneal cavity of BALB/c nude mice, as well as to quantify gross tumor burden. Hypericin based cytology of peritoneal lavage samples, using both one and two photon laser confocal microscopy, demonstrated more than a two-fold increase in fluorescence emission of pancreatic tumor cells as compared to control samples. In vitro treatment of pancreatic cancer cells with hypericin based photodynamic therapy showed tumor cell cytotoxicity in a drug dose, incident laser power, and time dependent manner. For these experiments, a continuous wavelength solid-state laser source (532 nm) was operated at power levels in the range of 100-400 mW. Potential applications of hypericin in tumor diagnosis, staging, and therapy will be presented.

Lavu, Harish; Geary, Kevin; Fetterman, Harold R.; Saxton, Romaine E.



10 CFR Appendix W to Subpart B of... - Uniform Test Method for Measuring the Energy Consumption of Medium Base Compact Fluorescent Lamps  

Code of Federal Regulations, 2010 CFR

... Uniform Test Method for Measuring the Energy Consumption of Medium Base Compact Fluorescent Lamps W Appendix...Uniform Test Method for Measuring the Energy Consumption of Medium Base Compact Fluorescent Lamps 1....



10 CFR Appendix W to Subpart B of... - Uniform Test Method for Measuring the Energy Consumption of Medium Base Compact Fluorescent Lamps  

Code of Federal Regulations, 2010 CFR

... Uniform Test Method for Measuring the Energy Consumption of Medium Base Compact Fluorescent Lamps W Appendix...Uniform Test Method for Measuring the Energy Consumption of Medium Base Compact Fluorescent Lamps 1....



Solution and Fluorescence Properties of Symmetric Dipicolylamine-Containing Dichlorofluorescein-Based Zn2+ Sensors  

PubMed Central

The mechanism by which dipicolylamine (DPA) chelate-appended fluorophores respond to zinc was investigated by the synthesis and study of five new analogs of the 2?,7?-dichlorofluorescein-based Zn2+ sensor Zinpyr-1 (ZP1). With the use of absorption and emission spectroscopy in combination with potentiometric titrations, a detailed molecular picture has emerged of the Zn2+ and H+ binding properties of the ZP1 family of sensors. The two separate N3O donor atom sets on ZP1 each converge to form binding pockets in which all four heteroatoms participate in coordination to either Zn2+ or protons. The position of the pyridyl group nitrogen atom, 2-pyridyl or 4-pyridyl, has a large impact on the fluorescence response of the dyes to protons despite relatively small changes in pKa values. The fluorescence quenching effects of such multifunctional electron-donating units are often taken as a whole. Despite the structural complexity of ZP1, however, we provide evidence that the pyridyl arms of the DPA appendages participate in the quenching process, in addition to the contribution from the tertiary nitrogen amine atom. Potentiometric titrations reveal ZP1 dissociation constants (Kd) for Zn2+ of 0.04 pM and 1.2 nM for binding to the first and second binding pockets of the ligand, respectively, the second of which correlates with the value observed by fluorescence titration. This result demonstrates that both binding pockets of this symmetric, ditopic sensor need to be occupied in order for full fluorescence turn-on to be achieved. These results have significant implications for the design and implementation of fluorescent sensors for studies of mobile zinc ions in biology.

Wong, Brian A.; Friedle, Simone; Lippard, Stephen J.



Community-Based Prevention Using Simple, Low-Cost, Evidence-Based Kernels and Behavior Vaccines  

ERIC Educational Resources Information Center

A paradox exists in community prevention of violence and drugs. Good research now exists on evidence-based programs, yet extensive expenditures on prevention have not produced community-level results. Various multiproblems are quite prevalent in the United States, such as violence, Attention Deficit Hyperactivity Disorder (ADHD), conduct problems,…

Embry, Dennis D.



A new fluorescent quantitative PCR-based in vitro neutralization assay for white spot syndrome virus.  


A fluorescent quantitative PCR (FQ-PCR) assay utilizing SYBR green I dye is described for quantitation of white spot syndrome virus (WSSV) particles isolated from infected crayfish, Cambarus clarkii. For this assay, a primer set was designed which amplifies, with high efficiency and specificity, a 129bp target sequence within ORF167 of the WSSV genome. Conveniently, WSSV particles can be added into the FQ-PCR assay with a simple and convenient method to release its DNA. To establish the basis for an in vitro neutralization test, primary cultures of shrimp cells were challenged with WSSV that had been incubated with a polyclonal anti-WSSV serum or with control proteins. The number of WSSV particles released from the cells after these treatments were assayed by FQ-PCR. This test may serve as a method to screen monoclonal antibody pools or recombinant antibody pools for neutralizing activity prior to in vivo animal experiments. PMID:17645951

Yuan, Li; Zhang, Xiaohua; Chang, Mingxian; Jia, Chensong; Hemmingsen, Sean M; Dai, Heping



Chemosensors and biosensors based on polyelectrolyte microcapsules containing fluorescent dyes and enzymes.  


The concept of enzyme-assisted substrate sensing based on use of fluorescent markers to detect the products of enzymatic reaction has been investigated by fabrication of micron-scale polyelectrolyte capsules containing enzymes and dyes in one entity. Microcapsules approximately 5 ?m in size entrap glucose oxidase or lactate oxidase, with peroxidase, together with the corresponding markers Tris(4,7-diphenyl-1,10-phenanthroline)ruthenium(II) dichloride (Ru(dpp)) complex and dihydrorhodamine 123 (DHR123), which are sensitive to oxygen and hydrogen peroxide, respectively. These capsules are produced by co-precipitation of calcium carbonate particles with the enzyme followed by layer-by-layer assembly of polyelectrolytes over the surface of the particles and incorporation of the dye in the capsule interior or in the multilayer shell. After dissolution of the calcium carbonate the enzymes and dyes remain in the multilayer capsules. In this study we produced enzyme-containing microcapsules sensitive to glucose and lactate. Calibration curves based on fluorescence intensity of Ru(dpp) and DHR123 were linearly dependent on substrate concentration, enabling reliable sensing in the millimolar range. The main advantages of using these capsules with optical recording is the possibility of building single capsule-based sensors. The response from individual capsules was observed by confocal microscopy as increasing fluorescence intensity of the capsule on addition of lactate at millimolar concentrations. Because internalization of the micron-sized multi-component capsules was feasible, they could be further optimized for in-situ intracellular sensing and metabolite monitoring on the basis of fluorescence reporting. PMID:22968684

Kazakova, Lyubov I; Shabarchina, Lyudmila I; Anastasova, Salzitsa; Pavlov, Anton M; Vadgama, Pankaj; Skirtach, Andre G; Sukhorukov, Gleb B



Fluorescence detection of single-nucleotide polymorphisms with two simple and low cost methods: a double-DNA-probe method and a bulge form method.  


