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Sample records for fluorescent lipid probes

  1. Fluorescent lipid probes : some properties and applications (a review)

    NARCIS (Netherlands)

    Maier, O; Oberle, [No Value; Hoekstra, D

    Odd as it may seem, experimental challenges in lipid research are often hampered by the simplicity of the lipid structure. Since, as in protein research. mutants or overexpression of lipids are not realistic, a considerable amount of lipid research relies on the use Of tagged lipid analogues.

  2. Fluorescent molecular probes based on excited state prototropism in lipid bilayer membrane

    Science.gov (United States)

    Mohapatra, Monalisa; Mishra, Ashok K.

    2012-03-01

    Excited state prototropism (ESPT) is observed in molecules having one or more ionizable protons, whose proton transfer efficiency is different in ground and excited states. The interaction of various ESPT molecules like naphthols and intramolecular ESPT (ESIPT) molecules like hydroxyflavones etc. with different microheterogeneous media have been studied in detail and excited state prototropism as a probe concept has been gaining ground. The fluorescence of different prototropic forms of such molecules, on partitioning to an organized medium like lipid bilayer membrane, often show sensitive response to the local environment with respect to the local structure, physical properties and dynamics. Our recent work using 1-naphthol as an ESPT fluorescent molecular probe has shown that the incorporation of monomeric bile salt molecules into lipid bilayer membranes composed from dipalmitoylphosphatidylcholine (DPPC, a lung surfactant) and dimyristoylphosphatidylcholine (DMPC), in solid gel and liquid crystalline phases, induce appreciable wetting of the bilayer up to the hydrocarbon core region, even at very low (fisetin, an ESIPT molecule having antioxidant properties, in lipid bilayer membrane has been sensitively monitored from its intrinsic fluorescence behaviour.

  3. DHA-fluorescent probe is sensitive to membrane order and reveals molecular adaptation of DHA in ordered lipid microdomains☆

    Science.gov (United States)

    Teague, Heather; Ross, Ron; Harris, Mitchel; Mitchell, Drake C.; Shaikh, Saame Raza

    2012-01-01

    Docosahexaenoic acid (DHA) disrupts the size and order of plasma membrane lipid microdomains in vitro and in vivo. However, it is unknown how the highly disordered structure of DHA mechanistically adapts to increase the order of tightly packed lipid microdomains. Therefore, we studied a novel DHA-Bodipy fluorescent probe to address this issue. We first determined if the DHA-Bodipy probe localized to the plasma membrane of primary B and immortal EL4 cells. Image analysis revealed that DHA-Bodipy localized into the plasma membrane of primary B cells more efficiently than EL4 cells. We then determined if the probe detected changes in plasma membrane order. Quantitative analysis of time-lapse movies established that DHA-Bodipy was sensitive to membrane molecular order. This allowed us to investigate how DHA-Bodipy physically adapted to ordered lipid microdomains. To accomplish this, we employed steady-state and time-resolved fluorescence anisotropy measurements in lipid vesicles of varying composition. Similar to cell culture studies, the probe was highly sensitive to membrane order in lipid vesicles. Moreover, these experiments revealed, relative to controls, that upon incorporation into highly ordered microdomains, DHA-Bodipy underwent an increase in its fluorescence lifetime and molecular order. In addition, the probe displayed a significant reduction in its rotational diffusion compared to controls. Altogether, DHA-Bodipy was highly sensitive to membrane order and revealed for the first time that DHA, despite its flexibility, could become ordered with less rotational motion inside ordered lipid microdomains. Mechanistically, this explains how DHA acyl chains can increase order upon formation of lipid microdomains in vivo. PMID:22841541

  4. Fluorescence of nitrobenzoxadiazole (NBD) labeled lipids in model membranes is connected not to lipid mobility, but to probe location

    Czech Academy of Sciences Publication Activity Database

    Amaro, Mariana; Filipe, H. A.; Ramalho, J. P. P.; Hof, Martin; Loura, L. M. S.

    2016-01-01

    Roč. 18, č. 10 (2016), s. 7042-7054 ISSN 1463-9076 R&D Projects: GA ČR GBP208/12/G016 Institutional support: RVO:61388955 Keywords : fluorescence * nitrobenzoxadiazole Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 4.123, year: 2016

  5. Nine New Fluorescent Probes

    Science.gov (United States)

    Lin, Tsung-I.; Jovanovic, Misa V.; Dowben, Robert M.

    1989-06-01

    Absorption and fluorescence spectroscopic studies are reported here for nine new fluorescent probes recently synthesized in our laboratories: four pyrene derivatives with substituents of (i) 1,3-diacetoxy-6,8-dichlorosulfonyl, (ii) 1,3-dihydroxy-6,8-disodiumsulfonate, (iii) 1,3-disodiumsulfonate, and (iv) l-ethoxy-3,6,8-trisodiumsulfonate groups, and five [7-julolidino] coumarin derivatives with substituents of (v) 3-carboxylate-4-methyl, (vi) 3- methylcarboxylate, (vii) 3-acetate-4-methyl, (viii) 3-propionate-4-methyl, and (ix) 3-sulfonate-4-methyl groups. Pyrene compounds i and ii and coumarin compounds v and vi exhibit interesting absorbance and fluorescence properties: their absorption maxima are red shifted compared to the parent compound to the blue-green region, and the band width broadens considerably. All four blue-absorbing dyes fluoresce intensely in the green region, and the two pyrene compounds emit at such long wavelengths without formation of excimers. The fluorescence properties of these compounds are quite environment-sensitive: considerable spectral shifts and fluorescence intensity changes have been observed in the pH range from 3 to 10 and in a wide variety of polar and hydrophobic solvents with vastly different dielectric constants. The high extinction and fluorescence quantum yield of these probes make them ideal fluorescent labeling reagents for proteins, antibodies, nucleic acids, and cellular organelles. The pH and hydrophobicity-dependent fluorescence changes can be utilized as optical pH and/or hydrophobicity indicators for mapping environmental difference in various cellular components in a single cell. Since all nine probes absorb in the UV, but emit at different wavelengths in the visible, these two groups of compounds offer an advantage of utilizing a single monochromatic light source (e.g., a nitrogen laser) to achieve multi-wavelength detection for flow cytometry application. As a first step to explore potential application in

  6. Probing lipid membrane electrostatics

    Science.gov (United States)

    Yang, Yi

    The electrostatic properties of lipid bilayer membranes play a significant role in many biological processes. Atomic force microscopy (AFM) is highly sensitive to membrane surface potential in electrolyte solutions. With fully characterized probe tips, AFM can perform quantitative electrostatic analysis of lipid membranes. Electrostatic interactions between Silicon nitride probes and supported zwitterionic dioleoylphosphatidylcholine (DOPC) bilayer with a variable fraction of anionic dioleoylphosphatidylserine (DOPS) were measured by AFM. Classical Gouy-Chapman theory was used to model the membrane electrostatics. The nonlinear Poisson-Boltzmann equation was numerically solved with finite element method to provide the potential distribution around the AFM tips. Theoretical tip-sample electrostatic interactions were calculated with the surface integral of both Maxwell and osmotic stress tensors on tip surface. The measured forces were interpreted with theoretical forces and the resulting surface charge densities of the membrane surfaces were in quantitative agreement with the Gouy-Chapman-Stern model of membrane charge regulation. It was demonstrated that the AFM can quantitatively detect membrane surface potential at a separation of several screening lengths, and that the AFM probe only perturbs the membrane surface potential by external field created by the internai membrane dipole moment. The analysis yields a dipole moment of 1.5 Debye per lipid with a dipole potential of +275 mV for supported DOPC membranes. This new ability to quantitatively measure the membrane dipole density in a noninvasive manner will be useful in identifying the biological effects of the dipole potential. Finally, heterogeneous model membranes were studied with fluid electric force microscopy (FEFM). Electrostatic mapping was demonstrated with 50 nm resolution. The capabilities of quantitative electrostatic measurement and lateral charge density mapping make AFM a unique and powerful

  7. Interface-Targeting Strategy Enables Two-Photon Fluorescent Lipid Droplet Probes for High-Fidelity Imaging of Turbid Tissues and Detecting Fatty Liver.

    Science.gov (United States)

    Guo, Lifang; Tian, Minggang; Feng, Ruiqing; Zhang, Ge; Zhang, Ruoyao; Li, Xuechen; Liu, Zhiqiang; He, Xiuquan; Sun, Jing Zhi; Yu, Xiaoqiang

    2018-04-04

    Lipid droplets (LDs) with unique interfacial architecture not only play crucial roles in protecting a cell from lipotoxicity and lipoapoptosis but also closely relate with many diseases such as fatty liver and diabetes. Thus, as one of the important applied biomaterials, fluorescent probes with ultrahigh selectivity for in situ and high-fidelity imaging of LDs in living cells and tissues are critical to elucidate relevant physiological and pathological events as well as detect related diseases. However, available probes only utilizing LDs' waterless neutral cores but ignoring the unique phospholipid monolayer interfaces exhibit low selectivity. They cannot differentiate neutral cores of LDs from intracellular other lipophilic microenvironments, which results in extensively cloud-like background noise and severely limited their bioapplications. Herein, to design LD probes with ultrahigh selectivity, the exceptional interfacial architecture of LDs is considered adequately and thus an interface-targeting strategy is proposed for the first time. According to the novel strategy, we have developed two amphipathic fluorescent probes (N-Cy and N-Py) by introducing different cations into a lipophilic fluorophore (nitrobenzoxadiazole (NBD)). Consequently, their cationic moiety precisely locates the interfaces through electrostatic interaction and simultaneously NBD entirely embeds into the waterless core via hydrophobic interaction. Thus, high-fidelity and background-free fluorescence imaging of LDs are expectably realized in living cells in situ. Moreover, LDs in turbid tissues like skeletal muscle slices have been clearly imaged (up to 82 μm depth) by a two-photon microscope. Importantly, using N-Cy, we not only intuitively monitored the variations of LDs in number, size, and morphology but also clearly revealed their abnormity in hepatic tissues resulting from fatty liver. Therefore, these unique probes provide excellent imaging tools for elucidating LD

  8. Application of lipid peroxidation and protein oxidation biomarkers for oxidative damage in mammalian cells. A comparison with two fluorescent probes

    NARCIS (Netherlands)

    Orhan, H.; Gurer-Orhan, H.; Vriese, E.; Vermeulen, N.P.E.; Meerman, J.H.N.

    2006-01-01

    We recently developed two biomarker sets for oxidative damage: one for determination of lipid peroxidation (LPO) degradation products; acetaldehyde, propanal, butanal, pentanal, hexanal, heptanal, octanal, nonanal, malondialdehyde and acetone, by a gas chromatography-electron capture detection

  9. A fluorescent probe for ecstasy.

    Science.gov (United States)

    Masseroni, D; Biavardi, E; Genovese, D; Rampazzo, E; Prodi, L; Dalcanale, E

    2015-08-18

    A nanostructure formed by the insertion in silica nanoparticles of a pyrene-derivatized cavitand, which is able to specifically recognize ecstasy in water, is presented. The absence of effects from interferents and an efficient electron transfer process occurring after complexation of ecstasy, makes this system an efficient fluorescent probe for this popular drug.

  10. Lateral distribution of NBD-PC fluorescent lipid analogs in membranes probed by molecular dynamics-assisted analysis of Förster Resonance Energy Transfer (FRET) and fluorescence quenching.

    Science.gov (United States)

    Loura, Luís M S

    2012-11-08

    Förster resonance energy transfer (FRET) is a powerful tool used for many problems in membrane biophysics, including characterization of the lateral distribution of lipid components and other species of interest. However, quantitative analysis of FRET data with a topological model requires adequate choices for the values of several input parameters, some of which are difficult to obtain experimentally in an independent manner. For this purpose, atomistic molecular dynamics (MD) simulations can be potentially useful as they provide direct detailed information on transverse probe localization, relative probe orientation, and membrane surface area, all of which are required for analysis of FRET data. This is illustrated here for the FRET pairs involving 1,6-diphenylhexatriene (DPH) as donor and either 1-palmitoyl,2-(6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino] hexanoyl)- sn-glycero-3-phosphocholine (C6-NBD-PC) or 1-palmitoyl,2-(12-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]dodecanoyl)-sn-glycero-3-phosphocholine (C12-NBD-PC) as acceptors, in fluid vesicles of 1,2-dipalmitoyl-sn-3-glycerophosphocholine (DPPC, 50 °C). Incorporation of results from MD simulations improves the statistical quality of model fitting to the experimental FRET data. Furthermore, the decay of DPH in the presence of moderate amounts of C12-NBD-PC (>0.4 mol%) is consistent with non-random lateral distribution of the latter, at variance with C6-NBD-PC, for which aggregation is ruled out up to 2.5 mol% concentration. These conclusions are supported by analysis of NBD-PC fluorescence self-quenching. Implications regarding the relative utility of these probes in membrane studies are discussed.

  11. New Fluorescence Probes for Biomolecules

    Directory of Open Access Journals (Sweden)

    Katarzyna Jurek

    2015-07-01

    Full Text Available Steady state fluorescence measurements have been used for the investigation of interaction between the bovine serum albumin (BSA and fluorescence probes: 3-hydroxy-2,4- bis[(3-methyl-1,3-benzoxazol-2(3H-ylidenemethyl]cyclobut-2-en-1-one (SQ6, 3-hydroxy- 2,4-bis[(3-methyl-1,3-benzothiazol-2(3H-ylidenemethyl]cyclobut-2-en-1-one (SQ7 and 3-hydroxy-2,4-bis[(1,3,3-trimethyl-1,3-dihydro-2H-indol-2-ylidenemethyl]cyclobut-2-en-1-one (SQ8. The binding constant between bovine serum albumin and squarine dyes has been determined by using both the Benesi-Hildebrand and Stern-Volmer equations. The negative value of free energy change indicates the existence of a spontaneous complexation process of BSA with squarine dyes.

  12. Flow cytometry, fluorescent probes, and flashing bacteria

    NARCIS (Netherlands)

    Bunthof, C.J.

    2002-01-01


    Key words: fluorescent probes, flow cytometry, CSLM, viability, survival, microbial physiology, lactic acid bacteria, Lactococcus lactis , Lactobacillus plantarum , cheese, milk,

  13. Cell-based lipid flippase assay employing fluorescent lipid derivatives

    DEFF Research Database (Denmark)

    Jensen, Maria Stumph; Costa, Sara; Günther-Pomorski, Thomas

    2016-01-01

    P-type ATPases in the P4 subfamily (P4-ATPases) are transmembrane proteins unique for eukaryotes that act as lipid flippases, i.e., to translocate phospholipids from the exofacial to the cytofacial monolayer of cellular membranes. While initially characterized as aminophospholipid translocases, s...... flippase activities in the plasma membrane of cells, using yeast as an example.......P-type ATPases in the P4 subfamily (P4-ATPases) are transmembrane proteins unique for eukaryotes that act as lipid flippases, i.e., to translocate phospholipids from the exofacial to the cytofacial monolayer of cellular membranes. While initially characterized as aminophospholipid translocases......, studies of individual P4-ATPase family members from fungi, plants, and animals show that P4-ATPases differ in their substrate specificities and mediate transport of a broader range of lipid substrates. Here, we describe an assay based on fluorescent lipid derivatives to monitor and characterize lipid...

  14. Genetically encoded fluorescent probe to visualize phosphatidylinositol

    Czech Academy of Sciences Publication Activity Database

    Eisenreichová, Andrea; Humpolíčková, Jana; Bouřa, Evžen

    2017-01-01

    Roč. 284, Suppl 1 (2017), s. 364-365 ISSN 1742-464X. [FEBS Congress /42./ From Molecules to Cells and Back. 10.09.2017-14.09.2017, Jerusalem] R&D Projects: GA ČR GJ15-21030Y; GA MŠk LO1302 Institutional support: RVO:61388963 Keywords : phosphatidylinositol * fluorescent probe Subject RIV: CE - Biochemistry

  15. Azaphthalocyanines: Red Fluorescent Probes for Cations

    Czech Academy of Sciences Publication Activity Database

    Nováková, V.; Lochman, L.; Zajícová, I.; Kopecký, K.; Miletin, M.; Lang, Kamil; Kirakci, Kaplan; Zimcik, P.

    2013-01-01

    Roč. 19, č. 16 (2013), s. 5025-5028 ISSN 0947-6539 R&D Projects: GA ČR GAP208/10/1678 Institutional support: RVO:61388980 Keywords : crown compounds * fluorescent probes * phthalocyanine s * potassium * sodium Subject RIV: CA - Inorganic Chemistry Impact factor: 5.696, year: 2013

  16. Recent developments in multimodality fluorescence imaging probes

    Directory of Open Access Journals (Sweden)

    Jianhong Zhao

    2018-05-01

    Full Text Available Multimodality optical imaging probes have emerged as powerful tools that improve detection sensitivity and accuracy, important in disease diagnosis and treatment. In this review, we focus on recent developments of optical fluorescence imaging (OFI probe integration with other imaging modalities such as X-ray computed tomography (CT, magnetic resonance imaging (MRI, positron emission tomography (PET, single-photon emission computed tomography (SPECT, and photoacoustic imaging (PAI. The imaging technologies are briefly described in order to introduce the strengths and limitations of each techniques and the need for further multimodality optical imaging probe development. The emphasis of this account is placed on how design strategies are currently implemented to afford physicochemically and biologically compatible multimodality optical fluorescence imaging probes. We also present studies that overcame intrinsic disadvantages of each imaging technique by multimodality approach with improved detection sensitivity and accuracy. KEY WORDS: Optical imaging, Fluorescence, Multimodality, Near-infrared fluorescence, Nanoprobe, Computed tomography, Magnetic resonance imaging, Positron emission tomography, Single-photon emission computed tomography, Photoacoustic imaging

  17. A simple and sensitive fluorescent probe for specific detection of ...

    Indian Academy of Sciences (India)

    Yan-Fei Kang

    A fluorescent probe, with simplicity of structure and convenience of synthesis, is capable of detecting ... Yan-Fei Kang et al. .... Pastore A, Federici G, Bertini E and Ptemonte F 2003 ... Urano Y 2010 New Strategies for Fluorescent Probe.

  18. Characterization of a new series of fluorescent probes for imaging membrane order.

    Directory of Open Access Journals (Sweden)

    Joanna M Kwiatek

    Full Text Available Visualization and quantification of lipid order is an important tool in membrane biophysics and cell biology, but the availability of environmentally sensitive fluorescent membrane probes is limited. Here, we present the characterization of the novel fluorescent dyes PY3304, PY3174 and PY3184, whose fluorescence properties are sensitive to membrane lipid order. In artificial bilayers, the fluorescence emission spectra are red-shifted between the liquid-ordered and liquid-disordered phases. Using ratiometric imaging we demonstrate that the degree of membrane order can be quantitatively determined in artificial liposomes as well as live cells and intact, live zebrafish embryos. Finally, we show that the fluorescence lifetime of the dyes is also dependent on bilayer order. These probes expand the current palate of lipid order-sensing fluorophores affording greater flexibility in the excitation/emission wavelengths and possibly new opportunities in membrane biology.

  19. The interaction of new piroxicam analogues with lipid bilayers--a calorimetric and fluorescence spectroscopic study.

    Science.gov (United States)

    Maniewska, Jadwiga; Szczęśniak-Sięga, Berenika; Poła, Andrzej; Sroda-Pomianek, Kamila; Malinka, Wiesław; Michalak, Krystyna

    2014-01-01

    The purpose of the present paper was to assess the ability of new piroxicam analogues to interact with the lipid bilayers. The results of calorimetric and fluorescence spectroscopic experiments of two new synthesized analogues of piroxicam, named PR17 and PR18 on the phase behavior of phospholipid bilayers and fluorescence quenching of fluorescent probes (Laurdan and Prodan), which molecular location within membranes is known with certainty, are shown in present work. The presented results revealed that, depending on the details of chemical structure, the studied compounds penetrated the lipid bilayers.

  20. Use of a Novel Rover-mounted Fluorescence Imager and Fluorescent Probes to Detect Biological Material in the Atacama Desert in Daylight

    Science.gov (United States)

    Weinstein, S.; Pane, D.; Warren-Rhodes, K.; Cockell, C.; Ernst, L. A.; Minkley, E.; Fisher, G.; Emani, S.; Wettergreen, D. S.; Wagner, M.

    2005-01-01

    We have developed an imaging system, the Fluorescence Imager (FI), for detecting fluorescence signals from sparse microorganisms and biofilms during autonomous rover exploration. The fluorescence signals arise both from naturally occurring chromophores, such as chlorophyll of cyanobacteria and lichens, and from fluorescent probes applied to soil and rocks. Daylight imaging has been accomplished by a novel use of a high-powered flashlamp synchronized to a CCD camera. The fluorescent probes are cell permanent stains that have extremely low intrinsic fluorescence (quantum yields less than 0.01) and a large fluorescence enhancement (quantum yields greater than 0.4) when bound to the target. Each probe specifically targets either carbohydrates, proteins, nucleic acids or membrane lipids, the four classes of macromolecules found in terrestrial life. The intent of the probes is to interrogate the environment for surface and endolithic life forms.

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

    Science.gov (United States)

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

    2009-01-01

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

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

    International Nuclear Information System (INIS)

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

    2000-01-01

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

  3. Fluorescent Lipids: Functional Parts of Fusogenic Liposomes and Tools for Cell Membrane Labeling and Visualization

    Directory of Open Access Journals (Sweden)

    Christian Kleusch

    2012-01-01

    Full Text Available In this paper a rapid and highly efficient method for controlled incorporation of fluorescent lipids into living mammalian cells is introduced. Here, the fluorescent molecules have two consecutive functions: First, they trigger rapid membrane fusion between cellular plasma membranes and the lipid bilayers of their carrier particles, so called fusogenic liposomes, and second, after insertion into cellular membranes these molecules enable fluorescence imaging of cell membranes and membrane traffic processes. We tested the fluorescent derivatives of the following essential membrane lipids for membrane fusion: Ceramide, sphingomyelin, phosphocholine, phosphatidylinositol-bisphosphate, ganglioside, cholesterol, and cholesteryl ester. Our results show that all probed lipids could more efficiently be incorporated into the plasma membrane of living cells than by using other methods. Moreover, labeling occurred in a gentle manner under classical cell culture conditions reducing cellular stress responses. Staining procedures were monitored by fluorescence microscopy and it was observed that sphingolipids and cholesterol containing free hydroxyl groups exhibit a decreased distribution velocity as well as a longer persistence in the plasma membrane compared to lipids without hydroxyl groups like phospholipids or other artificial lipid analogs. After membrane staining, the fluorescent molecules were sorted into membranes of cell organelles according to their chemical properties and biological functions without any influence of the delivery system.

  4. Benzothiadiazole Derivatives as Fluorescence Imaging Probes: Beyond Classical Scaffolds.

    Science.gov (United States)

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

    2015-06-16

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

  5. Assay of flippase activity in proteoliposomes using fluorescent lipid derivatives

    DEFF Research Database (Denmark)

    Marek, Magdalena; Günther-Pomorski, Thomas

    2016-01-01

    Specific membrane proteins, termed lipid flippases, play a central role in facilitating the movement of lipids across cellular membranes. In this protocol, we describe the reconstitution of ATP-driven lipid flippases in liposomes and the analysis of their in vitro flippase activity based on the use...... of fluorescent lipid derivatives. Working with purified and reconstituted systems provides a well-defined experimental setup and allows to directly characterize these membrane proteins at the molecular level....

  6. Flow Cytometric Applicability of Fluorescent Vitality Probes on Phytoplankton

    NARCIS (Netherlands)

    Peperzak, L.; Brussaard, C.P.D.

    2011-01-01

    The applicability of six fluorescent probes (four esterase probes: acetoxymethyl ester of Calcein [Calcein-AM], 5-chloromethylfluorescein diacetate [CMFDA], fluorescein diacetate [FDA], and 2',7'-dichlorofluorescein diacetate [H(2)DCFDA]; and two membrane probes: bis-(1,3-dibutylbarbituric acid)

  7. Synthesis and validation of novel cholesterol-based fluorescent lipids designed to observe the cellular trafficking of cationic liposomes.

    Science.gov (United States)

    Kim, Bieong-Kil; Seu, Young-Bae; Choi, Jong-Soo; Park, Jong-Won; Doh, Kyung-Oh

    2015-09-15

    Cholesterol-based fluorescent lipids with ether linker were synthesized using NBD (Chol-E-NBD) or Rhodamine B (Chol-E-Rh), and the usefulnesses as fluorescent probes for tracing cholesterol-based liposomes were validated. The fluorescent intensities of liposomes containing these modified lipids were measured and observed under a microscope. Neither compound interfered with the expression of GFP plasmid, and live cell images were obtained without interferences. Changes in the fluorescent intensity of liposomes containing Chol-E-NBD were followed by flow cytometry for up to 24h. These fluorescent lipids could be useful probes for trafficking of cationic liposome-mediated gene delivery. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. A cell-surface-anchored ratiometric fluorescent probe for extracellular pH sensing.

    Science.gov (United States)

    Ke, Guoliang; Zhu, Zhi; Wang, Wei; Zou, Yuan; Guan, Zhichao; Jia, Shasha; Zhang, Huimin; Wu, Xuemeng; Yang, Chaoyong James

    2014-09-10

    Accurate sensing of the extracellular pH is a very important yet challenging task in biological and clinical applications. This paper describes the development of an amphiphilic lipid-DNA molecule as a simple yet useful cell-surface-anchored ratiometric fluorescent probe for extracellular pH sensing. The lipid-DNA probe, which consists of a hydrophobic diacyllipid tail and a hydrophilic DNA strand, is modified with two fluorescent dyes; one is pH-sensitive as pH indicator and the other is pH-insensitive as an internal reference. The lipid-DNA probe showed sensitive and reversible response to pH change in the range of 6.0-8.0, which is suitable for most extracellular studies. In addition, based on simple hydrophobic interactions with the cell membrane, the lipid-DNA probe can be easily anchored on the cell surface with negligible cytotoxicity, excellent stability, and unique ratiometric readout, thus ensuring its accurate sensing of extracellular pH. Finally, this lipid-DNA-based ratiometric pH indicator was successfully used for extracellular pH sensing of cells in 3D culture environment, demonstrating the potential applications of the sensor in biological and medical studies.

  9. Protein recognition by a pattern-generating fluorescent molecular probe

    Science.gov (United States)

    Pode, Zohar; Peri-Naor, Ronny; Georgeson, Joseph M.; Ilani, Tal; Kiss, Vladimir; Unger, Tamar; Markus, Barak; Barr, Haim M.; Motiei, Leila; Margulies, David

    2017-12-01

    Fluorescent molecular probes have become valuable tools in protein research; however, the current methods for using these probes are less suitable for analysing specific populations of proteins in their native environment. In this study, we address this gap by developing a unimolecular fluorescent probe that combines the properties of small-molecule-based probes and cross-reactive sensor arrays (the so-called chemical 'noses/tongues'). On the one hand, the probe can detect different proteins by generating unique identification (ID) patterns, akin to cross-reactive arrays. On the other hand, its unimolecular scaffold and selective binding enable this ID-generating probe to identify combinations of specific protein families within complex mixtures and to discriminate among isoforms in living cells, where macroscopic arrays cannot access. The ability to recycle the molecular device and use it to track several binding interactions simultaneously further demonstrates how this approach could expand the fluorescent toolbox currently used to detect and image proteins.

  10. [Development of a Fluorescence Probe for Live Cell Imaging].

    Science.gov (United States)

    Shibata, Aya

    2017-01-01

    Probes that detect specific biological materials are indispensable tools for deepening our understanding of various cellular phenomena. In live cell imaging, the probe must emit fluorescence only when a specific substance is detected. In this paper, we introduce a new probe we developed for live cell imaging. Glutathione S-transferase (GST) activity is higher in tumor cells than in normal cells and is involved in the development of resistance to various anticancer drugs. We previously reported the development of a general strategy for the synthesis of probes for detection of GST enzymes, including fluorogenic, bioluminogenic, and 19 F-NMR probes. Arylsulfonyl groups were used as caging groups during probe design. The fluorogenic probes were successfully used to quantitate very low levels of GST activity in cell extracts and were also successfully applied to the imaging of microsomal MGST1 activity in living cells. The bioluminogenic and 19 F-NMR probes were able to detect GST activity in Escherichia coli cells. Oligonucleotide-templated reactions are powerful tools for nucleic acid sensing. This strategy exploits the target strand as a template for two functionalized probes and provides a simple molecular mechanism for multiple turnover reactions. We developed a nucleophilic aromatic substitution reaction-triggered fluorescent probe. The probe completed its reaction within 30 s of initiation and amplified the fluorescence signal from 0.5 pM target oligonucleotide by 1500 fold under isothermal conditions. Additionally, we applied the oligonucleotide-templated reaction for molecular releasing and peptide detection.

  11. Polarization of fluorescence: a probe of molecular autoionization

    International Nuclear Information System (INIS)

    Leroi, G.E.; Dehmer, J.L.; Parr, A.C.; Poliakoff, E.D.

    1983-01-01

    The polarization of fluorescence from excited-state molecular photoions provides a direct probe of the photoionization dynamics and the symmetry signatures of autoionizing resonances. Measurements on CO 2 and CS 2 are presented as examples

  12. Using fluorescent lipids in live zebrafish larvae: From imaging whole animal physiology to subcellular lipid trafficking.

    Science.gov (United States)

    Anderson, J L; Carten, J D; Farber, S A

    2016-01-01

    Lipids serve essential functions in cells as signaling molecules, membrane components, and sources of energy. Defects in lipid metabolism are implicated in a number of pandemic human diseases, including diabetes, obesity, and hypercholesterolemia. Many aspects of how fatty acids and cholesterol are absorbed and processed by intestinal cells remain unclear and present a hurdle to developing approaches for disease prevention and treatment. Numerous studies have shown that the zebrafish is an excellent model for vertebrate lipid metabolism. In this chapter, we review commercially available fluorescent lipids that can be deployed in live zebrafish to better understand lipid signaling and metabolism. In this chapter, we present criteria one should consider when selecting specific fluorescent lipids for the study of digestive physiology or lipid metabolism in larval zebrafish. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Phenylethynylpyrene excimer forming hybridization probes for fluorescence SNP detection

    DEFF Research Database (Denmark)

    Prokhorenko, Igor A.; Astakhova, Irina V.; Momynaliev, Kuvat T.

    2009-01-01

    Excimer formation is a unique feature of some fluorescent dyes (e.g., pyrene) which can be used for probing the proximity of biomolecules. Pyrene excimer fluorescence has previously been used for homogeneous detection of single nucleotide polymorphism (SNP) on DNA. 1-Phenylethynylpyrene (1-1-PEPy...

  14. Probing Lipid Bilayers under Ionic Imbalance.

    Science.gov (United States)

    Lin, Jiaqi; Alexander-Katz, Alfredo

    2016-12-06

    Biological membranes are normally under a resting transmembrane potential (TMP), which originates from the ionic imbalance between extracellular fluids and cytosols, and serves as electric power storage for cells. In cell electroporation, the ionic imbalance builds up a high TMP, resulting in the poration of cell membranes. However, the relationship between ionic imbalance and TMP is not clearly understood, and little is known about the effect of ionic imbalance on the structure and dynamics of biological membranes. In this study, we used coarse-grained molecular dynamics to characterize a dipalmitoylphosphatidylcholine bilayer system under ionic imbalances ranging from 0 to ∼0.06 e charges per lipid (e/Lip). We found that the TMP displayed three distinct regimes: 1) a linear regime between 0 and 0.045 e/Lip, where the TMP increased linearly with ionic imbalance; 2) a yielding regime between ∼0.045 and 0.060 e/Lip, where the TMP displayed a plateau; and 3) a poration regime above ∼0.060 e/Lip, where we observed pore formation within the sampling time (80 ns). We found no structural changes in the linear regime, apart from a nonlinear increase in the area per lipid, whereas in the yielding regime the bilayer exhibited substantial thinning, leading to an excess of water and Na + within the bilayer, as well as significant misalignment of the lipid tails. In the poration regime, lipid molecules diffused slightly faster. We also found that the fluid-to-gel phase transition temperature of the bilayer dropped below the normal value with increased ionic imbalances. Our results show that a high ionic imbalance can substantially alter the essential properties of the bilayer, making the bilayer more fluid like, or conversely, depolarization of a cell could in principle lead to membrane stiffening. Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  15. NOVEL FLUORESCENT PROBES FOR THE DOPAMINE TRANSPORTER

    DEFF Research Database (Denmark)

    Cha, J; Vægter, Christian Bjerggaard; Adkins, Erica

    -reactive rhodamine red derivatives. The resulting N-substituted (JHC 1-64) and 2-substituted (JHC 1-53) ligands showed high affinity binding to DAT expressed in HEK 293 cells (Ki= 6.4 and 29 nM, respectively). Their ability to selectively label the DAT was demonstrated by confocal laser scanning microscopy of HEK......To enable visualization of the dopamine transporter (DAT) through fluorescence technologies we have synthesized a novel series of fluorescently tagged analogs of cocaine. Previous structure-activity relationship (SAR) studies have demonstrated that the dopamine transporter (DAT) can tolerate...... in untransfected control cells. The possibility of using these ligands for direct labeling of the DAT in living cells represents a new and important approach for understanding cellular targeting and trafficking of the DAT. Moreover, these fluorescent ligands might also provide the molecular tools...

  16. Analysis of Cholesterol Trafficking with Fluorescent Probes

    DEFF Research Database (Denmark)

    Maxfield, Frederick R.; Wustner, Daniel

    2012-01-01

    Cholesterol plays an important role in determining the biophysical properties of biological membranes, and its concentration is tightly controlled by homeostatic processes. The intracellular transport of cholesterol among organelles is a key part of the homeostatic mechanism, but sterol transport...... that can bind to cholesterol to reveal its distribution in cells. We also discuss the use of intrinsically fluorescent sterols that closely mimic cholesterol, as well as some minimally modified fluorophore-labeled sterols. Methods for imaging these sterols by conventional fluorescence microscopy...... and by multiphoton microscopy are described. Some label-free methods for imaging cholesterol itself are also discussed briefly....

  17. Fluorescence correction in electron probe microanalysis

    International Nuclear Information System (INIS)

    Castellano, Gustavo; Riveros, J.A.

    1987-01-01

    In this work, several expressions for characteristic fluorescence corrections are computed, for a compilation of experimental determinations on standard samples. Since this correction does not take significant values, the performance of the different models is nearly the same; this fact suggests the use of the simplest available expression. (Author) [es

  18. Fluorescence spectral studies on interaction of fluorescent probes with Bovine Serum Albumin (BSA)

    Energy Technology Data Exchange (ETDEWEB)

    Ghosh, Kaushik, E-mail: ghoshfcy@iitr.ac.in; Rathi, Sweety; Arora, Deepshikha

    2016-07-15

    Interaction of 2-(1-(naphthale-1-ylimino)ethyl)phenol (1), 2-methoxy-4-(((4-methoxyphenyl)imino)methyl)phenol (2) and 2-methoxy-4-((naphthalene-1-ylimino)methyl)phenol (3) with Bovine Serum Albumin (BSA) was examined. Fluorescence spectral data were obtained from the probes by varying the concentration of BSA as well as from BSA by varying the concentration of probes. Synchronous fluorescence measurements were performed and binding constants of the probes were calculated. To understand mode of quenching, Stern–Volmer plot, absorption spectral studies and life time measurements were performed. Förster resonance energy transfer (FRET) was also scrutinized. - Highlights: • Schiff bases with pendant phenolato function and interaction with BSA. • Synchronous fluorescence studies and a preferred interaction with tryptophan. • Probable interaction of probes with Trp-213 residue in the hydrophobic cavity. • 1:1 binding stoichiometry of probes and BSA in Benesi–Hildebrand graph.

  19. Probing protein-lipid interactions by FRET between membrane fluorophores

    Science.gov (United States)

    Trusova, Valeriya M.; Gorbenko, Galyna P.; Deligeorgiev, Todor; Gadjev, Nikolai

    2016-09-01

    Förster resonance energy transfer (FRET) is a powerful fluorescence technique that has found numerous applications in medicine and biology. One area where FRET proved to be especially informative involves the intermolecular interactions in biological membranes. The present study was focused on developing and verifying a Monte-Carlo approach to analyzing the results of FRET between the membrane-bound fluorophores. This approach was employed to quantify FRET from benzanthrone dye ABM to squaraine dye SQ-1 in the model protein-lipid system containing a polycationic globular protein lysozyme and negatively charged lipid vesicles composed of phosphatidylcholine and phosphatidylglycerol. It was found that acceptor redistribution between the lipid bilayer and protein binding sites resulted in the decrease of FRET efficiency. Quantification of this effect in terms of the proposed methodology yielded both structural and binding parameters of lysozyme-lipid complexes.

  20. Photoswitchable non-fluorescent thermochromic dye-nanoparticle hybrid probes

    OpenAIRE

    Harrington, Walter N.; Haji, Mwafaq R.; Galanzha, Ekaterina I.; Nedosekin, Dmitry A.; Nima, Zeid A.; Watanabe, Fumiya; Ghosh, Anindya; Biris, Alexandru S.; Zharov, Vladimir P.

    2016-01-01

    Photoswitchable fluorescent proteins with controllable light?dark states and spectral shifts in emission in response to light have led to breakthroughs in the study of cell biology. Nevertheless, conventional photoswitching is not applicable for weakly fluorescent proteins and requires UV light with low depth penetration in bio-tissue. Here we introduce a novel concept of photoswitchable hybrid probes consisting of thermochromic dye and absorbing nanoparticles, in which temperature-sensitive ...

  1. Fluorescent and Colorimetric Molecular Recognition Probe for Hydrogen Bond Acceptors

    OpenAIRE

    Pike, Sarah Jane; Hunter, Christopher Alexander

    2018-01-01

    The association constants for formation of 1 : 1 complexes between a H-bond donor, 1-naphthol, and a diverse range of charged and neutral H-bond acceptors have been measured using UV/vis absorption and fluorescence emission titrations. The performance of 1-naphthol as a dual colorimetric and fluorescent molecular recognition probe for determining the H-bond acceptor (HBA) parameters of charged and neutral solutes has been investigated in three solvents. The data were employed to establish sel...

  2. In Situ Live-Cell Nucleus Fluorescence Labeling with Bioinspired Fluorescent Probes.

    Science.gov (United States)

    Ding, Pan; Wang, Houyu; Song, Bin; Ji, Xiaoyuan; Su, Yuanyuan; He, Yao

    2017-08-01

    Fluorescent imaging techniques for visualization of nuclear structure and function in live cells are fundamentally important for exploring major cellular events. The ideal cellular labeling method is capable of realizing label-free, in situ, real-time, and long-term nucleus labeling in live cells, which can fully obtain the nucleus-relative information and effectively alleviate negative effects of alien probes on cellular metabolism. However, current established fluorescent probes-based strategies (e.g., fluorescent proteins-, organic dyes-, fluorescent organic/inorganic nanoparticles-based imaging techniques) are unable to simultaneously realize label-free, in situ, long-term, and real-time nucleus labeling, resulting in inevitable difficulties in fully visualizing nuclear structure and function in live cells. To this end, we present a type of bioinspired fluorescent probes, which are highly efficacious for in situ and label-free tracking of nucleus in long-term and real-time manners. Typically, the bioinspired polydopamine (PDA) nanoparticles, served as fluorescent probes, can be readily synthesized in situ within live cell nucleus without any further modifications under physiological conditions (37 °C, pH ∼7.4). Compared with other conventional nuclear dyes (e.g., propidium iodide (PI), Hoechst), superior spectroscopic properties (e.g., quantum yield of ∼35.8% and high photostability) and low cytotoxicity of PDA-based probes enable long-term (e.g., 3 h) fluorescence tracking of nucleus. We also demonstrate the generality of this type of bioinspired fluorescent probes in different cell lines and complex biological samples.

  3. Newly synthesized benzanthrone derivatives as prospective fluorescent membrane probes

    International Nuclear Information System (INIS)

    Zhytniakivska, Olga; Trusova, Valeriya; Gorbenko, Galyna; Kirilova, Elena; Kalnina, Inta; Kirilov, Georgiy; Kinnunen, Paavo

    2014-01-01

    Fluorescence spectral properties of a series of novel benzanthrone derivatives have been explored in lipid bilayers composed of zwitterionic lipid phosphatidylcholine (PC) and its mixtures with cholesterol (Chol) and anionic phospholipid cardiolipin (CL). Analysis of partition coefficients showed that all the examined compounds possess rather high lipid-associating ability, with the amidino derivatives exhibiting stronger membrane partitioning compared with the aminobenzanthrones. To understand how benzanthrone partition properties correlate with their structure, quantitative structure property relationship (QSPR) analysis was performed involving a range of quantum chemical molecular descriptors. -- Highlights: • Benzanthrone partitioning into lipid bilayer correlates with lipophilicity of the dyes. • Partition properties of benzanthrones depend on the dye dipole moment. • Amidino derivatives exhibit higher membrane affinity than aminobenzanthrones

  4. Lipid diffusion in planar membranes investigated by fluorescence correlation spectroscopy

    Czech Academy of Sciences Publication Activity Database

    Macháň, Radek; Hof, Martin

    2010-01-01

    Roč. 1798, č. 7 (2010), s. 1377-1391 ISSN 0005-2736 R&D Projects: GA ČR GA203/08/0114; GA AV ČR GEMEM/09/E006 Institutional research plan: CEZ:AV0Z40400503 Keywords : supported lipid bilayer * giant unilamellar vesicle * fluorescence recovery after photobleaching Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 4.647, year: 2010

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

    Science.gov (United States)

    Mayer-Cumblidge, M. Uljana; Cao, Haishi

    2013-01-15

    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.

  6. New fluorescent probes for detection and characterization of amyloid fibrils

    Science.gov (United States)

    Gorbenko, Galyna; Trusova, Valeriya; Kirilova, Elena; Kirilov, Georgiy; Kalnina, Inta; Vasilev, Aleksey; Kaloyanova, Stefka; Deligeorgiev, Todor

    2010-08-01

    The applicability of the novel fluorescent probes, aminoderivative of benzanthrone ABM, squaraine dye SQ-1 and polymethine dye V2 to identification and structural analysis of amyloid fibrils has been evaluated using the lysozyme model system in which fibrillar aggregates have been formed in concentrated ethanol solution. The association constant, binding stoichiometry and molar fluorescence of the bound dye have been determined. ABM was found to surpass classical amyloid marker ThT in the sensitivity to the presence of fibrillar aggregates. Resonance energy transfer measurements involving ABM-SQ-1 and SQ-1-V2 donor-acceptor pairs yielded the limits for fractal-like dimension of lysozyme fibrils.

  7. Fluorescent Probes for Analysis and Imaging of Monoamine Oxidase Activity

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Dokyoung; Jun, Yong Woong; Ahn, Kyo Han [POSTECH, Pohang (Korea, Republic of)

    2014-05-15

    Monoamine oxidases catalyze the oxidative deamination of dietary amines and amine neurotransmitters, and assist in maintaining the homeostasis of the amine neurotransmitters in the brain. Dysfunctions of these enzymes can cause neurological and behavioral disorders including Parkinson's and Alzheimer's diseases. To understand their physiological roles, efficient assay methods for monoamine oxidases are essential. Reviewed in this Perspective are the recent progress in the development of fluorescent probes for monoamine oxidases and their applications to enzyme assays in cells and tissues. It is evident that still there is strong need for a fluorescent probe with desirable substrate selectivity and photophysical properties to challenge the much unsolved issues associated with the enzymes and the diseases.

  8. Monitoring membrane hydration with 2-(dimethylamino)-6-acylnaphtalenes fluorescent probes

    DEFF Research Database (Denmark)

    Bagatolli, Luis

    2015-01-01

    of LAURDAN and PRODAN are exquisitely sensitive to cholesterol effects, allowing interpretations that correlate changes in membrane packing with membrane hydration. Different membrane model systems as well as innate biological membranes have been studied with this family of probes allowing interesting...... comparative studies. This chapter presents a short historical overview about these fluorescent reporters, discusses on different models proposed to explain their sensitivity to membrane hydration, and includes relevant examples from experiments performed in artificial and biological membranes....

  9. Fluorescent nanodiamonds as non-photobleachable responsive probes

    Czech Academy of Sciences Publication Activity Database

    Cígler, Petr

    2016-01-01

    Roč. 14, č. 5 (2016), s. 232 ISSN 2336-7202. [Sjezd českých a slovenských chemických společností /68./. 04.09.2016-07.09.2016, Praha] R&D Projects: GA ČR(CZ) GA16-16336S Institutional support: RVO:61388963 Keywords : fluorescent nanodiamonds * FNDs * probes Subject RIV: CG - Electrochemistry

  10. Local mobility in lipid domains of supported bilayers characterized by atomic force microscopy and fluorescence correlation spectroscopy.

    Energy Technology Data Exchange (ETDEWEB)

    Frankel, Daniel J.; Buranda, T. (University of New Mexico, Albuquerque, NM); Burns, Alan Richard

    2005-01-01

    Fluorescence correlation spectroscopy (FCS) is used to examine mobility of labeled probes at specific sites in supported bilayers consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid domains in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). Those sites are mapped beforehand with simultaneous atomic force microscopy and submicron confocal fluorescence imaging, allowing characterization of probe partitioning between gel DPPC and disordered liquid DOPC domains with corresponding topography of domain structure. We thus examine the relative partitioning and mobility in gel and disordered liquid phases for headgroup- and tailgroup-labeled GM1 ganglioside probes and for headgroup- and tailgroup-labeled phospholipid probes. For the GM1 probes, large differences in mobility between fluid and gel domains are observed; whereas unexpected mobility is observed in submicron gel domains for the phospholipid probes. We attribute the latter to domain heterogeneities that could be induced by the probe. Furthermore, fits to the FCS data for the phospholipid probes in the DOPC fluid phase require two components (fast and slow). Although proximity to the glass substrate may be a factor, local distortion of the probe by the fluorophore could also be important. Overall, we observe nonideal aspects of phospholipid probe mobility and partitioning that may not be restricted to supported bilayers.

  11. A Dansyl Fluorescence-Based Assay for Monitoring Kinetics of Lipid Extraction and Transfer

    Science.gov (United States)

    Ran, Yong

    2008-01-01

    Lipid transfer proteins (LTPs) play important roles in cellular biology, and fluorescence spectroscopy has found wide range use as a facile means for time-resolved monitoring of protein-lipid interactions[1]. Here, we show how the fluorescence emission properties of dansyl-DHPE can be exploited to characterize lipid extraction and lipid transfer kinetics. The GM2 activator protein serves as an example LTP where the ability to independently characterize lipid extraction from donor vesicles, formation of a protein:lipid complex in solution, and release of lipid from the complex to acceptor liposomes is crucial for full kinetic characterization of lipid transfer. PMID:18694718

  12. Probing Zeolite Crystal Architecture and Structural Imperfections using Differently Sized Fluorescent Organic Probe Molecules.

    Science.gov (United States)

    Hendriks, Frank C; Schmidt, Joel E; Rombouts, Jeroen A; Lammertsma, Koop; Bruijnincx, Pieter C A; Weckhuysen, Bert M

    2017-05-05

    A micro-spectroscopic method has been developed to probe the accessibility of zeolite crystals using a series of fluorescent 4-(4-diethylaminostyryl)-1-methylpyridinium iodide (DAMPI) probes of increasing molecular size. Staining large zeolite crystals with MFI (ZSM-5) topology and subsequent mapping of the resulting fluorescence using confocal fluorescence microscopy reveal differences in structural integrity: the 90° intergrowth sections of MFI crystals are prone to develop structural imperfections, which act as entrance routes for the probes into the zeolite crystal. Polarization-dependent measurements provide evidence for the probe molecule's alignment within the MFI zeolite pore system. The developed method was extended to BEA (Beta) crystals, showing that the previously observed hourglass pattern is a general feature of BEA crystals with this morphology. Furthermore, the probes can accurately identify at which crystal faces of BEA straight or sinusoidal pores open to the surface. The results show this method can spatially resolve the architecture-dependent internal pore structure of microporous materials, which is difficult to assess using other characterization techniques such as X-ray diffraction. © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

  13. Hemoglobin detection using carbon dots as a fluorescence probe.

    Science.gov (United States)

    Barati, Ali; Shamsipur, Mojtaba; Abdollahi, Hamid

    2015-09-15

    Herein, we have described the application of high fluorescent carbon dots (CDs) without any surface modification as a simple and fast responding fluorescence probe for sensitive and selective determination of hemoglobin (Hb) in the presence of H2O2. Although Hb itself was able to quench the fluorescence of CDs, based on the inner filter effect (IFE) of the protein that affects both excitation and emission spectra of CDs, the presence of H2O2 resulted in further improvement of the sensitivity of Hb detection. The assay is based on the reaction of Hb with H2O2 that generates reactive oxygen species including hydroxyl (OH•) and superoxide (O2(•-)) radicals under heme degradation and/or iron release from Hb and the subsequent reaction of hydroxyl radicals, as strong oxidizing agents, with CDs resulting in high fluorescence quenching. The proposed probe was used for determination of Hb in concentration range of 1-100 nM with a detection limit of 0.4 nM. The method was successfully applied to the determination of Hb in human blood samples. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. FLOW CYTOMETRIC APPLICABILITY OF FLUORESCENT VITALITY PROBES ON PHYTOPLANKTON1.

    Science.gov (United States)

    Peperzak, Louis; Brussaard, Corina P D

    2011-06-01

    The applicability of six fluorescent probes (four esterase probes: acetoxymethyl ester of Calcein [Calcein-AM], 5-chloromethylfluorescein diacetate [CMFDA], fluorescein diacetate [FDA], and 2',7'-dichlorofluorescein diacetate [H 2 DCFDA]; and two membrane probes: bis-(1,3-dibutylbarbituric acid) trimethine oxonol [DiBAC 4 (3)] and SYTOX-Green) as vitality stains was tested on live and killed cells of 40 phytoplankton strains in exponential and stationary growth phases, belonging to 12 classes and consisting of four cold-water, 26 temperate, and four warm-water species. The combined live/dead ratios of all six probes indicated significant differences between the 12 plankton classes (P live/dead ratios of FDA and CMFDA were not significantly different from each other, and both performed better than Calcein-AM and H 2 DCFDA (P live/dead ratios) among all six probes belonged to nine genera from six classes of phytoplankton. In conclusion, FDA, CMFDA, DIBAC 4 (3), and SYTOX-Green represent a wide choice of vitality probes in the study of phytoplankton ecology, applicable in many species from different algal classes, originating from different regions and at different stages of growth. © 2011 Phycological Society of America.

  15. Design and Evaluation of Novel Polymyxin Fluorescent Probes

    Directory of Open Access Journals (Sweden)

    Bo Yun

    2017-11-01

    Full Text Available Polymyxins (polymyxin B and colistin are cyclic lipopeptide antibiotics that serve as a last-line defence against Gram-negative “superbugs”. In the present study, two novel fluorescent polymyxin probes were designed through regio-selective modifications of the polymyxin B core structure at the N-terminus and the hydrophobic motif at positions 6 and 7. The resulting probes, FADDI-285 and FADDI-286 demonstrated comparable antibacterial activity (MICs 2–8 mg/L to polymyxin B and colistin (MICs 0.5–8 mg/L against a panel of gram-negative clinical isolates of Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. These probes should prove to be of considerable utility for imaging cellular uptake and mechanistic investigations of these important last-line antibiotics.

  16. Sensitive determination of nucleic acids using organic nanoparticle fluorescence probes

    Science.gov (United States)

    Zhou, Yunyou; Bian, Guirong; Wang, Leyu; Dong, Ling; Wang, Lun; Kan, Jian

    2005-06-01

    This paper describes the preparation of organic nanoparticles by reprecipitation method under sonication and vigorous stirring. Transmission electron microscopy (TEM) was used to characterize the size and size distribution of the luminescent nanoparticles. Their average diameter was about 25 nm with a size variation of ±18%. The fluorescence decay lifetime of the nanoparticles also was determined on a self-equipped fluorospectrometer with laser light source. The lifetime (˜0.09 μs) of nanoparticles is about three times long as that of the monomer. The nanoparticles were in abundant of hydrophilic groups, which increased their miscibility in aqueous solution. These organic nanoparticles have high photochemical stability, excellent resistance to chemical degradation and photodegradation, and a good fluorescence quantum yield (25%). The fluorescence can be efficiently quenched by nucleic acids. Based on the fluorescence quenching of nanoparticles, a fluorescence quenching method was developed for determination of microamounts of nucleic acids by using the nanoparticles as a new fluorescent probe. Under optimal conditions, maximum fluorescence quenching is produced, with maximum excitation and emission wavelengths of 345 and 402 nm, respectively. Under optimal conditions, the calibration graphs are linear over the range 0.4-19.0 μg ml -1 for calf thymus DNA (ct-DNA) and 0.3-19.0 μg ml -1 for fish sperm DNA (fs-DNA). The corresponding detection limits are 0.25 μg ml -1 for ct-DNA and 0.17 μg ml -1 for fs-DNA. The relative standard deviation of six replicate measurements is 1.3-2.1%. The method is simple, rapid and sensitive with wide linear range. The recovery and relative standard deviation are very satisfactory.

  17. Retarded Local Dynamics of Single Fluorescent Probes in Polymeric Glass due to Interaction Strengthening

    Science.gov (United States)

    Zhang, Hao; Yang, Jingfa; Zhao, Jiang

    The effect of strengthening of interaction between single fluorescent probes and polymer matrix to the probes dynamics is investigated using single molecule fluorescence defocus microscopy. By introducing multiple hydroxyl groups to the fluorescent probes, which builds up hydrogen bonds between the probe and polymer matrix, the dynamics is discovered to be retarded. This is evidenced by the lowering of the frequency of the vibrational modes in the power spectra of the rotation trajectories of individual fluorescent probes, and also by the lowering of population of rotating probes. The results show that by strengthening the probe-matrix interaction, the local dynamics detected by the probes is equivalent to that detected by a bigger probe, due to the enhanced friction between the probe and the polymer matrix. the National Basic Research Program of China (2012CB821500).

  18. Characterization of the Lateral Distribution of Fluorescent Lipid in Binary-Constituent Lipid Monolayers by Principal Component Analysis

    Directory of Open Access Journals (Sweden)

    István P. Sugár

    2010-01-01

    Full Text Available Lipid lateral organization in binary-constituent monolayers consisting of fluorescent and nonfluorescent lipids has been investigated by acquiring multiple emission spectra during measurement of each force-area isotherm. The emission spectra reflect BODIPY-labeled lipid surface concentration and lateral mixing with different nonfluorescent lipid species. Using principal component analysis (PCA each spectrum could be approximated as the linear combination of only two principal vectors. One point on a plane could be associated with each spectrum, where the coordinates of the point are the coefficients of the linear combination. Points belonging to the same lipid constituents and experimental conditions form a curve on the plane, where each point belongs to a different mole fraction. The location and shape of the curve reflects the lateral organization of the fluorescent lipid mixed with a specific nonfluorescent lipid. The method provides massive data compression that preserves and emphasizes key information pertaining to lipid distribution in different lipid monolayer phases. Collectively, the capacity of PCA for handling large spectral data sets, the nanoscale resolution afforded by the fluorescence signal, and the inherent versatility of monolayers for characterization of lipid lateral interactions enable significantly enhanced resolution of lipid lateral organizational changes induced by different lipid compositions.

  19. ESIPT-Based Photoactivatable Fluorescent Probe for Ratiometric Spatiotemporal Bioimaging

    Directory of Open Access Journals (Sweden)

    Xiaohong Zhou

    2016-10-01

    Full Text Available Photoactivatable fluorophores have become an important technique for the high spatiotemporal resolution of biological imaging. Here, we developed a novel photoactivatable probe (PHBT, which is based on 2-(2-hydroxyphenylbenzothiazole (HBT, a small organic fluorophore known for its classic luminescence mechanism through excited-state intramolecular proton transfer (ESIPT with the keto form and the enol form. After photocleavage, PHBT released a ratiometric fluorophore HBT, which showed dual emission bands with more than 73-fold fluorescence enhancement at 512 nm in buffer and more than 69-fold enhancement at 452 nm in bovine serum. The probe displayed a high ratiometric imaging resolution and is believed to have a wide application in biological imaging.

  20. A Dansyl-Rhodamine Based Fluorescent Probe for Detection of Hg2+ and Cu2.

    Science.gov (United States)

    Yuan, Shizhuang; Su, Wei; Wang, Enju

    2017-09-01

    A novel fluorescent probe based on dansyl-appended rhodamine B was developed. The probe can selectively recognize and sense Hg2+ and Cu2+ from other common metal ions by showing unique fluorescence and absorption characteristics. In MeCN/HEPES buffer solution, the probe gives a ratiometric fluorescent response to Hg2+, which was ascribed to the fluorescence resonance energy transfer from dansyl moiety to the ring-opened rhodamine B moiety, while the presence of Cu2+ causes fluorescence quenching. Beside the fluorescence change, the presence of Cu2+ and Hg2+ can induce intensive absorption at about 555 nm, which resulted in a color change from colorless to pink.

  1. Fluorescent and colorimetric molecular recognition probe for hydrogen bond acceptors.

    Science.gov (United States)

    Pike, Sarah J; Hunter, Christopher A

    2017-11-22

    The association constants for formation of 1 : 1 complexes between a H-bond donor, 1-naphthol, and a diverse range of charged and neutral H-bond acceptors have been measured using UV/vis absorption and fluorescence emission titrations. The performance of 1-naphthol as a dual colorimetric and fluorescent molecular recognition probe for determining the H-bond acceptor (HBA) parameters of charged and neutral solutes has been investigated in three solvents. The data were employed to establish self-consistent H-bond acceptor parameters (β) for benzoate, azide, chloride, thiocyanate anions, a series of phosphine oxides, phosphate ester, sulfoxide and a tertiary amide. The results demonstrate both the transferability of H-bond parameters between different solvents and the utility of the naphthol-based dual molecular recognition probe to exploit orthogonal spectroscopic techniques to determine the HBA properties of neutral and charged solutes. The benzoate anion is the strongest HBA studied with a β parameter of 15.4, and the neutral tertiary amide is the weakest H-bond acceptor investigated with a β parameter of 8.5. The H-bond acceptor strength of the azide anion is higher than that of chloride (12.8 and 12.2 respectively), and the thiocyanate anion has a β value of 10.8 and thus is a significantly weaker H-bond acceptor than both the azide and chloride anions.

  2. Formulation of novel lipid-coated magnetic nanoparticles as the probe for in vivo imaging

    Directory of Open Access Journals (Sweden)

    Mou Chung-Yuan

    2009-09-01

    Full Text Available Abstract Background Application of superparamagnetic iron oxide nanoparticles (SPIOs as the contrast agent has improved the quality of magnetic resonance (MR imaging. Low efficiency of loading the commercially available iron oxide nanoparticles into cells and the cytotoxicity of previously formulated complexes limit their usage as the image probe. Here, we formulated new cationic lipid nanoparticles containing SPIOs feasible for in vivo imaging. Methods Hydrophobic SPIOs were incorporated into cationic lipid 1,2-dioleoyl-3-(trimethylammonium propane (DOTAP and polyethylene-glycol-2000-1,2-distearyl-3-sn-phosphatidylethanolamine (PEG-DSPE based micelles by self-assembly procedure to form lipid-coated SPIOs (L-SPIOs. Trace amount of Rhodamine-dioleoyl-phosphatidylethanolamine (Rhodamine-DOPE was added as a fluorescent indicator. Particle size and zeta potential of L-SPIOs were determined by Dynamic Light Scattering (DLS and Laser Doppler Velocimetry (LDV, respectively. HeLa, PC-3 and Neuro-2a cells were tested for loading efficiency and cytotoxicity of L-SPIOs using fluorescent microscopy, Prussian blue staining and flow cytometry. L-SPIO-loaded CT-26 cells were tested for in vivo MR imaging. Results The novel formulation generates L-SPIOs particle with the average size of 46 nm. We showed efficient cellular uptake of these L-SPIOs with cationic surface charge into HeLa, PC-3 and Neuro-2a cells. The L-SPIO-loaded cells exhibited similar growth potential as compared to unloaded cells, and could be sorted by a magnet stand over ten-day duration. Furthermore, when SPIO-loaded CT-26 tumor cells were injected into Balb/c mice, the growth status of these tumor cells could be monitored using optical and MR images. Conclusion We have developed a novel cationic lipid-based nanoparticle of SPIOs with high loading efficiency, low cytotoxicity and long-term imaging signals. The results suggested these newly formulated non-toxic lipid-coated magnetic

  3. Complementary studies of lipid membrane dynamics using iSCAT and super-resolved fluorescence correlation spectroscopy

    Science.gov (United States)

    Reina, Francesco; Galiani, Silvia; Shrestha, Dilip; Sezgin, Erdinc; de Wit, Gabrielle; Cole, Daniel; Lagerholm, B. Christoffer; Kukura, Philipp; Eggeling, Christian

    2018-06-01

    Observation techniques with high spatial and temporal resolution, such as single-particle tracking based on interferometric scattering (iSCAT) microscopy, and fluorescence correlation spectroscopy applied on a super-resolution STED microscope (STED-FCS), have revealed new insights of the molecular organization of membranes. While delivering complementary information, there are still distinct differences between these techniques, most prominently the use of fluorescent dye tagged probes for STED-FCS and a need for larger scattering gold nanoparticle tags for iSCAT. In this work, we have used lipid analogues tagged with a hybrid fluorescent tag–gold nanoparticle construct, to directly compare the results from STED-FCS and iSCAT measurements of phospholipid diffusion on a homogeneous supported lipid bilayer (SLB). These comparative measurements showed that while the mode of diffusion remained free, at least at the spatial (>40 nm) and temporal (50  ⩽  t  ⩽  100 ms) scales probed, the diffussion coefficient was reduced by 20- to 60-fold when tagging with 20 and 40 nm large gold particles as compared to when using dye tagged lipid analogues. These FCS measurements of hybrid fluorescent tag–gold nanoparticle labeled lipids also revealed that commercially supplied streptavidin-coated gold nanoparticles contain large quantities of free streptavidin. Finally, the values of apparent diffusion coefficients obtained by STED-FCS and iSCAT differed by a factor of 2–3 across the techniques, while relative differences in mobility between different species of lipid analogues considered were identical in both approaches. In conclusion, our experiments reveal that large and potentially cross-linking scattering tags introduce a significant slow-down in diffusion on SLBs but no additional bias, and our labeling approach creates a new way of exploiting complementary information from STED-FCS and iSCAT measurements.

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

    DEFF Research Database (Denmark)

    Okholm, Anders; Kjems, Jørgen; Astakhova, Kira

    2014-01-01

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

  5. Highly stable lipid-encapsulation of fluorescent nanodiamonds for bioimaging applications.

    Science.gov (United States)

    Sotoma, Shingo; Hsieh, Feng-Jen; Chen, Yen-Wei; Tsai, Pei-Chang; Chang, Huan-Cheng

    2018-01-23

    Highly stable lipid-encapsulated fluorescent nanodiamonds (FNDs) are produced by photo-crosslinking of diacetylene-containing lipids physically attached to the FND surface. Not only is this coating method simple and fast, but also it gives the FND-lipid hybrids favorable properties for bioapplications. The hybrids are useful as fluorescent biolabels as well as fiducial markers for correlative light and electron microscopy.

  6. Rotational and translational diffusions of fluorescent probes during gelation process

    Science.gov (United States)

    Hattori, Yusuke; Panizza, Pascal; Letamendia, Louis; Ushiki, Hideharu

    2006-04-01

    Gelation process has been investigated by using light scattering techniques in recent years. We measured both of rotational and translational motions of fluorescent probes during gelation process. The measurements were performed after the temperature quenched at 30 °C. As the results, rotational diffusion coefficient of fluorescein was decreased after 6.0 × 10 4 s and energy transfer rate was reduced after 2.0 × 10 4 s. We sorted the gelation process into the following three parts, (I) pre-gelation, (II) reduction of translational diffusion (aging), and (III) reduction of rotational diffusion with saturating translational diffusion (post-gelation). The time scale of the process was completely different from the results of other methods.

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

    Science.gov (United States)

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

    2016-03-01

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

  8. The lipid dependence of melittin action investigated by dual-color fluorescence burst analysis

    NARCIS (Netherlands)

    Bogaart, Geert van den; Mika, Jacek T.; Krasnikov, Viktor; Poolman, Bert

    Dual-color fluorescence-burst analysis was used to study melittin-induced leakage of macromolecules from liposomes of various lipid compositions. To perform dual-color fluorescence-burst analysis, fluorescently labeled size-marker molecules were encapsulated into liposomes, labeled with a second

  9. Accumulation of polyunsaturated aldehydes in the gonads of the copepod Acartia tonsa revealed by tailored fluorescent probes.

    Science.gov (United States)

    Wolfram, Stefanie; Nejstgaard, Jens C; Pohnert, Georg

    2014-01-01

    Polyunsaturated aldehydes (PUAs) are released by several diatom species during predation. Besides other attributed activities, these oxylipins can interfere with the reproduction of copepods, important predators of diatoms. While intensive research has been carried out to document the effects of PUAs on copepod reproduction, little is known about the underlying mechanistic aspects of PUA action. Especially PUA uptake and accumulation in copepods has not been addressed to date. To investigate how PUAs are taken up and interfere with the reproduction in copepods we developed a fluorescent probe containing the α,β,γ,δ-unsaturated aldehyde structure element that is essential for the activity of PUAs as well as a set of control probes. We developed incubation and monitoring procedures for adult females of the calanoid copepod Acartia tonsa and show that the PUA derived fluorescent molecular probe selectively accumulates in the gonads of this copepod. In contrast, a saturated aldehyde derived probe of an inactive parent molecule was enriched in the lipid sac. This leads to a model for PUAs' teratogenic mode of action involving accumulation and covalent interaction with nucleophilic moieties in the copepod reproductive tissue. The teratogenic effect of PUAs can therefore be explained by a selective targeting of the molecules into the reproductive tissue of the herbivores, while more lipophilic but otherwise strongly related structures end up in lipid bodies.

  10. Accumulation of polyunsaturated aldehydes in the gonads of the copepod Acartia tonsa revealed by tailored fluorescent probes.

    Directory of Open Access Journals (Sweden)

    Stefanie Wolfram

    Full Text Available Polyunsaturated aldehydes (PUAs are released by several diatom species during predation. Besides other attributed activities, these oxylipins can interfere with the reproduction of copepods, important predators of diatoms. While intensive research has been carried out to document the effects of PUAs on copepod reproduction, little is known about the underlying mechanistic aspects of PUA action. Especially PUA uptake and accumulation in copepods has not been addressed to date. To investigate how PUAs are taken up and interfere with the reproduction in copepods we developed a fluorescent probe containing the α,β,γ,δ-unsaturated aldehyde structure element that is essential for the activity of PUAs as well as a set of control probes. We developed incubation and monitoring procedures for adult females of the calanoid copepod Acartia tonsa and show that the PUA derived fluorescent molecular probe selectively accumulates in the gonads of this copepod. In contrast, a saturated aldehyde derived probe of an inactive parent molecule was enriched in the lipid sac. This leads to a model for PUAs' teratogenic mode of action involving accumulation and covalent interaction with nucleophilic moieties in the copepod reproductive tissue. The teratogenic effect of PUAs can therefore be explained by a selective targeting of the molecules into the reproductive tissue of the herbivores, while more lipophilic but otherwise strongly related structures end up in lipid bodies.

  11. Hoechst tagging: a modular strategy to design synthetic fluorescent probes for live-cell nucleus imaging.

    Science.gov (United States)

    Nakamura, Akinobu; Takigawa, Kazumasa; Kurishita, Yasutaka; Kuwata, Keiko; Ishida, Manabu; Shimoda, Yasushi; Hamachi, Itaru; Tsukiji, Shinya

    2014-06-11

    We report a general strategy to create small-molecule fluorescent probes for the nucleus in living cells. Our strategy is based on the attachment of the DNA-binding Hoechst compound to a fluorophore of interest. Using this approach, simple fluorescein, BODIPY, and rhodamine dyes were readily converted to novel turn-on fluorescent nucleus-imaging probes.

  12. Instant live-cell super-resolution imaging of cellular structures by nanoinjection of fluorescent probes.

    Science.gov (United States)

    Hennig, Simon; van de Linde, Sebastian; Lummer, Martina; Simonis, Matthias; Huser, Thomas; Sauer, Markus

    2015-02-11

    Labeling internal structures within living cells with standard fluorescent probes is a challenging problem. Here, we introduce a novel intracellular staining method that enables us to carefully control the labeling process and provides instant access to the inner structures of living cells. Using a hollow glass capillary with a diameter of <100 nm, we deliver functionalized fluorescent probes directly into the cells by (di)electrophoretic forces. The label density can be adjusted and traced directly during the staining process by fluorescence microscopy. We demonstrate the potential of this technique by delivering and imaging a range of commercially available cell-permeable and nonpermeable fluorescent probes to cells.

  13. Fluorescent probes and nanoparticles for intracellular sensing of pH values

    Science.gov (United States)

    Shi, Wen; Li, Xiaohua; Ma, Huimin

    2014-12-01

    Intracellular pH regulates a number of cell metabolism processes and its sensing is thus of great importance for cell studies. Among various methods, fluorescent probes have been widely used for sensing intracellular pH values because of their high sensitivity and spatiotemporal resolution capability. In this article, the development of fluorescent probes with good practicability in sensing intracellular pH values and pH variation during 2009 - 2014 is reviewed. These fluorescence probes are divided into two kinds: small molecules and nanoparticles. Photophysical properties, advantages/disadvantages and applications of the two kinds of probes are discussed in detail.

  14. Fluorescent probes and nanoparticles for intracellular sensing of pH values

    International Nuclear Information System (INIS)

    Shi, Wen; Li, Xiaohua; Ma, Huimin

    2014-01-01

    Intracellular pH regulates a number of cell metabolism processes and its sensing is thus of great importance for cell studies. Among various methods, fluorescent probes have been widely used for sensing intracellular pH values because of their high sensitivity and spatiotemporal resolution capability. In this article, the development of fluorescent probes with good practicability in sensing intracellular pH values and pH variation during 2009 − 2014 is reviewed. These fluorescence probes are divided into two kinds: small molecules and nanoparticles. Photophysical properties, advantages/disadvantages and applications of the two kinds of probes are discussed in detail. (topical review)

  15. Measuring reactive oxygen and nitrogen species with fluorescent probes: challenges and limitations

    Science.gov (United States)

    Kalyanaraman, Balaraman; Darley-Usmar, Victor; Davies, Kelvin J.A.; Dennery, Phyllis A.; Forman, Henry Jay; Grisham, Matthew B.; Mann, Giovanni E.; Moore, Kevin; Roberts, L. Jackson; Ischiropoulos, Harry

    2013-01-01

    The purpose of this position paper is to present a critical analysis of the challenges and limitations of the most widely used fluorescent probes for detecting and measuring reactive oxygen and nitrogen species. Where feasible, we have made recommendations for the use of alternate probes and appropriate analytical techniques that measure the specific products formed from the reactions between fluorescent probes and reactive oxygen and nitrogen species. We have proposed guidelines that will help present and future researchers with regard to the optimal use of selected fluorescent probes and interpretation of results. PMID:22027063

  16. Detection of Pathogenic Biofilms with Bacterial Amyloid Targeting Fluorescent Probe, CDy11

    DEFF Research Database (Denmark)

    Kim, Jun Young; Sahu, Srikanta; Yau, Yin Hoe

    2016-01-01

    Bacterial biofilms are responsible for a wide range of persistent infections. In the clinic, diagnosis of biofilm-associated infections relies heavily on culturing methods, which fail to detect nonculturable bacteria. Identification of novel fluorescent probes for biofilm imaging will greatly...... facilitate diagnosis of pathogenic bacterial infection. Herein, we report a novel fluorescent probe, CDy11 (compound of designation yellow 11), which targets amyloid in the Pseudomonas aeruginosa biofilm matrix through a diversity oriented fluorescent library approach (DOFLA). CDy11 was further demonstrated...

  17. Highly selective and rapidly responsive fluorescent probe for hydrogen sulfide detection in wine.

    Science.gov (United States)

    Wang, Hao; Wang, Jialin; Yang, Shaoxiang; Tian, Hongyu; Liu, Yongguo; Sun, Baoguo

    2018-08-15

    A new fluorescent probe 6-(2, 4-dinitrophenoxy)-2-naphthonitrile (probe 1) was designed and synthesized for the selective detection of hydrogen sulfide (H 2 S). The addition of H 2 S to a solution of probe 1 resulted in a marked fluorescence turn-on alongside a visual color change from colorless to light yellow. Importantly, this distinct color response indicated that probe 1 could be used as a visual sensor for H 2 S. Moreover, probe 1 was successfully used as a signal tool to determine the H 2 S levels in beer and red wine. Copyright © 2018 Elsevier Ltd. All rights reserved.

  18. Fluorescent probe based on heteroatom containing styrylcyanine: pH-sensitive properties and bioimaging in vivo

    International Nuclear Information System (INIS)

    Yang, Xiaodong; Gao, Ya; Huang, Zhibing; Chen, Xiaohui; Ke, Zhiyong; Zhao, Peiliang; Yan, Yichen; Liu, Ruiyuan; Qu, Jinqing

    2015-01-01

    A novel fluorescent probe based on heteroatom containing styrylcyanine is synthesized. The fluorescence of probe is bright green in basic and neutral media but dark orange in strong acidic environments, which could be reversibly switched. Such behavior enables it to work as a fluorescent pH sensor in the solution state and a chemosensor for detecting acidic and basic volatile organic compounds. Analyses by NMR spectroscopy confirm that the protonation or deprotonation of pyridinyl moiety is responsible for the sensing process. In addition, the fluorescent microscopic images of probe in live cells and zebrafish are achieved successfully, suggesting that the probe has good cell membrane permeability and low cytotoxicity. - Graphical abstract: A novel styrylcyanine-based fluorescent pH probe was designed and synthesized, the fluorescence of which is bright green in basic and neutral media but dark orange in strong acidic environments. Such behavior enables it to work as a fluorescent pH sensor in solution states, and a chemosensor for detecting volatile organic compounds with high acidity and basicity in solid state. In addition, it can be used for fluorescent imaging in living cell and living organism. - Highlights: • Bright green fluorescence was observed in basic and neutral media. • Dark orange fluorescence was found in strong acidic environments. • Volatile organic compounds with high acidity and basicity could be detected. • Bioimaging in living cell and living organism was achieved successfully

  19. Inference of protein diffusion probed via fluorescence correlation spectroscopy

    Science.gov (United States)

    Tsekouras, Konstantinos

    2015-03-01

    Fluctuations are an inherent part of single molecule or few particle biophysical data sets. Traditionally, ``noise'' fluctuations have been viewed as a nuisance, to be eliminated or minimized. Here we look on how statistical inference methods - that take explicit advantage of fluctuations - have allowed us to draw an unexpected picture of single molecule diffusional dynamics. Our focus is on the diffusion of proteins probed using fluorescence correlation spectroscopy (FCS). First, we discuss how - in collaboration with the Bustamante and Marqusee labs at UC Berkeley - we determined using FCS data that individual enzymes are perturbed by self-generated catalytic heat (Riedel et al, Nature, 2014). Using the tools of inference, we found how distributions of enzyme diffusion coefficients shift in the presence of substrate revealing that enzymes performing highly exothermic reactions dissipate heat by transiently accelerating their center of mass following a catalytic reaction. Next, when molecules diffuse in the cell nucleus they often appear to diffuse anomalously. We analyze FCS data - in collaboration with Rich Day at the IU Med School - to propose a simple model for transcription factor binding-unbinding in the nucleus to show that it may give rise to apparent anomalous diffusion. Here inference methods extract entire binding affinity distributions for the diffusing transcription factors, allowing us to precisely characterize their interactions with different components of the nuclear environment. From this analysis, we draw key mechanistic insight that goes beyond what is possible by simply fitting data to ``anomalous diffusion'' models.

  20. A highly sensitive fluorescent probe based on BODIPY for Hg2+ in aqueous solution

    Directory of Open Access Journals (Sweden)

    ZHAO Junwei

    2016-12-01

    Full Text Available A highly sensitive fluorescent probe based on BODIPY and hydrazine for Hg2+ was designed and synthesized.This probe could detect mercury ions in aqueous solutions within 5 min.With the increase of Hg2+ mole concentration,an obvious red shift of UV-Vis absorption wavelength was observed and the fluorescence intensity significantly enhanced.It was found that the fluorescence intensity of an aqueous solution containing 0.1 μmol/L Hg2+ is much stronger than that of blank solution,which indicats that the fluorescent probe has high sensitivity.In addition,other metal ions could not cause the change of fluorescent spectra,which means this probe has good selectivity,as well.

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

    International Nuclear Information System (INIS)

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

    2008-01-01

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

  2. An environmentally-friendly fluorescent method for quantification of lipid contents in yeast

    DEFF Research Database (Denmark)

    Severo Poli, Jandora; Lützhøft, Hans-Christian Holten; Karakashev, Dimitar Borisov

    2014-01-01

    lipid and the calibration curve showed linearity (R2 = 0.994) between 0.50 and 25 mg/L. Compared with traditional gravimetric analysis, the developed method is much faster and uses less organic solvents. Lipid contents determined by fluorescence and gravimetry were the same for some strains......This study aimed at developing an efficient, fast and environmentally-friendly method to quantify neutral lipid contents in yeast. After optimising the fluorescence instrument parameters and influence of organic solvent concentrations, a new method to quantify neutral lipids in yeast based......, but for other strains the lipid contents determined by fluorescence were less. This new method will therefore be suitable for fast screening purposes....

  3. A Fluorescent Probe for Sensitive Detection of Hydrazine and Its Application in Red Wine and Water.

    Science.gov (United States)

    Wang, Jialin; Wang, Hao; Yang, Shaoxiang; Tian, Hongyu; Liu, Yongguo; Hao, Yanfeng; Zhang, Jie; Sun, Baoguo

    2018-01-01

    A fluorescent probe, 7-(diethylamino)-2-oxo-2H-chromene-4-carbaldehyde (probe 1), was designed and synthesized for the sensitive detection of hydrazine. The addition of N 2 H 4 caused the fluorescence intensity of probe 1 to decrease. The probe's fluorescence was turn-off after adding N 2 H 4 , which could be observed under UV light at 365 nm. Moreover, once treated with different concentrations N 2 H 4 solutions, the solution color change could be distinguished, which indicates that probe 1 could be used as a visual sensor for hydrazine. Moreover, probe 1 can be used as a signal tool to determine hydrazine levels in solutions, such as red wine and water.

  4. A novel dansyl-based fluorescent probe for highly selective detection of ferric ions.

    Science.gov (United States)

    Yang, Min; Sun, Mingtai; Zhang, Zhongping; Wang, Suhua

    2013-02-15

    A novel dansyl-based fluorescent probe was synthesized and characterized. It exhibits high selectivity and sensitivity towards Fe(3+) ion. This fluorescent probe is photostable, water soluble and pH insensitive. The limit of detection is found to be 0.62 μM. These properties make it a good fluorescent probe for Fe(3+) ion detection in both chemical and biological systems. Spike recovery test confirms its practical application in tap water samples. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. Imaging lipid domains in cell membranes: the advent of super-resolution fluorescence microscopy

    Directory of Open Access Journals (Sweden)

    Dylan Myers Owen

    2013-12-01

    Full Text Available The lipid bilayer of model membranes, liposomes reconstituted from cell lipids, and plasma membrane vesicles and spheres can separate into two distinct liquid phases to yield lipid domains with liquid-ordered and liquid-disordered properties. These observations are the basis of the lipid raft hypothesis that postulates the existence of cholesterol-enriched ordered-phase lipid domains in cell membranes that could regulate protein mobility, localization and interaction. Here we review the evidence that nano-scaled lipid complexes and meso-scaled lipid domains exist in cell membranes and how new fluorescence microscopy techniques that overcome the diffraction limit provide new insights into lipid organization in cell membranes.

  6. Application of locked nucleic acid-based probes in fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Fontenete, Sílvia; Carvalho, Daniel R; Guimarães, Nuno

    2016-01-01

    of nucleic acid mimics used as mixmers in LNA-based probes strongly influence the efficiency of detection. LNA probes with 10 to 15 mers showed the highest efficiency. Additionally, the combination of 2′-OMe RNA with LNA allowed an increase on the fluorescence intensities of the probes. Overall......Fluorescence in situ hybridization (FISH) employing nucleic acid mimics as probes is becoming an emerging molecular tool in the microbiology area for the detection and visualization of microorganisms. However, the impact that locked nucleic acid (LNA) and 2′-O-methyl (2′-OMe) RNA modifications have...

  7. A benzothiazole-based fluorescent probe for hypochlorous acid detection and imaging in living cells

    Science.gov (United States)

    Nguyen, Khac Hong; Hao, Yuanqiang; Zeng, Ke; Fan, Shengnan; Li, Fen; Yuan, Suke; Ding, Xuejing; Xu, Maotian; Liu, You-Nian

    2018-06-01

    A benzothiazole-based turn-on fluorescent probe with a large Stokes shift (190 nm) has been developed for hypochlorous acid detection. The probe displays prompt fluorescence response for HClO with excellent selectivity over other reactive oxygen species as well as a low detection limit of 0.08 μM. The sensing mechanism involves the HClO-induced specific oxidation of oxime moiety of the probe to nitrile oxide, which was confirmed by HPLC-MS technique. Furthermore, imaging studies demonstrated that the probe is cell permeable and can be applied to detect HClO in living cells.

  8. Can pyrene probes be used to measure lateral pressure profiles of lipid membranes? Perspective through atomistic simulations

    DEFF Research Database (Denmark)

    Franova, M. D.; Vattulainen, I.; Ollila, O. H. S.

    2014-01-01

    The lateral pressure profile of lipid bilayers has gained a lot of attention, since changes in the pressure profile have been suggested to shift the membrane protein conformational equilibrium. This relation has been mostly studied with theoretical methods, especially with molecular dynamics....../monomer fluorescence ratio has been assumed to represent the lateral pressure in the location of the pyrene moieties. Here, we consider the validity of this assumption through atomistic molecular dynamics simulations in a DOPC (dioleoylphosphatidylcholine) membrane, which hosts di-pyr-PC probes with different acyl...... simulations, since established methods to measure the lateral pressure profile experimentally have not been available. The only experiments that have attempted to gauge the lateral pressure profile have been done by using di-pyrenyl-phosphatidylcholine (di-pyr-PC) probes. In these experiments, the excimer...

  9. A small molecular pH-dependent fluorescent probe for cancer cell imaging in living cell.

    Science.gov (United States)

    Ma, Junbao; Li, Wenqi; Li, Juanjuan; Shi, Rongguang; Yin, Gui; Wang, Ruiyong

    2018-05-15

    A novel pH-dependent two-photon fluorescent molecular probe ABMP has been prepared based on the fluorophore of 2, 4, 6-trisubstituted pyridine. The probe has an absorption wavelength at 354 nm and corresponding emission wavelength at 475 nm with the working pH range from 2.20 to 7.00, especially owning a good liner response from pH = 2.40 to pH = 4.00. ABMP also has excellent reversibility, photostability and selectivity which promotes its ability in analytical application. The probe can be excited with a two-photon fluorescence microscopy and the fluorescence cell imaging indicated that the probe can distinguish Hela cancer cells out of normal cells with a two-photon fluorescence microscopy which suggested its potential application in tumor cell detection. Copyright © 2018 Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

    Mayer-Cumblidge, M Uljana [Richland, WA; Cao, Haishi [Richland, WA

    2010-08-17

    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.