Two 10-mer DNA probes, or one 20-mer DNA probe, respectively, hybridize with a 21-mer target DNA to form a vacancy or bulge opposite the target nucleotide. The former double-DNA-probe method and the latter bulge form method are applicable to the detection of single-nucleotide polymorphisms (SNPs). A small fluorescent dye enters into the vacancy or bulge and binds with a target nucleotide via a hydrogen bonding interaction, which causes fluorescence quenching. The interaction between fluorescent dye and the target nucleotide is confirmed by measuring the melting temperature and fluorescence spectra. The fluorescent dye, ADMND (2-amino-5,7-dimethyl-1,8-naphthyridine), is found to selectively bind with C over A or G. The methods proposed here are economic, convenient, and effective for the fluorescence detection of SNPs. Finally, the double-DNA-probe method and bulge form method are successfully applied to the detection of C/G and C/A mutations in the estrogen receptor 2 gene and progesterone receptor gene using ADMND. PMID:17658318

Li, Na; Mei, Ling; Xiang, Yu; Tong, Aijun; Nishizawa, Seiichi; Teramae, Norio



Fluorescence immunosensor based on p-acid-encapsulated silica nanoparticles for tumor marker detection.  


A novel, simple and highly sensitive amplified fluorescence label-free immunosensor by using p-acid-encapsulated silica nanomaterials has been developed for the first time. 4,4'-(2,5-Dimethoxy-1,4-phenylene)bis(ethyne-2,1-diyl)dibenzoic acid (p-acid) and p-acid-encapsulated silica were prepared, and characterized by Fourier transform infrared spectroscopy, nuclear magnetic resonance, ultraviolet visible spectroscopy (UV-vis) and fluorescence spectroscopy. In layer-by-layer self-assembling processes using (3-aminopropyl)triethoxysilane, p-acid@SiO(2) was assembled on the glass substrate. Antibody was immobilized on the surface of p-acid@SiO(2) with N,N'-carbonyldiimidazole. The functional nanomaterials present good analytical properties with a calibration range of 0.1-100 ng mL(-1), and allow the detection of carcinoembryonic antigen (CEA) at a concentration as low as 0.04 ng mL(-1). What is important is that the as-synthesized p-acid@SiO(2) nanomaterials could be further extended for the detection of other biomarkers or biocompounds. PMID:22558632

Yan, Mei; Ge, Shenguang; Gao, Weiqiang; Chu, Chengchao; Yu, Jinghua; Song, Xianrang



Simple thermal imaging system based on hollow glass waveguides or silver halide fibers as scanning elements for medical applications  

Microsoft Academic Search

A simple thermal imaging system based on silver halide fibers or glass hollow waveguides was constructed. The proximal end of each type of waveguide was fixed and attached to an infrared detector. The distal end of each waveguide was scanning in two directions. Such a device was used to construct a simple imaging system. The thermal image of a warm

B. Dekel; Abraham Katzir; Alexandra Inberg; Nathan I. Croitoru



Pencil-drawn paper supported electrodes as simple electrochemical detectors for paper-based fluidic devices.  


A simple procedure for preparing inexpensive paper-based three-electrode electrochemical cells is described here. They consist of small circular pads of hydrophilic paper defined by hydrophobic barriers printed on paper with wax-based ink. The back face of these pads is insulated by thermally laminating a polyethylene layer and working, reference and counter electrodes are drawn on paper by using commercial pencil leads. At last, a controlled volume of sample containing a supporting electrolyte was laid to soak in paper channels. Their performance was evaluated by assaying these devices as both simple cells suitable for recording voltammograms on static samples and low-cost detectors for flowing systems. Voltammetric tests, conducted by using potassium hexacyanoferrate(II) as model prototype, were also exploited for identifying the brand and softness of graphite sticks enabling paper to be marked with lines displaying the best conductivity. By taking advantage of the satisfactory information thus gained, pencil drawn electrodes were tested as amperometric detectors for the separation of ascorbic acid and sunset yellow, which were chosen as prototype electroactive analytes because they are frequently present concomitantly in several food matrices, such as soft drinks and fruit juices. This separation was performed by planar thin layer chromatography conducted on microfluidic paper-based devices prepared by patterning on filter paper two longitudinal hydrophobic barriers, once again printed with wax-based ink. Factors affecting both separation and electrochemical detection were examined and optimised, with best performance achieved by using a 20 mM acetate running buffer (pH 4.5) and by applying a detection potential of 0.9 V. Under these optimum conditions, the target analytes could be separated and detected within 6 min. The recorded peaks were well separated and characterized by good repeatability and fairly good sensitivity, thus proving that this approach is indeed suitable for rapidly assembling inexpensive and reliable electrochemical detectors for flow analysis systems. PMID:23161669

Dossi, Nicolò; Toniolo, Rosanna; Pizzariello, Andrea; Impellizzieri, Flavia; Piccin, Evandro; Bontempelli, Gino



High resolution single-mode-fiber-based sensor for intravascular detection of fluorescent molecular probes  

NASA Astrophysics Data System (ADS)

Early detection of coronary atherosclerosis is an unmet clinical challenge. The detection system has to be highly sensitive and possess high spacial resolution, in order to provide precise information of the vulnerable plaque location and size. Recently molecular fluorescence probes have been identified as efficient inflammation biomarkers for the inflammation process within vulnerable plaques1 and being used in the proposed application to detect inflamed lesions in the blood vessel wall. The general principle of the proposed solution is based on a sensor whose head is guided by an intravascular catheter to the region of interest (coronary artery). When the sensor illuminates an activated fluorescent probe, located in inflamed areas of vulnerable plaques, the fluorescence is excited and light is emitted with a slightly shifted spectrum. The emitted light is being collected by the same sensor head, guided through the optical fiber and finally detected by photo-detectors. In this way, by detecting emitted fluorescence one can obtain information about the location of vulnerable plaques. The localization resolution is critically depending on the spot size of the illuminating light beam. Moreover, for a high signal to noise ratio in the detection electronics, as much fluorescent light as possible has to be collected from the plaque location. It has been already demonstrated that using single-mode fibers in combination with graded index fibers, a Gaussian beam, with adjustable waist position and diameter can be formed, representing the fundamental limit of achievable spot size2. However, when using single mode fibers in this application, the collection efficiency would be very low due to the small core diameter of this fiber and thus signal to noise ratio would be strongly reduced. In this work, we present a solution to this challenge, combining both principles. A single mode fiber in combination with a graded index fiber is used for illumination purposes, while the fluorescence light is collected by the same fiber, but employing the cladding/coating total reflection to form a multimode fiber for the backwards propagating light. Thus, a narrow spot size can be obtained allowing high resolution images, with high signal to noise ratio due to the multimodal collection scheme. We show preliminary results of spot size and beam diameter measurements from the sensor head and discuss the implication for the improvement of the current catheter-based detection systems.

Razansky, R. Nika; Mueller, Mathias S.; Borisov, Alexander; Koch, Alexander W.; Jaffer, Farouc A.; Ntziachristos, Vasilis



Multiplex immunoassays of equine virus based on fluorescent encoded magnetic composite nanoparticles.  


A new detection format for multiplexed analysis based on fluorescent encoded magnetic composite nanoparticles is presented. Two kinds of virus were analyzed by this new method: equine influenza virus (EIV) and equine infectious anemia virus (EIAV). Firstly, EIV antigen and EIAV antigen were conjugated to two kinds of fluorescent encoded magnetic composite nanoparticles, while the green-emitting CdTe quantum dots (QDs) were attached to the antibody of EIV and EIAV. Then both green-emitting CdTe QD-labeled antibodies and antigens labeled with fluorescent encoded magnetic composite nanoparticles were used to form an immunoassay system for the detection of EIV and EIAV antigens. The method is time-saving and has higher sensitivity (1.3 ng mL(-1) for EIV antigens and 1.2 ng mL(-1) for EIAV antigens) than the conventional methods. A competitive immunoassay method based on this analysis system was used to detect EIV and EIAV antigens in spiked serum samples with satisfactory results. PMID:20652548

Wang, Guannan; Gao, Yuan; Huang, Hui; Su, Xingguang



Prism-based spectral imaging of single-molecule fluorescence from gold-nanoparticle/fluorophore complex.  


A wavelength-calibration method for prism-based spectral imaging of single-molecule (SM) fluorescence was developed. With this method, a wavelength reference is provided by photoluminescence from 50-nm-diameter gold nanoparticles (AuNPs) binding with fluorophores. The AuNPs each bound with a SM fluorophore, either Alexa488 or Cy3, to form AuNP/fluorophore complexes in tris-HCl buffer. Each complex was immobilized on a silica slide and then excited by total-internal-reflection illumination to make it emit SM fluorescence and AuNP photoluminescence. The portion of the AuNP photoluminescence transmitted by a band-pass filter gives the wavelength reference. A spectral-imaging system composed of a prism-based spectroscope (with a reciprocal dispersion of about 4 nm/?m) and a charge-coupled device with 6.45-?m-square pixels was used to obtain an SM-fluorescence spectrum and a wavelength-reference spectrum. Through smoothed differentiation of these two spectra, the peak location of the former in relation to the latter was determined with subpixel precision. After that, the SM fluorophore was classified as either Alexa488 or Cy3 according to the peak location. The error rate of the classification was estimated to be only 0.3%. PMID:21384172

Sonehara, Tsuyoshi; Sakai, Tomoyuki; Haga, Takanobu; Fujita, Takeshi; Takahashi, Satoshi



Fabrication of holographic diffractive optical elements for enhancing light collection from fluorescence-based biochips.  


An approach to enhancing the light collection from fluorescence emitters bound on the surface of a biochip is presented. It is based on the integration of diffractive optical elements on the underside of the chip that are essentially thick volume holograms written into a layer of photopolymer recording material. The high diffractive efficiency and angular selectivity of these types of diffractive elements make them very effective collectors of the spatially anisotropic light emitted by surface-bound fluorophores. The holographic lithography setup used to fabricate the diffractive elements is described. Results obtained using both a focused laser and light from a fluorescence spot to characterize the performance of the diffractive optical elements are presented. PMID:19015685

Macko, Peter; Whelan, Maurice P



Quantitative analysis results of CE-1 X-ray fluorescence spectrometer ground base experiment  

NASA Astrophysics Data System (ADS)

As the nearest celestial body to the earth, the moon has become a hot spot again in astronomy field recently. The element analysis is a much important subject in many lunar projects. Remote X-ray spectrometry plays an important role in the geochemical exploration of the solar bodies. Because of the quasi-vacuum atmosphere on the moon, which has no absorption of X-ray, the X-ray fluorescence analysis is an effective way to determine the elemental abundance of lunar surface. The CE-1 X-ray fluorescence spectrometer (CE-1/XFS) aims to map the major elemental compositions on the lunar surface. This paper describes a method for quantitative analysis of elemental compositions. A series of ground base experiments are done to examine the capability of XFS. The obtained results, which show a reasonable agreement with the certified values at a 30% uncertainty level for major elements, are presented.

Cui, Xing-Zhu; Wang, Huan-Yu; Zhang, Cheng-Mo; Chen, Yong; Zhang, Jia-Yu; Peng, Wen-Xi; Cao, Xue-Lei; Liang, Xiao-Hua; Wang, Jin-Zhou; Gao, Min; Yang, Jia-Wei



One new dicoumarol-based fluorescent compound: Synthesis, crystal structure and metal ions recognition  

NASA Astrophysics Data System (ADS)

A new dicoumarol-based compound (C6H4)[CH2NHCO(C9O2H4)N(C2H5)2]21 was synthesized and characterized by IR, UV spectroscopy and single-crystal X-ray diffraction analysis. The structure of 1 exhibits a transoid formation with the two coumarin-containing arms sited on the two sides of the center benzene ring. In the crystal packing the molecule further interact with each other and form a three-dimensional framework through ?-? stacking interactions and multiform hydrogen bonds. The compound 1 shows the main emission peak at 540 nm corresponding to the green hue in the solid state. The fluorescence recognition behaviors for various metal ions were investigated and 1 exhibits a highly fluorescence-quenching selectivity for FeIII ion in the mixed CH3CNsbnd H2O solvent.

Bai, Yan; Yu, Ke; Pan, Hui; Dang, Dong-Bin



Rapid screening of DOM's metal-binding ability using a fluorescence-based microplate assay.  


This note describes the development of a method to rapidly quantify the metal-binding ability of low-volume DOM samples based on their fluorescence properties. The method uses 96-well microplates to screen the fluorescence quenching observed when increasing concentrations of various metals are added to the pH-buffered DOM sample. Only 1.6 mL of DOM sample is required to quantify the binding affinity for each metal and the result is obtained in a few minutes. This study presents results for a surface soil DOM sample for which binding was assessed for Cu2+, Fe2+, Ni2+ at pH 5 and 9. PMID:16081142

Dudal, Y; Holgado, R; Maestri, G; Guillon, E; Dupont, L



Structure-guided engineering enhances a phytochrome-based infrared fluorescent protein.  


Phytochrome is a multidomain dimeric red light photoreceptor that utilizes a chromophore-binding domain (CBD), a PHY domain, and an output module to induce cellular changes in response to light. A promising biotechnology tool emerged when a structure-based substitution at Asp-207 was shown to be an infrared fluorophore that uses a biologically available tetrapyrrole chromophore. We report multiple crystal structures of this D207H variant of the Deinococcus radiodurans CBD, in which His-207 is observed to form a hydrogen bond with either the tetrapyrrole A-ring oxygen or the Tyr-263 hydroxyl. Based on the implications of this duality for fluorescence properties, Y263F was introduced and shown to have stronger fluorescence than the original D207H template. Our structures are consistent with the model that the Y263F change prevents a red light-induced far-red light absorbing phytochrome chromophore configuration. With the goal of decreasing size and thereby facilitating use as a fluorescent tag in vivo, we also engineered a monomeric form of the CBD. Unexpectedly, photoconversion was observed in the monomer despite the lack of a PHY domain. This observation underscores an interplay between dimerization and the photochemical properties of phytochrome and suggests that the monomeric CBD could be used for further studies of the photocycle. The D207H substitution on its own in the monomer did not result in fluorescence, whereas Y263F did. Combined, the D207H and Y263F substitutions in the monomeric CBD lead to the brightest of our variants, designated Wisconsin infrared phytofluor (Wi-Phy). PMID:22210774

Auldridge, Michele E; Satyshur, Kenneth A; Anstrom, David M; Forest, Katrina T



Optimization of a Concanavalin A-based glucose sensor using fluorescence anisotropy.  