  11. A chromenoquinoline-based fluorescent off-on thiol probe for bioimaging.

    Science.gov (United States)

    Kand, Dnyaneshwar; Kalle, Arunasree Marasanapalli; Varma, Sreejith Jayasree; Talukdar, Pinaki

    2012-03-11

    A new chromenoquinoline-based fluorescent off-on thiol probe 2 is reported. In aqueous buffer solutions at physiological pH, the probe exhibited 223-fold enhancement in fluorescence intensity by a Michael addition of cysteine to the maleimide appended to a chromenoquinoline. Cell permeability and live cell imaging of thiols are also demonstrated. This journal is © The Royal Society of Chemistry 2012

  12. Visualizing tributyltin (TBT) in bacterial aggregates by specific rhodamine-based fluorescent probes.

    Science.gov (United States)

    Jin, Xilang; Hao, Likai; She, Mengyao; Obst, Martin; Kappler, Andreas; Yin, Bing; Liu, Ping; Li, Jianli; Wang, Lanying; Shi, Zhen

    2015-01-01

    Here we present the first examples of fluorescent and colorimetric probes for microscopic TBT imaging. The fluorescent probes are highly selective and sensitive to TBT and have successfully been applied for imaging of TBT in bacterial Rhodobacter ferrooxidans sp. strain SW2 cell-EPS-mineral aggregates and in cell suspensions of the marine cyanobacterium Synechococcus PCC 7002 by using confocal laser scanning microscopy. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. In situ hybridization of phytoplankton using fluorescently labeled rRNA probes

    OpenAIRE

    Groben, R.; Medlin, Linda

    2005-01-01

    Fluorescently-labelled molecular probes were used to identify and characterise phytoplankton species using in situ hybridisation coupled with fluorescence microscopy and flow cytometry. The application of this technique is sometimes problematic, because of the many different species with which this method is to be used. Problems that may occur are: probe penetration versus maintanance of cell stability, strong autofluorescence and/or cell lost during the sample processing. Here we present a m...

  14. Optical imaging of non-fluorescent nanoparticle probes in live cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Gufeng; Stender, Anthony S.; Sun, Wei; and Fang, Ning

    2009-12-17

    Precise imaging of cellular and subcellular structures and dynamic processes in live cells is crucial for fundamental research in life sciences and in medical applications. Non-fluorescent nanoparticles are an important type of optical probe used in live-cell imaging due to their photostability, large optical cross-sections, and low toxicity. Here, we provide an overview of recent developments in the optical imaging of non-fluorescent nanoparticle probes in live cells.

  15. A new fluorescent pH probe for extremely acidic conditions

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Yu [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Jiang, Zheng [School of Life Science, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Xiao, Yu [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Bi, Fu-Zhen [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Miao, Jun-Ying, E-mail: miaojy@sdu.edu.cn [School of Life Science, Shandong University, Jinan 250100 (China); Zhao, Bao-Xiang, E-mail: bxzhao@sdu.edu.cn [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China)

    2014-04-01

    A new coumarin-based fluorescent probe can detect highly acidic conditions in both solution and bacteria with high selectivity and sensitivity. Highlights: • A new fluorescence probe for very low pH was synthesized and characterized. • The probe can monitor pH in solution and bacteria. • The two-step protonation of N atoms of the probe leads to fluorescence quenching. Abstract: A novel turn-off fluorescent probe based on coumarin and imidazole moiety for extremely acidic conditions was designed and developed. The probe with pKa = 2.1 is able to respond to very low pH value (below 3.5) with high sensitivity relying on fluorescence quenching at 460 nm in fluorescence spectra or the ratios of absorbance maximum at 380 nm to that at 450 nm in UV–vis spectra. It can quantitatively detect pH value based on equilibrium equation, pH = pKa -log[(Ix - Ib)/(Ia - Ix)]. It had very short response time that was less than 1 min, good reversibility and nearly no interference from common metal ions. Moreover, using ¹H NMR analysis and theoretical calculation of molecular orbital, we verified that a two-step protonation process of two N atoms of the probe leaded to photoinduced electron transfer (PET), which was actually the mechanism of the fluorescence quenching phenomenon under strongly acidic conditions. Furthermore, the probe was also applied to imaging strong acidity in bacteria, E.coli and had good effect. This work illustrates that the new probe could be a practical and ideal pH indicator for strongly acidic conditions with good biological significance.

  16. Molecularly imprinted fluorescent probe based on FRET for selective and sensitive detection of doxorubicin

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Zhifeng, E-mail: 897061147@qq.com [College of Chemistry and Materials Science, Hengyang Normal University, Key Laboratory of Functional Organometallic Materials of Hunan Province University, Hengyang 421008 (China); Deng, Peihong; Li, Junhua [College of Chemistry and Materials Science, Hengyang Normal University, Key Laboratory of Functional Organometallic Materials of Hunan Province University, Hengyang 421008 (China); Xu, Li [Department of Applied Chemistry, College of Materials and Energy, South China Agricultural University, Guangzhou 510642 (China); Tang, Siping [College of Chemistry and Materials Science, Hengyang Normal University, Key Laboratory of Functional Organometallic Materials of Hunan Province University, Hengyang 421008 (China)

    2017-04-15

    Highlights: • FRET-based molecularly imprinted probe for detection of doxorubicin was prepared. • The detection limit of the probe was 13.8 nM for doxorubicin. • The FRET-based probe had a higher selectivity for the template than ordinary MIMs. - Abstract: In this work, a new type of fluorescent probe for detection of doxorubicin has been constructed by the combined use of fluorescence resonance energy transfer (FRET) technology and molecular imprinting technique (MIT). Using doxorubicin as the template, the molecularly imprinted polymer thin layer was fabricated on the surfaces of carbon dot (CD) modified silica by sol-gel polymerization. The excitation energy of the fluorescent donor (CDs) could be transferred to the fluorescent acceptor (doxorubicin). The FRET based fluorescent probe demonstrated high sensitivity and selectivity for doxorubicin. The detection limit was 13.8 nM. The fluorescent probe was successfully applied for detecting doxorubicin in doxorubicin-spiked plasmas with a recovery of 96.8–103.8%, a relative standard deviation (RSD) of 1.3–2.8%. The strategy for construction of FRET-based molecularly imprinted materials developed in this work is very promising for analytical applications.

  17. Fluoromodule-based reporter/probes designed for in vivo fluorescence imaging

    Science.gov (United States)

    Zhang, Ming; Chakraborty, Subhasish K.; Sampath, Padma; Rojas, Juan J.; Hou, Weizhou; Saurabh, Saumya; Thorne, Steve H.; Bruchez, Marcel P.; Waggoner, Alan S.

    2015-01-01

    Optical imaging of whole, living animals has proven to be a powerful tool in multiple areas of preclinical research and has allowed noninvasive monitoring of immune responses, tumor and pathogen growth, and treatment responses in longitudinal studies. However, fluorescence-based studies in animals are challenging because tissue absorbs and autofluoresces strongly in the visible light spectrum. These optical properties drive development and use of fluorescent labels that absorb and emit at longer wavelengths. Here, we present a far-red absorbing fluoromodule–based reporter/probe system and show that this system can be used for imaging in living mice. The probe we developed is a fluorogenic dye called SC1 that is dark in solution but highly fluorescent when bound to its cognate reporter, Mars1. The reporter/probe complex, or fluoromodule, produced peak emission near 730 nm. Mars1 was able to bind a variety of structurally similar probes that differ in color and membrane permeability. We demonstrated that a tool kit of multiple probes can be used to label extracellular and intracellular reporter–tagged receptor pools with 2 colors. Imaging studies may benefit from this far-red excited reporter/probe system, which features tight coupling between probe fluorescence and reporter binding and offers the option of using an expandable family of fluorogenic probes with a single reporter gene. PMID:26348895

  18. A fluorescent glycolipid-binding peptide probe traces cholesterol dependent microdomain-derived trafficking pathways.

    Directory of Open Access Journals (Sweden)

    Steffen Steinert

    Full Text Available BACKGROUND: The uptake and intracellular trafficking of sphingolipids, which self-associate into plasma membrane microdomains, is associated with many pathological conditions, including viral and toxin infection, lipid storage disease, and neurodegenerative disease. However, the means available to label the trafficking pathways of sphingolipids in live cells are extremely limited. In order to address this problem, we have developed an exogenous, non-toxic probe consisting of a 25-amino acid sphingolipid binding domain, the SBD, derived from the amyloid peptide Abeta, and conjugated by a neutral linker with an organic fluorophore. The current work presents the characterization of the sphingolipid binding and live cell trafficking of this novel probe, the SBD peptide. SBD was the name given to a motif originally recognized by Fantini et al in a number of glycolipid-associated proteins, and was proposed to interact with sphingolipids in membrane microdomains. METHODOLOGY/PRINCIPAL FINDINGS: In accordance with Fantini's model, optimal SBD binding to membranes depends on the presence of sphingolipids and cholesterol. In synthetic membrane binding assays, SBD interacts preferentially with raft-like lipid mixtures containing sphingomyelin, cholesterol, and complex gangliosides in a pH-dependent manner, but is less glycolipid-specific than Cholera toxin B (CtxB. Using quantitative time-course colocalization in live cells, we show that the uptake and intracellular trafficking route of SBD is unlike that of either the non-raft marker Transferrin or the raft markers CtxB and Flotillin2-GFP. However, SBD traverses an endolysosomal route that partially intersects with raft-associated pathways, with a major portion being diverted at a late time point to rab11-positive recycling endosomes. Trafficking of SBD to acidified compartments is strongly disrupted by cholesterol perturbations, consistent with the regulation of sphingolipid trafficking by cholesterol

  19. A dansyl-rhodamine ratiometric fluorescent probe for Hg2+ based on FRET mechanism.

    Science.gov (United States)

    Xie, Puhui; Guo, Fengqi; Wang, Lingyu; Yang, Sen; Yao, Denghui; Yang, Guoyu

    2015-03-01

    Based on resonance energy transfer (FRET) from dansyl to rhodamine 101, a new fluorescent probe (compound 1) containing rhodamine 101 and a dansyl unit was synthesized for detecting Hg(2+) through ratiometric sensing in DMSO aqueous solutions. This probe shows a fast, reversible and selective response toward Hg(2+) in a wide pH range. Hg(2+) induced ring-opening reactions of the spirolactam rhodamine moiety of 1, leading to the formation of fluorescent derivatives that can serve as the FRET acceptors. Very large stokes shift (220 nm) was observed in this case. About 97-fold increase in fluorescence intensity ratio was observed upon its binding with Hg(2+).

  20. Cu2+-labeled dansyl compounds as fluorescent and PET probes for imaging apoptosis.

    Science.gov (United States)

    Han, Junyan; Wang, Xukui; Yu, MeiXiang

    2016-11-15

    Compound DNSTT-Cu 2+ , a novel chelate of Cu 2+ with DOTA conjugated to a fluorescent dansyl fragment, is developed for imaging cell apoptosis. Apoptotic U-87MG cells could be selectively visualized by the fluorescence of DNSTT-Cu 2+ from cytoplasm of cells, confirmed by the fluorescence of apoptosis cells co-labeled with Alexa Fluor 568-labeled annexin V, a conventional probe for selectively labeling membranes of apoptosis cells. A radioactive 64 Cu 2 + analog, DNSTT- 64 Cu 2+ , was easily synthesized, providing a potential PET probe for imaging apoptosis in vivo. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. The influence of NBD fluorescent probe on model membranes containing POPC and DPPC.

    Science.gov (United States)

    Weng, Chi-Jung; Wu, Ju-Ping; Kuo, Ming-Yen; Hsueh, Ya-Wei

    2016-03-01

    To investigate the effect of fluorescent probe on the properties of membranes, we studied model membranes composed of 1,2- dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1-palmitoyl 2-oleoyl-sn-glycero-3-phosphocholine (POPC) in the presence and absence of fluorescent probe. The morphology of giant unilamellar vesicles (GUVs) has been observed as a function of temperature and composition by fluorescence microscopy using NBD-DOPE or C 6 -NBD-PC as the probe. The phase behavior of model membranes containing no fluorescent probe was investigated by 2 H-NMR spectroscopy. We found that the bright phase observed on GUVs was the fluid phase enriched in POPC and the dark phase was the gel phase enriched in DPPC. NBD-DOPE and C 6 -NBD-PC preferentially participated in the fluid-phase domains when GUVs were in the gel + fluid phase coexistence. Inclusion of both fluorescent probes (1 mol%) lowered the transition temperature of POPC/DPPC membranes. In addition, C 6 -NBD-PC exhibited a stronger effect than NBD-DOPE, which was considered to be associated with the structures of fluorescent molecules.

  2. Highly selective detection of glutathione using a NIP/Cu2+ complex fluorescent probe

    International Nuclear Information System (INIS)

    Liang Wenrui; Zhao Zhi; Zhang Yang; Wang Qiusheng; Zhao Xin; Ouyang Jie

    2012-01-01

    A novel fluorescent compound, 4-(trimethyl ammonium chloride)acetamide-2-(1H-naphtho[2,3-d]imidazol-2-yl)phenol (TMACA-NIP), was synthesized and used as a fluorescent probe for detecting glutathione reduced (GSH). The new NIP-based probe exhibited high fluorescence in water, which was quenched during the presence of copper (II) due to the complexation between TMACA-NIP and Cu 2+ . But after adding GSH into the TMACA-NIP and Cu 2+ system, the fluorescence of TMACA-NIP was recovered because the binding force between GSH and Cu 2+ is stronger than that between TMACA-NIP and Cu 2+ , which destroys the equilibrium between NIP and copper (II) ions and releases the fluorescence probe of TMACA-NIP. This three-component competing system of NIP/Cu 2+ /GSH can be used to detect GSH simply and rapidly. - Highlights: ► A novel fluorescence probe was developed to detect GSH that operates in aqueous solution. ► TMACA-NIP was synthesized and employed as “read-out” units of NIP/Cu 2+ /GSH. ► NIP-based probe shows high selectivity over other sulfhydryl compounds.

  3. Bifunctional fluorescent probes for detection of amyloid aggregates and reactive oxygen species.

    Science.gov (United States)

    Needham, Lisa-Maria; Weber, Judith; Fyfe, James W B; Kabia, Omaru M; Do, Dung T; Klimont, Ewa; Zhang, Yu; Rodrigues, Margarida; Dobson, Christopher M; Ghandi, Sonia; Bohndiek, Sarah E; Snaddon, Thomas N; Lee, Steven F

    2018-02-01

    Protein aggregation into amyloid deposits and oxidative stress are key features of many neurodegenerative disorders including Parkinson's and Alzheimer's disease. We report here the creation of four highly sensitive bifunctional fluorescent probes, capable of H 2 O 2 and/or amyloid aggregate detection. These bifunctional sensors use a benzothiazole core for amyloid localization and boronic ester oxidation to specifically detect H 2 O 2 . We characterized the optical properties of these probes using both bulk fluorescence measurements and single-aggregate fluorescence imaging, and quantify changes in their fluorescence properties upon addition of amyloid aggregates of α-synuclein and pathophysiological H 2 O 2 concentrations. Our results indicate these new probes will be useful to detect and monitor neurodegenerative disease.

  4. Bifunctional fluorescent probes for detection of amyloid aggregates and reactive oxygen species

    Science.gov (United States)

    Needham, Lisa-Maria; Weber, Judith; Fyfe, James W. B.; Kabia, Omaru M.; Do, Dung T.; Klimont, Ewa; Zhang, Yu; Rodrigues, Margarida; Dobson, Christopher M.; Ghandi, Sonia; Bohndiek, Sarah E.; Snaddon, Thomas N.; Lee, Steven F.

    2018-02-01

    Protein aggregation into amyloid deposits and oxidative stress are key features of many neurodegenerative disorders including Parkinson's and Alzheimer's disease. We report here the creation of four highly sensitive bifunctional fluorescent probes, capable of H2O2 and/or amyloid aggregate detection. These bifunctional sensors use a benzothiazole core for amyloid localization and boronic ester oxidation to specifically detect H2O2. We characterized the optical properties of these probes using both bulk fluorescence measurements and single-aggregate fluorescence imaging, and quantify changes in their fluorescence properties upon addition of amyloid aggregates of α-synuclein and pathophysiological H2O2 concentrations. Our results indicate these new probes will be useful to detect and monitor neurodegenerative disease.

  5. Development of Ultrasonic Modulation Probe for Fluorescence Tomography Based on Acousto-Optic Effect

    Directory of Open Access Journals (Sweden)

    Trinh Quang Duc

    2011-01-01

    Full Text Available We have developed an ultrasonic probe for fluorescence modulation to image fluorescence within biological tissues. The probe consists of a focused ultrasonic transducer mounted on actuators for mechanical fan scanning, which can be used in contact with the measuring object aiming for clinical application. The mechanical fan scanning employed in the probe has a beneficial feature of portability. As a result, fluorescent beads, which were localized with the diameter of 2 mm at 20 mm depth in a pork meat tissue, were detected with resolution of 3 mm. The system performance denotes the feasibility of development towards the final goal of ultrasonic fluorescence modulation tomography for clinical applications.

  6. Spectrally-resolved response properties of the three most advanced FRET based fluorescent protein voltage probes.

    Directory of Open Access Journals (Sweden)

    Hiroki Mutoh

    Full Text Available Genetically-encoded optical probes for membrane potential hold the promise of monitoring electrical signaling of electrically active cells such as specific neuronal populations in intact brain tissue. The most advanced class of these probes was generated by molecular fusion of the voltage sensing domain (VSD of Ci-VSP with a fluorescent protein (FP pair. We quantitatively compared the three most advanced versions of these probes (two previously reported and one new variant, each involving a spectrally distinct tandem of FPs. Despite these different FP tandems and dissimilarities within the amino acid sequence linking the VSD to the FPs, the amplitude and kinetics of voltage dependent fluorescence changes were surprisingly similar. However, each of these fluorescent probes has specific merits when considering different potential applications.

  7. A coumarin-based colorimetric fluorescent probe for hydrogen sulfide

    Indian Academy of Sciences (India)

    with nitric oxide (NO) and carbon monoxide (CO), which mediate ... esterification, no emission of the fluorophore would be found due to ... Fluorescence emission spectra were obtained ... of excessive organic solvent might restrict the practical.

  8. Determination of paraquat in water samples using a sensitive fluorescent probe titration method.

    Science.gov (United States)

    Yao, Feihu; Liu, Hailong; Wang, Guangquan; Du, Liming; Yin, Xiaofen; Fu, Yunlong

    2013-06-01

    Paraquat (PQ), a nonselective herbicide, is non-fluorescent in aqueous solutions. Thus, its determination through direct fluorescent methods is not feasible. The supramolecular inclusion interaction of PQ with cucurbit[7]uril was studied by a fluorescent probe titration method. Significant quenching of the fluorescence intensity of the cucurbit[7]uril-coptisine fluorescent probe was observed with the addition of PQ. A new fluorescent probe titration method with high selectivity and sensitivity at the ng/mL level was developed to determine PQ in aqueous solutions with good precision and accuracy based on the significant quenching of the supramolecular complex fluorescence intensity. The proposed method was successfully used in the determination of PQ in lake water, tap water, well water, and ditch water in an agricultural area, with recoveries of 96.73% to 105.77%. The fluorescence quenching values (deltaF) showed a good linear relationship with PQ concentrations from 1.0 x 10(-8) to 1.2 x 10(-5) mol/L with a detection limit of 3.35 x 10(-9) mol/L. In addition, the interaction models of the supramolecular complexes formed between the host and the guest were established using theoretical calculations. The interaction mechanism between the cucurbit[7]uril and PQ was confirmed by 1H NMR spectroscopy.

  9. Probing the binding of cationic lipids with dendrimers.

    Science.gov (United States)

    Mandeville, J S; Bourassa, P; Tajmir-Riahi, H A

    2013-01-14

    Polycationic polymers are used extensively in biology to disrupt cell membranes and thus enhance the transport of materials into the cell. We report the bindings of several lipids cholesterol (Chol), 1,2-dioleoyl-3-trimethylammonium-propane(DOTAP), dioctadecyldimethylammoniumbromide (DDAB), and dioleoylphosphatidylethanolamine (DOPE) to dendrimers of different compositions such as mPEG-PAMAM (G3), mPEG-PAMAM (G4), and PAMAM (G4) under physiological conditions. FTIR, UV-visible spectroscopic, methods and molecular modeling were used to analyze the lipid binding mode, the binding constant, and the effects of lipid complexation on the dendrimer structure. The structural analysis showed that lipids bind dendrimers through both hydrophilic and hydrophobic contacts with overall binding constants of K(chol-mPEG-G3) = 1.7 × 10(3) M(-1), K(chol-mPEG-PAMAM-G4) = 2.7 × 10(3) M(-1), K(chol-PAMAM-G4) = 1.0 × 10(3) M(-1), K(DOPE-mPEG-G3) = 1.5 × 10(3) M(-1), K(DOPE-mPEG-PAMAM-G4) = 1.6 × 10(3) M(-1), K(DOPE-PAMAM-G4) = 5.3 × 10(2) M(-1), K(DDAB-mPEG-G3) = 1.5 × 10(3) M(-1), K(DDAB-mPEG-PAMAM-G4) = 1.9 × 10(2) M(-1), K(DDAB-PAMAM-G4) = 7.0 × 10(2) M(-1), K(DOTAP-mPEG-G3) = 1.9 × 10(3) M(-1), K(DOTAP-mPEG-PAMAM-G4) = 1.5 × 10(3) M(-1), and K(DOTAP-PAMAM-G4) = 5.7 × 10(2) M(-1). Weaker interaction was observed as dendrimer cationic charges increased. The free binding energies from docking were -5.15 (cholesterol), -5.79 (DDAB), and -5.36 kcal/mol (DOTAP) with the order of stability DDAB-PAMAM-G-4 > DOTAP-PAMAM-G4 > cholesterol-PAMAM-G4, consistent with the spectroscopic results. Dendrimers might act as carriers to transport lipids in vitro.

  10. An effective colorimetric and ratiometric fluorescent probe for bisulfite in aqueous solution

    International Nuclear Information System (INIS)

    Dai, Xi; Zhang, Tao; Du, Zhi-Fang; Cao, Xiang-Jian; Chen, Ming-Yu; Hu, Sheng-Wen; Miao, Jun-Ying; Zhao, Bao-Xiang

    2015-01-01

    We have developed the first two-photon colorimetric and ratiometric fluorescent probe, BICO, for the detection of bisulfite (HSO 3 − ) in aqueous solution. The probe contains coumarin and benzimidazole moieties and can detect HSO 3 − based on the Michael addition reaction with a limit of detection 5.3 × 10 −8  M in phosphate-buffered saline solution. The probe was used to detect bisulfite in tap water, sugar and dry white wine. Moreover, test strips were made and used easily. We successfully applied the probe to image living cells, using one-photon fluorescence imaging. BICO overcomes the limitations in sensitivity of previously reported probes and the solvation effect of bisulfite, which demonstrates its excellent value in practical application. - Highlights: • A colorimetric and ratiometric fluorescent probe was developed. • The probe could detect bisulfite in PBS buffer solution and real samples. • Bisulfite test paper was made to naked-eye detect bisulfite. • This probe successfully used to living cell imaging in ratiometric manner

  11. An effective colorimetric and ratiometric fluorescent probe for bisulfite in aqueous solution

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Xi [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Zhang, Tao [Institute of Developmental Biology, School of Life Science, Shandong University, Jinan 250100 (China); Du, Zhi-Fang; Cao, Xiang-Jian; Chen, Ming-Yu [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Hu, Sheng-Wen [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Miao, Jun-Ying, E-mail: miaojy@sdu.edu.cn [Institute of Developmental Biology, School of Life Science, Shandong University, Jinan 250100 (China); Zhao, Bao-Xiang, E-mail: bxzhao@sdu.edu.cn [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China)

    2015-08-12

    We have developed the first two-photon colorimetric and ratiometric fluorescent probe, BICO, for the detection of bisulfite (HSO{sub 3}{sup −}) in aqueous solution. The probe contains coumarin and benzimidazole moieties and can detect HSO{sub 3}{sup −} based on the Michael addition reaction with a limit of detection 5.3 × 10{sup −8} M in phosphate-buffered saline solution. The probe was used to detect bisulfite in tap water, sugar and dry white wine. Moreover, test strips were made and used easily. We successfully applied the probe to image living cells, using one-photon fluorescence imaging. BICO overcomes the limitations in sensitivity of previously reported probes and the solvation effect of bisulfite, which demonstrates its excellent value in practical application. - Highlights: • A colorimetric and ratiometric fluorescent probe was developed. • The probe could detect bisulfite in PBS buffer solution and real samples. • Bisulfite test paper was made to naked-eye detect bisulfite. • This probe successfully used to living cell imaging in ratiometric manner.

  12. The interaction of antibodies with lipid membranes unraveled by fluorescence methodologies

    Science.gov (United States)

    Figueira, Tiago N.; Veiga, Ana Salomé; Castanho, Miguel A. R. B.

    2014-12-01

    The interest and investment in antibody therapies has reached an overwhelming scale in the last decade. Yet, little concern has been noticed among the scientific community to unravel important interactions of antibodies with biological structures other than their respective epitopes. Lipid membranes are particularly relevant in this regard as they set the stage for protein-protein recognition, a concept potentially inclusive of antibody-antigen recognition. Fluorescence techniques allow experimental monitoring of protein partition between aqueous and lipid phases, deciphering events of adsorption, insertion and diffusion. This review focuses on the available fluorescence spectroscopy methodologies directed to the study of antibody-membrane interactions.

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

    Science.gov (United States)

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

    2014-03-01

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

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

    Science.gov (United States)

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

    2012-07-01

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

  15. Mercury speciation with fluorescent gold nanocluster as a probe.

    Science.gov (United States)

    Yang, Jian-Yu; Yang, Ting; Wang, Xiao-Yan; Chen, Ming-Li; Yu, Yong-Liang; Wang, Jian-Hua

    2018-05-11

    Fluorescent nanoparticles are widely used for sensing biologically significant species. However, it is rarely reported for the discrimination or speciation of metal species. In this work, we report for the first time the speciation of mercury (Hg 2+ ) and methylmercury (CH 3 Hg + ) by taking advantage of the fluorescence feature of folic acid-capped gold nanoclusters (FA-AuNCs). FA-Au NCs exhibit an average size of 2.08±0.15 nm and a maximum emission at λ ex /λ em = 280/440 nm with a quantum yield of 27.3%. It is interesting that Hg 2+ causes a significant quench on the fluorescence of FA-Au NCs, whereas CH 3 Hg + leads to a remarkable fluorescence enhancement. Based on this discriminative fluorescent response between Hg 2+ and CH 3 Hg + , a novel nanosensor for the speciation of CH 3 Hg + and Hg 2+ was developed, providing limits of detection (LOD) of 28 nM for Hg 2+ and 25 nM for CH 3 Hg + within 100-1000 nM. This sensing system is highly selective to mercury. Its practical applications were further demonstrated by the analysis of CH 3 Hg + and the speciation of mercury (CH 3 Hg + and Hg 2+ ) in environmental water and fish samples.

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

    Directory of Open Access Journals (Sweden)

    Zhou Han

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

  17. Selective imaging of cancer cells with a pH-activatable lysosome-targeting fluorescent probe.

    Science.gov (United States)

    Shi, Rongguang; Huang, Lu; Duan, Xiaoxue; Sun, Guohao; Yin, Gui; Wang, Ruiyong; Zhu, Jun-Jie

    2017-10-02

    Fluorescence imaging with tumor-specific fluorescent probe has emerged as a tool to aid surgeons in the identification and removal of tumor tissue. We report here a new lysosome-targeting fluorescent probe (NBOH) with BODIPY fluorephore to distinguish tumor tissue out of normal tissue based on different pH environment. The probe exhibited remarkable pH-dependent fluorescence behavior in a wide pH range from 3.0 to 11.0, especially a sensitive pH-dependent fluorescence change at pH range between 3.5 and 5.5, corresponding well to the acidic microenvironment of tumor cells, in aqueous solution. The response time of NBOH was extremely short and the photostability was proved to be good. Toxicity test and fluorescence cell imaging together with a sub-cellular localization study were carried out revealing its low biotoxicity and good cell membrane permeability. And NBOH was successfully applied to the imaging of tumor tissue in tumor-bearing mice suggesting potential application to surgery as a tumor-specific probe. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Synergistic Combination of Unquenching and Plasmonic Fluorescence Enhancement in Fluorogenic Nucleic Acid Hybridization Probes.

    Science.gov (United States)

    Vietz, Carolin; Lalkens, Birka; Acuna, Guillermo P; Tinnefeld, Philip

    2017-10-11

    Fluorogenic nucleic acid hybridization probes are widely used for detecting and quantifying nucleic acids. The achieved sensitivity strongly depends on the contrast between a quenched closed form and an unquenched opened form with liberated fluorescence. So far, this contrast was improved by improving the quenching efficiency of the closed form. In this study, we modularly combine these probes with optical antennas used for plasmonic fluorescence enhancement and study the effect of the nanophotonic structure on the fluorescence of the quenched and the opened form. As quenched fluorescent dyes are usually enhanced more by fluorescence enhancement, a detrimental reduction of the contrast between closed and opened form was anticipated. In contrast, we could achieve a surprising increase of the contrast with full additivity of quenching of the dark form and fluorescence enhancement of the bright form. Using single-molecule experiments, we demonstrate that the additivity of the two mechanisms depends on the perfect quenching in the quenched form, and we delineate the rules for new nucleic acid probes for enhanced contrast and absolute brightness. Fluorogenic hybridization probes optimized not only for quenching but also for the brightness of the open form might find application in nucleic acid assays with PCR avoiding detection schemes.

  19. Multiplex fluorescence melting curve analysis for mutation detection with dual-labeled, self-quenched probes.

    Directory of Open Access Journals (Sweden)

    Qiuying Huang

    2011-04-01

    Full Text Available Probe-based fluorescence melting curve analysis (FMCA is a powerful tool for mutation detection based on melting temperature generated by thermal denaturation of the probe-target hybrid. Nevertheless, the color multiplexing, probe design, and cross-platform compatibility remain to be limited by using existing probe chemistries. We hereby explored two dual-labeled, self-quenched probes, TaqMan and shared-stem molecular beacons, in their ability to conduct FMCA. Both probes could be directly used for FMCA and readily integrated with closed-tube amplicon hybridization under asymmetric PCR conditions. Improved flexibility of FMCA by using these probes was illustrated in three representative applications of FMCA: mutation scanning, mutation identification and mutation genotyping, all of which achieved improved color-multiplexing with easy probe design and versatile probe combination and all were validated with a large number of real clinical samples. The universal cross-platform compatibility of these probes-based FMCA was also demonstrated by a 4-color mutation genotyping assay performed on five different real-time PCR instruments. The dual-labeled, self-quenched probes offered unprecedented combined advantage of enhanced multiplexing, improved flexibility in probe design, and expanded cross-platform compatibility, which would substantially improve FMCA in mutation detection of various applications.

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

    Directory of Open Access Journals (Sweden)

    Gregor P. C. Drummen

    2012-11-01

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

  1. Copper nanoclusters as probes for turn-on fluorescence sensing of L-lysine.

    Science.gov (United States)

    Zhang, Mingming; Qiao, Juan; Zhang, Shufeng; Qi, Li

    2018-05-15

    Herein, a unique protocol based on copper nanoclusters (CuNCs) probe for turn-on fluorescence sensing of L-lysine was developed. The fluorescent CuNCs with ovalbumin as the stabilizer was prepared by a simple, one-step and green method. When 370 nm was used as the excitation wavelength, the resultant CuNCs exhibited a pale blue fluorescence with the maximum emission at 440 nm. Interestingly, existence of L-lysine evoked the obvious fluorescence intensity increase of CuNCs. The detection limit of the proposed method for L-lysine was 5.5 μM, with a good linear range from 10.0 μM to 1.0 mM (r 2 = 0.999). Moreover, the possible mechanism for enhanced fluorescence intensity of CuNCs by addition of L-lysine was explored and discussed briefly. Further, the as-prepared fluorescent CuNCs was successfully applied in detection of L-lysine in urine. Our results demonstrated that L-lysine could be monitored by the probe, providing new path for construction of CuNCs as fluorescent probes and showing great potential in quantification of L-lysine in real samples. Copyright © 2018 Elsevier B.V. All rights reserved.

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

    International Nuclear Information System (INIS)

    Badugu, Ramachandram; Lakowicz, Joseph R.; Geddes, Chris D.

    2004-01-01

    We characterize six new fluorescent probes that show both intensity and lifetime changes in the presence of free uncomplexed aqueous cyanide, allowing for fluorescence based cyanide sensing up to physiological safeguard levels, i.e. 2 to the anionic R-B - (CN) 3 form, a new cyanide binding mechanism which we have recently reported. The presence of an electron deficient quaternary heterocyclic nitrogen nucleus, and the electron rich cyanide bound form, provides for the intensity changes observed. We have determined the disassociation constants of the probes to be in the range ∼15-84 μM 3 . In addition we have synthesized control compounds which do not contain the boronic acid moiety, allowing for a rationale of the cyanide responses between the probe isomers to be made. The lifetime of the cyanide bound probes are significantly shorter than the free R-B(OH) 2 probe forms, providing for the opportunity of lifetime based cyanide sensing up to physiologically lethal levels. Finally, while fluorescent probes containing the boronic acid moiety have earned a well-deserved reputation for monosaccharide sensing, we show that strong bases such as CN - and OH - preferentially bind as compared to glucose, enabling the potential use of these probes for cyanide safeguard and determination in physiological fluids, especially given that physiologies do not experience any notable changes in pH

  3. LASER FLUORESCENCE EEM PROBE FOR CONE PENETROMETER POLLUTION ANALYSIS

    Science.gov (United States)

    A fiber optic LIF (Laser induced fluorescence) EEM (Excitation emission matrix) instrument for CPT deployment has been successfully developed and field tested. The system employs a Nd: YAG laser and Raman shifter as a rugged field portable excitation source. This excitation sou...

  4. The interaction of fluorescent nanodiamond probes with cellular media

    NARCIS (Netherlands)

    Hemelaar, Simon R; Nagl, Andreas; Bigot, François; Rodriquez Garcia, Melissa; de Vries, Marcel P; Chipaux, Mayeul; Schirhagl, Romana

    Fluorescent nanodiamonds (FNDs) are promising tools to image cells, bioanalytes and physical quantities such as temperature, pressure, and electric or magnetic fields with nanometer resolution. To exploit their potential for intracellular applications, the FNDs have to be brought into contact with

  5. Optical probing of single fluorescent molecules and proteins

    NARCIS (Netherlands)

    Garcia Parajo, M.F.; Veerman, J.A.; Bouwhuis, R.; Bouwhuis, Rudo; van Hulst, N.F.; Vallée, R.A.L.

    2001-01-01

    Single-molecule detection and analysis of organic fluorescent molecules and proteins are presented, with emphasis o­n the underlying principles methodology and the application of single-molecule analysis at room temperature. This Minireview is mainly focused o­n the application of confocal and

  6. A new rhodamine-based fluorescent probe for the discrimination of Fe"3"+ from Fe"2"+

    International Nuclear Information System (INIS)

    You, Qi Hua; Huang, Hua Bin; Zhuang, Zhi Xia; Wang, Xiao Ru; Chan, Wing Hong

    2016-01-01

    A new rhodamine-based fluorescent probe for the discrimination of Fe"3"+ from Fe"2"+ has been designed and investigated. The probe shows an immediate visual color change in response to Fe"3"+ and Cu"2"+, while only Fe"3"+ triggers the fluorescent change of the probe. The existence of large amount of other metal ions shows negligible interference in the detection of Fe"3"+. The association constant K_a_s_s of 4.64 × 10"8 M"-"2 (R"2 = 0.994) and 5.38 × 10"8 M"-"2 (R"2 = 0.991) of the complex was derived from UV/Vis and fluorescence titration assuming 1:2 stoichiometry of probe–Fe"3"+ complex, respectively

  7. Study of protein-probe complexation equilibria and protein-surfactant interaction using charge transfer fluorescence probe methyl ester of N,N-dimethylamino naphthyl acrylic acid

    Energy Technology Data Exchange (ETDEWEB)

    Mahanta, Subrata; Balia Singh, Rupashree; Bagchi, Arnab [Department of Chemistry University of Calcutta 92, A.P.C. Road, Kolkata 700009 (India); Nath, Debnarayan [Department of Physical Chemistry, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 700 032 (India); Guchhait, Nikhil, E-mail: nguchhait@yahoo.co [Department of Chemistry University of Calcutta 92, A.P.C. Road, Kolkata 700009 (India)

    2010-06-15

    In this paper, we demonstrate the interaction between intramolecular charge transfer (ICT) probe-Methyl ester of N,N-dimethylamino naphthyl acrylic acid (MDMANA) with bovine serum albumin (BSA) using absorption and fluorescence emission spectroscopy. The nature of probe protein binding interaction, fluorescence resonance energy transfer from protein to probe and time resolved fluorescence decay measurement predict that the probe molecule binds strongly to the hydrophobic cavity of the protein. Furthermore, the interaction of the anionic surfactant sodium dodecyl sulphate (SDS) with water soluble protein BSA has been investigated using MDMANA as fluorescenece probe. The changes in the spectral characteristics of charge transfer fluorescence probe MDMANA in BSA-SDS environment reflects well the nature of the protein-surfactant binding interaction such as specific binding, non-cooperative binding, cooperative binding and saturation binding.

  8. Lipid domains in giant unilamellar vesicles and their correspondence with equilibrium thermodynamic phases: A quantitative fluorescence microscopy imaging approach

    DEFF Research Database (Denmark)

    Fidorra, Matthias; Garcia, Alejandra; Ipsen, John Hjort

    2009-01-01

    We report a novel analytical procedure to measure the surface areas of coexisting lipid domains in giant unilamellar vesicles (GUVs) based on image processing of 3D fluorescence microscopy data. The procedure involves the segmentation of lipid domains from fluorescent image stacks...

  9. Thermodynamic and fluorescence studies of the underlying factors in benzyl alcohol-induced lipid interdigitated phase.

    Science.gov (United States)

    Chen, C H; Hoye, K; Roth, L G

    1996-09-15

    To further investigate factors contributing to the action of alcohol in the solute-induced lipid interdigitation phase, thermodynamic and fluorescence polarization measurements were carried out to study the interaction of benzyl alcohol with dipalmitoyl phosphatidylcholine bilayer vesicles. The obtained results were compared with those previously reported for ethanol and cyclohexanol (L. G. Roth and C-H. Chen, Arch. Biochem. Biophys. 296, 207, 1992). Similar to ethanol, benzyl alcohol was found to exhibit a biphasic effect on the enthalpy (delta Hm) and the temperature (tm) of the lipid-phase transition and the steady-state fluorescence polarization (P) monitored by 1,6-diphenyl-1,3,5-hexatriene. At a total concentration of benzyl alcohol delta Hm and P, which were correlated with the formation of a lipid interdigitated phase, as evidenced by reported X-ray diffraction data. Combining the results with benzyl alcohol and ethanol suggested that simultaneously large changes in delta Hm and P can be used as an indication of the occurrence of a solute-induced lipid interdigitated phase. The overall interacting force in the formation of this lipid phase, as derived from the interactions of the hydroxyl portion of an alcohol with the lipid phosphate head group and the hydrophobic portion of an alcohol with the lipid hydrocarbon chains, may or may not be dominated by hydrophobic interaction. Although lipid/water partition coefficients and the contribution of hydrophobic interaction to the overall interacting force were comparable between benzyl alcohol and cyclohexanol, benzyl alcohol induced lipid interdigitated phase, but not for cyclohexanol. This was due to the ability of benzyl alcohol to be more effective than cyclohexanol in simultaneously interacting with the phosphate head group and the hydrocarbon chains of lipid.

  10. Pulsed Laser Deposition of Polymers Doped with Fluorescent Probes. Application to Environmental Sensors

    International Nuclear Information System (INIS)

    Rebollar, E; Villavieja, Mm; Gaspard, S; Oujja, M; Corrales, T; Georgiou, S; Domingo, C; Bosch, P; Castillejo, M

    2007-01-01

    Pulsed laser deposition (PLD) has been used to obtain thin films of poly(methyl methacrylate) and polystyrene doped with fluorescent probes, amino aromatic compounds S5 and S6, that could be used to sense the presence of contaminating environmental agents. These dopants both in solution and inserted in polymeric films are sensitive to changes in pH, viscosity and polarity, increasing their fluorescence emission and/or modifying the position of their emission band. Films deposits on quartz substrates, obtained by irradiating targets with a Ti:Sapphire laser (800 nm, 120 fs pulse) were analyzed by optical and Environmental Scanning Electron Microscopy, Fluorescence Microscopy, Laser-Induced Fluorescence, Micro Raman Spectroscopy and Flow Injection Analysis-Mass Spectrometry. The transfer of the polymer and the probe to the substrate is observed to be strongly dependent on the optical absorption coefficient of the polymeric component of the target at the irradiation wavelength

  11. A sensitive fluorescent probe for the polar solvation dynamics at protein-surfactant interfaces.

    Science.gov (United States)

    Singh, Priya; Choudhury, Susobhan; Singha, Subhankar; Jun, Yongwoong; Chakraborty, Sandipan; Sengupta, Jhimli; Das, Ranjan; Ahn, Kyo-Han; Pal, Samir Kumar

    2017-05-17

    Relaxation dynamics at the surface of biologically important macromolecules is important taking into account their functionality in molecular recognition. Over the years it has been shown that the solvation dynamics of a fluorescent probe at biomolecular surfaces and interfaces account for the relaxation dynamics of polar residues and associated water molecules. However, the sensitivity of the dynamics depends largely on the localization and exposure of the probe. For noncovalent fluorescent probes, localization at the region of interest in addition to surface exposure is an added challenge compared to the covalently attached probes at the biological interfaces. Here we have used a synthesized donor-acceptor type dipolar fluorophore, 6-acetyl-(2-((4-hydroxycyclohexyl)(methyl)amino)naphthalene) (ACYMAN), for the investigation of the solvation dynamics of a model protein-surfactant interface. A significant structural rearrangement of a model histone protein (H1) upon interaction with anionic surfactant sodium dodecyl sulphate (SDS) as revealed from the circular dichroism (CD) studies is nicely corroborated in the solvation dynamics of the probe at the interface. The polarization gated fluorescence anisotropy of the probe compared to that at the SDS micellar surface clearly reveals the localization of the probe at the protein-surfactant interface. We have also compared the sensitivity of ACYMAN with other solvation probes including coumarin 500 (C500) and 4-(dicyanomethylene)-2-methyl-6-(p-dimethylamino-styryl)-4H-pyran (DCM). In comparison to ACYMAN, both C500 and DCM fail to probe the interfacial solvation dynamics of a model protein-surfactant interface. While C500 is found to be delocalized from the protein-surfactant interface, DCM becomes destabilized upon the formation of the interface (protein-surfactant complex). The timescales obtained from this novel probe have also been compared with other femtosecond resolved studies and molecular dynamics simulations.