To date, the dependent nature of the recognition and transduction mechanisms in optical glucose sensors based upon Concanavalin A (ConA) has tended to prevent the sensors' full potential from being realized. In this paper, these mechanisms are independently optimized for a given assay configuration in order to decrease the predictive error of a ConA-based glucose sensor and to give a more accurate demonstration of its potential. To this end, we used fluorescence anisotropy as the transduction mechanism to determine the binding of ConA to 4 kDa FITC-dextran by measuring the change in the rotational correlation lifetime between the bound and unbound populations. By tracking the fluorescence anisotropy of this ligand, the ranges of ConA and 4 kDa FITC-dextran concentrations capable of being explored were not limited by the transduction mechanism. Using predetermined association constants, the binding responses to physiological glucose concentrations were predicted for different assay configurations, and experimentally collected fluorescence anisotropy data displayed the predicted trends for these assay configurations. From the experimental results, a calibration fit was generated for the optimized assay configuration to predict the glucose concentrations using the fluorescence anisotropy. This optimized assay displayed a mean standard error of prediction of 7.5 mg/dL (0-300 mg/dL), and 100% of the data points fell within clinically acceptable zones (A and B) upon the Clarke Error Grid Analysis. This indicates that, by independently optimizing the recognition and transduction mechanisms for the final assay configuration, the sensitivity of a competitive binding chemistry using ConA can be appropriately configured for continuous glucose monitoring applications. PMID:23627407

Cummins, Brian M; Garza, Javier T; Coté, Gerard L



Piezoresistor-equipped fluorescence-based cantilever probe for near-field scanning.  


Scanning near-field optical microscopes (SNOMs) with fluorescence-based probes are promising tools for evaluating the optical characteristics of nanoaperture devices used for biological investigations, and this article reports on the development of a microfabricated fluorescence-based SNOM probe with a piezoresistor. The piezoresistor was built into a two-legged root of a 160-microm-long cantilever. To improve the displacement sensitivity of the cantilever, the piezoresistor's doped area was shallowly formed on the cantilever surface. A fluorescent bead, 500 nm in diameter, was attached to the bottom of the cantilever end as a light-intensity-sensitive material in the visible-light range. The surface of the scanned sample was simply detected by the probe's end being displaced by contact with the sample. Measuring displacements piezoresistively is advantageous because it eliminates the noise arising from the use of the optical-lever method and is free of any disturbance in the absorption or the emission spectrum of the fluorescent material at the probe tip. The displacement sensitivity was estimated to be 6.1 x 10(-6) nm(-1), and the minimum measurable displacement was small enough for near-field measurement. This probe enabled clear scanning images of the light field near a 300 x 300 nm(2) aperture to be obtained in the near-field region where the tip-sample distance is much shorter than the light wavelength. This scanning result indicates that the piezoresistive way of tip-sample distance regulation is effective for characterizing nanoaperture optical devices. PMID:17764312

Kan, Tetsuo; Matsumoto, Kiyoshi; Shimoyama, Isao



Fluorescence-based high-throughput screening assay for drug interactions with UGT1A6.  


The increasing awareness and the rising importance of UDP-glucuronosyltransferases (UGTs) in the pharmacokinetics of drugs have evoked a need to develop more powerful tools for studying the role of UGTs in the metabolism of drug candidates. To this end, we have developed a fluorescent high-throughput screening assay for screening potential inhibitors and/or substrates for recombinant human UGTs-here, for the UGT1A6. The assay is based on the increase in fluorescence intensity when 1-naphthol is glucuronidated. The formation of the highly fluorescent product, 1-naphthylglucuronide, is followed at excitation wavelengths of 295 and 300 nm with fixed emission (335 nm) in real time directly from the reaction mixture. A probe concentration of 5 ?M with 2.5 ?g of total protein in phosphate buffer at pH 7.4 with 5% dimethyl sulfoxide resulted in optimal linearity and acceptable signal separation (signal-to-base, 3.0) for the probe reaction. The interactions of test compounds with the enzyme are detected as lower rate of 1-naphthylglucuronide formation and thus lower rate of fluorescence increase. The success of the assay was first demonstrated with the known UGT1A6 substrates 4-hydroxyindole and scopoletin (Z' factor ?0.5) and later with nonsteroidal anti-inflammatory drugs and salicylate derivatives. Diclofenac, 5-methylsalicylic acid, 5-bromosalicylic acid, 5-chlorosalicylic acid, and 5-fluorosalicylic acid decreased the probe glucuronidation rate at 500??M by >50%. Further, the results gained with the high-throughput screening assay correlated well with the results obtained, in parallel, with the reference high-performance liquid chromatography method. PMID:21438674

Soikkeli, Anne; Kurkela, Mika; Hirvonen, Jouni; Yliperttula, Marjo; Finel, Moshe



Promising fluorescent dye for solar energy conversion based on a perylene perinone.  


We describe the synthesis of a dye based on a perylene perinone and evaluate its potential as the functional material for use in the luminescent solar concentrator (LSC). The dye extends the absorption wavelength of LSCs using the perylene-based dye Lumogen Red 305 by more than ~50 nm, translating into the collection of potentially 25% more photons at a reasonable fluorescent quantum yield and photostability. When the new perinone is used in a two-waveguide LSC in conjunction with Red 305, the integrated edge emission of the total LSC system may be increased more than 24% when compared to the Red 305 dye alone. PMID:21221140

Debije, Michael G; Verbunt, Paul P C; Nadkarni, Pradeep J; Velate, Suresh; Bhaumik, Kankan; Nedumbamana, Sankaran; Rowan, Brenda C; Richards, Bryce S; Hoeks, Theo L



Simple generation of cationic aluminum alkyls and alkoxides based on the pendant arm tridentate schiff base.  


The prepared in situ methyl(chloro)aluminum complex (2) from Me2AlCl and the pendant arm tridentate Schiff base (H-SchNMe2) was used to generate the methylaluminum cationic species [(SchNMe2)AlMe]+ in further reaction with 1 equiv of AlCl3 or NaBPh4 as the chloride abstracting reagents. The exposure of the resulting methylaluminum cationic species to an excess of dry dioxygen at 0 degrees C afforded the alkoxyaluminum cationic species, [(SchNMe2)AlOMe]+ or [(SchNMe2)AlOPh]+. The alkoxylaluminum cations proved to be a very efficient catalyst in the polymerization of epsilon-caprolactone. PMID:15360224

Lewi?ski, Janusz; Horeglad, Pawe?; Dranka, Maciej; Justyniak, Iwona



Combination of Adhesive-tape-based Sampling and Fluorescence in situ Hybridization for Rapid Detection of Salmonella on Fresh Produce  

PubMed Central

This protocol describes a simple approach for adhesive-tape-based sampling of tomato and other fresh produce surfaces, followed by on-tape fluorescence in situ hybridization (FISH) for rapid culture-independent detection of Salmonella spp. Cell-charged tapes can also be placed face-down on selective agar for solid-phase enrichment prior to detection. Alternatively, low-volume liquid enrichments (liquid surface miniculture) can be performed on the surface of the tape in non-selective broth, followed by FISH and analysis via flow cytometry. To begin, sterile adhesive tape is brought into contact with fresh produce, gentle pressure is applied, and the tape is removed, physically extracting microbes present on these surfaces. Tapes are mounted sticky-side up onto glass microscope slides and the sampled cells are fixed with 10% formalin (30 min) and dehydrated using a graded ethanol series (50, 80, and 95%; 3 min each concentration). Next, cell-charged tapes are spotted with buffer containing a Salmonella-targeted DNA probe cocktail and hybridized for 15 - 30 min at 55°C, followed by a brief rinse in a washing buffer to remove unbound probe. Adherent, FISH-labeled cells are then counterstained with the DNA dye 4',6-diamidino-2-phenylindole (DAPI) and results are viewed using fluorescence microscopy. For solid-phase enrichment, cell-charged tapes are placed face-down on a suitable selective agar surface and incubated to allow in situ growth of Salmonella microcolonies, followed by FISH and microscopy as described above. For liquid surface miniculture, cell-charged tapes are placed sticky side up and a silicone perfusion chamber is applied so that the tape and microscope slide form the bottom of a water-tight chamber into which a small volume (? 500 ?L) of Trypticase Soy Broth (TSB) is introduced. The inlet ports are sealed and the chambers are incubated at 35 - 37°C, allowing growth-based amplification of tape-extracted microbes. Following incubation, inlet ports are unsealed, cells are detached and mixed with vigorous back and forth pipetting, harvested via centrifugation and fixed in 10% neutral buffered formalin. Finally, samples are hybridized and examined via flow cytometry to reveal the presence of Salmonella spp. As described here, our "tape-FISH" approach can provide simple and rapid sampling and detection of Salmonella on tomato surfaces. We have also used this approach for sampling other types of fresh produce, including spinach and jalapeño peppers.

Bisha, Bledar; Brehm-Stecher, Byron F.



Simple fiber optic refractive index sensor based on evanescent higher order modes  

NASA Astrophysics Data System (ADS)

This paper presents an optical fiber refractive index sensor based on the evanescent higher order modes. Its structure and principle are quite simple. The sensor is composed of two segments of optical fibers that are spliced together. An ordinary multimode fiber with a core diameter of 50 ?m is used to input the light. The functions of a second multimode fiber with a core diameter of 200 ?m are twofold. In the region of the splice, a section of the cladding a few centimetre long is removed by an electrical discharge. This part works as a sensing element, and the rest of the fiber is used to output the light. Once the light travels though the input fiber and crosses the splice to enter the second fiber, numerous modes both guided and leaky are generated due to the abrupt increase of the core diameter. The evanescent light fields of these guided modes are sensitive to changes in the refractive index of the material surrounding the fiber cladding. The evanescent field change directly causes a change in the output light intensity. The developed sensor is compact in size, simple to fabricate, promising in performance, and has a high potential for practical applications.

Chen, Jiahua; Bock, Wojtek J.; Mikulic, Predrag



Application of biologically based computer modeling to simple or complex mixtures.  

PubMed Central

The complexity and the astronomic number of possible chemical mixtures preclude any systematic experimental assessment of toxicology of all potentially troublesome chemical mixtures. Thus, the use of computer modeling and mechanistic toxicology for the development of a predictive tool is a promising approach to deal with chemical mixtures. In the past 15 years or so, physiologically based pharmacokinetic/pharmacodynamic (PBPK/PD) modeling has been applied to the toxicologic interactions of chemical mixtures. This approach is promising for relatively simple chemical mixtures; the most complicated chemical mixtures studied so far using this approach contained five or fewer component chemicals. In this presentation we provide some examples of the utility of PBPK/PD modeling for toxicologic interactions in chemical mixtures. The probability of developing predictive tools for simple mixtures using PBPK/PD modeling is high. Unfortunately, relatively few attempts have been made to develop paradigms to consider the risks posed by very complex chemical mixtures such as gasoline, diesel, tobacco smoke, etc. However, recent collaboration between scientists at Colorado State University and engineers at Rutgers University attempting to use reaction network modeling has created hope for the possible development of a modeling approach with the potential of predicting the outcome of toxicology of complex chemical mixtures. We discuss the applications of reaction network modeling in the context of petroleum refining and its potential for elucidating toxic interactions with mixtures.

Liao, Kai H; Dobrev, Ivan D; Dennison, James E; Andersen, Melvin E; Reisfeld, Brad; Reardon, Kenneth F; Campain, Julie A; Wei, Wei; Klein, Michael T; Quann, Richard J; Yang, Raymond S H



A fluorescent probe directly detect peroxynitrite based on boronate oxidation and its applications for fluorescence imaging in living cells.  


We present the design, synthesis, spectroscopy, and biological applications of PyBor, a new type of fluorescent probe for peroxynitrite detection in aqueous solution and living cells. The probe employs pyrene as the fluorophore, and is equipped with a chemically responsive unit boronate. The fluorescent probe can selectively detect peroxynitrite with fluorimetric determination and high-performance liquid chromatography analyses in aqueous solution and RAW264.7 cells intracellular free extracts. We also study our probe to time dependent peroxynitrite release from 3-morpholinylsydnonimine hydrochloride. Confocal microscopy experiments using mouse macrophage cell line RAW264.7 show that PyBor is able to detect the different intracellular peroxynitrite levels. In addition, we have performed quantum chemical calculations with TD-DFT/M06/TZVP level with COSMO solvation model basis sets using a suite of Gaussian 09 programs to provide insights into the structure optical properties of PyBor and PyOH. PMID:22741158

Yu, Fabiao; Song, Ping; Li, Peng; Wang, Bingshuai; Han, Keli



Scenario based outdoor simulation in pre-hospital trauma care using a simple mannequin model  