  12. Fluorescent Reporters and Biosensors for Probing the Dynamic Behavior of Protein Kinases

    Directory of Open Access Journals (Sweden)

    Juan A. González-Vera

    2015-11-01

    Full Text Available Probing the dynamic activities of protein kinases in real-time in living cells constitutes a major challenge that requires specific and sensitive tools tailored to meet the particular demands associated with cellular imaging. The development of genetically-encoded and synthetic fluorescent biosensors has provided means of monitoring protein kinase activities in a non-invasive fashion in their native cellular environment with high spatial and temporal resolution. Here, we review existing technologies to probe different dynamic features of protein kinases and discuss limitations where new developments are required to implement more performant tools, in particular with respect to infrared and near-infrared fluorescent probes and strategies which enable improved signal-to-noise ratio and controlled activation of probes.

  13. A new fluorescent pH probe for imaging lysosomes in living cells.

    Science.gov (United States)

    Lv, Hong-Shui; Huang, Shu-Ya; Xu, Yu; Dai, Xi; Miao, Jun-Ying; Zhao, Bao-Xiang

    2014-01-15

    A new rhodamine B-based pH fluorescent probe has been synthesized and characterized. The probe responds to acidic pH with short response time, high selectivity and sensitivity, and exhibits a more than 20-fold increase in fluorescence intensity within the pH range of 7.5-4.1 with the pKa value of 5.72, which is valuable to study acidic organelles in living cells. Also, it has been successfully applied to HeLa cells, for its low cytotoxicity, brilliant photostability, good membrane permeability and no 'alkalizing effect' on lysosomes. The results demonstrate that this probe is a lysosome-specific probe, which can selectively stain lysosomes and monitor lysosomal pH changes in living cells. Copyright © 2013 Elsevier Ltd. All rights reserved.

  14. Probing plasma membrane microdomains in cowpea protoplasts using lipidated GFP-fusion proteins and multimode FRET microscopy

    NARCIS (Netherlands)

    Vermeer, J.E.M.; van Munster, E.B.; Vischer, N.O.; Gadella, T.

    2004-01-01

    Multimode fluorescence resonance energy transfer (FRET) microscopy was applied to study the plasma membrane organization using different lipidated green fluorescent protein (GFP)-fusion proteins co-expressed in cowpea protoplasts. Cyan fluorescent protein (CFP) was fused to the hyper variable region

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

    International Nuclear Information System (INIS)

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

    2008-01-01

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

  16. Gd(iii)-doped carbon dots as a dual fluorescent-MRI probe

    KAUST Repository

    Bourlinos, Athanasios B.; Bakandritsos, Aristides; Kouloumpis, Antonios; Gournis, Dimitrios; Krysmann, Marta; Giannelis, Emmanuel P.; Polakova, Katerina; Safarova, Klara; Hola, Katerina; Zboril, Radek

    2012-01-01

    We describe the synthesis of Gd(iii)-doped carbon dots as dual fluorescence-MRI probes for biomedical applications. The derived Gd(iii)-doped carbon dots show uniform particle size (3-4 nm) and gadolinium distribution and form stable dispersions in water. More importantly, they exhibit bright fluorescence, strong T1-weighted MRI contrast and low cytotoxicity. © The Royal Society of Chemistry 2012.

  17. Ultrasensitive near-infrared fluorescence-enhanced probe for in vivo nitroreductase imaging.

    Science.gov (United States)

    Li, Yuhao; Sun, Yun; Li, Jiachang; Su, Qianqian; Yuan, Wei; Dai, Yu; Han, Chunmiao; Wang, Qiuhong; Feng, Wei; Li, Fuyou

    2015-05-20

    Nitroreductase (NTR) can be overexpressed in hypoxic tumors, thus the selective and efficient detection of NTR is of great importance. To date, although a few optical methods have been reported for the detection of NTR in solution, an effective optical probe for NTR monitoring in vivo is still lacking. Therefore, it is necessary to develop a near-infrared (NIR) fluorescent detection probe for NTR. In this study, five NIR cyanine dyes with fluorescence reporting structure decorated with different nitro aromatic groups, Cy7-1-5, have been designed and explored for possible rapid detection of NTR. Our experimental results presented that only a para-nitro benzoate group modified cyanine probe (Cy7-1) could serve as a rapid NIR fluorescence-enhanced probe for monitoring and bioimaging of NTR. The structure-function relationship has been revealed by theoretical study. The linker connecting the detecting and fluorescence reporting groups and the nitro group position is a key factor for the formation of hydrogen bonds and spatial structure match, inducing the NTR catalytic ability enhancement. The in vitro response and mechanism of the enzyme-catalyzed reduction of Cy7-1 have been investigated through kinetic optical studies and other methods. The results have indicated that an electro-withdrawing group induced electron-transfer process becomes blocked when Cy7-1 is catalytically reduced to Cy7-NH2 by NTR, which is manifested in enhanced fluorescence intensity during the detection process. Confocal fluorescence imaging of hypoxic A549 cells has confirmed the NTR detection ability of Cy7-1 at the cellular level. Importantly, Cy7-1 can detect tumor hypoxia in a murine hypoxic tumor model, showing a rapid and significant enhancement of its NIR fluorescence characteristics suitable for fluorescence bioimaging. This method may potentially be used for tumor hypoxia diagnosis.

  18. A Molecular Probe for the Detection of Polar Lipids in Live Cells.

    Science.gov (United States)

    Bader, Christie A; Shandala, Tetyana; Carter, Elizabeth A; Ivask, Angela; Guinan, Taryn; Hickey, Shane M; Werrett, Melissa V; Wright, Phillip J; Simpson, Peter V; Stagni, Stefano; Voelcker, Nicolas H; Lay, Peter A; Massi, Massimiliano; Plush, Sally E; Brooks, Douglas A

    2016-01-01

    Lipids have an important role in many aspects of cell biology, including membrane architecture/compartment formation, intracellular traffic, signalling, hormone regulation, inflammation, energy storage and metabolism. Lipid biology is therefore integrally involved in major human diseases, including metabolic disorders, neurodegenerative diseases, obesity, heart disease, immune disorders and cancers, which commonly display altered lipid transport and metabolism. However, the investigation of these important cellular processes has been limited by the availability of specific tools to visualise lipids in live cells. Here we describe the potential for ReZolve-L1™ to localise to intracellular compartments containing polar lipids, such as for example sphingomyelin and phosphatidylethanolamine. In live Drosophila fat body tissue from third instar larvae, ReZolve-L1™ interacted mainly with lipid droplets, including the core region of these organelles. The presence of polar lipids in the core of these lipid droplets was confirmed by Raman mapping and while this was consistent with the distribution of ReZolve-L1™ it did not exclude that the molecular probe might be detecting other lipid species. In response to complete starvation conditions, ReZolve-L1™ was detected mainly in Atg8-GFP autophagic compartments, and showed reduced staining in the lipid droplets of fat body cells. The induction of autophagy by Tor inhibition also increased ReZolve-L1™ detection in autophagic compartments, whereas Atg9 knock down impaired autophagosome formation and altered the distribution of ReZolve-L1™. Finally, during Drosophila metamorphosis fat body tissues showed increased ReZolve-L1™ staining in autophagic compartments at two hours post puparium formation, when compared to earlier developmental time points. We concluded that ReZolve-L1™ is a new live cell imaging tool, which can be used as an imaging reagent for the detection of polar lipids in different intracellular

  19. A novel acidic pH fluorescent probe based on a benzothiazole derivative

    Science.gov (United States)

    Ma, Qiujuan; Li, Xian; Feng, Suxiang; Liang, Beibei; Zhou, Tiqiang; Xu, Min; Ma, Zhuoyi

    2017-04-01

    A novel acidic pH fluorescent probe 1 based on a benzothiazole derivative has been designed, synthesized and developed. The linear response range covers the acidic pH range from 3.44 to 6.46, which is valuable for pH researches in acidic environment. The evaluated pKa value of the probe 1 is 4.23. The fluorescence enhancement of the studied probe 1 with an increase in hydrogen ions concentration is based on the hindering of enhanced photo-induced electron transfer (PET) process. Moreover, the pH sensor possesses a highly selective response to H+ in the presence of metal ions, anions and other bioactive small molecules which would be interfere with its fluorescent pH response. Furthermore, the probe 1 responds to acidic pH with short response time that was less than 1 min. The probe 1 has been successfully applied to confocal fluorescence imaging in live HeLa cells and can selectively stain lysosomes. All of such good properties prove it can be used to monitoring pH fluctuations in acidic environment with high sensitivity, pH dependence and short response time.

  20. Selective turn-on fluorescent probes for imaging hydrogen sulfide in living cells.

    Science.gov (United States)

    Montoya, Leticia A; Pluth, Michael D

    2012-05-16

    Hydrogen sulfide (H(2)S) is an important biological messenger but few biologically-compatible methods are available for its detection. Here we report two bright fluorescent probes that are selective for H(2)S over cysteine, glutathione and other reactive sulfur, nitrogen, and oxygen species. Both probes are demonstrated to detect H(2)S in live cells. This journal is © The Royal Society of Chemistry 2012

  1. Lipid nanoparticle vectorization of indocyanine green improves fluorescence imaging for tumor diagnosis and lymph node resection.

    Science.gov (United States)

    Navarro, Fabrice P; Berger, Michel; Guillermet, Stéphanie; Josserand, Véronique; Guyon, Laurent; Neumann, Emmanuelle; Vinet, Françoise; Texier, Isabelle

    2012-10-01

    Fluorescence imaging is opening a new era in image-guided surgery and other medical applications. The only FDA approved contrast agent in the near infrared is IndoCyanine Green (ICG), which despites its low toxicity, displays poor chemical and optical properties for long-term and sensitive imaging applications in human. Lipid nanoparticles are investigated for improving ICG optical properties and in vivo fluorescence imaging sensitivity. 30 nm diameter lipid nanoparticles (LNP) are loaded with ICG. Their characterization and use for tumor and lymph node imaging are described. Nano-formulation benefits dye optical properties (6 times improved brightness) and chemical stability (>6 months at 4 degrees C in aqueous buffer). More importantly, LNP vectorization allows never reported sensitive and prolonged (>1 day) labeling of tumors and lymph nodes. Composed of human-use approved ingredients, this novel ICG nanometric formulation is foreseen to expand rapidly the field of clinical fluorescence imaging applications.

  2. Enhancement of growth and lipid production from microalgae using fluorescent paint under the solar radiation.

    Science.gov (United States)

    Seo, Yeong Hwan; Cho, Changsoon; Lee, Jung-Yong; Han, Jong-In

    2014-12-01

    Solar radiation has intensity that is too high to inhibit microalgae activity and is composed of wide light spectrum including ultraviolet (UV) range which cannot be utilized for microalgae. For these reasons, the modification of solar radiation is required for effective microalgae cultivation, and to do that, fluorescent paint was used for not only blocking excessive solar energy but also converting UV to visible light. With fluorescent aqueous layer, microalgae was protected from photoinhibition and could grow well, but there was difference in growth and lipid accumulation efficiencies depending on the color; maximum dry weight of 1.7 g/L was achieved in red paint, whereas best lipid content of 30% was obtained in blue one. This phenomenon was due to the different light spectrum made by colors. With simple process using fluorescent paint, modification of light was successfully done and allowing microalgae to grow under strong radiation such as solar radiation. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Studies of radiation induced membrane damage in lymphocytes using fluorescent probes

    International Nuclear Information System (INIS)

    Nikesch, W.

    1974-01-01

    The fluorescent probes perylene (PER), 1-anilino-8-naphthalene sulfonic acid (ANS), and fluorescein diacetate (FDA) were used to investigate membrane changes caused by ionizing radiation. Probe response to various other perturbations (variation of pH, temperature, and salt concentration, and treatment with phythohemagglutinin (PHA) and saponins) was also investigated to better understand membrane-probe interactions. ANS was used to probe the membrane surface, PER to probe the membrane interior, and FDA to investigate membrane integrity. Polarization of fluorescent light from ANS and PER was used to investigate the microviscosity and order of the membrane surface and interior respectively. Irradiated cells (600 R) were shown to have a decreased rate of hydrolysis of FDA probably due to cytoplasmic changes effecting the enzymatic reaction. Also evident was an increase in loss of intracellular fluorescein and a decrease in PER polarization indicating that the cells have a decreased membrane integrity, possibly the result of an increased disorganization of the phospholipid hydrocarbon chains in the membrane interior. Experiments with PHA link the decreased membrane integrity with the eventual interphase death of the cells. In general it is shown that the fluorescent probes ANS, PER, and FDA provide useful ways to investigate order and microviscosity in the cell membrane surface and interior, membrane surface charges, internal membrane polarity changes, and membrane integrity. (U.S.)

  4. An excited-state intramolecular photon transfer fluorescence probe for localizable live cell imaging of cysteine

    Science.gov (United States)

    Liu, Wei; Chen, Wen; Liu, Si-Jia; Jiang, Jian-Hui

    2017-03-01

    Small molecule probes suitable for selective and specific fluorescence imaging of some important but low-concentration intracellular reactive sulfur species such as cysteine (Cys) pose a challenge in chemical biology. We present a readily available, fast-response fluorescence probe CHCQ-Ac, with 2-(5‧-chloro-2-hydroxyl-phenyl)-6-chloro-4(3 H)-quinazolinone (CHCQ) as the fluorophore and acrylate group as the functional moiety, that enables high-selectivity and high-sensitivity for detecting Cys in both solution and biological system. After specifically reacted with Cys, the probe undergoes a seven-membered intramolecular cyclization and released the fluorophore CHCQ with excited-state intramolecular photon transfer effect. A highly fluorescent, insoluble aggregate was then formed to facilitate high-sensitivity and high-resolution imaging. The results showed that probe CHCQ-Ac affords a remarkably large Stokes shift and can detect Cys under physiological pH condition with no interference from other analytes. Moreover, this probe was proved to have excellent chemical stability, low cytotoxicity and good cell permeability. Our design of this probe provides a novel potential tool to visualize and localize cysteine in bioimaging of live cells that would greatly help to explore various Cys-related physiological and pathological cellular processes in cell biology and diagnostics.

  5. Poly(o-phenylenediamine) colloid-quenched fluorescent oligonucleotide as a probe for fluorescence-enhanced nucleic acid detection.

    Science.gov (United States)

    Tian, Jingqi; Li, Hailong; Luo, Yonglan; Wang, Lei; Zhang, Yingwei; Sun, Xuping

    2011-02-01

    In this Letter, we demonstrate that chemical oxidation polymerization of o-phenylenediamine (OPD) by potassium bichromate at room temperature results in the formation of submicrometer-scale poly(o-phenylenediamine) (POPD) colloids. Such colloids can absorb and quench dye-labeled single-stranded DNA (ssDNA) very effectively. In the presence of a target, a hybridization event occurs, which produces a double-stranded DNA (dsDNA) that detaches from the POPD surface, leading to recovery of dye fluorescence. With the use of an oligonucleotide (OND) sequence associated with human immunodeficiency virus (HIV) as a model system, we demonstrate the proof of concept that POPD colloid-quenched fluorescent OND can be used as a probe for fluorescence-enhanced nucleic acid detection with selectivity down to single-base mismatch.

  6. Fluorescent Bisphosphonate and Carboxyphosphonate Probes: A Versatile Imaging Toolkit for Applications in Bone Biology and Biomedicine.

    Science.gov (United States)

    Sun, Shuting; Błażewska, Katarzyna M; Kadina, Anastasia P; Kashemirov, Boris A; Duan, Xuchen; Triffitt, James T; Dunford, James E; Russell, R Graham G; Ebetino, Frank H; Roelofs, Anke J; Coxon, Fraser P; Lundy, Mark W; McKenna, Charles E

    2016-02-17

    A bone imaging toolkit of 21 fluorescent probes with variable spectroscopic properties, bone mineral binding affinities, and antiprenylation activities has been created, including a novel linking strategy. The linking chemistry allows attachment of a diverse selection of dyes fluorescent in the visible to near-infrared range to any of the three clinically important heterocyclic bisphosphonate bone drugs (risedronate, zoledronate, and minodronate or their analogues). The resultant suite of conjugates offers multiple options to "mix and match" parent drug structure, fluorescence emission wavelength, relative bone affinity, and presence or absence of antiprenylation activity, for bone-related imaging applications.

  7. Terbium fluorescence as a sensitive, inexpensive probe for UV-induced damage in nucleic acids

    International Nuclear Information System (INIS)

    El-Yazbi, Amira F.; Loppnow, Glen R.

    2013-01-01

    Graphical abstract: -- Highlights: •Simple, inexpensive, mix-and-read assay for positive detection of DNA damage. •Recognition of undamaged DNA via hybridization to a hairpin probe. •Terbium(III) fluorescence reports the amount of damage by binding to ssDNA. •Tb/hairpin is a highly selective and sensitive fluorescent probe for DNA damage. -- Abstract: Much effort has been focused on developing methods for detecting damaged nucleic acids. However, almost all of the proposed methods consist of multi-step procedures, are limited, require expensive instruments, or suffer from a high level of interferences. In this paper, we present a novel simple, inexpensive, mix-and-read assay that is generally applicable to nucleic acid damage and uses the enhanced luminescence due to energy transfer from nucleic acids to terbium(III) (Tb 3+ ). Single-stranded oligonucleotides greatly enhance the Tb 3+ emission, but duplex DNA does not. With the use of a DNA hairpin probe complementary to the oligonucleotide of interest, the Tb 3+ /hairpin probe is applied to detect ultraviolet (UV)-induced DNA damage. The hairpin probe hybridizes only with the undamaged DNA. However, the damaged DNA remains single-stranded and enhances the intrinsic fluorescence of Tb 3+ , producing a detectable signal directly proportional to the amount of DNA damage. This allows the Tb 3+ /hairpin probe to be used for sensitive quantification of UV-induced DNA damage. The Tb 3+ /hairpin probe showed superior selectivity to DNA damage compared to conventional molecular beacons probes (MBs) and its sensitivity is more than 2.5 times higher than MBs with a limit of detection of 4.36 ± 1.2 nM. In addition, this probe is easier to synthesize and more than eight times cheaper than MBs, which makes its use recommended for high-throughput, quantitative analysis of DNA damage

  8. Probing phosphate ion via the europium(III)-modulated fluorescence of gold nanoclusters

    International Nuclear Information System (INIS)

    Ding, Shou-Nian; Li, Chun-Mei; Gao, Bu-Hong; Kargbo, Osman; Zhou, Chan; Chen, Xi; Wan, Neng

    2014-01-01

    Fluorescent gold nanoclusters (Au-NCs) were synthesized by a one-pot method using 11-mercaptoundecanoic acid as a reducing and capping reagent. It is found that the red fluorescence of the Au-NCs is quenched by the introduction of Eu(III) at pH 7.0, but that fluorescence is restored on addition of phosphate. The Au-NCs were investigated by transmission electron microscopy and fluorescence photographs. The effect of pH on fluorescence was studied in the range from pH 6 to 10 and is found to be strong. Based on these findings, we have developed an assay for phosphate. Ions such as citrate, Fe(CN) 6 3− , SO 4 2− , S 2 O 8 2− , Cl − , HS − , Br − , AcO − , NO 2 − , SCN − , ClO 4 − , HCO 3 − , NO 3 − , Cd 2+ , Ba 2+ , Zn 2+ , Mg 2+ , and glutamate do not interfere, but ascorbate and Fe 3+ can quench Au-NCs fluorescence. The fluorescent nanocluster probe responds to phosphate in the range from 0.18 to 250 μM, and the detection limit is 180 nM. The probe also responds to pyrophosphate and ATP. (author)

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

    Science.gov (United States)

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

    2016-01-01

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

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

    International Nuclear Information System (INIS)

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

    1989-01-01

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

  11. Fluorescent porous silicon biological probes with high quantum efficiency and stability.

    Science.gov (United States)

    Tu, Chang-Ching; Chou, Ying-Nien; Hung, Hsiang-Chieh; Wu, Jingda; Jiang, Shaoyi; Lin, Lih Y

    2014-12-01

    We demonstrate porous silicon biological probes as a stable and non-toxic alternative to organic dyes or cadmium-containing quantum dots for imaging and sensing applications. The fluorescent silicon quantum dots which are embedded on the porous silicon surface are passivated with carboxyl-terminated ligands through stable Si-C covalent bonds. The porous silicon bio-probes have shown photoluminescence quantum yield around 50% under near-UV excitation, with high photochemical and thermal stability. The bio-probes can be efficiently conjugated with antibodies, which is confirmed by a standard enzyme-linked immunosorbent assay (ELISA) method.

  12. Characterization of the vitreous body of the human eye using a cyanine dye as a spectral and fluorescent probe

    Science.gov (United States)

    Panova, Ina G.; Tatikolov, Alexander S.

    2009-02-01

    We used one of cyanine dyes as a spectral and fluorescent probe in the study of the composition of the extracellular matrix of the human eye (its vitreous body). Owing to the unique ability of the dye to bind to collagens and human serum albumin, we revealed the simultaneous presence of both types of biomacromolecules in the vitreous body. The formation of the dye complex with human serum albumin leads to appearance of a long-wavelength absorption band (~612 nm) and a steep rise of fluorescence, whereas in the presence of collagens the dye forms J-aggregates with a longer-wavelength absorption band (640-660 nm) and moderate fluorescence. In this work we studied the composition of the human fetus vitreous body and its dynamics from 9 to 31 gestation weeks. On the basis of the data obtained by this method, we may assume that albumin, being a carrier protein, probably provides the vitreous body and surrounding tissues with necessary growth factors, hormones, lipids, vitamins, and some other biomolecules. The data show that the dye is promising not only for study of albumin functions in eye development, but also for characterization of some eye diseases and for analysis of other extracellular media.

  13. Mesoporous silica for drug delivery: Interactions with model fluorescent lipid vesicles and live cells.

    Science.gov (United States)

    Bardhan, Munmun; Majumdar, Anupa; Jana, Sayantan; Ghosh, Tapas; Pal, Uttam; Swarnakar, Snehasikta; Senapati, Dulal

    2018-01-01

    Formulated mesoporous silica nanoparticle (MSN) systems offer the best possible drug delivery system through the release of drug molecules from the accessible pores. In the present investigation, steady state and time resolved fluorescence techniques along with the fluorescence imaging were applied to investigate the interactions of dye loaded MSN with fluorescent unilamellar vesicles and live cells. Here 1,2-dimyristoyl-sn-glycero-3-phospocholine (DMPC) was used to prepare Small Unilamellar Vesicles (SUVs) as the model membrane with fluorescent 1,6-diphenyl-1,3,5-hexatriene (DPH) molecule incorporated inside the lipid bilayer. The interaction of DPH incorporated DMPC membrane with Fluorescein loaded MSN lead to the release of Fluorescein (Fl) dye from the interior pores of MSN systems. The extent of release of Fl and spatial distribution of the DPH molecule has been explored by monitoring steady-state fluorescence intensity and fluorescence lifetime at physiological condition. To investigate the fate of drug molecule released from MSN, fluorescence anisotropy has been used. The drug delivery efficiency of the MSN as a carrier for doxorubicin (DOX), a fluorescent chemotherapeutic drug, has also been investigated at physiological conditions. The study gives a definite confirmation for high uptake and steady release of DOX in primary oral mucosal non-keratinized squamous cells in comparison to naked DOX treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Photobleaching kinetics and time-integrated emission of fluorescent probes in cellular membranes

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Christensen, Tanja; Solanko, Lukasz Michal

    2014-01-01

    Since the pioneering work of Hirschfeld, it is known that time-integrated emission (TiEm) of a fluorophore is independent of fluorescence quantum yield and illumination intensity. Practical implementation of this important result for determining exact probe distribution in living cells is often h...

  15. A subnanomolar fluorescent probe for protein kinase CK2 interaction studies

    DEFF Research Database (Denmark)

    Enkvist, Erki; Viht, Kaido; Bischoff, Nils

    2012-01-01

    of the functions of CK2 could be facilitated by the application of small-molecule fluorescent probes that bind to the active site of the enzyme with high affinity and selectivity. We have used a bisubstrate approach for the development of a highly potent inhibitor of CK2. 4,5,6,7-Tetrabromo-1H-benzimidazole...

  16. Synthesis and application of a highly selective copper ions fluorescent probe based on the coumarin group

    Science.gov (United States)

    He, Guangjie; Liu, Xiangli; Xu, Jinhe; Ji, Liguo; Yang, Linlin; Fan, Aiying; Wang, Songjun; Wang, Qingzhi

    2018-02-01

    A highly selective copper ions fluorescent probe based on the coumarin-type Schiff base derivative 1 (probe) was produced by condensation reaction between coumarin carbohydrazide and 1H-indazole-3-carbaldehyde. The UV-vis spectroscopy showed that the maximum absorption peak of compound 1 appeared at 439 nm. In the presence of Cu2 + ions, the maximum peak decreased remarkably compared with other physiological important metal ions and a new absorption peak at 500 nm appeared. The job's plot experiments showed that complexes of 1:2 binding mode were formed in CH3CN:HEPES (3:2, v/v) solution. Compound 1 exhibited a strong blue fluorescence. Upon addition of copper ions, the fluorescence gradually decreased and reached a plateau with the fluorescence quenching rate up to 98.73%. The detection limit for Cu2 + ions was estimated to 0.384 ppm. Fluorescent microscopy experiments demonstrated that probe 1 had potential to be used to investigate biological processes involving Cu2 + ions within living cells.

  17. Molecular engineering of two-photon fluorescent probes for bioimaging applications

    Science.gov (United States)

    Liu, Hong-Wen; Liu, Yongchao; Wang, Peng; Zhang, Xiao-Bing

    2017-03-01

    During the past two decades, two-photon microscopy (TPM), which utilizes two near-infrared photons as the excitation source, has emerged as a novel, attractive imaging tool for biological research. Compared with one-photon microscopy, TPM offers several advantages, such as lowering background fluorescence in living cells and tissues, reducing photodamage to biosamples, and a photobleaching phenomenon, offering better 3D spatial localization, and increasing penetration depth. Small-molecule-based two-photon fluorescent probes have been well developed for the detection and imaging of various analytes in biological systems. In this review, we will give a general introduction of molecular engineering of two-photon fluorescent probes based on different fluorescence response mechanisms for bioimaging applications during the past decade. Inspired by the desired advantages of small-molecule two-photon fluorescent probes in biological imaging applications, we expect that more attention will be devoted to the development of new two-photon fluorophores and applications of TPM in areas of bioanalysis and disease diagnosis.

  18. Chromenoquinoline-based thiol probes: a study on the quencher position for controlling fluorescent Off-On characteristics.

    Science.gov (United States)

    Kand, Dnyaneshwar; Kalle, Arunasree Marasanapalli; Talukdar, Pinaki

    2013-02-13

    The design, synthesis and thiol sensing ability of chromenoquinoline-based fluorescent probes 4, 5 and 6 and are reported here. The relative position of the maleimide moiety was varied along the chromenoquinoline fluorophore to decrease the background fluorescence. Lower background fluorescence in probes 4 and 6 was rationalized by the smaller k(r)/k(nr) values compared to that of probe 5. An intramolecular charge transfer (ICT) mechanism was proposed for quenching and the extent was dependent on the position of the maleimide quencher. Fluorescent Off-On characteristics were evaluated by theoretical calculations. All probes were selective only towards thiol containing amino acids. Thiol sensing by probes 4 and 6 were much better compared to 5. Probe 4 displayed a better fluorescence response for less hindered thiol (185-, 223- and 156-fold for Hcy, Cys and GSH, respectively), while for probe 6, a higher enhancement in fluorescence was observed with more hindered thiols (180-, 205- and 245-fold for Hcy, Cys and GSH, respectively). The better response to bulkier thiol, GSH by probe 6 was attributed to the steric crowding at the C-4 position and bulkiness of the GSH group which force the succinimide unit to be in a nearly orthogonal conformation. This spatial arrangement was important in reducing the fluorescence quenching ability of the succinimide moiety. The application of probes 4, 5 and 6 was demonstrated by naked eye detection thiols using a 96-well plate system as well as by live-cell imaging.

  19. Mass amplifying probe for sensitive fluorescence anisotropy detection of small molecules in complex biological samples.

    Science.gov (United States)

    Cui, Liang; Zou, Yuan; Lin, Ninghang; Zhu, Zhi; Jenkins, Gareth; Yang, Chaoyong James

    2012-07-03

    Fluorescence anisotropy (FA) is a reliable and excellent choice for fluorescence sensing. One of the key factors influencing the FA value for any molecule is the molar mass of the molecule being measured. As a result, the FA method with functional nucleic acid aptamers has been limited to macromolecules such as proteins and is generally not applicable for the analysis of small molecules because their molecular masses are relatively too small to produce observable FA value changes. We report here a molecular mass amplifying strategy to construct anisotropy aptamer probes for small molecules. The probe is designed in such a way that only when a target molecule binds to the probe does it activate its binding ability to an anisotropy amplifier (a high molecular mass molecule such as protein), thus significantly increasing the molecular mass and FA value of the probe/target complex. Specifically, a mass amplifying probe (MAP) consists of a targeting aptamer domain against a target molecule and molecular mass amplifying aptamer domain for the amplifier protein. The probe is initially rendered inactive by a small blocking strand partially complementary to both target aptamer and amplifier protein aptamer so that the mass amplifying aptamer domain would not bind to the amplifier protein unless the probe has been activated by the target. In this way, we prepared two probes that constitute a target (ATP and cocaine respectively) aptamer, a thrombin (as the mass amplifier) aptamer, and a fluorophore. Both probes worked well against their corresponding small molecule targets, and the detection limits for ATP and cocaine were 0.5 μM and 0.8 μM, respectively. More importantly, because FA is less affected by environmental interferences, ATP in cell media and cocaine in urine were directly detected without any tedious sample pretreatment. Our results established that our molecular mass amplifying strategy can be used to design aptamer probes for rapid, sensitive, and selective

  20. Organization of fluorescent cholesterol analogs in lipid bilayers - lessons from cyclodextrin extraction.

    Science.gov (United States)

    Milles, Sigrid; Meyer, Thomas; Scheidt, Holger A; Schwarzer, Roland; Thomas, Lars; Marek, Magdalena; Szente, Lajos; Bittman, Robert; Herrmann, Andreas; Günther Pomorski, Thomas; Huster, Daniel; Müller, Peter

    2013-08-01

    To characterize the structure and dynamics of cholesterol in membranes, fluorescent analogs of the native molecule have widely been employed. The cholesterol content in membranes is in general manipulated by using water-soluble cyclodextrins. Since the interactions between cyclodextrins and fluorescent-labeled cholesterol have not been investigated in detail so far, we have compared the cyclodextrin-mediated membrane extraction of three different fluorescent cholesterol analogs (one bearing a NBD and two bearing BODIPY moieties). Extraction of these analogs was followed by measuring the Förster resonance energy transfer between a rhodamine moiety linked to phosphatidylethanolamine and the labeled cholesterol. The extraction kinetics revealed that the analogs are differently extracted from membranes. We examined the orientation of the analogs within the membrane and their influence on lipid condensation using NMR and EPR spectroscopies. Our data indicate that the extraction of fluorescent sterols from membranes is determined by several parameters, including their impact on lipid order, their hydrophobicity, their intermolecular interactions with surrounding lipids, their orientation within the bilayer, and their affinity with the exogenous acceptor. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. A Pyridazine-Based Fluorescent Probe Targeting Aβ Plaques in Alzheimer’s Disease

    Directory of Open Access Journals (Sweden)

    Yong Dae Park

    2018-01-01

    Full Text Available Accumulation of β-amyloid (Aβ plaques comprising Aβ40 and Aβ42 in the brain is the most significant factor in the pathogenesis of Alzheimer’s disease (AD. Thus, the detection of Aβ plaques has increasingly attracted interest in the context of AD diagnosis. In the present study, a fluorescent pyridazine-based dye that can detect and image Aβ plaques was designed and synthesized, and its optical properties in the presence of Aβ aggregates were evaluated. An approximately 34-fold increase in emission intensity was exhibited by the fluorescent probe after binding with Aβ aggregates, for which it showed high affinity (KD = 0.35 µM. Moreover, the reasonable hydrophobic properties of the probe (log P = 2.94 allow it to penetrate the blood brain barrier (BBB. In addition, the pyridazine-based probe was used in the histological costaining of transgenic mouse (APP/PS1 brain sections to validate the selective binding of the probe to Aβ plaques. The results suggest that the pyridazine-based compound has the potential to serve as a fluorescent probe for the diagnosis of AD.

  2. Detection of Dysplastic Intestinal Adenomas Using a Fluorescent Folate Imaging Probe

    Directory of Open Access Journals (Sweden)

    Wei-Tsung Chen

    2005-01-01

    Full Text Available Macrophages have long been recognized as a prominent component of tumors. Activated macrophages overexpress folate receptors and we used this phenomenon to image inflammatory reactions in colon dysplasia using a fluorescent folate probe (FFP. APCΔ468 mice injected with FFP showed fluorescent adenomas (target-to-background ratio, adenoma vs. adjacent normal mucosa, of 2.46 ± 0.41, significantly higher (p < .001 than adenomas in animals injected with a non-folate-containing control probe. Fluorescence-activated cell-sorting analysis revealed a 3-fold higher content of Mac1-positive cells in colonic adenomas compared with normal adjacent mucosa (6.8% vs. 2.2%, and confirmed the source of FFP-positive cells to be primarily an F4/80-positive macrophage subpopulation. Taken together, these results indicate that FFP potentially can be used to image dysplastic intestinal adenomas in vivo.

  3. Folate receptor targeting silica nanoparticle probe for two-photon fluorescence bioimaging

    Science.gov (United States)

    Wang, Xuhua; Yao, Sheng; Ahn, Hyo-Yang; Zhang, Yuanwei; Bondar, Mykhailo V.; Torres, Joseph A.; Belfield, Kevin D.

    2010-01-01

    Narrow dispersity organically modified silica nanoparticles (SiNPs), diameter ~30 nm, entrapping a hydrophobic two-photon absorbing fluorenyl dye, were synthesized by hydrolysis of triethoxyvinylsilane and (3-aminopropyl)triethoxysilane in the nonpolar core of Aerosol-OT micelles. The surface of the SiNPs were functionalized with folic acid, to specifically deliver the probe to folate receptor (FR) over-expressing Hela cells, making these folate two-photon dye-doped SiNPs potential candidates as probes for two-photon fluorescence microscopy (2PFM) bioimaging. In vitro studies using FR over-expressing Hela cells and low FR expressing MG63 cells demonstrated specific cellular uptake of the functionalized nanoparticles. One-photon fluorescence microscopy (1PFM) imaging, 2PFM imaging, and two-photon fluorescence lifetime microscopy (2P-FLIM) imaging of Hela cells incubated with folate-modified two-photon dye-doped SiNPs were demonstrated. PMID:21258480

  4. An easily Prepared Fluorescent pH Probe Based on Dansyl.

    Science.gov (United States)

    Sha, Chunming; Chen, Yuhua; Chen, Yufen; Xu, Dongmei

    2016-09-01

    A novel fluorescent pH probe from dansyl chloride and thiosemicarbazide was easily prepared and fully characterized by (1)H NMR, (13)C NMR, LC-MS, Infrared spectra and elemental analysis. The probe exhibited high selectivity and sensitivity to H(+) with a pK a value of 4.98. The fluorescence intensity at 510 nm quenched 99.5 % when the pH dropped from 10.88 to 1.98. In addition, the dansyl-based probe could respond quickly and reversibly to the pH variation and various common metal ions showed negligible interference. The recognition could be ascribed to the intramolecular charge transfer caused by the protonation of the nitrogen in the dimethylamino group.

  5. Sensitive and Selective Ratiometric Fluorescence Probes for Detection of Intracellular Endogenous Monoamine Oxidase A.

    Science.gov (United States)

    Wu, Xiaofeng; Li, Lihong; Shi, Wen; Gong, Qiuyu; Li, Xiaohua; Ma, Huimin

    2016-01-19

    Monoamine oxidase A (MAO-A) is known to widely exist in most cell lines in the body, and its dysfunction (unusually high or low levels of MAO-A) is thought to be responsible for several psychiatric and neurological disorders. Thus, a sensitive and selective method for evaluating the relative MAO-A levels in different live cells is urgently needed to better understand the function of MAO-A, but to our knowledge such a method is still lacking. Herein, we rationally design two new ratiometric fluorescence probes (1 and 2) that can sensitively and selectively detect MAO-A. The probes are constructed by incorporating a recognition group of propylamine into the fluorescent skeleton of 1,8-naphthalimide, and the detection mechanism is based on amine oxidation and β-elimination to release the fluorophore (4-hydroxy-N-butyl-1,8-naphthalimide), which is verified by HPLC analysis. Reaction of the probes with MAO-A produces a remarkable fluorescence change from blue to green, and the ratio of fluorescence intensity at 550 and 454 nm is directly proportional to the concentration of MAO-A in the ranges of 0.5-1.5 and 0.5-2.5 μg/mL with detection limits of 1.1 and 10 ng/mL (k = 3) for probes 1 and 2, respectively. Surprisingly, these probes show strong fluorescence responses to MAO-A but almost none to MAO-B (one of two isoforms of MAO), indicating superior ability to distinguish MAO-A from MAO-B. The high specificity of the probes for MAO-A over MAO-B is further supported by different inhibitor experiments. Moreover, probe 1 displays higher sensitivity than probe 2 and is thus investigated to image the relative MAO-A levels in different live cells, such as HeLa and NIH-3T3 cells. It is found that the concentration of endogenous MAO-A in HeLa cells is approximately 1.8 times higher than that in NIH-3T3 cells, which is validated by the result from an ELISA kit. Additionally, the proposed probes may find more uses in the specific detection of MAO-A between the two isoforms of MAO

  6. In Vivo Fluorescence Lifetime Imaging Monitors Binding of Specific Probes to Cancer Biomarkers

    Science.gov (United States)

    Ardeshirpour, Yasaman; Chernomordik, Victor; Zielinski, Rafal; Capala, Jacek; Griffiths, Gary; Vasalatiy, Olga; Smirnov, Aleksandr V.; Knutson, Jay R.; Lyakhov, Ilya; Achilefu, Samuel; Gandjbakhche, Amir; Hassan, Moinuddin

    2012-01-01

    One of the most important factors in choosing a treatment strategy for cancer is characterization of biomarkers in cancer cells. Particularly, recent advances in Monoclonal Antibodies (MAB) as primary-specific drugs targeting tumor receptors show that their efficacy depends strongly on characterization of tumor biomarkers. Assessment of their status in individual patients would facilitate selection of an optimal treatment strategy, and the continuous monitoring of those biomarkers and their binding process to the therapy would provide a means for early evaluation of the efficacy of therapeutic intervention. In this study we have demonstrated for the first time in live animals that the fluorescence lifetime can be used to detect the binding of targeted optical probes to the extracellular receptors on tumor cells in vivo. The rationale was that fluorescence lifetime of a specific probe is sensitive to local environment and/or affinity to other molecules. We attached Near-InfraRed (NIR) fluorescent probes to Human Epidermal Growth Factor 2 (HER2/neu)-specific Affibody molecules and used our time-resolved optical system to compare the fluorescence lifetime of the optical probes that were bound and unbound to tumor cells in live mice. Our results show that the fluorescence lifetime changes in our model system delineate HER2 receptor bound from the unbound probe in vivo. Thus, this method is useful as a specific marker of the receptor binding process, which can open a new paradigm in the “image and treat” concept, especially for early evaluation of the efficacy of the therapy. PMID:22384092

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

    Directory of Open Access Journals (Sweden)

    Yasaman Ardeshirpour

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

  8. Quantitative generalized ratiometric fluorescence spectroscopy for turbid media based on probe encapsulated by biologically localized embedding

    International Nuclear Information System (INIS)

    Yan, Xiu-Fang; Chen, Zeng-Ping; Cui, Yin-Yin; Hu, Yuan-Liang; Yu, Ru-Qin

    2016-01-01

    PEBBLE (probe encapsulated by biologically localized embedding) nanosensor encapsulating an intensity-based fluorescence indicator and an inert reference fluorescence dye inside the pores of stable matrix can be used as a generalized wavelength-ratiometric probe. However, the lack of an efficient quantitative model render the choices of inert reference dyes and intensity-based fluorescence indicators used in PEBBLEs based generalized wavelength-ratiometric probes rather limited. In this contribution, an extended quantitative fluorescence model was derived specifically for generalized wavelength-ratiometric probes based on PEBBLE technique (QFM GRP ) with a view to simplify the design of PEBBLEs and hence further extend their application potentials. The effectiveness of QFM GRP has been tested on the quantitative determination of free Ca 2+ in both simulated and real turbid media using a Ca 2+ sensitive PEBBLE nanosensor encapsulating Rhod-2 and eosin B inside the micropores of stable polyacrylamide matrix. Experimental results demonstrated that QFM GRP could realize precise and accurate quantification of free Ca 2+ in turbid samples, even though there is serious overlapping between the fluorescence excitation peaks of eosin B and Ca 2+ bound Rhod-2. The average relative predictive error value of QFM GRP for the test simulated turbid samples was 5.9%, about 2–4 times lower than the corresponding values of partial least squares calibration model and the empirical ratiometric model based on the ratio of fluorescence intensities at the excitation peaks of Ca 2+ bound Rhod-2 and eosin B. The recovery rates of QFM GRP for the real and spiked turbid samples varied from 93.1% to 101%, comparable to the corresponding results of atomic absorption spectrometry. - Highlights: • An advanced model was derived for generalized wavelength-ratiometric PEBBLEs. • The model can simplify the design of generalized wavelength-ratiometric PEBBLEs. • The model realized accurate

  9. Bodipy-FL-Verapamil: A Fluorescent Probe for the Study of Multidrug Resistance Proteins

    Directory of Open Access Journals (Sweden)

    Anna Rosati

    2004-01-01

    Full Text Available Most of the substances used as fluorescent probes to study drug transport and the effect of efflux blockers in multidrug resistant cells have many drawbacks, such as toxicity, unspecific background, accumulation in mitochondria. New fluorescent compounds, among which Bodipy‐FL‐verapamil (BV, have been therefore proposed as more useful tools. The uptake of BV has been evaluated by cytofluorimetry and fluorescence microscopy using cell lines that overexpress P‐glycoprotein (P388/ADR and LLC‐PK1/ADR or MRP (multidrug resistance‐related protein (PANC‐1 and clinical specimens from patients. The effect of specific inhibitors for P‐glycoprotein (verapamil and vinblastine or MRP (MK571 and probenecid has been also studied. BV intracellular concentrations were significantly lower in the two P‐glycoprotein overexpressing cell lines in comparison with the parental lines. In addition, verapamil and vinblastine increased the intracellular concentrations of the dye; MK571 and probenecid, two MRP inhibitors, increased BV levels in PANC‐1 cells, that express this protein. These findings were confirmed in clinical specimens from patients. Fluorescence microscopy revealed a faint fluorescence emission in P‐glycoprotein or MRP expressing cell lines; however, treatment with specific inhibitors significantly increased the fluorescence. BV is a useful tool for studying multidrug resistance proteins with different techniques such as cytofluorimetry and fluorescence microscopy, but does not discriminate between P‐glycoprotein and MRP. In comparison with other classic fluorescent probes, the assay with this dye is extremely rapid, simple, not toxic for cells, devoid of fluorescent background, and can be useful in the clinical settings.