PubMed Central

Introduction We describe a system of scenario-based training using simple mannequins under realistic circumstances for the training of pre-hospital care providers. Methods A simple intubatable mannequin or student volunteers are used together with a training version of the equipment used on a routine basis by the pre-hospital care team (doctor + paramedic). Training is conducted outdoors at the base location all year round. The scenarios are led by scenario facilitators who are predominantly senior physicians. Their role is to brief the training team and guide the scenario, results of patient assessment and the simulated responses to interventions and treatment. Pilots, fire-fighters and medical students are utilised in scenarios to enhance realism by taking up roles as bystanders, additional ambulance staff and police. These scenario participants are briefed and introduced to the scene in a realistic manner. After completion of the scenario, the training team would usually be invited to prepare and deliver a hospital handover as they would in a real mission. A formal structured debrief then takes place. Results This training method technique has been used for the training of all London Helicopter Emergency Medical Service (London HEMS) doctors and paramedics over the last 24 months. Informal participant feedback suggests that this is a very useful teaching method, both for improving motor skills, critical decision-making, scene management and team interaction. Although formal assessment of this technique has not yet taken place we describe how this type of training is conducted in a busy operational pre-hospital trauma service. Discussion The teaching and maintenance of pre-hospital care skills is essential to an effective pre-hospital trauma care system. Simple mannequin based scenario training is feasible on a day-to-day basis and has the advantages of low cost, rapid set up and turn around. The scope of scenarios is limited only by the imagination of the trainers. Significant effort is made to put the participants into "the Zone" - the psychological mindset, where they believe they are in a realistic setting and treating a real patient, so that they gain the most from each teaching session. The method can be used for learning new skills, communication and leadership as well as maintaining existing skills. Conclusion The method described is a low technology, low cost alternative to high technology simulation which may provide a useful adjunct to delivering effective training when properly prepared and delivered. We find this useful for both induction and regular training of pre-hospital trauma care providers.



A new selective colorimetric and fluorescent chemodosimeter for HSO4- based on hydrolysis of Schiff base  

NASA Astrophysics Data System (ADS)

Two new receptors 1 and 2 were prepared, and their chromogenic and fluorogenic behaviors toward various anions were investigated. Receptors 1 and 2 show exclusive response toward HSO4- ion and also distinguish HSO4- from other anions by different color changes in aqueous solution (CH3CN/H2O = 4/1, v/v). Between them receptor 1 selectively exhibits a pronounced HSO4--induced fluorescence enhancement. The detection limit for the HSO4- ion was determined as (0.24 ± 0.03 ?M). Thus, the receptor 1 can be used as a colorimetric and fluorescent sensor for the determination of HSO4- ion. The sensing mechanism has been suggested to proceed via a hydrolysis process. The hydrolysis product has been isolated and further identified by 1H NMR spectroscopy, ESI-MS analysis and X-ray diffraction.

Lin, Chi-Yung; Huang, Keh-Feng; Yen, Yao-Pin



A sensitive, homogeneous fluorescence assay for detection of thymine DNA glycosylase activity based on exonuclease-mediated amplification.  


A novel homogeneous fluorescence assay strategy for highly sensitive detection of thymine DNA glycosylase (TDG) enzyme activity based on the exonuclease-mediated signal amplification reaction was reported. PMID:23703157

Chen, Cuihua; Zhou, Dianming; Tang, Hao; Liang, Manfen; Jiang, Jianhui



Copper sensing based on the far-red fluorescent protein, HcRed, from Heteractis crispa.  


In this article, we report for the first time on the copper (Cu(2+)) binding characteristics of the far-red fluorescent protein, HcRed, and its application in the development of a reagentless sensing system for copper. The far-red emission of HcRed (lambda(max) = 645 nm) where background cellular fluorescence is low should prove to be advantageous in the development of the sensing system. In the studies performed in our laboratory, we found that the fluorescence of HcRed is quenched in the presence of copper ions (Cu(2+)). The results obtained through UV-visible and circular dichroism spectra generated in the presence and absence of copper, as well as Stern-Volmer plots at different temperatures, indicate static quenching of HcRed fluorescence in the presence of copper, possibly through the formation of a copper-protein complex. On the basis of this observation, we developed a reagentless sensing system for the detection of copper(II) based on HcRed as the biosensing element. A detection limit for Cu(2+) in the nanomolar range was obtained. HcRed was found to bind copper ions selectively when compared with other divalent ions. A dissociation constant of 3.6muM was observed for copper binding. Histidine and cysteine residues are commonly involved in copper binding within proteins; therefore, to investigate the role of these amino acids present in HcRed, we chemically modified Cys and His residues using iodoacetamide and diethyl pyrocarbonate, respectively. The effect of copper addition on the fluorescence of the chemically modified HcRed was investigated. The His modification of HcRed substantially affected copper ion binding, pointing to histidine as the possible amino acid residue involved in the binding of copper ions in HcRed. A purification strategy for HcRed was also developed based on a copper immobilized affinity column without the addition of any affinity tag on the protein. The HcRed-based copper sensing system can potentially be employed to perform intracellular copper detection by genetically encoding the biosensing element or can be employed in environmental sensing. PMID:17599800

Rahimi, Yasmeen; Shrestha, Suresh; Banerjee, Tanushree; Deo, Sapna K



A label-free fluorescent molecular beacon based on DNA-templated silver nanoclusters for detection of adenosine and adenosine deaminase.  


A simple and reliable fluorescent molecular beacon is developed utilizing DNA-templated silver nanoclusters as a signal indicator and adenosine triphosphate (ATP) and adenosine deaminase as mechanical activators. PMID:22543727

Zhang, Min; Guo, Su-Miao; Li, Ying-Ru; Zuo, Peng; Ye, Bang-Ce



Design and Implementation of a Simple Model Interface for Component Based Modeling  

NASA Astrophysics Data System (ADS)

Component based architectures offer an alternative approach for building large, complex hydrologic modeling systems. In contrast to more traditional coding structures (i.e. sequential and modular modeling approaches), component-based modeling allows individuals to construct autonomous computational units that can be linked together through the exchange of shared boundary conditions during run-time. One of the challenges in component-based modeling is designing simple yet robust component interface definitions that allow hydrologic processes to be quickly incorporated into a modeling system. In this study we address this challenge by presenting a new interface design that simplifies the process of implementing the Open Modeling Interface (OpenMI). A component is created by (1) authoring an xml-based configuration file that defines the component's core properties and (2) creating a class that implements the newly defined interface and its three methods: initialize, perform time step, and finish. We will present this approach for creating components and demonstrate how it can be used to create a hydrologic model.

Castronova, A. M.; Goodall, J. L.



Simple colorimetric bacterial detection and high-throughput drug screening based on a graphene-enzyme complex  

NASA Astrophysics Data System (ADS)

A simple, colorimetric, sensitive, cost-effective and high-throughput system based on a positively charged graphene oxide-enzyme complex was developed for bacterial detection and drug screening.A simple, colorimetric, sensitive, cost-effective and high-throughput system based on a positively charged graphene oxide-enzyme complex was developed for bacterial detection and drug screening. Electronic supplementary information (ESI) available: Experimental details and supporting figures and procedures. See DOI: 10.1039/c2nr32704j

Li, Juan; Wu, Ling-Jie; Guo, Shan-Shan; Fu, Hua-E.; Chen, Guo-Nan; Yang, Huang-Hao



Fluorescence-based fast diagnostics platform for the direct and indirect immunodiagnostic analysis methods  

NASA Astrophysics Data System (ADS)

VTT Technical Research Centre of Finland has developed two reader prototypes for immunodiagnostic tests. VTT has also developed a one-step, homogeneous noncompetitive immunoassay for small analytes using recombinant antibodies and morphine as the model analyte. VTT developed reader for lateral flow test. Lateral flow test is a strip, which has a sample area and a detection area. In the sample area there are antibodies attached to gold or fluorescence particles, which are captured into the detection area, if a sample has a desired analyte. The concentration of the measured sample is then calculated from the fluorescence detection or color change. The second developed prototype reader is based on Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET). In this reader samples are put on microwell array. There are two fluorophores in each of the wells and emission of both fluorophores is measured. The sample concentration is calculated from these emission signals. The optimization of homogenous FRET assays for morphine was included to this project. The first results obtained with the TR-FRET reader prototype show that the sensitivity of the current morphine test is clearly adequate.