  10. Two sugar-rhodamine "turn-on" fluorescent probes for the selective detection of Fe3 +

    Science.gov (United States)

    Chen, Qing; Fang, Zhijie

    2018-03-01

    Two new sugar-rhodamine fluorescent probes (RDG1 and RDG2) have been synthesized and characterized by 1H NMR, 13C NMR and HRMS. Their UV-Vis, fluorescence spectra and fluorescence-response to Fe3 + are investigated and discussed. RDG1 had a very nice linear relationship between UV absorbance and Fe3 + concentration with the correlation coefficient as high as 0.997 and the detection limit is 3.46 × 10- 6 M. Upon the addition of Fe3 +, the spirolactam ring of RDG1 was opened and a 1:1 metal ligand complex was formed from Job's plot. The results showed that RDG1 can be used as an effective fluorescent probe for selective detection of Fe3 + in water. RDG2 was incorporated the well-known rhodamine group and a water-soluble D-glucose group within one molecule and can be used for detecting Fe3 + in natural water as a selective fluorescent sensor. The addition of Fe3 + into RDG2 resulted in a strongly enhanced fluorescence as well as color change of solution from colorless to pink. Job's plot of RDG2 indicated 1:1 stoichiometry of RDG2-Fe3 +. RDG2 can serve as a probe for Fe3 + between pH = 4.0 to 7.0 and it's detection limit is 2.09 × 10- 6 M. The OFF-ON fluorescent mechanisms of RDG1-Fe3 + and RDG2-Fe3 + are proposed.

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

    Science.gov (United States)

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

    2018-05-01

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

  12. The interaction of fluorescent nanodiamond probes with cellular media.

    Science.gov (United States)

    Hemelaar, Simon R; Nagl, Andreas; Bigot, François; Rodríguez-García, Melissa M; de Vries, Marcel P; Chipaux, Mayeul; Schirhagl, Romana

    2017-01-01

    Fluorescent nanodiamonds (FNDs) are promising tools to image cells, bioanalytes and physical quantities such as temperature, pressure, and electric or magnetic fields with nanometer resolution. To exploit their potential for intracellular applications, the FNDs have to be brought into contact with cell culture media. The interactions between the medium and the diamonds crucially influence sensitivity as well as the ability to enter cells. The authors demonstrate that certain proteins and salts spontaneously adhere to the FNDs and may cause aggregation. This is a first investigation on the fundamental questions on how (a) FNDs interact with the medium, and (b) which proteins and salts are being attracted. A differentiation between strongly binding and weakly binding proteins is made. Not all proteins participate in the formation of FND aggregates. Surprisingly, some main components in the medium seem to play no role in aggregation. Simple strategies to prevent aggregation are discussed. These include adding the proteins, which are naturally present in the cell culture to the diamonds first and then inserting them in the full medium. Graphical abstractSchematic of the interaction of nanodiamonds with cell culture medium. Certain proteins and salts adhere to the diamond surface and lead to aggregation or to formation of a protein corona.

  13. Chemoproteomic profiling of targets of lipid-derived electrophiles by bioorthogonal aminooxy probe

    Directory of Open Access Journals (Sweden)

    Ying Chen

    2017-08-01

    Full Text Available Redox imbalance in cells induces lipid peroxidation and generates a class of highly reactive metabolites known as lipid-derived electrophiles (LDEs that can modify proteins and affects their functions. Identifying targets of LDEs is critical to understand how such modifications are functionally implicated in oxidative-stress associated diseases. Here we report a quantitative chemoproteomic method to globally profile protein targets and sites modified by LDEs. In this strategy, we designed and synthesized an alkyne-functionalized aminooxy probe to react with LDE-modified proteins for imaging and proteomic profiling. Using this probe, we successfully quantified >4000 proteins modified by 4-hydroxy-2-nonenal (HNE of high confidence in mammalian cell lysate and combined with a tandem-orthogonal proteolysis activity-based protein profiling (TOP-ABPP strategy, we identified ~400 residue sites targeted by HNE including reactive cysteines in peroxiredoxins, an important family of enzymes with anti-oxidant roles. Our method expands the toolbox to quantitatively profile protein targets of endogenous electrophiles and the enlarged inventory of LDE-modified proteins and sites will contribute to functional elucidation of cellular pathways affected by oxidative stress. Keywords: Lipid-derived electrophile, 4-hydroxy-2-nonenal, Chemoproteomics, Aminooxy probe, Activity-based protein profiling

  14. Rapid probing of photocatalytic activity on titania-based self-cleaning materials using 7-hydroxycoumarin fluorescent probe

    International Nuclear Information System (INIS)

    Guan Huimin; Zhu Lihua; Zhou Hehui; Tang Heqing

    2008-01-01

    Self-cleaning materials are widely applied, but the available methods for determining their photocatalytic activity are time consuming. A simple analysis method was proposed to evaluate rapidly the photocatalytic activity of self-cleaning materials. This method is based on monitoring of a highly fluorescent product generated by the self-cleaning materials after illumination. Under UV irradiation, holes photo-induced on the surface of self-cleaning materials can oxidize water molecules (or hydroxide ions) adsorbed on the surface to produce hydroxyl radicals, which then quantitatively oxidize coumarin to highly fluorescent 7-hydroxycoumarin. It was observed that the fluorescence intensity of photo-generated 7-hydroxycoumarin at 456 nm (excited at 346 nm) linearly increased with irradiation time, and the fluorescence intensity at a given irradiation time was linearly proportional to the photocatalytic activity of self-cleaning materials. Consequently, the photocatalytic activity of self-cleaning materials was able to be probed simply by using this new method, which requires an analysis time of 40 min, being much less than 250 min required for a dye method

  15. Spontaneous charged lipid transfer between lipid vesicles.

    Science.gov (United States)

    Richens, Joanna L; Tyler, Arwen I I; Barriga, Hanna M G; Bramble, Jonathan P; Law, Robert V; Brooks, Nicholas J; Seddon, John M; Ces, Oscar; O'Shea, Paul

    2017-10-03

    An assay to study the spontaneous charged lipid transfer between lipid vesicles is described. A donor/acceptor vesicle system is employed, where neutrally charged acceptor vesicles are fluorescently labelled with the electrostatic membrane probe Fluoresceinphosphatidylethanolamine (FPE). Upon addition of charged donor vesicles, transfer of negatively charged lipid occurs, resulting in a fluorescently detectable change in the membrane potential of the acceptor vesicles. Using this approach we have studied the transfer properties of a range of lipids, varying both the headgroup and the chain length. At the low vesicle concentrations chosen, the transfer follows a first-order process where lipid monomers are transferred presumably through the aqueous solution phase from donor to acceptor vesicle. The rate of transfer decreases with increasing chain length which is consistent with energy models previously reported for lipid monomer vesicle interactions. Our assay improves on existing methods allowing the study of a range of unmodified lipids, continuous monitoring of transfer and simplified experimental procedures.

  16. Selective and sensitive fluorescence-shift probes based on two dansyl groups for mercury(ii) ion detection.

    Science.gov (United States)

    Ma, Li-Jun; Liu, Jialun; Deng, Lefang; Zhao, Meili; Deng, Zhifu; Li, Xutian; Tang, Jian; Yang, Liting

    2014-11-01

    Two probes ( and ) bearing two dansyl fluorophores were synthesized and applied to the detection of mercury(ii) ions in aqueous solution. These probes exhibited a selective response to Hg(2+) in a buffered solution, with high sensitivity and a unique fluorescence response signal which displayed a blue-shift effect in the fluorescence emission peak. The Hg(2+) recognition mechanisms of the probes were determined by NMR spectroscopy, ESI-MS and UV-vis spectroscopy. The results showed that probe and mercury(ii) ions formed an unusual 2:2 stoichiometric ratio complex, while probe and Hg(2+) formed a multidentate complex with a stoichiometric ratio of 2:1.

  17. A new fluorescence-based method identifies protein phosphatases regulating lipid droplet metabolism.

    Directory of Open Access Journals (Sweden)

    Bruno L Bozaquel-Morais

    Full Text Available In virtually every cell, neutral lipids are stored in cytoplasmic structures called lipid droplets (LDs and also referred to as lipid bodies or lipid particles. We developed a rapid high-throughput assay based on the recovery of quenched BODIPY-fluorescence that allows to quantify lipid droplets. The method was validated by monitoring lipid droplet turnover during growth of a yeast culture and by screening a group of strains deleted in genes known to be involved in lipid metabolism. In both tests, the fluorimetric assay showed high sensitivity and good agreement with previously reported data using microscopy. We used this method for high-throughput identification of protein phosphatases involved in lipid droplet metabolism. From 65 yeast knockout strains encoding protein phosphatases and its regulatory subunits, 13 strains revealed to have abnormal levels of lipid droplets, 10 of them having high lipid droplet content. Strains deleted for type I protein phosphatases and related regulators (ppz2, gac1, bni4, type 2A phosphatase and its related regulator (pph21 and sap185, type 2C protein phosphatases (ptc1, ptc4, ptc7 and dual phosphatases (pps1, msg5 were catalogued as high-lipid droplet content strains. Only reg1, a targeting subunit of the type 1 phosphatase Glc7p, and members of the nutrient-sensitive TOR pathway (sit4 and the regulatory subunit sap190 were catalogued as low-lipid droplet content strains, which were studied further. We show that Snf1, the homologue of the mammalian AMP-activated kinase, is constitutively phosphorylated (hyperactive in sit4 and sap190 strains leading to a reduction of acetyl-CoA carboxylase activity. In conclusion, our fast and highly sensitive method permitted us to catalogue protein phosphatases involved in the regulation of LD metabolism and present evidence indicating that the TOR pathway and the SNF1/AMPK pathway are connected through the Sit4p-Sap190p pair in the control of lipid droplet biogenesis.

  18. Diffusion studies on permeable nitroxyl spin probes through bilayer lipid membranes: A low frequency ESR study

    International Nuclear Information System (INIS)

    Meenakumari, V.; Benial, A. Milton Franklin; Utsumi, Hideo; Ichikawa, Kazuhiro; Yamada, Ken-ichi; Hyodo, Fuminori; Jawahar, A.

    2015-01-01

    Electron spin resonance (ESR) studies were carried out for permeable 2mM 14 N-labeled deutrated 3 Methoxy carbonyl-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl (MC-PROXYL) in pure water and 1mM, 2mM, 3mM, 4mM concentration of 14N-labeled deutrated MC-PROXYL in 400mM concentration of liposomal solution by using a 300 MHz ESR spectrometer. The ESR parameters such as linewidth, hyperfine coupling constant, g-factor, partition parameter and permeability were reported for these samples. The line broadening was observed for the nitroxyl spin probe in the liposomal solution. The line broadening indicates that the high viscous nature of the liposomal solution. The partition parameter and permeability values indicate the maximum diffusion of nitroxyl spin probes in the bilayer lipid membranes at 2 mM concentration of nitroxyl radical. This study illustrates that ESR can be used to differentiate between the intra and extra- membrane water by loading the liposome vesicles with a lipid-permeable nitroxyl spin probe. From the ESR results, the spin probe concentration was optimized as 2mM in liposomal solution for ESR phantom studies/imaging, invivo and invitro experiments

  19. Sensitive fluorescence on-off probes for the fast detection of a chemical warfare agent mimic.

    Science.gov (United States)

    Khan, Muhammad Shar Jhahan; Wang, Ya-Wen; Senge, Mathias O; Peng, Yu

    2018-01-15

    Two highly sensitive probes bearing a nucleophilic imine moiety have been utilized for the selective detection of chemical warfare agent (CWA) mimics. Diethyl chlorophosphate (DCP) was used as mimic CWAs. Both iminocoumarin-benzothiazole-based probes not only demonstrated a remarkable fluorescence ON-OFF response and good recognition, but also exhibited fast response times (10s) along with color changes upon addition of DCP. Limits of detection for the two sensors 1 and 2 were calculated as 0.065μM and 0.21μM, respectively, which are much lower than most other reported probes. These two probes not only show high sensitivity and selectivity in solution, but can also be applied for the recognition of DCP in the gas state, with significant color changes easily observed by the naked eye. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Comparison of N-alkyl acridine orange dyes as fluorescence probes for the determination of cardiolipin

    Energy Technology Data Exchange (ETDEWEB)

    Kaewsuya, P.; Miller, J.D. [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States); Danielson, N.D. [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States)], E-mail: danielnd@muohio.edu; Sanjeevi, J.; James, P.F. [Department of Zoology, Miami University, Oxford, OH 45056 (United States)

    2008-09-26

    The phospholipid (PL), cardiolipin (CL), is found almost exclusively in the inner membrane of mitochondria and loss of CL is considered as an important indication of cell apoptosis. Previously, 10-N-nonyl acridine orange (NAO) has been used as a fluorescent probe for the visualization of CL in mitochondrial cell membranes and in solution. In this work for the determination of CL, we have synthesized two new fluorescent probes, n-tetradecyl acridine orange (C14-AO), and n-octadecyl acridine orange (C18-AO) by reacting acridine orange with the corresponding n-alkyl bromide. Using excitation and emission wavelengths at about 500 and 525 nm and varying the percentage of methanol in water as the solvent, no interaction between CL and the fluorescent probes at 75% is noted but a proportional quenching of the fluorescence signal by CL is observed at 50% or less for C14-AO and 60% or less for C18-AO. Binding efficiency of these fluorescent probes to CL is compared using dye concentrations of 5, 10, and 20 {mu}M. C18-AO shows a better sensitivity than C14-AO and NAO, respectively, but is less selective. For C14-AO, the detection limit and limit of quantitation are 0.07 and 0.21 {mu}M, respectively, which are better than those previously reported for NAO. One anionic PL, phosphatidic acid, shows some quenching interference to both the C14 and C18 dyes but only at concentrations above the working range for sample analysis. The CL in mitochondrial membrane samples is determined by standard addition using C14-AO. The level of CL in the outer mitochondrial membrane compared to the inner membrane is significantly increased due to the addition of cadmium chloride into the cells causing cell apoptosis.

  1. Piezoresistor-equipped fluorescence-based cantilever probe for near-field scanning.

    Science.gov (United States)

    Kan, Tetsuo; Matsumoto, Kiyoshi; Shimoyama, Isao

    2007-08-01

    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.

  2. Intrinsically Labeled Fluorescent Oligonucleotide Probes on Quantum Dots for Transduction of Nucleic Acid Hybridization.

    Science.gov (United States)

    Shahmuradyan, Anna; Krull, Ulrich J

    2016-03-15

    Quantum dots (QDs) have been widely used in chemical and biosensing due to their unique photoelectrical properties and are well suited as donors in fluorescence resonance energy transfer (FRET). Selective hybridization interactions of oligonucleotides on QDs have been determined by FRET. Typically, the QD-FRET constructs have made use of labeled targets or have implemented labeled sandwich format assays to introduce dyes in proximity to the QDs for the FRET process. The intention of this new work is to explore a method to incorporate the acceptor dye into the probe molecule. Thiazole orange (TO) derivatives are fluorescent intercalating dyes that have been used for detection of double-stranded nucleic acids. One such dye system has been reported in which single-stranded oligonucleotide probes were doubly labeled with adjacent thiazole orange derivatives. In the absence of the fully complementary (FC) oligonucleotide target, the dyes form an H-aggregate, which results in quenching of fluorescence emission due to excitonic interactions between the dyes. The hybridization of the FC target to the probe provides for dissociation of the aggregate as the dyes intercalate into the double stranded duplex, resulting in increased fluorescence. This work reports investigation of the dependence of the ratiometric signal on the type of linkage used to conjugate the dyes to the probe, the location of the dye along the length of the probe, and the distance between adjacent dye molecules. The limit of detection for 34mer and 90mer targets was found to be identical and was 10 nM (2 pmol), similar to analogous QD-FRET using labeled oligonucleotide target. The detection system could discriminate a one base pair mismatch (1BPM) target and was functional without substantial compromise of the signal in 75% serum. The 1BPM was found to reduce background signal, indicating that the structure of the mismatch affected the environment of the intercalating dyes.

  3. Glucose oxidase-functionalized fluorescent gold nanoclusters as probes for glucose

    International Nuclear Information System (INIS)

    Xia, Xiaodong; Long, Yunfei; Wang, Jianxiu

    2013-01-01

    Highlights: ► A glucose oxidase/gold nanocluster conjugates formed by etching chemistry. ► Integration of the bioactivities and fluorescence properties within a single unit. ► These conjugates serve as novel fluorescent probe for glucose. -- Abstract: Creation and application of noble metal nanoclusters have received continuous attention. By integrating enzyme activity and fluorescence for potential applications, enzyme-capped metal clusters are more desirable. This work demonstrated a glucose oxidase (an enzyme for glucose)-functionalized gold cluster as probe for glucose. Under physiological conditions, such bioconjugate was successfully prepared by an etching reaction, where tetrakis (hydroxylmethyl) phosphonium-protected gold nanoparticle and thioctic acid-modified glucose oxidase were used as precursor and etchant, respectively. These bioconjugates showed unique fluorescence spectra (λ em max = 650 nm, λ ex max = 507 nm) with an acceptable quantum yield (ca. 7%). Moreover, the conjugated glucose oxidase remained active and catalyzed reaction of glucose and dissolved O 2 to produce H 2 O 2 , which quenched quantitatively the fluorescence of gold clusters and laid a foundation of glucose detection. A linear range of 2.0 × 10 −6 –140 × 10 −6 M and a detection limit of 0.7 × 10 −6 M (S/N = 3) were obtained. Also, another horseradish peroxidase/gold cluster bioconjugate was produced by such general synthesis method. Such enzyme/metal cluster bioconjugates represented a promising class of biosensors for biologically important targets in organelles or cells

  4. Hairpin-like fluorescent probe for imaging of NF-κB transcription factor activity.

    Science.gov (United States)

    Metelev, Valeri; Zhang, Surong; Tabatadze, David; Bogdanov, Alexei

    2011-04-20

    Three oligodeoxyribonucleotides (ODN) covalently labeled with near-infrared (NIR) fluorochromes were synthesized and characterized with a goal of comparing in vitro a hairpin-based and a duplex-based FRET probe designed for the detection of human recombinant NF-κB p50/p65 heterodimer binding to DNA. Using deoxyguanosine phosphoramidite with a phosphorus-linked aminoethylene (diethylene glycol) hydrophilic linker, we synthesized ODNs with internucleoside reactive sites. The hairpin loop amino linker was modified with IRDye 800CW (FRET acceptor), and the 3'-end was modified with Cy5.5 (FRET donor) using a dithio-linker. To obtain a duplex probe, we conjugated Cy5.5 and 800CW to complementary strands at the distance of ten base pairs in the resultant duplex. No quenching of dyes was observed in either probe. The FRET efficiency was higher in the duplex (71%) than in the hairpin (56%) due to a more favorable distance between the donor and the acceptor. However, the hairpin design allowed more precise ratiometric measurement of fluorescence intensity changes as a result of NF-κB p50/p65 binding to the probe. We determined that as a result of binding there was a statistically significant increase of fluorescence intensity of Cy5.5 (donor) due to a decrease of FRET if normalized by 800CW intensity measured independently of FRET. We conclude that the hairpin based probe design allows for the synthesis of a dual fluorescence imaging probe that renders signal changes that are simple to interpret and stoichiometrically correct for detecting transcription factor-DNA interactions.

  5. An off-on fluorescence probe targeting mitochondria based on oxidation-reduction response for tumor cell and tissue imaging

    Science.gov (United States)

    Yao, Hanchun; Cao, Li; Zhao, Weiwei; Zhang, Suge; Zeng, Man; Du, Bin

    2017-10-01

    In this study, a tumor-targeting poly( d, l-lactic-co-glycolic acid) (PLGA) loaded "off-on" fluorescent probe nanoparticle (PFN) delivery system was developed to evaluate the region of tumor by off-on fluorescence. The biodegradability of the nanosize PFN delivery system readily released the probe under tumor acidic conditions. The probe with good biocompatibility was used to monitor the intracellular glutathione (GSH) of cancer cells and selectively localize to mitochondria for tumor imaging. The incorporated tumor-targeting probe was based on the molecular photoinduced electron transfer (PET) mechanism preventing fluorescence ("off" state) and could be easily released under tumor acidic conditions. However, the released tumor-targeting fluorescence probe molecule was selective towards GSH with high selectivity and an ultra-sensitivity for the mitochondria of cancer cells and tissues significantly increasing the probe molecule fluorescence signal ("on" state). The tumor-targeting fluorescence probe showed sensitivity to GSH avoiding interference from cysteine and homocysteine. The PFNs could enable fluorescence-guided cancer imaging during cancer therapy. This work may expand the biological applications of PFNs as a diagnostic reagent, which will be beneficial for fundamental research in tumor imaging. [Figure not available: see fulltext.

  6. A rhodamine chromene-based turn-on fluorescence probe for selectively imaging Cu2+ in living cell

    Science.gov (United States)

    Liu, Wei-Yong; Li, Hai-Ying; Lv, Hong-Shui; Zhao, Bao-Xiang; Miao, Jun-Ying

    We describe the development of a rhodamine chromene-based turn-on fluorescence probe to monitor the intracellular Cu2+ level in living cells. The new fluorescent probe with a chlorine group in chromene moiety exhibits good membrane-permeable property than previous reported because the predicted lipophilicity of present probe 4 is stronger than that of methoxyl substituted probe in our previous work (CLogP of 4: 8.313, CLogP of methoxyl substituted probe: 7.706), and a fluorescence response toward Cu2+ under physiological conditions with high sensitivity and selectivity, and facilitates naked-eye detection of Cu2+. The fluorescence intensity was remarkably increased upon the addition of Cu2+ within 1 or 2 min, while the other sixteen metal ions caused no significant effect.

  7. Conception, synthesis and evaluation of fluorescent probes and PET radioligands for the oxytocin and vasopressin receptors

    International Nuclear Information System (INIS)

    Karpenko, Iuliia

    2014-01-01

    In order to better understand the role of OTR and AVPR in ASD, to reveal new features in its pharmacology and signaling and to establish high-throughput screening method on wild-type G protein-coupled receptors, we developed imaging probes for the oxytocin-vasopressin receptors family, namely radiotracers for positron emission tomography and optical probes for fluorescence detection and imaging. The fluorescent ligands have been used to establish TR-FRET binding assay for OTR and to initiate the development the screening assay for the wild-type oxytocin receptor. The PET radiotracers will be shortly tested in mice and monkeys to evaluate their potency in detecting the central oxytocin receptors. (author)

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

    Science.gov (United States)

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

    2012-03-01

    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.

  9. Sensing for intracellular thiols by water-insoluble two-photon fluorescent probe incorporating nanogel

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Xudong; Zhang, Xin; Wang, Shuangqing; Li, Shayu [Beijing National Laboratory for Molecular Sciences, Key laboratory of Photochemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Hu, Rui, E-mail: hurui@iccas.ac.cn [Beijing National Laboratory for Molecular Sciences, Key laboratory of Photochemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Li, Yi, E-mail: yili@mail.ipc.ac.cn [Key Laboratory of Photochemical Conversion and Optoelectronic Materials, Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Yang, Guoqiang, E-mail: gqyang@iccas.ac.cn [Beijing National Laboratory for Molecular Sciences, Key laboratory of Photochemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China)

    2015-04-15

    Highlights: • A novel “turn-on” two-photon fluorescent probe based on a π-conjugated triarylboron luminogen was designed and synthesized. • Fast, selective and sensitive detection of biothiols in 100% aqueous solution by simply loaded on a nanogel. • Single-photon and two-photon fluorescent bioimaging of biothiols in NIH/3T3 fibroblasts. - Abstract: A novel “turn-on” two-photon fluorescent probe containing a π-conjugated triarylboron luminogen and a maleimide moiety DMDP-M based on the photo-induced electron transfer (PET) mechanism for biothiol detection was designed and synthesized. By simply loading the hydrophobic DMDP-M on a cross-linked Pluronic{sup ®} F127 nanogel (CL-F127), a probing system DMDP-M/CL-F127 was established, which shows quick response, high selectivity and sensitivity to cysteine (Cys), homocysteine (Hcy) and glutathione (GSH) in aqueous phase. The DMDP-M/CL-F127 system presented the fastest response to Cys with a rate constant of 0.56 min{sup −1}, and the detection limit to Cys was calculated to be as low as 0.18 μM. The DMDP-M/CL-F127 system has been successfully applied to the fluorescence imaging of biothiols in NIH/3T3 fibroblasts either with single-photon or two-photon excitation because of its high biocompatibility and cell-membrane permeability. The present work provides a general, simple and efficient strategy for the application of hydrophobic molecules to sensing biothiols in aqueous phase, and a novel sensing system for intracellular biothiols fitted for both single-photon and two-photon fluorescence imaging.

  10. [A cell-based detection of ciguatoxin using sodium fluorescence probe].

    Science.gov (United States)

    Yuan, Jian-hui; Yang, Hui; Tang, Huan-wen; Huang, Wei; Xu, Xin-yun; Liu, Jian-jun; Ke, Yue-bin; Cheng, Jin-quan; Zhuang, Zhi-xiong

    2011-04-01

    To establish a cell-based detection method of ciguatoxin using fluorescence assay. Mouse neuroblastoma N-2A cells were exposed to ouabain and veratridine and different concentrations of standard ciguatoxin samples (P-CTX-1) to establish the curvilinear relationship between the toxin dosage and fluorescence intensity using the sodium fluorescence probe CoroNaTM Green. The toxicity curvilinear relationship was also generated between the toxin dosage and cell survival using CCK-8 method. Based on these standard curves, the presence of ciguatoxin was detected in 33 samples of deep-sea coral fish. A correlation was found between the detection results of cell-based fluorescence assay and cytotoxicity assay, whose detection limit reached 103 g/ml and 1012 g/ml, respectively. The cell-based fluorescent assay sensitivity showed a higher sensitivity than cytotoxicity assay with a 2-4 h reduction of the detection time. The cell-based fluorescent assay can quickly and sensitively detect ciguatoxin and may serve as a good option for preliminary screening of the toxin.

  11. Two colorimetric and ratiometric fluorescence probes for hydrogen sulfide based on AIE strategy of α-cyanostilbenes

    Science.gov (United States)

    Zhao, Baoying; Yang, Binsheng; Hu, Xiangquan; Liu, Bin

    2018-06-01

    Aggregation-induced emission (AIE) active fluorescent probes have attracted great potential in biological sensors. In this paper two cyanostilbene based fluorescence chemoprobe Cya-NO2 (1) and Cya-N3 (2) were developed and evaluated for the selective and sensitive detection of hydrogen sulfide (H2S). Both of these probes behave aggression-induced emission (AIE) activity which fluoresces in the red region with a large Stokes shift. They exhibit rapid response to H2S with enormous colorimetric and ratiometric fluorescent changes. They are readily employed for assessing intracellular H2S levels.

  12. Fluorescent probe encapsulated hydrogel microsphere for selective and reversible detection of Hg{sup 2+}

    Energy Technology Data Exchange (ETDEWEB)

    Song, Zhenhu; Wang, Fang; Qiang, Jian; Zhang, Zhijie; Chen, Yahui; Wang, Yong; Zhang, Wei; Chen, Xiaoqiang

    2017-03-15

    We developed a simple and sensitive hydrogel sensor in the form of microspheres by using fluorescence probe encapsulated within a hydrogel matrix for the detection of Hg{sup 2+}. The traditional fluorescence probes suspended in solution are not transportable and recoverable. To overcome these disadvantages, we devised poly(ethylene glycol) diacrylate-based hydrogel microspheres in which fluorescence probe (R19S) was embedded at high density. The functionalized hydrogel microspheres were prepared by combining a microfluidic device with UV light. The hydrogel microspheres-based sensor exhibited good selectivity to Hg{sup 2+} among various metal ions and high sensitivity with a detection limit of 90 nM. Furthermore, after binding with Hg{sup 2+}, the R19S encapsulated hydrogel microspheres can be separated from testing samples easily and treated with the solution containing KI to remove Hg{sup 2+} and realize reusable detection. The current work may offer a new method for Hg{sup 2+} recognition with a more efficient manner.

  13. Colorimetric and Fluorescent Bimodal Ratiometric Probes for pH Sensing of Living Cells.

    Science.gov (United States)

    Liu, Yuan-Yuan; Wu, Ming; Zhu, Li-Na; Feng, Xi-Zeng; Kong, De-Ming

    2015-06-01

    pH measurement is widely used in many fields. Ratiometric pH sensing is an important way to improve the detection accuracy. Herein, five water-soluble cationic porphyrin derivatives were synthesized and their optical property changes with pH value were investigated. Their pH-dependent assembly/disassembly behaviors caused significant changes in both absorption and fluorescence spectra, thus making them promising bimodal ratiometric probes for both colorimetric and fluorescent pH sensing. Different substituent identity and position confer these probes with different sensitive pH-sensing ranges, and the substituent position gives a larger effect. By selecting different porphyrins, different signal intensity ratios and different fluorescence excitation wavelengths, sensitive pH sensing can be achieved in the range of 2.1-8.0. Having demonstrated the excellent reversibility, good accuracy and low cytotoxicity of the probes, they were successfully applied in pH sensing inside living cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. An Amidochlorin-Based Colorimetric Fluorescent Probe for Selective Cu2+ Detection

    Directory of Open Access Journals (Sweden)

    Wenting Li

    2016-01-01

    Full Text Available The design and synthesis of selective and sensitive chemosensors for the quantification of environmentally and biologically important ionic species has attracted widespread attention. Amidochlorin p6 (ACP; an effective colorimetric and fluorescent probe for copper ions (Cu2+ in aqueous solution derived from methyl pheophorbide-a (MPa was designed and synthesized. A remarkable color change from pale yellow to blue was easily observed by the naked eye upon addition of Cu2+; and a fluorescence quenching was also determined. The research of fluorescent quenching of ACP-Cu2+ complexation showed the detection limit was 7.5 × 10−8 mol/L; which suggested that ACP can act as a high sensitive probe for Cu2+ and can be used to quantitatively detect low levels of Cu2+ in aqueous solution. In aqueous solution the probe exhibits excellent selectivity and sensitivity toward Cu2+ ions over other metal ions (M = Zn2+; Ni2+; Ba2+; Ag+; Co2+; Na+; K+; Mg2+; Cd2+; Pb2+; Mn2+; Fe3+; and Ca2+. The obvious change from pale yellow to blue upon the addition of Cu2+ could make it a suitable “naked eye” indicator for Cu2+.

  15. Fluorescence detection of glutathione and oxidized glutathione in blood with a NIR-excitable cyanine probe

    Science.gov (United States)

    Liu, Chang-hui; Qi, Feng-pei; Wen, Fu-bin; Long, Li-ping; Liu, Ai-juan; Yang, Rong-hua

    2018-04-01

    Cyanine has been widely utilized as a near infrared (NIR) fluorophore for detection of glutathione (GSH). However, the excitation of most of the reported cyanine-based probes was less than 800 nm, which inevitably induce biological background absorption and lower the sensitivity, limiting their use for detection of GSH in blood samples. To address this issue, here, a heptamethine cyanine probe (DNIR), with a NIR excitation wavelength at 804 nm and a NIR emission wavelength at 832 nm, is employed for the detection of GSH and its oxidized form (GSSG) in blood. The probe displays excellent selectivity for GSH over GSSG and other amino acids, and rapid response to GSH, in particular a good property for indirect detection of GSSG in the presence of enzyme glutathione reductase and the reducing agent nicotinamideadenine dinucleotide phosphate, without further separation prior to fluorescent measurement. To the best of our knowledge, this is the first attempt to explore NIR fluorescent approach for the simultaneous assay of GSH and GSSG in blood. As such, we expect that our fluorescence sensors with both NIR excitation and NIR emission make this strategy suitable for the application in complex physiological systems.

  16. Quinoline Fluorescent Probes for Zinc - from Diagnostic to Therapeutic Molecules in Treating Neurodegenerative Diseases.

    Science.gov (United States)

    Czaplinska, Barbara; Spaczynska, Ewelina; Musiol, Robert

    2018-01-01

    Fluorescent compounds had gained strong attention due to their wide and appealing applications. Microscopic techniques and visualization are good examples among others. Introduction of fluorescent dyes into microbiology opens the possibility to observe tissues, organisms or organelle with exceptional sensitivity and resolution. Probes for detection of biologically relevant metals as zinc, iron or copper seems to be particularly important for drug design and pharmaceutical sciences. Quinoline derivatives are well known for their good metal affinity and wide spectrum of biological activity. In this regard, molecular sensors built on this scaffold may be useful not only as analytical but also as therapeutic agents. In the present review, application of quinoline moiety in designing of novel fluorescent probes for zinc is presented and discussed. Zinc cations are relevant for vast majority of processes and recently attract a great deal of attention for their role in neurodegenerative diseases. Compounds interacting with Zn2+ may be used for early diagnosis of such disorders, for example the Alzheimer disease. Quinoline-based zinc probes may exert some beneficial role in organism acting as theranostic agents. First preliminary drugs for Alzheimer therapy that are based on quinoline moiety are good example of this trend. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  17. Tissue classification and diagnostics using a fiber probe for combined Raman and fluorescence spectroscopy

    Science.gov (United States)

    Cicchi, Riccardo; Anand, Suresh; Crisci, Alfonso; Giordano, Flavio; Rossari, Susanna; De Giorgi, Vincenzo; Maio, Vincenza; Massi, Daniela; Nesi, Gabriella; Buccoliero, Anna Maria; Guerrini, Renzo; Pimpinelli, Nicola; Pavone, Francesco S.

    2015-07-01

    Two different optical fiber probes for combined Raman and fluorescence spectroscopic measurements were designed, developed and used for tissue diagnostics. Two visible laser diodes were used for fluorescence spectroscopy, whereas a laser diode emitting in the NIR was used for Raman spectroscopy. The two probes were based on fiber bundles with a central multimode optical fiber, used for delivering light to the tissue, and 24 surrounding optical fibers for signal collection. Both fluorescence and Raman spectra were acquired using the same detection unit, based on a cooled CCD camera, connected to a spectrograph. The two probes were successfully employed for diagnostic purposes on various tissues in a good agreement with common routine histology. This study included skin, brain and bladder tissues and in particular the classification of: malignant melanoma against melanocytic lesions and healthy skin; urothelial carcinoma against healthy bladder mucosa; brain tumor against dysplastic brain tissue. The diagnostic capabilities were determined using a cross-validation method with a leave-one-out approach, finding very high sensitivity and specificity for all the examined tissues. The obtained results demonstrated that the multimodal approach is crucial for improving diagnostic capabilities. The system presented here can improve diagnostic capabilities on a broad range of tissues and has the potential of being used for endoscopic inspections in the near future.

  18. Design of mitochondria-targeted colorimetric and ratiometric fluorescent probes for rapid detection of SO2 derivatives in living cells

    Science.gov (United States)

    Yang, Yutao; Zhou, Tingting; Bai, Bozan; Yin, Caixia; Xu, Wenzhi; Li, Wei

    2018-05-01

    Two mitochondria-targeted colorimetric and ratiometric fluorescent probes for SO2 derivatives were constructed based on the SO2 derivatives-triggered Michael addition reaction. The probes exhibit high specificity toward HSO3-/SO32- by interrupting their conjugation system resulting in a large ratiometric blue shift of 46-121 nm in their emission spectrum. The two well-resolved emission bands can ensure accurate detection of HSO3-. The detection limits were calculated to be 1.09 and 1.35 μM. Importantly, probe 1 and probe 2 were successfully used to fluorescence ratiometric imaging of endogenous HSO3- in BT-474 cells.

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

    Science.gov (United States)

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

    2014-09-01

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

  20. A simple rhodamine hydrazide-based turn-on fluorescent probe for HOCl detection.

    Science.gov (United States)

    Zhang, Zhen; Zou, Yuan; Deng, Chengquan; Meng, Liesu

    2016-06-01

    Hypochlorous acid (HOCl) plays a crucial role in daily life and mediates a variety of physiological processes, however, abnormal levels of HOCl have been associated with numerous human diseases. It is therefore of significant interest to establish a simple, selective, rapid and sensitive fluorogenic method for the detection of HOCl in environmental and biological samples. A hydrazide-containing fluorescent probe based on a rhodamine scaffold was facilely developed that could selectively detect HOCl over other biologically relevant reactive oxygen species, reactive nitrogen species and most common metal ions in vitro. Via an irreversible oxidation-hydrolysis mechanism, and upon HOCl-triggered opening of the intramolecular spirocyclic ring during detection, the rhodamine hydrazide-based probe exhibited large fluorescence enhancement in the emission spectra with a fast response, low detection limit and comparatively wide pH detection range in aqueous media. The probe was further successfully applied to monitoring trace HOCl in tap water and imaging both exogenous and endogenous HOCl within living cells. It is anticipated that this simple and useful probe might be an efficient tool with which to facilitate more HOCl-related chemical and biological research. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  1. Hyaluronan functionalizing QDs as turn-on fluorescent probe for targeted recognition CD44 receptor

    Science.gov (United States)

    Zhou, Shang; Huo, Danqun; Hou, Changjun; Yang, Mei; Fa, Huanbao

    2017-09-01

    The recognition of tumor markers in living cancer cells has attracted increasing interest. In the present study, the turn-on fluorescence probe was designed based on the fluorescence of thiolated chitosan-coated CdTe QDs (CdTe/TCS QDs) quenched by hyaluronan, which could provide the low background signal for sensitive cellular imaging. This system is expected to offer specific recognition of CD44 receptor over other substances owing to the specific affinity of hyaluronan and CD44 receptor ( 8-9 kcal/mol). The probe is stable in aqueous and has little toxicity to living cells; thus, it can be utilized for targeted cancer cell imaging. The living lung cancer cell imaging experiments further demonstrate its value in recognizing cell-surface CD44 receptor with turn-on mode. In addition, the probe can be used to recognize and differentiate the subtypes of lung cancer cells based on the difference of CD44 expression on the surface of lung cancer cells. And, the western blot test further confirmed that the expression level of the CD44 receptor in lung cancer cells is different. Therefore, this probe may be potentially applied in recognizing lung cancer cells with higher contrast and sensitivity and provide new tools for cancer prognosis and therapy. [Figure not available: see fulltext.

  2. CdTe/ZnS quantum dots as fluorescent probes for ammonium determination.

    Science.gov (United States)

    Yi, Kui-Yu

    2016-06-01

    Novel CdTe/ZnS quantum dot (QD) probes based on the quenching effect were proposed for the simple, rapid, and specific determination of ammonium in aqueous solutions. The QDs were modified using 3-mercaptopropionic acid, and the fluorescence responses of the CdTe/ZnS QD probes to ammonium were detected through regularity quenching. The quenching levels of the CdTe/ZnS QDs and ammonium concentration showed a good linear relationship between 4.0 × 10(-6) and 5.0 × 10(-4) mol/L; the detection limit was 3.0 × 10(-7) mol/L. Ammonium contents in synthetic explosion soil samples were measured to determine the practical applications of the QD probes and a probable quenching mechanism was described. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  3. Lipid bilayer regulation of membrane protein function: gramicidin channels as molecular force probes

    DEFF Research Database (Denmark)

    Lundbæk, Jens August; Collingwood, S.A.; Ingolfsson, H.I.

    2010-01-01

    with collective physical properties (e.g. thickness, intrinsic monolayer curvature or elastic moduli). Studies in physico-chemical model systems have demonstrated that changes in bilayer physical properties can regulate membrane protein function by altering the energetic cost of the bilayer deformation associated...... with a protein conformational change. This type of regulation is well characterized, and its mechanistic elucidation is an interdisciplinary field bordering on physics, chemistry and biology. Changes in lipid composition that alter bilayer physical properties (including cholesterol, polyunsaturated fatty acids...... channels as molecular force probes for studying this mechanism, with a unique ability to discriminate between consequences of changes in monolayer curvature and bilayer elastic moduli....

  4. Synchronous fluorescence based biosensor for albumin determination by cooperative binding of fluorescence probe in a supra-biomolecular host-protein assembly.

    Science.gov (United States)

    Patra, Digambara

    2010-01-15

    A synchronous fluorescence probe based biosensor for estimation of albumin with high sensitivity and selectivity was developed. Unlike conventional fluorescence emission or excitation spectral measurements, synchronous fluorescence measurement offered exclusively a new synchronous fluorescence peak in the shorter wavelength range upon binding of chrysene with protein making it an easy identification tool for albumin determination. The cooperative binding of a fluorescence probe, chrysene, in a supramolecular host-protein assembly during various albumin assessments was investigated. The presence of supramolecular host molecules such as beta-cyclodextrin, curucurbit[6]uril or curucurbit[7]uril have little influence on sensitivity or limit of detection during albumin determination but reduced dramatically interference from various coexisting metal ion quenchers/enhancers. Using the present method the limit of detection for BSA and gamma-Globulin was found to be 0.005 microM which is more sensitive than reported values. Copyright 2009 Elsevier B.V. All rights reserved.

  5. In-vivo fluorescence detection of breast cancer growth factor receptors by fiber-optic probe

    Science.gov (United States)

    Bustamante, Gilbert; Wang, Bingzhi; DeLuna, Frank; Sun, LuZhe; Ye, Jing Yong

    2018-02-01

    Breast cancer treatment options often include medications that target the overexpression of growth factor receptors, such as the proto-oncogene human epidermal growth factor receptor 2 (HER2/neu) and epidermal growth factor receptor (EGFR) to suppress the abnormal growth of cancerous cells and induce cancer regression. Although effective, certain treatments are toxic to vital organs, and demand assurance that the pursued receptor is present at the tumor before administration of the drug. This requires diagnostic tools to provide tumor molecular signatures, as well as locational information. In this study, we utilized a fiber-optic probe to characterize in vivo HER2 and EGFR overexpressed tumors through the fluorescence of targeted dyes. HER2 and EGFR antibodies were conjugated with ICG-Sulfo-OSu and Alexa Fluor 680, respectively, to tag BT474 (HER2+) and MDA-MB-468 (EGFR+) tumors. The fiber was inserted into the samples via a 30-gauge needle. Different wavelengths of a supercontinuum laser were selected to couple into the fiber and excite the corresponding fluorophores in the samples. The fluorescence from the dyes was collected through the same fiber and quantified by a time-correlated single photon counter. Fluorescence at different antibody-dye concentrations was measured for calibration. Mice with subcutaneous HER2+ and/or EGFR+ tumors received intravenous injections of the conjugates and were later probed at the tumor sites. The measured fluorescence was used to distinguish between tumor types and to calculate the concentration of the antibody-dye conjugates, which were detectable at levels as low as 40 nM. The fiber-optic probe presents a minimally invasive instrument to characterize the molecular signatures of breast cancer in vivo.

  6. [1,10]Phenanthroline based cyanine dyes as fluorescent probes for ribonucleic acids in live cells

    Science.gov (United States)

    Kovalska, Vladyslava; Kuperman, Marina; Varzatskii, Oleg; Kryvorotenko, Dmytro; Kinski, Elisa; Schikora, Margot; Janko, Christina; Alexiou, Christoph; Yarmoluk, Sergiy; Mokhir, Andriy

    2017-12-01

    A series of monomethine, trimethine- and styrylcyanine dyes based on a [1,10]phenanthroline moiety was synthesized, characterized and investigated as potential fluorescent probes for nucleic acids in cell free settings and in cells. The dyes were found to be weakly fluorescent in the unbound state, whereas upon the binding to dsDNA or RNA their emission intensity raised up to 50 times (for monomethine benzothiazole derivative FT1 complexed with RNA). The strongest fluorescence intensity in assemblies with dsDNA and RNA was observed for the trimethine benzothiazole derivative FT4. The quantum yield of FT4 fluorescence in its complex with dsDNA was found to be 1.5% and the binding constant (K b) was estimated to be 7.9 × 104 M-1 that is a typical value for intercalating molecules. The FT4 dye was found to be cell membrane permeable. It stains RNA rich components—the nucleoli and most probably the cytoplasmic RNA. FT4 bound to RNAs delivers a very strong fluorescence signal, which makes this easily accessible dye a potentially useful alternative to known RNA stains, e.g. expensive SYTO® 83. The advantage of FT4 is its easy synthetic access including no chromatographic purification steps, which will be reflected in its substantially lower price.

  7. Measurement of fluorescent probes concentration ratio in the cerebrospinal fluid for early detection of Alzheimer's disease

    Science.gov (United States)

    Harbater, Osnat; Gannot, Israel

    2014-03-01

    The pathogenic process of Alzheimer's Disease (AD), characterized by amyloid plaques and neurofibrillary tangles in the brain, begins years before the clinical diagnosis. Here, we suggest a novel method which may detect AD up to nine years earlier than current exams, minimally invasive, with minimal risk, pain and side effects. The method is based on previous reports which relate the concentrations of biomarkers in the Cerebrospinal Fluid (CSF) (Aβ and Tau proteins) to the future development of AD in mild cognitive impairment patients. Our method, which uses fluorescence measurements of the relative concentrations of the CSF biomarkers, replaces the lumbar puncture process required for CSF drawing. The process uses a miniature needle coupled trough an optical fiber to a laser source and a detector. The laser radiation excites fluorescent probes which were prior injected and bond to the CSF biomarkers. Using the ratio between the fluorescence intensities emitted from the two biomarkers, which is correlated to their concentration ratio, the patient's risk of developing AD is estimated. A theoretical model was developed and validated using Monte Carlo simulations, demonstrating the relation between fluorescence emission and biomarker concentration. The method was tested using multi-layered tissue phantoms simulating the epidural fat, the CSF in the sub-arachnoid space and the bone. These phantoms were prepared with different scattering and absorption coefficients, thicknesses and fluorescence concentrations in order to simulate variations in human anatomy and in the needle location. The theoretical and in-vitro results are compared and the method's accuracy is discussed.

  8. Development of tumor-targeted near infrared probes for fluorescence guided surgery.

    Science.gov (United States)

    Kelderhouse, Lindsay E; Chelvam, Venkatesh; Wayua, Charity; Mahalingam, Sakkarapalayam; Poh, Scott; Kularatne, Sumith A; Low, Philip S

    2013-06-19

    Complete surgical resection of malignant disease is the only reliable method to cure cancer. Unfortunately, quantitative tumor resection is often limited by a surgeon's ability to locate all malignant disease and distinguish it from healthy tissue. Fluorescence-guided surgery has emerged as a tool to aid surgeons in the identification and removal of malignant lesions. While nontargeted fluorescent dyes have been shown to passively accumulate in some tumors, the resulting tumor-to-background ratios are often poor, and the boundaries between malignant and healthy tissues can be difficult to define. To circumvent these problems, our laboratory has developed high affinity tumor targeting ligands that bind to receptors that are overexpressed on cancer cells and deliver attached molecules selectively into these cells. In this study, we explore the use of two tumor-specific targeting ligands (i.e., folic acid that targets the folate receptor (FR) and DUPA that targets prostate specific membrane antigen (PSMA)) to deliver near-infrared (NIR) fluorescent dyes specifically to FR and PSMA expressing cancers, thereby rendering only the malignant cells highly fluorescent. We report here that all FR- and PSMA-targeted NIR probes examined bind cultured cancer cells in the low nanomolar range. Moreover, upon intravenous injection into tumor-bearing mice with metastatic disease, these same ligand-NIR dye conjugates render receptor-expressing tumor tissues fluorescent, enabling their facile resection with minimal contamination from healthy tissues.

  9. Determination of ATP as a fluorescence probe with europium(III)-doxycycline.

    Science.gov (United States)

    Hou, Faju; Wang, Xiaolei; Jiang, Chongqiu

    2005-03-01

    A new spectrofluorimetric method has been developed for the determination of adenosine disodium triphosphate (ATP). We studied the interactions between the doxycycline (DC)-Eu3+ complex and adenosine disodium triphosphate (ATP) by using UV-visible absorption and fluorescence spectra. Using doxycycline (DC)-Eu3+ as a fluorescence probe, under the optimum conditions, ATP could remarkably enhance the fluorescence intensity of the DC-Eu3+ complex at lambda = 612 nm. The enhanced fluorescence intensity of the Eu3+ ion was in proportion to the concentration of ATP. The optimum conditions for the determination of ATP were also investigated. The linear ranges for ATP were 1.00 x 10(-7) - 2.00 x 10(-6) mol L(-1) with detection limits of 4.07 x 10(-8) mol L(-1). This method is simple, practical and relatively free of interference from coexisting substances, and can be successfully applied to the determination of ATP in samples. The mechanism of fluorescence enhancement between the doxycycline (DC)-Eu3+ complex and ATP was also studied.

  10. Determination of adenosine disodium triphosphate using prulifloxacin-terbium(III) as a fluorescence probe by spectrofluorimetry

    International Nuclear Information System (INIS)

    Yu Fengshan; Li Lin; Chen Fang

    2008-01-01

    A new spectrofluorimetric method is developed for determination of adenosine disodium triphosphate (ATP). The interactions between prulifloxacin (PUFX)-Tb 3+ complex and adenosine disodium triphosphate has been studied by using UV-vis absorption and fluorescence spectra. Using prulifloxacin-Tb 3+ as a fluorescence probe, under the optimum conditions, ATP can remarkably enhance the fluorescence intensity of the prulifloxacin-Tb 3+ complex at λ = 545 nm and the enhanced fluorescence intensity is in proportion to the concentration of ATP. Optimum conditions for the determination of ATP were also investigated. The dynamic range for the determination of ATP is 4.0 x 10 -7 to 2.0 x 10 -5 mol L -1 , and the detection limit (3 σ/k) is 1.7 x 10 -8 mol L -1 . This method is simple, practical and relatively free interference from coexisting substances and can be successfully applied to determination of ATP in real pharmaceutical samples. The mechanism of fluorescence enhancement of prulifloxacin-Tb 3+ complex by ATP was also discussed

  11. Fluorescent probes as a tool for labelling and tracking the amphibian chytrid fungus Batrachochytrium dendrobatidis.

    Science.gov (United States)

    Herbert, Sarah M; Leung, Tommy L F; Bishop, Phillip J

    2011-09-09

    The dissemination of the virulent pathogen Batrachochytrium dendrobatidis (Bd) has contributed to the decline and extinction of many amphibian species worldwide. Several different strains have been identified, some of which are sympatric. Interactions between co-infecting strains of a pathogen can have significant influences on disease epidemiology and evolution; therefore the dynamics of multi-strain infections is an important area of research. We stained Bd cells with 2 fluorescent BODIPY fatty acid probes to determine whether these can potentially be used to distinguish and track Bd cell lines in multi-strain experiments. Bd cells in broth culture were stained with 5 concentrations of green-fluorescent BODIPY FL and red-fluorescent BODIPY 558/568 and visualised under an epifluorescent microscope for up to 16 d post-dye. Dyed strains were also assessed for growth inhibition. The most effective concentration for both dyes was 10 pM. This concentration of dye produced strong fluorescence for 12 to 16 d in Bd cultures held at 23 degrees C (3 to 4 generations), and did not inhibit Bd growth. Cells dyed with BODIPY FL and BODIPY 558/568 can be distinguished from each other on the basis of their fluorescence characteristics. Therefore, it is likely that this technique will be useful for research into multi-strain dynamics of Bd infections.

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

    Directory of Open Access Journals (Sweden)

    Wu H

    2018-03-01

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

  13. Combining reflectometry and fluorescence microscopy: an assay for the investigation of leakage processes across lipid membranes.

    Science.gov (United States)

    Stephan, Milena; Mey, Ingo; Steinem, Claudia; Janshoff, Andreas

    2014-02-04

    The passage of solutes across a lipid membrane plays a central role in many cellular processes. However, the investigation of transport processes remains a serious challenge in pharmaceutical research, particularly the transport of uncharged cargo. While translocation reactions of ions across cell membranes is commonly measured with the patch-clamp, an equally powerful screening method for the transport of uncharged compounds is still lacking. A combined setup for reflectometric interference spectroscopy (RIfS) and fluorescence microscopy measurements is presented that allows one to investigate the passive exchange of uncharged compounds across a free-standing membrane. Pore-spanning lipid membranes were prepared by spreading giant 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) vesicles on porous anodic aluminum oxide (AAO) membranes, creating sealed attoliter-sized compartments. The time-resolved leakage of different dye molecules (pyranine and crystal violet) as well as avidin through melittin induced membrane pores and defects was investigated.

  14. Glucose oxidase-functionalized fluorescent gold nanoclusters as probes for glucose

    Energy Technology Data Exchange (ETDEWEB)

    Xia, Xiaodong [College of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China); School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201 (China); Long, Yunfei, E-mail: l_yunfei927@163.com [School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201 (China); Wang, Jianxiu, E-mail: jxiuwang@csu.edu.cn [College of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China)

    2013-04-15

    Highlights: ► A glucose oxidase/gold nanocluster conjugates formed by etching chemistry. ► Integration of the bioactivities and fluorescence properties within a single unit. ► These conjugates serve as novel fluorescent probe for glucose. -- Abstract: Creation and application of noble metal nanoclusters have received continuous attention. By integrating enzyme activity and fluorescence for potential applications, enzyme-capped metal clusters are more desirable. This work demonstrated a glucose oxidase (an enzyme for glucose)-functionalized gold cluster as probe for glucose. Under physiological conditions, such bioconjugate was successfully prepared by an etching reaction, where tetrakis (hydroxylmethyl) phosphonium-protected gold nanoparticle and thioctic acid-modified glucose oxidase were used as precursor and etchant, respectively. These bioconjugates showed unique fluorescence spectra (λ{sub em} {sub max} = 650 nm, λ{sub ex} {sub max} = 507 nm) with an acceptable quantum yield (ca. 7%). Moreover, the conjugated glucose oxidase remained active and catalyzed reaction of glucose and dissolved O{sub 2} to produce H{sub 2}O{sub 2}, which quenched quantitatively the fluorescence of gold clusters and laid a foundation of glucose detection. A linear range of 2.0 × 10{sup −6}–140 × 10{sup −6} M and a detection limit of 0.7 × 10{sup −6} M (S/N = 3) were obtained. Also, another horseradish peroxidase/gold cluster bioconjugate was produced by such general synthesis method. Such enzyme/metal cluster bioconjugates represented a promising class of biosensors for biologically important targets in organelles or cells.

  15. Fluorescent peptide biosensor for probing the relative abundance of cyclin-dependent kinases in living cells.

    Directory of Open Access Journals (Sweden)

    Laetitia Kurzawa

    Full Text Available Cyclin-dependant kinases play a central role in coordinating cell growth and division, and in sustaining proliferation of cancer cells, thereby constituting attractive pharmacological targets. However, there are no direct means of assessing their relative abundance in living cells, current approaches being limited to antigenic and proteomic analysis of fixed cells. In order to probe the relative abundance of these kinases directly in living cells, we have developed a fluorescent peptide biosensor with biligand affinity for CDKs and cyclins in vitro, that retains endogenous CDK/cyclin complexes from cell extracts, and that bears an environmentally-sensitive probe, whose fluorescence increases in a sensitive fashion upon recognition of its targets. CDKSENS was introduced into living cells, through complexation with the cell-penetrating carrier CADY2 and applied to assess the relative abundance of CDK/Cyclins through fluorescence imaging and ratiometric quantification. This peptide biosensor technology affords direct and sensitive readout of CDK/cyclin complex levels, and reports on differences in complex formation when tampering with a single CDK or cyclin. CDKSENS further allows for detection of differences between different healthy and cancer cell lines, thereby enabling to distinguish cells that express high levels of these heterodimeric kinases, from cells that present decreased or defective assemblies. This fluorescent biosensor technology provides information on the overall status of CDK/Cyclin complexes which cannot be obtained through antigenic detection of individual subunits, in a non-invasive fashion which does not require cell fixation or extraction procedures. As such it provides promising perspectives for monitoring the response to therapeutics that affect CDK/Cyclin abundance, for cell-based drug discovery strategies and fluorescence-based cancer diagnostics.

  16. An intramolecular charge transfer process based fluorescent probe for monitoring subtle pH fluctuation in living cells.

    Science.gov (United States)

    Sun, Mingtai; Du, Libo; Yu, Huan; Zhang, Kui; Liu, Yang; Wang, Suhua

    2017-01-01

    It is crucial to monitor intracellular pH values and their fluctuation since the organelles of cells have different pH distribution. Herein we construct a new small molecule fluorescent probe HBT-O for monitoring the subtle pH values within the scope of neutral to acid in living cells. The probe exhibited good water solubility, a marked turquoise to olivine emission color change in response to pH, and tremendous fluorescence hypochromatic shift of ∼50nm (1718cm -1 ) as well as the increased fluorescence intensity when the pH value changed from neutral to acid. Thus, the probe HBT-O can distinguish the subtle changes in the range of normal pH values from neutral to acid with significant fluorescence changes. These properties can be attributed to the intramolecular charge transfer (ICT) process of the probe upon protonation in buffer solutions at varied pH values. Moreover, the probe was reversible and nearly non-toxic for living cells. Then the probe was successfully used to detect pH fluctuation in living cells by exhibiting different fluorescence colors and intensity. These findings demonstrate that the probe will find useful applications in biology and biomedical research. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Chromatic aberration correction and deconvolution for UV sensitive imaging of fluorescent sterols in cytoplasmic lipid droplets

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Faergeman, Nils J

    2008-01-01

    adipocyte differentiation. DHE is targeted to transferrin-positive recycling endosomes in preadipocytes but associates with droplets in mature adipocytes. Only in adipocytes but not in foam cells fluorescent sterol was confined to the droplet-limiting membrane. We developed an approach to visualize...... macrophage foam cells and in adipocytes. We used deconvolution microscopy and developed image segmentation techniques to assess the DHE content of lipid droplets in both cell types in an automated manner. Pulse-chase studies and colocalization analysis were performed to monitor the redistribution of DHE upon...

  18. A novel fluorescence probe based on triphenylamine Schiff base for bioimaging and responding to pH and Fe3+

    International Nuclear Information System (INIS)

    Wang, Lei; Yang, Xiaodong; Chen, Xiuli; Zhou, Yuping; Lu, Xiaodan; Yan, Chenggong; Xu, Yikai; Liu, Ruiyuan; Qu, Jinqing

    2017-01-01

    A novel fluorescence probe 1 based on triphenylamine was synthesized and characterized by NMR, IR, high resolution mass spectrometry and elemental analysis. Its fluorescence was quenched when pH below 2. There was a linear relationship between the fluorescence intensity and pH value ranged from 2 to 7. And its fluorescence emission was reversibility in acidic and alkaline solution. Furthermore, it exhibited remarkable selectivity and high sensitivity to Fe 3+ and was able to detect Fe 3+ in aqueous solution with low detection limit of 0.511 μM. Job plot showed that the binding stoichiometry of 1 with Fe 3+ was 1:1. Further observations of 1 H NMR titration suggested that coordination interaction between Fe 3+ and nitrogen atom on C =N bond promoted the intramolecular charge transfer (ICT) or energy transfer process causing fluorescence quenching. Additionally, 1 was also able to be applied for detecting Fe 3+ in living cell and bioimaging. - Graphical abstract: Triphenylamine based fluorescence probe can detect pH and Fe 3+ simultaneously in aqueous solution and be applied for detecting Fe 3+ in living cell and bioimaging. - Highlights: • The fluorescence probe is sensitive to pH in strong acid conditions. • The fluorescence chemosensor can detect pH and Fe 3+ simultaneously. • The recognition is able to carry out in aqueous solution. • The probe can also be applied for detecting Fe 3+ in living cell and bioimaging. • The sensor is synthesized easily with one step.

  19. Single-component supported lipid bilayers probed using broadband nonlinear optics.

    Science.gov (United States)

    Olenick, Laura L; Chase, Hilary M; Fu, Li; Zhang, Yun; McGeachy, Alicia C; Dogangun, Merve; Walter, Stephanie R; Wang, Hong-Fei; Geiger, Franz M

    2018-01-31

    Broadband SFG spectroscopy is shown to offer considerable advantages over scanning systems in terms of signal-to-noise ratios when probing well-formed single-component supported lipid bilayers formed from zwitterionic lipids with PC headgroups. The SFG spectra obtained from bilayers formed from DOPC, POPC, DLPC, DMPC, DPPC and DSPC show a common peak at ∼2980 cm -1 , which is subject to interference between the C-H and the O-H stretches from the aqueous phase, while membranes having transition temperatures above the laboratory temperature produce SFG spectra with at least two additional peaks, one at ∼2920 cm -1 and another at ∼2880 cm -1 . The results validate spectroscopic and structural data from SFG experiments utilizing asymmetric bilayers in which one leaflet differs from the other in the extent of deuteration. Differences in H 2 O-D 2 O exchange experiments reveal that the lineshapes of the broadband SFG spectra are significantly influenced by interference from OH oscillators in the aqueous phase, even when those oscillators are not probed by the incident infrared light in our broadband setup. In the absence of spectral interference from the OH stretches of the solvent, the alkyl chain terminal methyl group of the bilayer is found to be tilted at an angle of 15° to 35° from the surface normal.

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

    International Nuclear Information System (INIS)

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

    2012-01-01

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

  1. Blue emitting copper nanoclusters as colorimetric and fluorescent probe for the selective detection of bilirubin

    Science.gov (United States)

    R. S., Aparna; J. S., Anjali Devi; John, Nebu; Abha, K.; S. S., Syamchand; George, Sony

    2018-06-01

    Hurdles to develop point of care diagnostic methods restrict the translation of progress in the health care sector from bench side to bedside. In this article a simple, cost effective fluorescent as well as colorimetric nanosensor was developed for the early and easy detection of hyperbilirubinemia. A stable, water soluble bovine serum albumin stabilised copper nanocluster (BSA CuNC) was used as the fluorescent probe which exhibited strong blue emission (404 nm) upon 330 nm excitation. The fluorescence of the BSA CuNC can be effectively quenched by the addition of bilirubin by the formation of copper-bilirubin complex. Meanwhile the copper-bilirubin complex resulted in an observable colour change from pale violet to green facilitating colorimetric detection. The prepared sensor displayed good selectivity and sensitivity over other co-existing molecules, and can be used for quantifying bilirubin with a detection limit down to 257 fM. Additionally, the as-prepared probe was coated on a paper strip to develop a portable paper strip sensor of bilirubin. Moreover, the method was successfully applied in real sample analysis and obtained promising result.

  2. Cellular Activation of the Self-Quenched Fluorescent Reporter Probe in Tumor Microenvironment

    Directory of Open Access Journals (Sweden)

    Alexei A. Bogdanov, Jr.

    2002-01-01

    Full Text Available The effect of intralysosomal proteolysis of near-infrared fluorescent (NIRF self-quenched macromolecular probe (PGC-Cy5.5 has been previously reported and used for tumor imaging. Here we demonstrate that proteolysis can be detected noninvasively in vivo at the cellular level. A codetection of GFP fluorescence (using two-photon excitation and NIRF was performed in tumor-bearing animals injected with PGC-Cy5.5. In vivo microscopy of tumor cells in subdermal tissue layers (up to 160 μm showed a strong Cy5.5 dequenching effect in GFP-negative cells. This observation was corroborated by flow cytometry, sorting, and reverse transcription polymerase chain reaction analysis of tumor-isolated cells. Both GFP-positive (81% total and GFP-negative (19% total populations contained Cy5.5-positive cells. The GFP-negative cells were confirmed to be host mouse cells by the absence of rat cathepsin mRNA signal. The subfraction of GFPnegative cells (2.5-3.0% had seven times higher NIRF intensity than the majority of GFP-positive or GFPnegative cells (372 and 55 AU, respectively. Highly NIRF-positive, FP-negative cells were CD45-and MAC3-positive. Our results indicate that: 1 intracellular proteolysis can be imaged in vivo at the cellular level using cathepsin-sensitive probes; 2 tumor-recruited cells of hematopoetic origin participate most actively in uptake and degradation of long-circulating macromolecular probes.

  3. A quick response fluorescent probe based on coumarin and quinone for glutathione and its application in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Xi [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Du, Zhi-Fang [Taishan College, Shandong University, Jinan 250100 (China); Wang, Li-Hong; Miao, Jun-Ying [Institute of Developmental Biology, School of Life Science, Shandong University, Jinan 250100 (China); Zhao, Bao-Xiang, E-mail: bxzhao@sdu.edu.cn [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China)

    2016-05-30

    We have designed and synthesized a simple but effective fluorescent probe for sensing glutathione (GSH) by PET process based on coumarin and quinone, which worked as fluorophore and reaction site, respectively. The probe could discriminate GSH from cysteine and homocysteine within 1 min in PBS-buffered solution. The sensing mechanism was confirmed by density functional theory (DFT), viscosity test, fluorescence spectrum analysis and HRMS, respectively. The probe has a low limit of detection (0.1 μM) and finally been used in cell imaging successfully. - Highlights: • This probe can discriminate glutathione from sulfhydryl compound by PET process. • This probe can be used to determine glutathione in aqueous solution within 1 min. • This probe has been successfully applied in living cell image.

  4. A quick response fluorescent probe based on coumarin and quinone for glutathione and its application in living cells

    International Nuclear Information System (INIS)

    Dai, Xi; Du, Zhi-Fang; Wang, Li-Hong; Miao, Jun-Ying; Zhao, Bao-Xiang

    2016-01-01

    We have designed and synthesized a simple but effective fluorescent probe for sensing glutathione (GSH) by PET process based on coumarin and quinone, which worked as fluorophore and reaction site, respectively. The probe could discriminate GSH from cysteine and homocysteine within 1 min in PBS-buffered solution. The sensing mechanism was confirmed by density functional theory (DFT), viscosity test, fluorescence spectrum analysis and HRMS, respectively. The probe has a low limit of detection (0.1 μM) and finally been used in cell imaging successfully. - Highlights: • This probe can discriminate glutathione from sulfhydryl compound by PET process. • This probe can be used to determine glutathione in aqueous solution within 1 min. • This probe has been successfully applied in living cell image.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1997-04-01

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

  6. Smart lanthanide coordination polymer fluorescence probe for mercury(II) determination

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Baoxia [Henan Key Laboratory Cultivation Base of Nanobiological Analytical Chemistry, College of Chemistry and Chemical Engineering, Shangqiu Normal University, Shangqiu 476000 (China); Huang, Yankai [Henan Key Laboratory Cultivation Base of Nanobiological Analytical Chemistry, College of Chemistry and Chemical Engineering, Shangqiu Normal University, Shangqiu 476000 (China); College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou 450001 (China); Zhu, Xu; Hao, Yuanqiang [Henan Key Laboratory Cultivation Base of Nanobiological Analytical Chemistry, College of Chemistry and Chemical Engineering, Shangqiu Normal University, Shangqiu 476000 (China); Ding, Yujie [College of Biochemical Engineering, Anhui Polytechnic University, Wuhu 241000 (China); Wei, Wei; Wang, Qi [Henan Key Laboratory Cultivation Base of Nanobiological Analytical Chemistry, College of Chemistry and Chemical Engineering, Shangqiu Normal University, Shangqiu 476000 (China); Qu, Peng, E-mail: qupeng0212@163.com [Henan Key Laboratory Cultivation Base of Nanobiological Analytical Chemistry, College of Chemistry and Chemical Engineering, Shangqiu Normal University, Shangqiu 476000 (China); College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou 450001 (China); Xu, Maotian, E-mail: xumaotian@sqnc.edu.cn [Henan Key Laboratory Cultivation Base of Nanobiological Analytical Chemistry, College of Chemistry and Chemical Engineering, Shangqiu Normal University, Shangqiu 476000 (China); College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou 450001 (China)

    2016-03-17

    Lanthanide coordination polymers (LCPs) have recently emerged as attractive biosensor materials due to their flexible components, high tailorable properties and unique luminescence features. In this work, we designed a smart LCP probe of Tb-CIP/AMP {(CIP, ciprofloxacin) (AMP, adenosine monophosphate)} for Hg{sup 2+} detection by using lanthanide ions as metal nodes, CIP as ligand molecule, and AMP as bridging linker and recognition unit. Tb-CIP/AMP emits strong green luminescence due to the inclusion of AMP, which withdraws the coordinated water molecules and shields Tb{sup 3+} from the quenching effect of O–H vibration in water molecules. The subsequent addition of Hg{sup 2+} into Tb-CIP/AMP can strongly quench the fluorescence because of the specific coordination interaction between AMP and Hg{sup 2+}. As a kind of Hg{sup 2+} nanosensor, the probe exhibited excellent selectivity for Hg{sup 2+} and high sensitivity with detection limit of 0.16 nM. In addition, the probe has long fluorescence lifetime up to millisecond and has been applied to detect Hg{sup 2+} in drinking water and human urine samples with satisfactory results. We envision that our strategy, in the future, could be extended to the designation of other LCP-based hypersensitive time-gated luminescence assays in biological media and biomedical imaging. - Highlights: • Lanthanide coordination polymer of Tb-CIP/AMP was synthesized via a simple self-assembly process. • AMP was employed as a bifunctional molecule for both fluorescence sensitization and target recognition. • Hypersensitive detection of Hg{sup 2+} was achieved based on time-resolved spectroscopy.

  7. Smart lanthanide coordination polymer fluorescence probe for mercury(II) determination

    International Nuclear Information System (INIS)

    Liu, Baoxia; Huang, Yankai; Zhu, Xu; Hao, Yuanqiang; Ding, Yujie; Wei, Wei; Wang, Qi; Qu, Peng; Xu, Maotian

    2016-01-01

    Lanthanide coordination polymers (LCPs) have recently emerged as attractive biosensor materials due to their flexible components, high tailorable properties and unique luminescence features. In this work, we designed a smart LCP probe of Tb-CIP/AMP {(CIP, ciprofloxacin) (AMP, adenosine monophosphate)} for Hg"2"+ detection by using lanthanide ions as metal nodes, CIP as ligand molecule, and AMP as bridging linker and recognition unit. Tb-CIP/AMP emits strong green luminescence due to the inclusion of AMP, which withdraws the coordinated water molecules and shields Tb"3"+ from the quenching effect of O–H vibration in water molecules. The subsequent addition of Hg"2"+ into Tb-CIP/AMP can strongly quench the fluorescence because of the specific coordination interaction between AMP and Hg"2"+. As a kind of Hg"2"+ nanosensor, the probe exhibited excellent selectivity for Hg"2"+ and high sensitivity with detection limit of 0.16 nM. In addition, the probe has long fluorescence lifetime up to millisecond and has been applied to detect Hg"2"+ in drinking water and human urine samples with satisfactory results. We envision that our strategy, in the future, could be extended to the designation of other LCP-based hypersensitive time-gated luminescence assays in biological media and biomedical imaging. - Highlights: • Lanthanide coordination polymer of Tb-CIP/AMP was synthesized via a simple self-assembly process. • AMP was employed as a bifunctional molecule for both fluorescence sensitization and target recognition. • Hypersensitive detection of Hg"2"+ was achieved based on time-resolved spectroscopy.

  8. Metal-organic framework based highly selective fluorescence turn-on probe for hydrogen sulphide

    Science.gov (United States)

    Nagarkar, Sanjog S.; Saha, Tanmoy; Desai, Aamod V.; Talukdar, Pinaki; Ghosh, Sujit K.

    2014-11-01

    Hydrogen sulphide (H2S) is known to play a vital role in human physiology and pathology which stimulated interest in understanding complex behaviour of H2S. Discerning the pathways of H2S production and its mode of action is still a challenge owing to its volatile and reactive nature. Herein we report azide functionalized metal-organic framework (MOF) as a selective turn-on fluorescent probe for H2S detection. The MOF shows highly selective and fast response towards H2S even in presence of other relevant biomolecules. Low cytotoxicity and H2S detection in live cells, demonstrate the potential of MOF towards monitoring H2S chemistry in biological system. To the best of our knowledge this is the first example of MOF that exhibit fast and highly selective fluorescence turn-on response towards H2S under physiological conditions.

  9. A turn-on fluorescent probe for endogenous formaldehyde in the endoplasmic reticulum of living cells

    Science.gov (United States)

    Tang, Yonghe; Ma, Yanyan; Xu, An; Xu, Gaoping; Lin, Weiying

    2017-06-01

    As the simplest aldehyde compounds, formaldehyde (FA) is implicated in nervous system diseases and cancer. Endoplasmic reticulum is an organelle that plays important functions in living cells. Accordingly, the development of efficient methods for FA detection in the endoplasmic reticulum (ER) is of great biomedical importance. In this work, we developed the first ER-targeted fluorescent FA probe Na-FA-ER. The detection is based on the condensation reaction of the hydrazine group and FA to suppress the photo-induced electron transfer (PET) pathway, resulting in a fluorescence increase. The novel Na-FA-ER showed high sensitivity to FA. In addition, the Na-FA-ER enabled the bio-imaging of exogenous and endogenous FA in living HeLa cells. Most significantly, the new Na-FA-ER was employed to visualize the endogenous FA in the ER in living cells for the first time.

  10. A benzothiazole-based fluorescent probe for distinguishing and bioimaging of Hg{sup 2+} and Cu{sup 2+}

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Biao; Huang, Liyan; Su, Wei; Duan, Xiaoli; Li, Haitao, E-mail: haitao-li@hunnu.edu.cn; Yao, Shouzhuo

    2017-02-15

    A new benzothiazole-based fluorescent probe 2-(benzo[d]thiazol-2-yl)-4-(1,3- dithian-2-yl)phenol (BT) with two different reaction sites, a thioacetal group (site 1 for Hg{sup 2+}), and O and N atoms of the benzothiazole dye (site 2 for Cu{sup 2+}), was designed and synthesized. The probe BT showed ratiometric fluorescent response to Hg{sup 2+} and fluorescence quenching behavior to Cu{sup 2+}, which induces naked-eye fluorescent color changes from green to blue and colorless, respectively. Moreover, it displayed highly sensitivity and selectivity toward Hg{sup 2+} and Cu{sup 2+} without interference from other metal ions. The sensing mechanisms were also confirmed by {sup 1}H NMR titration, mass spectrum and Job's plot analyses. Finally, probe BT was successfully used for fluorescent imaging of Hg{sup 2+} and Cu{sup 2+} in living cells, demonstrating its potential applications in biological science. - Highlights: • A benzothiazole-based probe for multiple metal ions has been firstly developed. • The differential sensing mechanisms of Hg{sup 2+} and Cu{sup 2+} relied on different reaction. • The probe could be used to monitor Hg{sup 2+} and Cu{sup 2+}in vitro and in vivo with distinct fluorescence changes.

  11. Nitrogen-Doped Carbon Quantum Dots as Fluorescent Probes for Sensitive and Selective Detection of Nitrite

    Directory of Open Access Journals (Sweden)

    Zhibiao Feng

    2017-11-01

    Full Text Available Nitrites are the upstream precursors of the carcinogenic nitrosamines, which are widely found in the natural environment and many food products. It is important to develop a simple and sensitive sensor for detecting nitrites. In this work, a fluorescence probe based on nitrogen-doped carbon quantum dots (N-CQDs was developed for the sensitive and selective determination of nitrites. At pH 2, the fluorescence of N-CQDs can be selectively quenched by nitrite due to the fact N-nitroso compounds can be formed in the reaction of amide groups with nitrous acid, which results in fluorescence static quenching. Under optimal conditions, fluorescence intensity quenching upon addition of nitrite gives a satisfactory linear relationship covering the linear range of 0.2–20 μM, and the limit of detection (LOD is 40 nM. Moreover, this method has been successfully applied to the determination of nitrites in tap water, which indicates its great potential for monitoring of nitrites in environmental samples.

  12. Fluorescent Sterols and Cholesteryl Esters as Probes for Intracellular Cholesterol Transport

    Science.gov (United States)

    Solanko, Katarzyna A.; Modzel, Maciej; Solanko, Lukasz M.; Wüstner, Daniel

    2015-01-01

    Cholesterol transport between cellular organelles comprised vesicular trafficking and nonvesicular exchange; these processes are often studied by quantitative fluorescence microscopy. A major challenge for using this approach is producing analogs of cholesterol with suitable brightness and structural and chemical properties comparable with those of cholesterol. This review surveys currently used fluorescent sterols with respect to their behavior in model membranes, their photophysical properties, as well as their transport and metabolism in cells. In the first part, several intrinsically fluorescent sterols, such as dehydroergosterol or cholestatrienol, are discussed. These polyene sterols (P-sterols) contain three conjugated double bonds in the steroid ring system, giving them slight fluorescence in ultraviolet light. We discuss the properties of P-sterols relative to cholesterol, outline their chemical synthesis, and explain how to image them in living cells and organisms. In particular, we show that P-sterol esters inserted into low-density lipoprotein can be tracked in the fibroblasts of Niemann–Pick disease using high-resolution deconvolution microscopy. We also describe fluorophore-tagged cholesterol probes, such as BODIPY-, NBD-, Dansyl-, or Pyrene-tagged cholesterol, and eventual esters of these analogs. Finally, we survey the latest developments in the synthesis and use of alkyne cholesterol analogs to be labeled with fluorophores by click chemistry and discuss the potential of all approaches for future applications. PMID:27330304

  13. Excited state hydrogen bonding fluorescent probe: Role of structure and environment

    Energy Technology Data Exchange (ETDEWEB)

    Dey, Debarati, E-mail: debaratidey07@gmail.com [Department of Chemistry, Vidyasagar College, 39 Sankar Ghosh Lane, Kolkata 700006 (India); Sarangi, Manas Kumar [Chemical Sciences Division, Saha Institute of Nuclear Physics, 1/AF, Bidhannagar, Kolkata 700064 (India); Ray, Angana; Bhattacharyya, Dhananjay [Computational Science Division, Saha Institute of Nuclear Physics, 1/AF, Bidhannagar, Kolkata 700064 (India); Maity, Dilip Kumar [Department of Chemistry, University College of Science and Technology, 92 A.P.C. Road, Kolkata 700009 (India)

    2016-05-15

    An environment sensitive fluorescent probe, 11-benzoyl-dibenzo[a,c]phenazine (BDBPZ), has been synthesized and characterized that acts via excited state hydrogen bonding (ESHB). On interaction with hydrogen bond donating solvents the fluorescence intensity of BDBPZ increases abruptly with a concomitant bathochromic shift. The extent of fluorescence increment and the red-shift of λ{sub max} depend on hydrogen bond donating ability of the solvent associated. ESHB restricts the free rotation of the benzoyl group and hence blocks the non-radiative deactivation pathway. BDBPZ forms an exciplex with organic amine in nonpolar medium that readily disappears on increasing the polarity of the solvent. In polar environment the fluorescence of both the free molecule and excited state hydrogen bonded species are quenched on addition of amine unlike its parent dibenzo[a,c]phenazine (DBPZ), that remains very much inaccessible towards the solvent as well as quencher molecules due to its structure. This newly synthesized derivative BDBPZ is much more interactive due to the benzoyl group that is flanked outside the skeletal aromatic rings of DBPZ, which helps to sense the environment properly and thus shows better ESHB capacity than DBPZ.

  14. A new simple phthalimide-based fluorescent probe for highly selective cysteine and bioimaging for living cells

    Science.gov (United States)

    Shen, Youming; Zhang, Xiangyang; Zhang, Youyu; Zhang, Chunxiang; Jin, Junling; Li, Haitao

    2017-10-01

    A new turn-on phthalimide fluorescent probe has designed and synthesized for sensing cysteine (Cys) based on excited state intramolecular proton transfer (ESIPT) process. It is consisted of a 3-hydroxyphthalimide derivative moiety as the fluorophore and an acrylic ester group as a recognition receptor. The acrylic ester acts as an ESIPT blocking agent. Upon addition of cystein, intermolecular nucleophilic attack of cysteine on acrylic ester releases the fluorescent 3-hydroxyphthalimide derivative, thereby enabling the ESIPT process and leading to enhancement of fluorescence. The probe displays high sensitivity, excellent selectivity and with large Stokes shift toward cysteine. The linear interval range of the fluorescence titration ranged from 0 to 1.0 × 10- 5 M and detection limit is low (6 × 10- 8 M). In addition, the probe could be used for bio-imaging in living cells.

  15. Fluorescent eosin probe in investigations of structural changes in glycated proteins

    Science.gov (United States)

    Pravdin, A. B.; Kochubey, V. I.; Mel'Nikov, A. G.

    2010-08-01

    The possibility of using the luminescent-kinetic probe method to investigate structural changes in bovine serum albumin (BSA) upon nonenzymatic thermal glycation is studied. An increase in the glycation time lead to a decrease in the intensity of the probe (eosin) fluorescence and to a long-wavelength shift of its maximum, as well as to an increase in the eosin phosphorescence intensity, which indicates that eosin binds to hydrophobic regions of protein at any times of incubation of BSA with glucose. From a decrease in the rate constant of the triplet-triplet energy transfer between the donor (eosin) and acceptor (anthracene) bound to proteins, it is found that the changes observed in the spectral characteristics of eosin are caused by structural changes in albumin globules as a result of glycosylation.

  16. Pentamethinium fluorescent probes: The impact of molecular structure on photophysical properties and subcellular localization

    Czech Academy of Sciences Publication Activity Database

    Bříza, T.; Rimpelová, S.; Králová, Jarmila; Záruba, K.; Kejík, Z.; Ruml, T.; Martásek, P.; Král, V.