Mannila, Rami; Pulli, Timo; Saari, Heikki; Tappura, Kirsi; Tuppurainen, Jussi; Välimäki, Hannu; Niskanen, Aimo



Desktop computer-based image analysis of cell surface fluorescence patterning from a photographic source.  


We report the use of standard computer-based image analysis technology to analyze, from a photographic source, individual cell surface receptor site patterns using fluorochrome labeling. The model used in this study was a Strongylocentrotus purpuratus sea urchin embryo labeled with fluorescein isothiocyanate-conjugated wheat germ agglutinin (FITC-WGA) (0.5 mg/ml for 5 min at 15 degrees C incubated with fertilization membrane free embryos). Image capture was performed using desktop-type digital scanning, and the images were imported into Adobe Photoshop for study. All images analyzed were derived from a single photographic negative: 1) the actual micrograph printed from the negative and scanned into a Macintosh IIx computer; 2) the scanned negative itself; and, 3) a high resolution scanning process used with a Kodak Photo CD. Patterns were analyzed using the densitometry feature of Photoshop, and were similar enough from all 3 scanned images to indicate that any of the 3 scanning processes can be used for fluorescence patterning analysis. Adobe Photoshop was also used to highlight, for closer analysis, the fluorescence patterns by producing 3-D effects, border mapping and transition area detailing. The desktop image analysis procedures described here to study fluorescence patterning require no expensive scientific hardware or software. PMID:8863858

Latham, V H; Latham, L E; Oppenheimer, S B



Fluorescence-based proxies for lignin in freshwater dissolved organic matter  

USGS Publications Warehouse

Lignin phenols have proven to be powerful biomarkers in environmental studies; however, the complexity of lignin analysis limits the number of samples and thus spatial and temporal resolution in any given study. In contrast, spectrophotometric characterization of dissolved organic matter (DOM) is rapid, noninvasive, relatively inexpensive, requires small sample volumes, and can even be measured in situ to capture fine-scale temporal and spatial detail of DOM cycling. Here we present a series of cross-validated Partial Least Squares models that use fluorescence properties of DOM to explain up to 91% of lignin compositional and concentration variability in samples collected seasonally over 2 years in the Sacramento River/San Joaquin River Delta in California, United States. These models were subsequently used to predict lignin composition and concentration from fluorescence measurements collected during a diurnal study in the San Joaquin River. While modeled lignin composition remained largely unchanged over the diurnal cycle, changes in modeled lignin concentrations were much greater than expected and indicate that the sensitivity of fluorescence-based proxies for lignin may prove invaluable as a tool for selecting the most informative samples for detailed lignin characterization. With adequate calibration, similar models could be used to significantly expand our ability to study sources and processing of DOM in complex surface water systems.

Hernes, Peter J.; Bergamaschi, Brian A.; Eckard, Robert S.; Spencer, Robert G. M.



An ultrasensitive fluorescence assay for protein detection by hybridization chain reaction-based DNA nanotags.  


An ultrasensitive fluorescence method for determination of protein is developed based on hybridization chain reaction (HCR). In this assay, the streptavidin-magnetic nanobeads were conjugated to biotinylated initiators and biotinylated anti-IgG. In the presence of human IgG, the magnetic nanobeads were fixed on the substrate and the carried initiators propagated the chain reaction of hybridization to form the nicked polymers. Because the nanobead probe carries with a large number of oligonucleotides per protein binding event, there is obvious amplification in the nicked polymers. Then, numerous SYBR Green I molecules were intercalated into the grooves of the long dsDNA polymers, generating a substantially apparent increase in the corresponding fluorescence intensity. With HCR amplification and magenetic nanobead to preamplify the fluorescence signal and reduce the background signal, the detection limit of this assay was 14aM. Compared with the reported protein detection methods, our method exhibited ultrahigh sensitivity. In addition, the proposed method possessed excellent selectivity and low matrix effect. What is more, the assay was also studied for clinical application in human serum with a satisfactory and reliable result. PMID:24001583

Dai, Shuang; Xue, Qingwang; Zhu, Jing; Ding, Yongshun; Jiang, Wei; Wang, Lei



Tandem Fluorescent Proteins as Enhanced FRET-based Substrates for Botulinum Neurotoxin Activity  

PubMed Central

The light chain of botulinum neurotoxin A (BoNT/A-LC) is a zinc-metalloprotease that requires two extended exosites for optimal substrate binding and recognition of its intracellular target SNAP25. CFP and YFP connected through SNAP25 peptide (141–206) containing both exosites (CsY) has been used in a FRET-based assay for BoNT/A. To further improve the FRET efficiency in this BoNT/A substrate for in vitro high-throughput assays, we explored the feasibility of enhancing the capture of CFP emission by doubling the number of YFP acceptors. In comparison to CsY, the tandem fluorescence substrates CsYY and YsCsY enhanced the ratiometric fluorescence signal between YFP and CFP. YsCsY, containing two substrate sites, offered the greatest fluorometric change upon toxin-catalyzed cleavage. In addition to known approaches for enhancing fluorescence yield through various mutations, this alternative tandem substrate approach can boost the FRET signal and is particularly useful for substrates requiring extensive exosite recognition for specificity.

Pires-Alves, Melissa; Ho, Mengfei; Aberle, Karla K.; Janda, Kim D.; Wilson, Brenda A.



Fiber optic based fluorescence detection system for in vivo studies of exogenous chromophore pharmacokinetics  

NASA Astrophysics Data System (ADS)

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.

Doiron, Daniel R.; Dunn, J. B.; Mitchell, W. L.; Dalton, Brian K.; Garbo, Greta M.; Warner, Jon A.



The fluorescence-based acetylation assay using thiol-sensitive probes.  


Lysine acetyltransferases (KATs) catalyze the acetylation of specific lysine residues in histone and nonhistone proteins. The enzymatic activities of KATs are involved in a broad spectrum of cellular processes. Thus far, the reaction of KAT catalysis has been studied by various bioanalytical methods such as radioisotopic labeling, spectrophotometric and fluorometric measurements, and antibody-dependent immunosorbent assays. In particular, the fluorescent method has the advantage of simplicity for implementation, fast assay speed, fine signal to noise ratio, and superior sensitivity. We describe here the technical protocols of using thiol-sensitive fluorogenic probes for the fluorescent analysis of enzymatic activities of KATs, with males on the first (MOF) as an exemplary KAT enzyme. 7-Diethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin (CPM) is selected as the KAT probe owing to its fast reaction kinetics with coenzyme A (CoA) and excellent fluorogenicity upon thiol conjugation. The fluorescence-based acetylation assay is well suited for both kinetic characterization of KAT catalysis and KAT inhibitor investigation. PMID:23381866

Gao, Tielong; Yang, Chao; Zheng, Yujun George



Rapid image-based cytometry for comparison of fluorescent viability staining methods.  


The ability to accurately measure cell viability is important for any cell-based research. Traditionally, viability measurements have been performed using trypan blue exclusion method on hemacytometer, which allowed researchers to visually distinguish viable from nonviable cells. However, the trypan blue method is often limited to only cell lines or primary cells that have been rigorously purified. In the recent years, small desktop image-based cell counters have been developed for rapid cell concentration and viability measurement due to advances in imaging and optics technologies as well as novel fluorescent stains. In this work, we employed the Cellometer image-based cytometer to demonstrate the ability to simplify viability detection compared to the current methods. We compared various fluorescence viability detection methods using single- or dual-staining technique. Single-staining method using nucleic acid stains including ethidium bromide, propidium iodide, 7AAD, DAPI, Sytox Green and Sytox Red, and enzymatic stains including CFDA and Calcein AM were performed. All stains produced comparable results to trypan blue exclusion method for cell line samples. Dual-staining method using AO/PI, CFDA/PI, Calcein AM/PI and Hoechst 33342/PI that enumerates viable and non-viable cells was tested on primary cell samples with high debris contents. This method allowed exclusion of cellular debris and non-nucleated cells from analysis, which can eliminate the need to perform purification step during sample preparation, and improves the efficiency of viability detection method. Overall, these image-based fluorescent cell counters can simplify assay procedures as well as capture images for visual confirmation. PMID:22718197

Chan, Leo L; Wilkinson, Alisha R; Paradis, Benjamin D; Lai, Ning



Compact and cost effective instrument for detecting drug precursors in different environments based on fluorescence polarization  

NASA Astrophysics Data System (ADS)

Several techniques for detecting chemical drug precursors have been developed in the last decade. Most of them are able to identify molecules at very low concentration under lab conditions. Other commercial devices are able to detect a fixed number and type of target substances based on a single detection technique providing an absence of flexibility with respect to target compounds. The construction of compact and easy to use detection systems providing screening for a large number of compounds being able to discriminate them with low false alarm rate and high probability of detection is still an open concern. Under CUSTOM project, funded by the European Commission within the FP7, a stand-alone portable sensing device based on multiple techniques is being developed. One of these techniques is based on the LED induced fluorescence polarization to detect Ephedrine and Benzyl Methyl Keton (BMK) as a first approach. This technique is highly selective with respect to the target compounds due to the generation of properly engineered fluorescent proteins which are able to bind the target analytes, as it happens in an "immune-type reaction". This paper deals with the advances in the design, construction and validation of the LED induced fluorescence sensor to detect BMK analytes. This sensor includes an analysis module based on high performance LED and PMT detector, a fluidic system to dose suitable quantities of reagents and some printed circuit boards, all of them fixed in a small structure (167mm × 193mm × 228mm) with the capability of working as a stand-alone application.

Antolín-Urbaneja, J. C.; Eguizabal, I.; Briz, N.; Dominguez, A.; Estensoro, P.; Secchi, A.; Varriale, A.; Di Giovanni, S.; D'Auria, S.



Efficient near-infrared organic light-emitting devices based on low-gap fluorescent oligomers  

NASA Astrophysics Data System (ADS)

We report efficient near-infrared (NIR) organic light-emitting devices (OLEDs) based on fluorescent donor-acceptor-donor conjugated oligomers. The energies of the highest occupied and lowest unoccupied molecular orbitals of these oligomers are controlled by the donor and acceptor components, respectively; hence the energy gap and therefore the emission wavelength can be tuned by changing the strengths of the donor and acceptor components. External quantum efficiencies (EQEs) up to 1.6% and power efficiencies up to 7.0 mW/W are achieved in NIR OLEDs based on 4,9-bis(2,3-dihydrothieno[3,4-b][1,4]dioxin-5-yl)-6,7-dimethyl-[1,2,5]thiadiazolo[3,4-g]-quinoxaline (BEDOT-TQMe2), in which the electroluminescence peaks at a wavelength of 692 nm but extends to well above 800 nm. With a stronger acceptor in the oligomer, 4,8-bis(2,3-dihydrothieno-[3,4-b][1,4]dioxin-5-yl)benzo[1,2-c4,5-c']bis [1,2,5]thiadiazole (BEDOT-BBT) based devices show longer wavelength emission peaked at 815 nm, although the maximum EQE is reduced to 0.51% due to the lower fluorescent quantum yield of the NIR emitter. The efficiencies of these NIR OLEDs are further increased by two to three times by using the sensitized fluorescent device structure, leading to a maximum EQE of 3.1% for BEDOT-TQMe2 and 1.6% for BEDOT-BBT based devices.

Yang, Yixing; Farley, Richard T.; Steckler, Timothy T.; Eom, Sang-Hyun; Reynolds, John R.; Schanze, Kirk S.; Xue, Jiangeng



Simple approach to approximate quantum error correction based on the transpose channel  

SciTech Connect

We demonstrate that there exists a universal, near-optimal recovery map--the transpose channel--for approximate quantum error-correcting codes, where optimality is defined using the worst-case fidelity. Using the transpose channel, we provide an alternative interpretation of the standard quantum error correction (QEC) conditions and generalize them to a set of conditions for approximate QEC (AQEC) codes. This forms the basis of a simple algorithm for finding AQEC codes. Our analytical approach is a departure from earlier work relying on exhaustive numerical search for the optimal recovery map, with optimality defined based on entanglement fidelity. For the practically useful case of codes encoding a single qubit of information, our algorithm is particularly easy to implement.

Ng, Hui Khoon; Mandayam, Prabha [Institute for Quantum Information, California Institute of Technology, Pasadena, California 91125 (United States)



Characterisation of simple sequence repeats from human ESTs and creation of a comprehensive data base.  


The aim of the present analysis is to create a complete database of Simple Sequence Repeats (SSRs) occurring in the Expressed Sequence Tags (ESTs) of Human Genome. For the recognition of SSRs, various Bioinformatics Tools, Databases and Softwares are employed. The data are analysed statistically for Mean, Density and Frequency distribution of the identified Repeats. A consolidated Database is created and maintained in SAS Environment. This database can provide comprehensive information regarding: 1) the complete sequence of microsatellites/tandem repeats; 2) the number of times each one is repeated; 3) the starting and ending position of each repeat; 4) its length. In addition the database can also provide the location of the gene and also its function. As these microsatellites are known to be widely used in a variety of fundamental and applied fields of life and medical sciences, the availability and accessibility to such comprehensive data base would greatly help the researchers. PMID:20615836