    2014-01-01

    Roč. 107, August 2014 (2014), s. 51-59 ISSN 0143-7208 R&D Projects: GA AV ČR KAN200100801; GA ČR(CZ) GAP303/11/1291; GA MŠk(CZ) CZ.01.05/2.1.00/01.00.30; GA MŠk(CZ) CZ.1.07/2.300/30.0060 Institutional support: RVO:68378050 Keywords : Pentamethinium salts * Fluorescent probes * Mitochondria * Cardiolipin Photostability * Organelle imaging Subject RIV: CE - Biochemistry Impact factor: 3.966, year: 2014

  17. Synthesis of improved upconversion nanoparticles as ultrasensitive fluorescence probe for mycotoxins

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Quansheng, E-mail: q.s.chen@hotmail.com; Hu, Weiwei; Sun, Cuicui; Li, Huanhuan; Ouyang, Qin

    2016-09-28

    Rare earth-doped upconversion nanoparticles (UCNPs) have promising potentials in biodetection due to their unique frequency upconverting capability and high detection sensitivity. This paper reports an improved UCNPs-based fluorescence probe for dual-sensing of Aflatoxin B1 (AFB1) and Deoxynivalenol (DON) using a magnetism-induced separation and the specific formation of antibody-targets complex. Herein, the improved UCNPs, which were namely NaYF{sub 4}:Yb/Ho/Gd and NaYF{sub 4}:Yb/Tm/Gd, were systematically studied based on the optimization of reaction time, temperature and the concentration of dopant ions with simultaneous phase and size controlled NaYF{sub 4} nanoparticles; and the targets were detected using the pattern of competitive combination assay. Under an optimized condition, the advanced fluorescent probes revealed stronger fluorescent properties, broader biological applications and better storage stabilities compared to traditional UCNPs-based ones; and ultrasensitive determinations of AFB1 and DON were achieved under a wide sensing range of 0.001–0.1 ng ml{sup −1} with the limit of detection (LOD) of 0.001 ng ml{sup −1}. Additionally, the applicability of the improved nanosensor for the detection of mycotoxins was also confirmed in adulterated oil samples. - Highlights: • Improved rare earth-doped upconversion nanoparticles were prepared with detailed optimizations. • Setup of an upconversion fluorescence spectrometer. • An advanced UCNPs-based immunosensor for dual-sensing mycotoxins was developed with a LOD of 0.001 ng ml{sup −1}. • Application of this biosensor to detect targets in real samples were confirmed with satisfied results.

  18. Non-Covalent Fluorescent Labeling of Hairpin DNA Probe Coupled with Hybridization Chain Reaction for Sensitive DNA Detection.

    Science.gov (United States)

    Song, Luna; Zhang, Yonghua; Li, Junling; Gao, Qiang; Qi, Honglan; Zhang, Chengxiao

    2016-04-01

    An enzyme-free signal amplification-based assay for DNA detection was developed using fluorescent hairpin DNA probes coupled with hybridization chain reaction (HCR). The hairpin DNAs were designed to contain abasic sites in the stem moiety. Non-covalent labeling of the hairpin DNAs was achieved when a fluorescent ligand was bound to the abasic sites through hydrogen bonding with the orphan cytosine present on the complementary strand, accompanied by quench of ligand fluorescence. As a result, the resultant probes, the complex formed between the hairpin DNA and ligand, showed almost no fluorescence. Upon hybridization with target DNA, the probe underwent a dehybridization of the stem moiety containing an abasic site. The release of ligand from the abasic site to the solution resulted in an effective fluorescent enhancement, which can be used as a signal. Compared with a sensing system without HCR, a 20-fold increase in the sensitivity was achieved using the sensing system with HCR. The fluorescent intensity of the sensing system increased with the increase in target DNA concentration from 0.5 nM to 100 nM. A single mismatched target ss-DNA could be effectively discriminated from complementary target DNA. Genotyping of a G/C single-nucleotide polymorphism of polymerase chain reaction (PCR) products was successfully demonstrated with the sensing system. Therefore, integrating HCR strategy with non-covalent labeling of fluorescent hairpin DNA probes provides a sensitive and cost-effective DNA assay. © The Author(s) 2016.

  19. A flavone-based turn-on fluorescent probe for intracellular cysteine/homocysteine sensing with high selectivity.

    Science.gov (United States)

    Zhang, Jian; Lv, Yanlin; Zhang, Wei; Ding, Hui; Liu, Rongji; Zhao, Yongsheng; Zhang, Guangjin; Tian, Zhiyuan

    2016-01-01

    A new type of flavone-based fluorescent probe (DMAF) capable of cysteine (Cys)/homocysteine (Hcy) sensing with high selectivity over other amino acids was developed. Such type of probe undergoes Cys/Hcy-mediated cyclization reaction with the involvement of its aldehyde group, which suppresses of the photoinduced electron transfer (PET) process of the probe molecule and consequently leads to the enhancement of fluorescence emission upon excitation using visible light. The formation of product of the Cys/Hcy-mediated cyclization reaction was confirmed and the preliminary fluorescence imaging experiments revealed the biocompatibility of the as-prepared probe and validated its practicability for intracellular Cys/Hcy sensing. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. “Turn-on” fluorescence probe integrated polymer nanoparticles for sensing biological thiol molecules

    Science.gov (United States)

    Ang, Chung Yen; Tan, Si Yu; Lu, Yunpeng; Bai, Linyi; Li, Menghuan; Li, Peizhou; Zhang, Quan; Selvan, Subramanian Tamil; Zhao, Yanli

    2014-11-01

    A ``turn-on'' thiol-responsive fluorescence probe was synthesized and integrated into polymeric nanoparticles for sensing intracellular thiols. There is a photo-induced electron transfer process in the off state of the probe, and this process is terminated upon the reaction with thiol compounds. Configuration interaction singles (CIS) calculation was performed to confirm the mechanism of this process. A series of sensing studies were carried out, showing that the probe-integrated nanoparticles were highly selective towards biological thiol compounds over non-thiolated amino acids. Kinetic studies were also performed to investigate the relative reaction rate between the probe and the thiolated amino acids. Subsequently, the Gibbs free energy of the reactions was explored by means of the electrochemical method. Finally, the detection system was employed for sensing intracellular thiols in cancer cells, and the sensing selectivity could be further enhanced with the use of a cancer cell-targeting ligand in the nanoparticles. This development paves a path for the sensing and detection of biological thiols, serving as a potential diagnostic tool in the future.

  1. Far-red fluorescent probes for canonical and non-canonical nucleic acid structures: current progress and future implications.

    Science.gov (United States)

    Suseela, Y V; Narayanaswamy, Nagarjun; Pratihar, Sumon; Govindaraju, Thimmaiah

    2018-02-05

    The structural diversity and functional relevance of nucleic acids (NAs), mainly deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), are indispensable for almost all living organisms, with minute aberrations in their structure and function becoming causative factors in numerous human diseases. The standard structures of NAs, termed canonical structures, are supported by Watson-Crick hydrogen bonding. Under special physiological conditions, NAs adopt distinct spatial organisations, giving rise to non-canonical conformations supported by hydrogen bonding other than the Watson-Crick type; such non-canonical structures have a definite function in controlling gene expression and are considered as novel diagnostic and therapeutic targets. Development of molecular probes for these canonical and non-canonical DNA/RNA structures has been an active field of research. Among the numerous probes studied, probes with turn-on fluorescence in the far-red (600-750 nm) region are highly sought-after due to minimal autofluorescence and cellular damage. Far-red fluorescent probes are vital for real-time imaging of NAs in live cells as they provide good resolution and minimal perturbation of the cell under investigation. In this review, we present recent advances in the area of far-red fluorescent probes of DNA/RNA and non-canonical G-quadruplex structures. For the sake of continuity and completeness, we provide a brief overview of visible fluorescent probes. Utmost importance is given to design criteria, characteristic properties and biological applications, including in cellulo imaging, apart from critical discussion on limitations of the far-red fluorescent probes. Finally, we offer current and future prospects in targeting canonical and non-canonical NAs specific to cellular organelles, through sequence- and conformation-specific far-red fluorescent probes. We also cover their implications in chemical and molecular biology, with particular focus on decoding various disease

  2. Activation of 5-[125I]iodonaphthyl-1-azide via excitation of fluorescent (N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)) lipid analogs in living cells. A potential tool for identification of compartment-specific proteins and proteins involved in intracellular transport and metabolism of lipids

    International Nuclear Information System (INIS)

    Rosenwald, A.G.; Pagano, R.E.; Raviv, Y.

    1991-01-01

    We describe a new technique for analysis of proteins located near fluorescent lipid analogs in intact living cells using the membrane-permeant, photoactivatable probe, 5-[ 125 I]iodonaphthyl-1-azide ([ 125 I]INA). [ 125 I] INA can be activated directly with UV light or indirectly through excitation of adjacent fluorophores (photosensitizers) with visible light to modify nearby proteins covalently with 125 I. In this report we demonstrate that fluorescent phospholipids and sphingolipids containing N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-6-aminocaproic acid serve as appropriate photosensitizers for [ 125 I]INA. Using Chinese hamster ovary fibroblasts, we optimized the labeling conditions with respect to lipid concentration and time of irradiation and then examined the profiles of cellular proteins that were labeled when fluorescent analogs of ceramide, sphingomyelin, and phosphatidic acid were used as photosensitizers in living cells. The use of different fluorescent lipids, which label different subcellular compartments of cells as determined by fluorescence microscopy, derivatized different sets of cellular proteins with 125 I. The labeled proteins were subsets of the total set of proteins available for derivatization as determined by direct activation of [ 125 I]INA. Most proteins labeled by this procedure were pelleted by centrifugation of cell lysates at high speed (260,000 x g), but several soluble proteins were also labeled under these conditions. The implications of using this technique for identification of compartment-specific proteins and proteins involved in lipid metabolism and transport are discussed

  3. A homogeneous and “off–on” fluorescence aptamer-based assay for chloramphenicol using vesicle quantum dot-gold colloid composite probes

    Energy Technology Data Exchange (ETDEWEB)

    Miao, Yang-Bao [State Key Laboratory Base of Novel Functional Materials and Preparation Science, Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo 315211 (China); Ren, Hong-Xia [Key Laboratory of Asymmetric Synthesis and Chirotechnology of Sichuan Province, Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); University of Chinese Academy of Sciences, Beijing 10049 (China); Gan, Ning, E-mail: ganning@nbu.edu.cn [State Key Laboratory Base of Novel Functional Materials and Preparation Science, Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo 315211 (China); Zhou, You [State Key Laboratory Base of Novel Functional Materials and Preparation Science, Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo 315211 (China); Cao, Yuting, E-mail: caoyuting@nbu.edu.cn [State Key Laboratory Base of Novel Functional Materials and Preparation Science, Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo 315211 (China); Li, Tianhua [State Key Laboratory Base of Novel Functional Materials and Preparation Science, Faculty of Materials Science and Chemical Engineering, Ningbo University, Ningbo 315211 (China); Chen, Yinji [Faculty of Food Science and Engineering, Nanjing University of Finance and Economics, Nanjing 210000 (China)

    2016-07-27

    In this work, a novel homogeneous and signal “off–on” aptamer based fluorescence assay was successfully developed to detect chloramphenicol (CAP) residues in food based on the fluorescence resonance energy transfer (FRET). The vesicle nanotracer was prepared through labeling single stranded DNA binding protein (SSB) on limposome-CdSe/ZnS quantum dot (SSB/L-QD) complexes. It was worth mentioning that the signal tracer (SSB/L-QD) with vesicle shape, which was fabricated being encapsulated with a number of quantum dots and SSB. The nanotracer has excellent signal amplification effects. The vesicle composite probe was formed by combining aptamer labeled nano-gold (Au-Apt) and SSB/L-QD. Which based on SSB's specific affinity towards aptamer. This probe can't emit fluoresce which is in “off” state because the signal from SSB/L-QD as donor can be quenched by the Au-aptas acceptor. When CAP was added in the composite probe solution, the aptamer on the Au-Apt can be preferentially bounded with CAP then release from the composite probe, which can turn the “off” signal of SSB/L-QD tracer into “on” state. The assay indicates excellent linear response to CAP from 0.001 nM to 10 nM and detection limit down to 0.3 pM. The vesicle probes with size of 88 nm have strong signal amplification. Because a larger number of QDs can be labeled inside the double phosphorus lipid membrane. Besides, it was employed to detect CAP residues in the milk samples with results being agreed well with those from ELISA, verifying its accuracy and reliability. - Highlights: • Homogeneous and “off–on” fluorescence aptamer-based assay was developed to detect chloramphenicol (CAP) residues in food. • This probe was fabricated based on a vesicle QDs signal tracer (SSB/L-QD) combining with Au-Aptamer. • The detection mechanism was based on FRET with high specificity. • The results for CAP detection in the milk samples agreed well with those from ELISA, while

  4. Lateral mobility of plasma membrane lipids in Xenopus eggs: Regional differences related to animal/vegetal polarity

    NARCIS (Netherlands)

    Laat, S.W. de; Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van; Tetteroo, P.A.T.; Tertoolen, L.G.J.

    1984-01-01

    Regional differences in the lateral mobility properties of plasma membrane lipids were studied in unfertilized and fertilized Xenopus eggs by fluorescence photobleaching recovery (FPR) measurements. Out of a variety of commonly used lipid probes only the aminofluorescein- -1abelled fatty

  5. In vivo fluorescence imaging of hepatocellular carcinoma using a novel GPC3-specific aptamer probe

    Science.gov (United States)

    Zhao, Menglong; Dong, Lili; Liu, Zhuang; Yang, Shuohui

    2018-01-01

    Background Glypican-3 (GPC3) is highly expressed in most of the hepatocellular carcinomas (HCCs), even in small HCCs. It may be used as a potential biomarker for early detection of HCC. The aptamer is a promising targeting agent with unique advantages over antibody. This study was to introduce a novel GPC3 specific aptamer (AP613-1), to verify its specific binding property in vitro, and to evaluate its targeting efficiency in vivo by performing near-infrared (NIR) fluorescence imaging on an HCC xenograft model. Methods AP613-1 was generated from the systematic evolution of ligands by exponential enrichment. Flow cytometry and aptamer-based immunofluorescence imaging were performed to verify the binding affinity of AP613-1 to GPC3 in vitro. NIR Fluorescence images of nude mice with unilateral (n=12) and bilateral (n=4) subcutaneous xenograft tumors were obtained. Correlation between the tumor fluorescence intensities in vivo and ex vivo was analyzed. Results AP613-1 could specifically bind to GPC3 in vitro. In vivo and ex vivo tumors, fluorescence intensities were in excellent correlation (Pfluorescence intensity is significantly higher in tumors given Alexa Fluor 750 (AF750) labeled AP613-1 than in those given AF750 labeled initial ssDNA library both in vivo (Pfluorescence intensities than A549 tumors both in vivo (P=0.016) and ex vivo (P=0.004). Conclusions AP613-1 displays a specific binding affinity to GPC3 positive HCC. Fluorescently labeled AP613-1 could be used as an imaging probe to subcutaneous HCC in xenograft models. PMID:29675356

  6. Fluorescence probes for real-time remote cyanobacteria monitoring: A review of challenges and opportunities.

    Science.gov (United States)

    Bertone, Edoardo; Burford, Michele A; Hamilton, David P

    2018-05-10

    In recent years, there has been a widespread deployment of submersible fluorescence sensors by water utilities. They are used to measure diagnostic pigments and estimate algae and cyanobacteria abundance in near real-time. Despite being useful and promising tools, operators and decision-makers often rely on the data provided by these probes without a full understanding of their limitations. As a result, this may lead to wrong and misleading estimations which, in turn, means that researchers and technicians distrust these sensors. In this review paper, we list and discuss the main limitations of such probes, as well as identifying the effect of environmental factors on pigment production, and in turn, the conversion to cyanobacteria abundance estimation. We argue that a comprehensive calibration approach to obtain reliable readings goes well beyond manufacturers' recommendations, and should involve several context-specific experiments. We also believe that if such a comprehensive set of experiments is conducted, the data collected from fluorescence sensors could be used in artificial intelligence modelling approaches to reliably predict, in near real-time, the presence and abundance of different cyanobacteria species. This would have significant benefits for both drinking and recreational water management, given that cyanobacterial toxicity, and taste and odour compounds production, are species-dependent. Copyright © 2018 Elsevier Ltd. All rights reserved.

  7. Challenges for Super-Resolution Localization Microscopy and Biomolecular Fluorescent Nano-Probing in Cancer Research

    Science.gov (United States)

    Ilić, Nataša; Pilarczyk, Götz; Lee, Jin-Ho; Logeswaran, Abiramy; Borroni, Aurora Paola; Krufczik, Matthias; Theda, Franziska; Waltrich, Nadine; Bestvater, Felix; Hildenbrand, Georg; Cremer, Christoph; Blank, Michael

    2017-01-01

    Understanding molecular interactions and regulatory mechanisms in tumor initiation, progression, and treatment response are key requirements towards advanced cancer diagnosis and novel treatment procedures in personalized medicine. Beyond decoding the gene expression, malfunctioning and cancer-related epigenetic pathways, investigations of the spatial receptor arrangements in membranes and genome organization in cell nuclei, on the nano-scale, contribute to elucidating complex molecular mechanisms in cells and tissues. By these means, the correlation between cell function and spatial organization of molecules or molecular complexes can be studied, with respect to carcinogenesis, tumor sensitivity or tumor resistance to anticancer therapies, like radiation or antibody treatment. Here, we present several new applications for bio-molecular nano-probes and super-resolution, laser fluorescence localization microscopy and their potential in life sciences, especially in biomedical and cancer research. By means of a tool-box of fluorescent antibodies, green fluorescent protein (GFP) tagging, or specific oligonucleotides, we present tumor relevant re-arrangements of Erb-receptors in membranes, spatial organization of Smad specific ubiquitin protein ligase 2 (Smurf2) in the cytosol, tumor cell characteristic heterochromatin organization, and molecular re-arrangements induced by radiation or antibody treatment. The main purpose of this article is to demonstrate how nano-scaled distance measurements between bio-molecules, tagged by appropriate nano-probes, can be applied to elucidate structures and conformations of molecular complexes which are characteristic of tumorigenesis and treatment responses. These applications open new avenues towards a better interpretation of the spatial organization and treatment responses of functionally relevant molecules, at the single cell level, in normal and cancer cells, offering new potentials for individualized medicine. PMID:28956810

  8. Challenges for Super-Resolution Localization Microscopy and Biomolecular Fluorescent Nano-Probing in Cancer Research

    Directory of Open Access Journals (Sweden)

    Michael Hausmann

    2017-09-01

    Full Text Available Understanding molecular interactions and regulatory mechanisms in tumor initiation, progression, and treatment response are key requirements towards advanced cancer diagnosis and novel treatment procedures in personalized medicine. Beyond decoding the gene expression, malfunctioning and cancer-related epigenetic pathways, investigations of the spatial receptor arrangements in membranes and genome organization in cell nuclei, on the nano-scale, contribute to elucidating complex molecular mechanisms in cells and tissues. By these means, the correlation between cell function and spatial organization of molecules or molecular complexes can be studied, with respect to carcinogenesis, tumor sensitivity or tumor resistance to anticancer therapies, like radiation or antibody treatment. Here, we present several new applications for bio-molecular nano-probes and super-resolution, laser fluorescence localization microscopy and their potential in life sciences, especially in biomedical and cancer research. By means of a tool-box of fluorescent antibodies, green fluorescent protein (GFP tagging, or specific oligonucleotides, we present tumor relevant re-arrangements of Erb-receptors in membranes, spatial organization of Smad specific ubiquitin protein ligase 2 (Smurf2 in the cytosol, tumor cell characteristic heterochromatin organization, and molecular re-arrangements induced by radiation or antibody treatment. The main purpose of this article is to demonstrate how nano-scaled distance measurements between bio-molecules, tagged by appropriate nano-probes, can be applied to elucidate structures and conformations of molecular complexes which are characteristic of tumorigenesis and treatment responses. These applications open new avenues towards a better interpretation of the spatial organization and treatment responses of functionally relevant molecules, at the single cell level, in normal and cancer cells, offering new potentials for individualized medicine.

  9. Diffusion studies on permeable nitroxyl spin probe through lipid bilayer membrane

    International Nuclear Information System (INIS)

    Benial, A. Milton Franklin; Meenakumari, V.; Ichikawa, Kazuhiro; Yamada, Ken-ichi; Utsumi, Hideo; Hyodo, Fuminori; Jawahar, A.

    2014-01-01

    Electron spin resonance (ESR) studies were carried out for 2mM 14 N labeled deutrated permeable 3- methoxycarbonyl-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl (MC-PROXYL) in pure water, 1 mM, 2 mM, 3 mM and 4 mM concentration of MC-PROXYL in 300 mM concentration of liposomal solution by using a L-band ESR spectrometer. The ESR parameters such as linewidth, hyperfine coupling constant, g-factor, partition parameter and permeability were reported. The partition parameter and permeability values indicate the maximum spin distribution in the lipid phase at 2 mM concentration. This study illustrates that ESR can be used to differentiate between the intra and extra-membrane water by loading the liposome vesicles with a lipid-permeable nitroxyl spin probe. From the ESR results, the radical concentration was optimized as 2 mM in liposomal solution for ESR phantom studies and experiments

  10. Probing GFP-actin diffusion in living cells using fluorescence correlation spectroscopy

    International Nuclear Information System (INIS)

    Engelke, Hanna; Heinrich, Doris; Rädler, Joachim O.

    2010-01-01

    The cytoskeleton of eukaryotic cells is continuously remodeled by polymerization and depolymerization of actin. Consequently, the relative content of polymerized filamentous actin (F-actin) and monomeric globular actin (G-actin) is subject to temporal and spatial fluctuations. Since fluorescence correlation spectroscopy (FCS) can measure the diffusion of fluorescently labeled actin it seems likely that FCS allows us to determine the dynamics and hence indirectly the structural properties of the cytoskeleton components with high spatial resolution. To this end we investigate the FCS signal of GFP-actin in living Dictyostelium discoideum cells and explore the inherent spatial and temporal signatures of the actin cytoskeleton. Using the free green fluorescent protein (GFP) as a reference, we find that actin diffusion inside cells is dominated by G-actin and slower than diffusion in diluted cell extract. The FCS signal in the dense cortical F-actin network near the cell membrane is probed using the cytoskeleton protein LIM and is found to be slower than cytosolic G-actin diffusion. Furthermore, we show that polymerization of the cytoskeleton induced by Jasplakinolide leads to a substantial decrease of G-actin diffusion. Pronounced fluctuations in the distribution of the FCS correlation curves can be induced by latrunculin, which is known to induce actin waves. Our work suggests that the FCS signal of GFP-actin in combination with scanning or spatial correlation techniques yield valuable information about the local dynamics and concomitant cytoskeletal properties

  11. Determination of the activity of telomerase in cancer cells by using BSA-protected gold nanoclusters as a fluorescent probe.

    Science.gov (United States)

    Xu, Yujuan; Zhang, Peng; Wang, Zhen; Lv, Shaoping; Ding, Caifeng

    2018-02-27

    Gold nanoclusters (AuNCs) protected with a bovine serum albumin (BSA) coating are known to emit red fluorescence (peaking at 650 nm) on photoexcitation with ultraviolet light (365 nm). On addition of Cu(II) ions, fluorescence is quenched because Cu(II) complexes certain amino acid units in the BSA chain. Fluorescence is, however, restored if pyrophosphate (PPi) is added because it will chelate Cu(II) and remove it from the BSA coating on the AuNCs. Because PPi is involved in the function of telomerase, the BSA@AuNCs loaded with Cu(II) can act as a fluorescent probe for determination of the activity of telomerase. A fluorescent assay was worked out for telomerase that is highly sensitive and has a wide linear range (10 nU to 10 fM per mL). The fluorescent probe was applied to the determination of telomerase activity in cervix carcinoma cells via imaging. It is shown that tumor cells can be well distinguished from normal cells by monitoring the differences in intracellular telomerase activity. Graphical abstract Gold nanoclusters (AuNCs) protected by bovine serum albumin (BSA) and displaying red photoluminescence were prepared as fluorescent probe for the determination of telomerase activity and used for imaging of cervix carcinoma (HeLa) cells.

  12. Direct fluorescence in situ hybridization on human metaphase chromosomes using quantum dot-platinum labeled DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Gyoyeon [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of); Lee, Hansol [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Lee, Jiyeon, E-mail: jylee@kist.re.kr [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of)

    2015-11-13

    The telomere shortening in chromosomes implies the senescence, apoptosis, or oncogenic transformation of cells. Since detecting telomeres in aging and diseases like cancer, is important, the direct detection of telomeres has been a very useful biomarker. We propose a telomere detection method using a newly synthesized quantum dot (QD) based probe with oligonucleotide conjugation and direct fluorescence in situ hybridization (FISH). QD-oligonucleotides were prepared with metal coordination bonding based on platinum-guanine binding reported in our previous work. The QD-oligonucleotide conjugation method has an advantage where any sequence containing guanine at the end can be easily bound to the starting QD-Pt conjugate. A synthesized telomeric oligonucleotide was bound to the QD-Pt conjugate successfully and this probe hybridized specifically on the telomere of fabricated MV-4-11 and MOLT-4 chromosomes. Additionally, the QD-telomeric oligonucleotide probe successfully detected the telomeres on the CGH metaphase slide. Due to the excellent photostability and high quantum yield of QDs, the QD-oligonucleotide probe has high fluorescence intensity when compared to the organic dye-oligonucleotide probe. Our QD-oligonucleotide probe, conjugation method of this QD probe, and hybridization protocol with the chromosomes can be a useful tool for chromosome painting and FISH. - Highlights: • We prepared a probe linked between QD and telomeric oligonucleotide with platinum-guanine bonding. • Telomeres were detected by our new telomere probes successfully in three different human metaphase chromosomes. • QDPt-DNA probe has high fluorescence intensity in comparison with organic dye-DNA probe.

  13. Electric dipole moments of the fluorescent probes Prodan and Laurdan: experimental and theoretical evaluations.

    Science.gov (United States)

    Vequi-Suplicy, Cíntia C; Coutinho, Kaline; Lamy, M Teresa

    2014-03-01

    Several experimental and theoretical approaches can be used for a comprehensive understanding of solvent effects on the electronic structure of solutes. In this review, we revisit the influence of solvents on the electronic structure of the fluorescent probes Prodan and Laurdan, focusing on their electric dipole moments. These biologically used probes were synthesized to be sensitive to the environment polarity. However, their solvent-dependent electronic structures are still a matter of discussion in the literature. The absorption and emission spectra of Prodan and Laurdan in different solvents indicate that the two probes have very similar electronic structures in both the ground and excited states. Theoretical calculations confirm that their electronic ground states are very much alike. In this review, we discuss the electric dipole moments of the ground and excited states calculated using the widely applied Lippert-Mataga equation, using both spherical and spheroid prolate cavities for the solute. The dimensions of the cavity were found to be crucial for the calculated dipole moments. These values are compared to those obtained by quantum mechanics calculations, considering Prodan in vacuum, in a polarizable continuum solvent, and using a hybrid quantum mechanics-molecular mechanics methodology. Based on the theoretical approaches it is evident that the Prodan dipole moment can change even in the absence of solute-solvent-specific interactions, which is not taken into consideration with the experimental Lippert-Mataga method. Moreover, in water, for electric dipole moment calculations, it is fundamental to consider hydrogen-bonded molecules.

  14. Nitroolefin-based BODIPY as a novel water-soluble ratiometric fluorescent probe for detection of endogenous thiols

    Science.gov (United States)

    Kang, Jin; Huo, Fangjun; Chao, Jianbin; Yin, Caixia

    2018-04-01

    Small molecule biothiols, including cysteine (Cys), homocysteine (Hcy), and glutathione (GSH), play many crucial roles in physiological processes. In this work, we have prepared a nitroolefin-based BODIPY fluorescent probe with excellent water solubility for detection thiols, which displayed ratiometric fluorescent signal for thiols. Incorporation of a nitroolefin unit to the BODIPY dye would transform it into a strong Michael acceptor, which would be highly susceptible to sulfhydryl nucleophiles. This probe shows an obvious ratio change upon response with thiols, an increase of the emission at 517 nm along with a concomitant decrease of fluorescence peak at 573 nm. Moreover, these successes of intracellular imaging experiments in A549 cells indicated that this probe is suitable for imaging of ex-/endogenous thiols in living cells.

  15. A new rhodamine-based fluorescent probe for the discrimination of Fe{sup 3+} from Fe{sup 2+}

    Energy Technology Data Exchange (ETDEWEB)

    You, Qi Hua; Huang, Hua Bin; Zhuang, Zhi Xia; Wang, Xiao Ru [Dept. of Science and Technology for Inspection, Xiamen Huaxia University, Xiamen (China); Chan, Wing Hong [Dept. of of Chemistry, Hong Kong Bap tist University, Hong Kong (China)

    2016-11-15

    A new rhodamine-based fluorescent probe for the discrimination of Fe{sup 3+} from Fe{sup 2+} has been designed and investigated. The probe shows an immediate visual color change in response to Fe{sup 3+} and Cu{sup 2+}, while only Fe{sup 3+} triggers the fluorescent change of the probe. The existence of large amount of other metal ions shows negligible interference in the detection of Fe{sup 3+}. The association constant K{sub ass} of 4.64 × 10{sup 8} M{sup -2} (R{sup 2} = 0.994) and 5.38 × 10{sup 8} M{sup -2} (R{sup 2} = 0.991) of the complex was derived from UV/Vis and fluorescence titration assuming 1:2 stoichiometry of probe–Fe{sup 3+} complex, respectively.

  16. Intravascular atherosclerotic imaging with combined fluorescence and optical coherence tomography probe based on a double-clad fiber combiner

    Science.gov (United States)

    Liang, Shanshan; Saidi, Arya; Jing, Joe; Liu, Gangjun; Li, Jiawen; Zhang, Jun; Sun, Changsen; Narula, Jagat; Chen, Zhongping

    2012-07-01

    We developed a multimodality fluorescence and optical coherence tomography probe based on a double-clad fiber (DCF) combiner. The probe is composed of a DCF combiner, grin lens, and micromotor in the distal end. An integrated swept-source optical coherence tomography and fluorescence intensity imaging system was developed based on the combined probe for the early diagnoses of atherosclerosis. This system is capable of real-time data acquisition and processing as well as image display. For fluorescence imaging, the inflammation of atherosclerosis and necrotic core formed with the annexin V-conjugated Cy5.5 were imaged. Ex vivo imaging of New Zealand white rabbit arteries demonstrated the capability of the combined system.

  17. A novel NBD-based fluorescent turn-on probe for the detection of cysteine and homocysteine in living cells

    Science.gov (United States)

    Wang, Jiamin; Niu, Linqiang; Huang, Jing; Yan, Zhijie; Wang, Jianhong

    2018-03-01

    Biothiols, such as cysteine (Cys), homocysteine (Hcy) and glutathione (GSH), are involved in a number of biological processes and play crucial roles in biological systems. Thus, the detection of biothiols is highly important for early diagnosis of diseases and evaluation of disease progression. Herein, we developed a new turn-on fluorescent probe 1 based on 7-nitro-2,1,3-benzoxadiazole (NBD) with high selectivity and sensitivity for Cys/Hcy on account of nucleophilic substitution and Smiles rearrangement reaction. The probe could sense Cys/Hcy rapidly, the intensity of fluorescence increased immediately within 1 min. Furthermore, the probe is low toxic and has been successfully applied to detect intracellular Cys/Hcy by cell fluorescence imaging in living normal and cancer cells.

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

    Directory of Open Access Journals (Sweden)

    Ji Yeon Hwang

    2016-11-01

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

  19. Mismatch discrimination of lipidated DNA and LNA-probes (LiNAs) in hybridization-controlled liposome assembly

    DEFF Research Database (Denmark)

    Jakobsen, Ulla; Vogel, Stefan

    2016-01-01

    Assays for mismatch discrimination and detection of single nucleotide variations by hybridization-controlled assembly of liposomes, which do not require tedious surface chemistry, are versatile for both DNA and RNA targets. We report herein a comprehensive study on different DNA and LNA (locked...... assay in the context of mismatch discrimination and SNP detection are presented. The advantages of membrane-anchored LiNA-probes compared to chemically attached probes on solid nanoparticles (e.g. gold nanoparticles) are described. Key functionalities such as non-covalent attachment of LiNA probes...... without the need for long spacers and the inherent mobility of membrane-anchored probes in lipid-bilayer membranes will be described for several different probe designs....

  20. Highly selective and sensitive detection of Cu2+ with lysine enhancing bovine serum albumin modified-carbon dots fluorescent probe.

    Science.gov (United States)

    Liu, Jia-Ming; Lin, Li-ping; Wang, Xin-Xing; Lin, Shao-Qin; Cai, Wen-Lian; Zhang, Li-Hong; Zheng, Zhi-Yong

    2012-06-07

    Based on the ability of lysine (Lys) to enhance the fluorescence intensity of bovine serum albumin modified-carbon dots (CDs-BSA) to decrease surface defects and quench fluorescence of the CDs-BSA-Lys system in the presence of Cu(2+) under conditions of phosphate buffer (PBS, pH = 5.0) at 45 °C for 10 min, a sensitive Lys enhancing CDs-BSA fluorescent probe was designed. The environment-friendly, simple, rapid, selective and sensitive fluorescent probe has been utilized to detect Cu(2+) in hair and tap water samples and it achieved consistent results with those obtained by inductively coupled plasma mass spectroscopy (ICP-MS). The mechanism of the proposed assay for the detection of Cu(2+) is discussed.

  1. Photophysical properties of a surfactive long-chain styryl merocyanine dye as fluorescent probe

    Energy Technology Data Exchange (ETDEWEB)

    Ismail, L.F.M., E-mail: Lailafmi@yahoo.com [Al-Azhar University, Faculty of Science, Chemistry Department, Nasr City, 12 Ibrahim El-Nagar, El-Hegaz Sq. Heliopolis, Cairo 11315 (Egypt)

    2012-09-15

    environment, which have often been used as models for more complex bioaggregates. The results point out to a possible use of this dye as a fluorescence probe for microenvironmental parameters as well as in some micellar systems.

  2. Design and Investigation of Optical Properties of N-(Rhodamine-B)-Lactam-Ethylenediamine (RhB-EDA) Fluorescent Probe.

    Science.gov (United States)

    Soršak, Eva; Volmajer Valh, Julija; Korent Urek, Špela; Lobnik, Aleksandra

    2018-04-14

    This study presents chemical modification of a Rhodamine B (RhB) sensor probe by ethylenediamine (EDA), and investigation of its spectral as well as sensor properties to the various metals. The synthesised N -(Rhodamine-B)-lactam-ethylenediamine (RhB-EDA) fluorescent probe shows interesting optical sensor properties, and high sensitivity and selectivity to Ag⁺ ions among all the tested metal ions (K⁺, Mg 2+ , Cu 2+ , Ni 2+ , Fe 2+ , Pb 2+ , Na⁺, Mn 2+ , Li⁺, Al 3+ , Co 2+ , Hg 2+ , Sr 2+ , Ca 2+ , Ag⁺, Cd 2+ and Zn 2+ ), while the well-known Rhodamine B (RhB) fluorescent probe shows much less sensitivity to Ag⁺ ions, but high sensitivity to Fe 2+ ions. The novel fluorescent sensor probe RhB-EDA has the capabilities to sense Ag⁺ ions up to µM ranges by using the fluorescence quenching approach. The probe displayed a dynamic response to Ag⁺ in the range of 0.43 × 10 -3 -10 -6 M with a detection limit of 0.1 μM. The sensing system of an RhB-EDA novel fluorescent probe was optimised according to the spectral properties, effect of pH and buffer, photostability, incubation time, sensitivity, and selectivity. Since all the spectral and sensing properties were tested in green aqueous media, although many other similar sensor systems rely on organic solvent solutions, the RhB-EDA sensing probe may be a good candidate for measuring Ag⁺ ions in real-life applications.

  3. Stratum corneum lipid organization as observed by atomic force, confocal and two-photon excitation fluorescence microscopy

    DEFF Research Database (Denmark)

    Norlén, Lars; Plasencia Gil, Maria Inés; Bagatolli, Luis

    2008-01-01

    -related biophysical techniques (e.g. atomic force microscopy and confocal/two-photon excitation fluorescence microscopy), it was recently shown that reconstituted membranes composed of extracted decontaminated human stratum corneum lipids do not form a fluid phase, but exclusively a single-gel phase that segregates...

  4. Organic liquids-responsive β-cyclodextrin-functionalized graphene-based fluorescence probe: label-free selective detection of tetrahydrofuran.

    Science.gov (United States)

    Hu, Huawen; Xin, John H; Hu, Hong; Wang, Xiaowen; Lu, Xinkun

    2014-06-06

    In this study, a label-free graphene-based fluorescence probe used for detection of volatile organic liquids was fabricated by a simple, efficient and low-cost method. To fabricate the probe, a bio-based β-cyclodextrin (β-CD) was firstly grafted on reduced graphene surfaces effectively and uniformly, as evidenced by various characterization techniques such as Ultraviolet/Visible spectroscopy, Fourier transform infrared spectroscopy, X-ray diffraction, thermogravimetric analysis, scanning electron microscopy and transmission electron microscopy. The subsequent inclusion of Rhodamine B (RhB) into the inner cavities of the β-CD grafted on the graphene surfaces was achieved easily by a solution mixing method, which yielded the graphene-based fluorescent switch-on probe. In addition, the gradual and controllable quenching of RhB by Fluorescence Resonance Energy Transfer from RhB to graphene during the process of stepwise accommodation of the RhB molecules into the β-CD-functionalized graphene was investigated in depth. A wide range of organic solvents was examined using the as-fabricated fluorescence probe, which revealed the highest sensitivity to tetrahydrofuran with the detection limit of about 1.7 μg/mL. Some insight into the mechanism of the different responsive behaviors of the fluorescence sensor to the examined targets was also described.

  5. Organic Liquids-Responsive β-Cyclodextrin-Functionalized Graphene-Based Fluorescence Probe: Label-Free Selective Detection of Tetrahydrofuran

    Directory of Open Access Journals (Sweden)

    Huawen Hu

    2014-06-01

    Full Text Available In this study, a label-free graphene-based fluorescence probe used for detection of volatile organic liquids was fabricated by a simple, efficient and low-cost method. To fabricate the probe, a bio-based β-cyclodextrin (β-CD was firstly grafted on reduced graphene surfaces effectively and uniformly, as evidenced by various characterization techniques such as Ultraviolet/Visible spectroscopy, Fourier transform infrared spectroscopy, X-ray diffraction, thermogravimetric analysis, scanning electron microscopy and transmission electron microscopy. The subsequent inclusion of Rhodamine B (RhB into the inner cavities of the β-CD grafted on the graphene surfaces was achieved easily by a solution mixing method, which yielded the graphene-based fluorescent switch-on probe. In addition, the gradual and controllable quenching of RhB by Fluorescence Resonance Energy Transfer from RhB to graphene during the process of stepwise accommodation of the RhB molecules into the β-CD-functionalized graphene was investigated in depth. A wide range of organic solvents was examined using the as-fabricated fluorescence probe, which revealed the highest sensitivity to tetrahydrofuran with the detection limit of about 1.7 μg/mL. Some insight into the mechanism of the different responsive behaviors of the fluorescence sensor to the examined targets was also described.

  6. Selective and Sensitive Detection of Cyanide Based on the Displacement Strategy Using a Water-Soluble Fluorescent Probe

    Science.gov (United States)

    La, Ming; Hao, Yuanqiang; Wang, Zhaoyang; Han, Guo-Cheng; Qu, Lingbo

    2016-01-01

    A water-soluble fluorescent probe (C-GGH) was used for the highly sensitive and selective detection of cyanide (CN−) in aqueous media based on the displacement strategy. Due to the presence of the recognition unit GGH (Gly-Gly-His), the probe C-GGH can coordinate with Cu2+ and consequently display ON-OFF type fluorescence response. Furthermore, the in situ formed nonfluorescent C-GGH-Cu2+ complex can act as an effective OFF-ON type fluorescent probe for sensing CN− anion. Due to the strong binding affinity of CN− to Cu2+, CN− can extract Cu2+ from C-GGH-Cu2+ complex, leading to the release of C-GGH and the recovery of fluorescent emission of the system. The probe C-GGH-Cu2+ allowed detection of CN− in aqueous solution with a LOD (limit of detection) of 0.017 μmol/L which is much lower than the maximum contaminant level (1.9 μmol/L) for CN− in drinking water set by the WHO (World Health Organization). The probe also displayed excellent specificity for CN− towards other anions, including F−, Cl−, Br−, I−, SCN−, PO4 3−, N3 −, NO3 −, AcO−, SO4 2−, and CO3 2−. PMID:26881185

  7. In vivo near-infrared fluorescence imaging of amyloid-β plaques with a dicyanoisophorone-based probe

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Jia-ying; Zhou, Lin-fu; Li, Yu-kun [School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, 510006 (China); Chen, Shuo-bin [School of Pharmaceutical Science, Sun Yat-sen University, Guangzhou, 510006 (China); Yan, Jin-wu, E-mail: yjw@scut.edu.cn [School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, 510006 (China); Zhang, Lei, E-mail: lzhangce@scut.edu.cn [School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, 510006 (China)

    2017-04-08

    A dicyanoisophorone-based probe with two-photon absorption and NIR emission was developed for the in vivo fluorescence imaging of amyloid-β plaques, which exhibited high selectivity toward Aβ aggregates over other intracellular proteins. The detection limit was calculated to be as low as 109 nM. In vivo imaging studies indicated that the probe could penetrate the blood–brain barrier and label Aβ plaques in the living transgenic mice, and its specific binding to cerebral Aβ plaques was further confirmed by one- and two-photon ex vivo fluorescence imaging. All these results featured its promising application prospects for amyloid-β sensing in basic research and biomedical research. - Highlights: • A two-photon probe (DCIP-1) with NIR emission based on dicyanoisophorone group, for the in vivo fluorescence imaging of amyloid-β plaques, was reported. • The probe showed turn-on fluorescence (13-fold) with a large Stokes shift upon inserting into the hydrophobic pockets of Aβ aggregates. • The in vivo imaging studies indicated that the probe can penetrate the blood–brain barrier efficiently and discriminate APP/PS1 transgenic mice from WT controls.

  8. Selective and Sensitive Detection of Cyanide Based on the Displacement Strategy Using a Water-Soluble Fluorescent Probe

    Directory of Open Access Journals (Sweden)

    Ming La

    2016-01-01

    Full Text Available A water-soluble fluorescent probe (C-GGH was used for the highly sensitive and selective detection of cyanide (CN− in aqueous media based on the displacement strategy. Due to the presence of the recognition unit GGH (Gly-Gly-His, the probe C-GGH can coordinate with Cu2+ and consequently display ON-OFF type fluorescence response. Furthermore, the in situ formed nonfluorescent C-GGH-Cu2+ complex can act as an effective OFF-ON type fluorescent probe for sensing CN− anion. Due to the strong binding affinity of CN− to Cu2+, CN− can extract Cu2+ from C-GGH-Cu2+ complex, leading to the release of C-GGH and the recovery of fluorescent emission of the system. The probe C-GGH-Cu2+ allowed detection of CN− in aqueous solution with a LOD (limit of detection of 0.017 μmol/L which is much lower than the maximum contaminant level (1.9 μmol/L for CN− in drinking water set by the WHO (World Health Organization. The probe also displayed excellent specificity for CN− towards other anions, including F−, Cl−, Br−, I−, SCN−, PO43-, N3-, NO3-, AcO−, SO42-, and CO32-.

  9. In vivo near-infrared fluorescence imaging of amyloid-β plaques with a dicyanoisophorone-based probe

    International Nuclear Information System (INIS)

    Zhu, Jia-ying; Zhou, Lin-fu; Li, Yu-kun; Chen, Shuo-bin; Yan, Jin-wu; Zhang, Lei

    2017-01-01

    A dicyanoisophorone-based probe with two-photon absorption and NIR emission was developed for the in vivo fluorescence imaging of amyloid-β plaques, which exhibited high selectivity toward Aβ aggregates over other intracellular proteins. The detection limit was calculated to be as low as 109 nM. In vivo imaging studies indicated that the probe could penetrate the blood–brain barrier and label Aβ plaques in the living transgenic mice, and its specific binding to cerebral Aβ plaques was further confirmed by one- and two-photon ex vivo fluorescence imaging. All these results featured its promising application prospects for amyloid-β sensing in basic research and biomedical research. - Highlights: • A two-photon probe (DCIP-1) with NIR emission based on dicyanoisophorone group, for the in vivo fluorescence imaging of amyloid-β plaques, was reported. • The probe showed turn-on fluorescence (13-fold) with a large Stokes shift upon inserting into the hydrophobic pockets of Aβ aggregates. • The in vivo imaging studies indicated that the probe can penetrate the blood–brain barrier efficiently and discriminate APP/PS1 transgenic mice from WT controls.

  10. Radiation-induced polymerization monitored in situ by time-resolved fluorescence of probe molecules in methyl methacrylate

    International Nuclear Information System (INIS)

    Frahn, Mark S.; Abellon, Ruben D.; Luthjens, Leonard H.; Vermeulen, Martien J.W.; Warman, John M.

    2003-01-01

    A technique is presented for monitoring radiation-induced polymerizations in situ based on the measurement of the fluorescence lifetime of molecular probes dissolved in the polymerizing medium. This method is illustrated with results on methyl methacrylate (MMA) using two fluorogenic probe molecules; N-(2-anthracene)methacrylamide (AnMA) and maleimido-fluoroprobe (MFP), a molecule which has a highly dipolar excited state

  11. Site-directed fluorescence labeling of a membrane protein with BADAN: probing protein topology and local environment

    NARCIS (Netherlands)

    Koehorst, R.B.M.; Spruijt, R.B.; Hemminga, M.A.

    2008-01-01

    We present a new and simple method based on site-directed fluorescence labeling using the BADAN label that allows to examine protein-lipid interactions in great detail. We apply this approach to a membrane-embedded mainly -helical reference protein, the M13 major coat protein, of which in a

  12. Spectroscopic quantification of 5-hydroxymethylcytosine in genomic DNA using boric acid-functionalized nano-microsphere fluorescent probes.

    Science.gov (United States)

    Chen, Hua-Yan; Wei, Jing-Ru; Pan, Jiong-Xiu; Zhang, Wei; Dang, Fu-Quan; Zhang, Zhi-Qi; Zhang, Jing

    2017-05-15

    5-hydroxymethylcytosine (5hmC) is the sixth base of DNA. It is involved in active DNA demethylation and can be a marker of diseases such as cancer. In this study, we developed a simple and sensitive 2-(4-boronophenyl)quinoline-4-carboxylic acid modified poly (glycidyl methacrylate (PBAQA-PGMA) fluorescent probe to detect the 5hmC content of genomic DNA based on T4 β-glucosyltransferase-catalyzed glucosylation of 5hmC. The fluorescence-enhanced intensity recorded from the DNA sample was proportional to its 5-hydroxymethylcytosine content and could be quantified by fluorescence spectrophotometry. The developed probe showed good detection sensitivity and selectivity and a good linear relationship between the fluorescence intensity and the concentration of 5 hmC within a 0-100nM range. Compared with other fluorescence detection methods, this method not only could determine trace amounts of 5 hmC from genomic DNA but also could eliminate the interference of fluorescent dyes and the need for purification. It also could avoid multiple labeling. Because the PBAQA-PGMA probe could enrich the content of glycosyl-5-hydroxymethyl-2-deoxycytidine from a complex ground substance, it will broaden the linear detection range and improve sensitivity. The limit of detection was calculated to be 0.167nM after enrichment. Furthermore, the method was successfully used to detect 5-hydroxymethylcytosine from mouse tissues. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. A quinoline-based Cu2 + ion complex fluorescence probe for selective detection of inorganic phosphate anion in aqueous solution and its application to living cells

    Science.gov (United States)

    Dai, Yanpeng; Wang, Peng; Fu, Jiaxin; Yao, Kun; Xu, Kuoxi; Pang, Xiaobin

    2017-08-01

    A quinaldine functionalized probe QP has been designed and synthesized. It exhibited selective turn-off fluorescence response toward Cu2 + ion over most of the biologically important ions at physiological pH. The binding ratio of the probe QP and Cu2 + ion was determined to be 1:1 through fluorescence titration, Job's plot and ESI-MS. The binding constant (K) of Cu2 + to probe QP was found to be 2.12 × 104 M- 1. Further, the Cu2 + ensemble of probe QP was found to respond H2PO4- and HPO42 - among other important biological anions via fluorescence turn-on response at physiological pH. Fluorescence microscopy imaging using living Hela cells showed that probe QP could be used as an effective fluorescent probe for detecting Cu2 + cation and H2PO4- and HPO42 - anions in living cells.

  14. A NBD-based simple but effective fluorescent pH probe for imaging of lysosomes in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Cao, Xiang-Jian [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Chen, Li-Na [Institute of Developmental Biology, School of Life Science, Shandong University, Jinan 250100 (China); Zhang, Xuan [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Liu, Jin-Ting [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Chen, Ming-Yu [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Wu, Qiu-Rong [Institute of Developmental Biology, School of Life Science, Shandong University, Jinan 250100 (China); Taishan College, Shandong University, Jinan 250100 (China); Miao, Jun-Ying, E-mail: miaojy@sdu.edu.cn [Institute of Developmental Biology, School of Life Science, Shandong University, Jinan 250100 (China); Zhao, Bao-Xiang, E-mail: bxzhao@sdu.edu.cn [Institute of Organic Chemistry, School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China)

    2016-05-12

    NBDlyso with lysosome-locating morpholine moiety has been developed as a high selective and sensitive fluorescent pH probe. This probe can respond to acidic pH (2.0–7.0) in a short time (less than 1 min) and not almost change after continuously illuminated for an extended period by ultraviolet light. The fluorescence intensity of NBDlyso enhanced 100-fold in acidic solution, with very good linear relationship (R{sup 2} = 0.996). The pK{sub a} of probe NBDlyso is 4.10. Therefore, NBDlyso was used to detect lysosomal pH changes successfully. Besides, X-ray crystallography was used to verify the structure of NBDlyso, and the recognition mechanism involving photo-induced electron transfer was interpreted theoretically by means of DFT and TDDFT calculations skillfully when NBDlyso comes into play under the acidic condition. This probe showed good ability to sense pH change in living cell image. - Highlights: • An effective NBD-based fluorescent pH probe was developed. • The sensing mechanism was interpreted by theoretical calculation. • This probe was successfully used to monitor lysosoml pH changes in Hela cells.

  15. NONINVASIVE OPTICAL IMAGING OF STAPHYLOCOCCUS AUREUS INFECTION IN VIVO USING AN ANTIMICROBIAL PEPTIDE FRAGMENT BASED NEAR-INFRARED FLUORESCENT PROBES

    Directory of Open Access Journals (Sweden)

    CUICUI LIU

    2013-07-01

    Full Text Available The diagnosis of bacterial infections remains a major challenge in medicine. Optical imaging of bacterial infection in living animals is usually conducted with genetic reporters such as light-emitting enzymes or fluorescent proteins. However, there are many circumstances where genetic reporters are not applicable, and there is an urgent need for exogenous synthetic probes that can selectively target bacteria. Optical imaging of bacteria in vivo is much less developed than methods such as radioimaging and MRI. Furthermore near-infrared (NIR dyes with emission wavelengths in the region of 650–900 nm can propagate through two or more centimeters of tissue and may enable deeper tissue imaging if sensitive detection techniques are employed. Here we constructed an antimicrobial peptide fragment UBI29-41-based near-infrared fluorescent imaging probe. The probe is composed of UBI29-41 conjugated to a near infrared dye ICG-Der-02. UBI29-41 is a cationic antimicrobial peptide that targets the anionic surfaces of bacterial cells. The probe allows detection of Staphylococcus aureus infection (5 × 107 cells in a mouse local infection model using whole animal near-infrared fluorescence imaging. Furthermore, we demonstrate that the UBI29-41-based imaging probe can selectively accumulate within bacteria. The significantly higher accumulation in bacterial infection suggests that UBI29-41-based imaging probe may be a promising imaging agent to detect bacterial infections.

  16. A NBD-based simple but effective fluorescent pH probe for imaging of lysosomes in living cells

    International Nuclear Information System (INIS)

    Cao, Xiang-Jian; Chen, Li-Na; Zhang, Xuan; Liu, Jin-Ting; Chen, Ming-Yu; Wu, Qiu-Rong; Miao, Jun-Ying; Zhao, Bao-Xiang

    2016-01-01

    NBDlyso with lysosome-locating morpholine moiety has been developed as a high selective and sensitive fluorescent pH probe. This probe can respond to acidic pH (2.0–7.0) in a short time (less than 1 min) and not almost change after continuously illuminated for an extended period by ultraviolet light. The fluorescence intensity of NBDlyso enhanced 100-fold in acidic solution, with very good linear relationship (R"2 = 0.996). The pK_a of probe NBDlyso is 4.10. Therefore, NBDlyso was used to detect lysosomal pH changes successfully. Besides, X-ray crystallography was used to verify the structure of NBDlyso, and the recognition mechanism involving photo-induced electron transfer was interpreted theoretically by means of DFT and TDDFT calculations skillfully when NBDlyso comes into play under the acidic condition. This probe showed good ability to sense pH change in living cell image. - Highlights: • An effective NBD-based fluorescent pH probe was developed. • The sensing mechanism was interpreted by theoretical calculation. • This probe was successfully used to monitor lysosoml pH changes in Hela cells.

  17. A fluorescent pH probe for acidic organelles in living cells.

    Science.gov (United States)

    Chen, Jyun-Wei; Chen, Chih-Ming; Chang, Cheng-Chung

    2017-09-26

    A water-soluble pH sensor, 2-(6-(4-aminostyryl)-1,3-dioxo-1H-benzo[de]isoquinolin-2(3H)-yl)-N, N-dimethylethanamine (ADA), was synthesized based on the molecular design of photoinduced electron transfer (PET) and intramolecular charge transfer (ICT). The fluorescence emission response against a pH value is in the range 3-6, which is suitable for labelling intracellular pH-dependent microenvironments. After biological evolution, ADA is more than a pH biosensor because it is also an endocytosis pathway tracking biosensor that labels endosomes, late endosomes, and lysosome pH gradients. From this, the emissive aggregates of ADA and protonated-ADA in these organs were evaluated to explore how this probe stresses emission colour change to cause these unique cellular images.

  18. Probing the photoluminescence properties of gold nanoclusters by fluorescence lifetime correlation spectroscopy

    International Nuclear Information System (INIS)

    Yuan, C. T.; Lin, T. N.; Shen, J. L.; Lin, C. A.; Chang, W. H.; Cheng, H. W.; Tang, J.

    2013-01-01

    Gold nanoclusters (Au NCs) have attracted much attention for promising applications in biological imaging owing to their tiny sizes and biocompatibility. So far, most efforts have been focused on the strategies for fabricating high-quality Au NCs and then characterized by conventional ensemble measurement. Here, a fusion single-molecule technique combining fluorescence correlation spectroscopy and time-correlated single-photon counting can be successfully applied to probe the photoluminescence (PL) properties for sparse Au NCs. In this case, the triplet-state dynamics and diffusion process can be observed simultaneously and the relevant time constants can be derived. This work provides a complementary insight into the PL mechanism at the molecular levels for Au NCs in solution

  19. Observing Fluorescent Probes in Living Cells using a Low-Cost LED Flashlight Retrofitted to a Common Vintage Light Microscope

    Directory of Open Access Journals (Sweden)

    G. A. Babbitt

    2013-03-01

    Full Text Available While the application of molecular biological techniques based upon fluorescent probes has rapidly expanded over recent decades, the equipment cost of fluorescent microscopy has largely prevented its adoption in the college and high school classroom. We offer a simple solution to this problem by describing in detail how to build with simple tools, a fluorescent microscope using a common brand of colored LED flashlights and second-hand components of vintage Nikon microscopes. This extremely low cost solution is qualitatively compared to an expensive modern Zeiss system.

  20. A Fluorescent Molecular Probe for the Detection of Hydrogen Based on Oxidative Addition Reactions with Crabtree-Type Hydrogenation Catalysts.

    Science.gov (United States)

    Kos, Pavlo; Plenio, Herbert

    2015-11-02

    A Crabtree-type Ir(I) complex tagged with a fluorescent dye (bodipy) was synthesized. The oxidative addition of H2 converts the weakly fluorescent Ir(I) complex (Φ=0.038) into a highly fluorescent Ir(III) species (Φ=0.51). This fluorogenic reaction can be utilized for the detection of H2 and to probe the oxidative addition step in the catalytic hydrogenation of olefins. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. The Effect of a Fluorophore Photo-Physics on the Lipid Vesicle Diffusion Coefficient Studied by Fluorescence Correlation Spectroscopy.

    Science.gov (United States)

    Drabik, Dominik; Przybyło, Magda; Sikorski, Aleksander; Langner, Marek

    2016-03-01

    Fluorescence Correlation Spectroscopy (FCS) is a technique, which allows determination of the diffusion coefficient and concentration of fluorescent objects suspended in the solution. The measured parameter is the fluctuation of the fluorescence signal emitted by diffusing molecules. When 100 nm DOPC vesicles labeled with various fluorescent dyes (Fluorescein-PE, NBD-PE, Atto488 DOPE or βBodipy FL) were measured, different values of diffusion coefficients have been obtained. These diffusion coefficients were different from the expected values measured using the dynamic light scattering method (DLS). The FCS was initially developed for solutions containing small fluorescent molecules therefore the observed inconsistency may result from the nature of vesicle suspension itself. The duration of the fluorescence signal may depend on the following factors: the exposure time of the labeled object to the excitation beam, the photo-physical properties (e.g., stability) of a fluorophore, the theoretical model used for the calculations of the diffusion coefficient and optical properties of the vesicle suspension. The diffusion coefficients determined for differently labeled liposomes show that its dependence on vesicle size and quantity of fluorescent probed used for labeling was significant demonstrating that the fluorescence properties of the fluorophore itself (bleaching and/or blinking) were critical factors for a correct outcome of FCS experiment. The new, based on combined FCS and DLS measurements, method for the determination of the focal volume prove itself to be useful for the evaluation of a fluorescence dye with respect to its applicability for FCS experiment.

  2. Synthesis and spectral properties of novel chlorinated pH fluorescent probes

    International Nuclear Information System (INIS)

    Wu Xianglong; Jin Xilang; Wang Yunxia; Mei Qibing; Li Jianli; Shi Zhen

    2011-01-01

    Eight chlorinated fluoresceins have been synthesized by the reaction of chlorinated resorcinols with 4, 5, 6, 7-tetrachlorophthalic anhydride or 3, 6-dichloro-4-carboxyphthalic anhydride in the presence of methanesulfonic acid. The spectral properties of the chlorinated fluoresceins were studied. It was found that they have absorption and emission maxima at long wavelengths and high fluorescence quantum yields. Emission spectra of chlorinated fluoresceins shifted towards long wavelength with increase in chlorine. pH-dependent properties of chlorinated fluoresceins were studied in detail. These compounds showed a strongly pH-sensitive range of 3.0-7.0. These chlorinated fluoresceins will be used as pH probes for pH measurement of the cell because of the high quantum yield and strong pH-sensitivity. - Research highlights: → Eight chlorinated fluoresceins have been synthesized in the presence of methanesulfonic acid. → Emission spectra of these compounds shifted towards long wavelength with increase in chlorine. → Eight chlorinated fluoresceins showed a strongly pH-sensitive range of 3.0-7.0. → They have emission maxima at long wavelengths and high fluorescence quantum yields.

  3. Synthesis of novel fluorescent probe Tb(III)-7-carboxymethoxy-4-methylcoumarin complex for sensing of DNA

    International Nuclear Information System (INIS)

    Hussein, Belal H.M.; Azab, Hassan A.; Fathalla, Walid; Ali, Sherin A.M.

    2013-01-01

    New fluorescent probe Tb(III) (7-carboxymethoxy-4-methylcoumarin)2(SCN) (C2H5OH)(H2O) was synthesized and characterized by spectroscopy and thermal analysis. The absorption and fluorescence spectra of 7-carboxymethoxy-4-methylcoumarin (CMMC) and Tb(III)–CMMC complex have been measured in different solvents. The interactions of Tb(III)–CMMC complex with calf thymus nucleic acid (CT-DNA) have been investigated using steady state fluorescence measurements. The changes in the fluorescence intensity have been used for the quantitative determination of DNA with LOD of 3.45 ng in methanol–water (9:1, v/v). The association constants of DNA with Tb(III)–CMMC complex was found to be 2.62×1010 M −1 . - Highlights: ► New fluorescent probe Terbium (III)-7-carboxy methoxy-4-methylcoumarin complex has been synthesized and characterized. ► FTIR spectrum of Tb(III)-complex shows a characteristic band for thiocyanate group. ► DNA interaction with Terbium (III)-7-carboxy methoxy-4-methylcoumarin has been studied by fluorescence techniques. ► The change in the fluorescence intensity has been used for the quantitative determination of DNA. ► The result was better than most of the well-known methods including the ethidium bromide method.

  4. Synthesis of novel fluorescent probe Tb(III)-7-carboxymethoxy-4-methylcoumarin complex for sensing of DNA

    Energy Technology Data Exchange (ETDEWEB)

    Hussein, Belal H.M., E-mail: belalhussein102@yahoo.com [Department of Chemistry, Faculty of Science, Suez Canal University, Ismailia (Egypt); Azab, Hassan A. [Department of Chemistry, Faculty of Science, Suez Canal University, Ismailia (Egypt); Fathalla, Walid [Department of Mathematical and Physical Sciences, Faculty of Engineering, Port-Said University, Port-Said (Egypt); Ali, Sherin A.M. [Department of Mathematical and Physical Sciences, Faculty of Engineering, Suez Canal University, Ismailia (Egypt)

    2013-02-15

    New fluorescent probe Tb(III) (7-carboxymethoxy-4-methylcoumarin)2(SCN) (C2H5OH)(H2O) was synthesized and characterized by spectroscopy and thermal analysis. The absorption and fluorescence spectra of 7-carboxymethoxy-4-methylcoumarin (CMMC) and Tb(III)-CMMC complex have been measured in different solvents. The interactions of Tb(III)-CMMC complex with calf thymus nucleic acid (CT-DNA) have been investigated using steady state fluorescence measurements. The changes in the fluorescence intensity have been used for the quantitative determination of DNA with LOD of 3.45 ng in methanol-water (9:1, v/v). The association constants of DNA with Tb(III)-CMMC complex was found to be 2.62 Multiplication-Sign 1010 M{sup -1}. - Highlights: Black-Right-Pointing-Pointer New fluorescent probe Terbium (III)-7-carboxy methoxy-4-methylcoumarin complex has been synthesized and characterized. Black-Right-Pointing-Pointer FTIR spectrum of Tb(III)-complex shows a characteristic band for thiocyanate group. Black-Right-Pointing-Pointer DNA interaction with Terbium (III)-7-carboxy methoxy-4-methylcoumarin has been studied by fluorescence techniques. Black-Right-Pointing-Pointer The change in the fluorescence intensity has been used for the quantitative determination of DNA. Black-Right-Pointing-Pointer The result was better than most of the well-known methods including the ethidium bromide method.

  5. Noninvasive imaging of multiple myeloma using near infrared fluorescent molecular probe

    Science.gov (United States)

    Hathi, Deep; Zhou, Haiying; Bollerman-Nowlis, Alex; Shokeen, Monica; Akers, Walter J.

    2016-03-01

    Multiple myeloma is a plasma cell malignancy characterized by monoclonal gammopathy and osteolytic bone lesions. Multiple myeloma is most commonly diagnosed in late disease stages, presenting with pathologic fracture. Early diagnosis and monitoring of disease status may improve quality of life and long-term survival for multiple myeloma patients from what is now a devastating and fatal disease. We have developed a near-infrared targeted fluorescent molecular probe with high affinity to the α4β1 integrin receptor (VLA-4)overexpressed by a majority of multiple myeloma cells as a non-radioactive analog to PET/CT tracer currently being developed for human diagnostics. A near-infrared dye that emits about 700 nm was conjugated to a high affinity peptidomimmetic. Binding affinity and specificity for multiple myeloma cells was investigated in vitro by tissue staining and flow cytometry. After demonstration of sensitivity and specificity, preclinical optical imaging studies were performed to evaluate tumor specificity in murine subcutaneous and metastatic multiple myeloma models. The VLA-4-targeted molecular probe showed high affinity for subcutaneous MM tumor xenografts. Importantly, tumor cells specific accumulation in the bone marrow of metastatic multiple myeloma correlated with GFP signal from transfected cells. Ex vivo flow cytometry of tumor tissue and bone marrow further corroborated in vivo imaging data, demonstrating the specificity of the novel agent and potential for quantitative imaging of multiple myeloma burden in these models.

  6. A single pH fluorescent probe for biosensing and imaging of extreme acidity and extreme alkalinity.

    Science.gov (United States)

    Chao, Jian-Bin; Wang, Hui-Juan; Zhang, Yong-Bin; Li, Zhi-Qing; Liu, Yu-Hong; Huo, Fang-Jun; Yin, Cai-Xia; Shi, Ya-Wei; Wang, Juan-Juan

    2017-07-04

    A simple tailor-made pH fluorescent probe 2-benzothiazole (N-ethylcarbazole-3-yl) hydrazone (Probe) is facilely synthesized by the condensation reaction of 2-hydrazinobenzothiazole with N-ethylcarbazole-3-formaldehyde, which is a useful fluorescent probe for monitoring extremely acidic and alkaline pH, quantitatively. The pH titrations indicate that Probe displays a remarkable emission enhancement with a pK a of 2.73 and responds linearly to minor pH fluctuations within the extremely acidic range of 2.21-3.30. Interestingly, Probe also exhibits strong pH-dependent characteristics with pK a 11.28 and linear response to extreme-alkalinity range of 10.41-12.43. In addition, Probe shows a large Stokes shift of 84 nm under extremely acidic and alkaline conditions, high selectivity, excellent sensitivity, good water-solubility and fine stability, all of which are favorable for intracellular pH imaging. The probe is further successfully applied to image extremely acidic and alkaline pH values fluctuations in E. coli cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Two rhodamine lactam modulated lysosome-targetable fluorescence probes for sensitively and selectively monitoring subcellular organelle pH change

    Energy Technology Data Exchange (ETDEWEB)

    Li, Hongmei [Ministry of Education Key Laboratory of Synthetic and Natural Functional Molecule Chemistry, College of Chemistry & Materials Science, Northwest University, Xi' an 710069 (China); Wang, Cuiling [Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, College of Life Science, Northwest University, Xi' an 710069 (China); She, Mengyao; Zhu, Yuelu; Zhang, Jidong; Yang, Zheng [Ministry of Education Key Laboratory of Synthetic and Natural Functional Molecule Chemistry, College of Chemistry & Materials Science, Northwest University, Xi' an 710069 (China); Liu, Ping, E-mail: liuping@nwu.edu.cn [Ministry of Education Key Laboratory of Synthetic and Natural Functional Molecule Chemistry, College of Chemistry & Materials Science, Northwest University, Xi' an 710069 (China); Wang, Yaoyu [Ministry of Education Key Laboratory of Synthetic and Natural Functional Molecule Chemistry, College of Chemistry & Materials Science, Northwest University, Xi' an 710069 (China); Li, Jianli, E-mail: lijianli@nwu.edu.cn [Ministry of Education Key Laboratory of Synthetic and Natural Functional Molecule Chemistry, College of Chemistry & Materials Science, Northwest University, Xi' an 710069 (China)

    2015-11-05

    Be a powerful technique for convenient detection of pH change in living cells, especially at subcellular level, fluorescent probes has attracted more and more attention. In this work, we designed and synthesized three rhodamine lactam modulated fluorescent probes RS1, RS2 and RS3, which all respond sensitively toward weak acidity (pH range 4–6) via the photophysical property in buffer solution without interference from the other metal ions, and they also show ideal pKa values and excellent reversibility. Particularly, by changing the lone pair electrons distribution of lactam-N atom with different conjugations, RS2 and RS3 exhibit high quantum yield, negligible cytotoxicity and excellent permeability. They are suitable to stain selectively lysosomes of tumor cells and monitor its pH changes sensitively via optical molecular imaging. The above findings suggest that the probes we designed could act as ideal and easy method for investigating the pivotal role of H{sup +} in lysosomes and are potential pH detectors in disease diagnosis through direct intracellular imaging. - Highlights: • Two probes for sensitively and selectively monitoring weak acidic pH change. • The pKa of the probes was highly suitable for staining lysosomes in tumor cells. • The properties of those probes were changed by different conjugate system. • These probes have negligible cytotoxicity and good sensitivity in vivo.

  8. Two rhodamine lactam modulated lysosome-targetable fluorescence probes for sensitively and selectively monitoring subcellular organelle pH change

    International Nuclear Information System (INIS)

    Li, Hongmei; Wang, Cuiling; She, Mengyao; Zhu, Yuelu; Zhang, Jidong; Yang, Zheng; Liu, Ping; Wang, Yaoyu; Li, Jianli

    2015-01-01

    Be a powerful technique for convenient detection of pH change in living cells, especially at subcellular level, fluorescent probes has attracted more and more attention. In this work, we designed and synthesized three rhodamine lactam modulated fluorescent probes RS1, RS2 and RS3, which all respond sensitively toward weak acidity (pH range 4–6) via the photophysical property in buffer solution without interference from the other metal ions, and they also show ideal pKa values and excellent reversibility. Particularly, by changing the lone pair electrons distribution of lactam-N atom with different conjugations, RS2 and RS3 exhibit high quantum yield, negligible cytotoxicity and excellent permeability. They are suitable to stain selectively lysosomes of tumor cells and monitor its pH changes sensitively via optical molecular imaging. The above findings suggest that the probes we designed could act as ideal and easy method for investigating the pivotal role of H + in lysosomes and are potential pH detectors in disease diagnosis through direct intracellular imaging. - Highlights: • Two probes for sensitively and selectively monitoring weak acidic pH change. • The pKa of the probes was highly suitable for staining lysosomes in tumor cells. • The properties of those probes were changed by different conjugate system. • These probes have negligible cytotoxicity and good sensitivity in vivo.

  9. Lateral mobility of plasma membrane lipids in dividing Xenopus eggs

    OpenAIRE

    Laat, S.W. de; Tetteroo, P.A.T.; Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van

    1984-01-01

    The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xaopus Levis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein (“HEDAF”) and 5-(N-tetradecanoyl)aminofluorescein (“TEDAF”) as probes. The preexisting plasma membrane of the animal side showed an inhomogeneous, dotted fluorescence pattern after labeling and the lateral mobility of both probes used was below the detection limits of the FP...

  10. A ratiometric fluorescent probe based on boron dipyrromethene and rhodamine Förster resonance energy transfer platform for hypochlorous acid and its application in living cells

    International Nuclear Information System (INIS)

    Liu, Ying; Zhao, Zhi-Min; Miao, Jun-Ying; Zhao, Bao-Xiang

    2016-01-01

    We have developed a ratiometric fluorescent probe BRT based on boron dipyrromethene (BODIPY) and rhodamine-thiohydrazide Förster resonance energy transfer (FRET) platform for sensing hypochlorous acid (HOCl) with high selectivity and sensitivity. The probe can detect HOCl in 15 s with the detection limit of 38 nM. Upon mixing with HOCl the fluorescence colour of probe BRT changed from green to orange. Moreover, probe BRT was applied to successfully monitor HOCl in living RAW 264.7 cells. - Highlights: • A probe based on BODIPY and rhodamine was developed for sensing HOCl. • The probe could sense HOCl in a ratiometric manner based on the FRET platform in PBS buffer solution. • The probe can detect HOCl in 15 s accompanied with a fluorescence colour change. • This probe was successfully used to monitor HOCl in living RAW 264.7 cells.

  11. FLUORESCENCE PROBING OF THE FORMATION OF HYDROPHOBIC MICRODOMAINS BY CROSS-LINKED POLY(ALKYLMETHYLDIALLYLAMMONIUM BROMIDES) IN AQUEOUS-SOLUTION

    NARCIS (Netherlands)

    WANG, GJ; ENGBERTS, J B F N

    Pyrene has been used as a fluorescence probe to investigate the conformational behavior of cross-linked poly(alkylmethyldiallylammonium bromides) in aqueous solutions. Binding of pyrene to hydrophobic microdomains, formed by the polysoaps, is reflected by a change in the ratio I-1/I-3 of the

  12. Fluorescent water-Soluble Probes Based on Ammonium Cation Peg Substituted Perylenepisimides: Synthesis, Photophysical Properties, and Live Cell Images

    Science.gov (United States)

    Yang, Wei; Cai, Jiaxuan; Zhang, Shuchen; Yi, Xuegang; Gao, Baoxiang

    2018-01-01

    To synthesize perylenbisimides (PBI) fluorescent probes that will improve the water-soluble ability and the cytocompatibility, the synthesis and properties of fluorescent water-soluble probes based on dendritic ammonium cation polyethylene glycol (PEG) substituted perylenebisimides(GPDIs) are presented. As we expected, with increased ammonium cation PEG, the aggregation of the PBI in an aqueous solution is completely suppressed by the hydrophilic ammonium cation PEG groups. And the fluorescence quantum yield increases from 25% for GPDI-1 to 62% for GPDI-2. When incubated with Hela cells for 48 h, the viabilities are 71% (for GPDI-1) and 76% (for GPDI-2). Live cell imaging shows that these probes are efficiently internalized by HeLa cells. The study of the photophysical properties indicated increasing the ammonium cation PEG generation can increase the fluorescence quantum yield. Live cell imaging shows that with the ammonium cation PEG chains of perylenebisimides has high biocompatibility. The exceptionally low cytotoxicity is ascribed to the ammonium cation PEG chains, which protect the dyes from nonspecifically interacting with the extracellular proteins. Live cell imaging shows that ammonium cations PEG chains can promote the internalization of these probes.

  13. Label-free fluorescence strategy for sensitive detection of adenosine triphosphate using a loop DNA probe with low background noise.

    Science.gov (United States)

    Lin, Chunshui; Cai, Zhixiong; Wang, Yiru; Zhu, Zhi; Yang, Chaoyong James; Chen, Xi

    2014-07-15

    A simple, rapid, label-free, and ultrasensitive fluorescence strategy for adenosine triphosphate (ATP) detection was developed using a loop DNA probe with low background noise. In this strategy, a loop DNA probe, which is the substrate for both ligation and digestion enzyme reaction, was designed. SYBR green I (SG I), a double-stranded specific dye, was applied for the readout fluorescence signal. Exonuclease I (Exo I) and exonuclease III (Exo III), sequence-independent nucleases, were selected to digest the loop DNA probe in order to minimize the background fluorescence signal. As a result, in the absence of ATP, the loop DNA was completely digested by Exo I and Exo III, leading to low background fluorescence owing to the weak electrostatic interaction between SG I and mononucleotides. On the other hand, ATP induced the ligation of the nicking site, and the sealed loop DNA resisted the digestion of Exo I and ExoIII, resulting in a remarkable increase of fluorescence response. Upon background noise reduction, the sensitivity of the ATP determination was improved significantly, and the detection limitation was found to be 1.2 pM, which is much lower than that in almost all the previously reported methods. This strategy has promise for wide application in the determination of ATP.

  14. Smart near-infrared fluorescence probes with donor-acceptor structure for in vivo detection of β-amyloid deposits.

    Science.gov (United States)

    Cui, Mengchao; Ono, Masahiro; Watanabe, Hiroyuki; Kimura, Hiroyuki; Liu, Boli; Saji, Hideo

    2014-03-05

    The deposition of β-amyloid (Aβ) plaques in the parenchymal and cortical brain is accepted as the main pathological hallmark of Alzheimer's disease (AD); however, early detection of AD still presents a challenge. With the assistance of molecular imaging techniques, imaging agents specifically targeting Aβ plaques in the brain may lead to the early diagnosis of AD. Herein, we report the design, synthesis, and evaluation of a series of smart near-infrared fluorescence (NIRF) imaging probes with donor-acceptor architecture bridged by a conjugated π-electron chain for Aβ plaques. The chemical structure of these NIRF probes is completely different from Congo Red and Thioflavin-T. Probes with a longer conjugated π system (carbon-carbon double bond) displayed maximum emission in PBS (>650 nm), which falls in the best range for NIRF probes. These probes were proved to have affinity to Aβ plaques in fluorescent staining of brain sections from an AD patient and double transgenic mice, as well as in an in vitro binding assay using Aβ(1-42) aggregates. One probe with high affinity (K(i) = 37 nM, K(d) = 27 nM) was selected for in vivo imaging. It can penetrate the blood-brain barrier of nude mice efficiently and is quickly washed out of the normal brain. Moreover, after intravenous injection of this probe, 22-month-old APPswe/PSEN1 mice exhibited a higher relative signal than control mice over the same period of time, and ex vivo fluorescent observations confirmed the existence of Aβ plaques. In summary, this probe meets most of the requirements for a NIRF contrast agent for the detection of Aβ plaques both in vitro and in vivo.

  15. Molecular Imaging of β-Amyloid Plaques with Near-Infrared Boron Dipyrromethane (BODIPY-Based Fluorescent Probes

    Directory of Open Access Journals (Sweden)

    Hiroyuki Watanabe

    2013-07-01

    Full Text Available The formation of β-amyloid (Aβ plaques is a critical neurodegenerative change in Alzheimer disease (AD. We designed and synthesized novel boron dipyrromethane (BODIPY-based Aβ probes (BAPs and evaluated their utility for near-infrared fluorescence imaging of Aβ plaques in the brain. In binding experiments in vitro, BAPs showed high affinity for synthetic Aβ aggregates (Kd = 18–149 nM. Furthermore, BAPs clearly stained Aβ plaques in sections of Tg2576 mice. In mouse brain tissue, BAPs showed sufficient uptake for optical imaging. In addition, ex vivo fluorescent staining of brain sections from Tg2576 mice after the injection of BAP-2 showed selective binding of Aβ plaques with little nonspecific binding. BAPs may be useful as a near-infrared fluorescent probe for imaging Aβ plaques.

  16. Imaging of Fluoride Ion in Living Cells and Tissues with a Two-Photon Ratiometric Fluorescence Probe

    Directory of Open Access Journals (Sweden)

    Xinyue Zhu

    2015-01-01

    Full Text Available A reaction-based two-photon (TP ratiometric fluorescence probe Z2 has been developed and successfully applied to detect and image fluoride ion in living cells and tissues. The Z2 probe was designed designed to utilize an ICT mechanism between n-butylnaphthalimide as a fluorophore and tert-butyldiphenylsilane (TBDPS as a response group. Upon addition of fluoride ion, the Si-O bond in the Z2 would be cleaved, and then a stronger electron-donating group was released. The fluorescent changes at 450 and 540 nm, respectively, made it possible to achieve ratiometric fluorescence detection. The results indicated that the Z2 could ratiometrically detect and image fluoride ion in living cells and tissues in a depth of 250 μm by two-photon microscopy (TPM.

  17. Evolution of group 14 rhodamines as platforms for near-infrared fluorescence probes utilizing photoinduced electron transfer.

    Science.gov (United States)

    Koide, Yuichiro; Urano, Yasuteru; Hanaoka, Kenjiro; Terai, Takuya; Nagano, Tetsuo

    2011-06-17

    The absorption and emission wavelengths of group 14 pyronines and rhodamines, which contain silicon, germanium, or tin at the 10 position of the xanthene chromophore, showed large bathochromic shifts compared to the original rhodamines, owing to stabilization of the LUMO energy levels by σ*-π* conjugation between group 14 atom-C (methyl) σ* orbitals and a π* orbital of the fluorophore. These group 14 pyronines and rhodamines retain the advantages of the original rhodamines, including high quantum efficiency in aqueous media (Φ(fl) = 0.3-0.45), tolerance to photobleaching, and high water solubility. Group 14 rhodamines have higher values of reduction potential than other NIR light-emitting original rhodamines, and therefore, we speculated their NIR fluorescence could be controlled through the photoinduced electron transfer (PeT) mechanism. Indeed, we found that the fluorescence quantum yield (Φ(fl)) of Si-rhodamine (SiR) and Ge-rhodamine (GeR) could be made nearly equal to zero, and the threshold level for fluorescence on/off switching lies at around 1.3-1.5 V for the SiRs. This is about 0.1 V lower than in the case of TokyoGreens, in which the fluorophore is well established to be effective for PeT-based probes. That is to say, the fluorescence of SiR and GeR can be drastically activated by more than 100-fold through a PeT strategy. To confirm the validity of this strategy for developing NIR fluorescence probes, we employed this approach to design two kinds of novel fluorescence probes emitting in the far-red to NIR region, i.e., a series of pH-sensors for use in acidic environments and a Zn(2+) sensor. We synthesized these probes and confirmed that they work well.

  18. A novel fluorescent probe (dtpa-bis(cytosine)) for detection of Eu(III) in rare earth metal ions

    Science.gov (United States)

    Yang, Fan; Ren, Peipei; Liu, Guanhong; Song, Youtao; Bu, Naishun; Wang, Jun

    2018-03-01

    In this paper, a novel fluorescent probe, dtpa-bis(cytosine), was designed and synthesized for detecting europium (Eu3 +) ion. Upon addition of Eu3 + ions into the dtpa-bis(cytosine) solution, the fluorescence intensity can strongly be enhanced. Conversely, adding other rare earth metal ions, such as Y3 +, Ce3 +, Pr3 +, Nd3 +, Sm3 +, Gd3 +, Tb3 +, Dy3 +, Ho3 +, Er3 +, Yb3 + and Lu3 +, into dtpa-bis(cytosine) solution, the fluorescence intensity is decreased slightly. Some parameters affecting the fluorescence intensity of dtpa-bis(cytosine) solution in the presence of Eu3 + ions were investigated, including solution pH value, Eu3 + ion concentration and interfering substances. The detection mechanism of Eu3 + ion using dtpa-bis(cytosine) as fluorescent probe was proposed. Under optimum conditions, the fluorescence emission intensities of EuIII-dtpa-bis(cytosine) at 375 nm in the concentration range of 0.50 × 10- 5 mol • L- 1-5.00 × 10- 5 mol • L- 1 of Eu3 + ion display a better linear relationship. The limit of detection (LOD) was determined as 8.65 × 10- 7 mol • L- 1 and the corresponding correlation coefficient (R2) of the linear equation is 0.9807. It is wished that the proposed method could be applied for sensitively and selectively detecting Eu3 + ion.

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

    Science.gov (United States)

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

    2013-08-01

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

  20. Single Molecule 3D Orientation in Time and Space: A 6D Dynamic Study on Fluorescently Labeled Lipid Membranes

    DEFF Research Database (Denmark)

    Börner, Richard; Ehrlich, Nicky; Hohlbein, Johannes

    2016-01-01

    Interactions between single molecules profoundly depend on their mutual three-dimensional orientation. Recently, we demonstrated a technique that allows for orientation determination of single dipole emitters using a polarization-resolved distribution of fluorescence into several detection channels...... interesting in non-isotropic environments such as lipid membranes, which are of great importance in biology. We used giant unilamellar vesicles (GUVs) labeled with fluorescent dyes down to a single molecule concentration as a model system for both, assessing the robustness of the orientation determination...

  1. Fluorescence and computational studies of thymidine phosphorylase affinity toward lipidated 5-FU derivatives

    Science.gov (United States)

    Lettieri, R.; D'Abramo, M.; Stella, L.; La Bella, A.; Leonelli, F.; Giansanti, L.; Venanzi, M.; Gatto, E.

    2018-04-01

    Thymidine phosphorylase (TP) is an enzyme that is up-regulated in a wide variety of solid tumors, including breast and colorectal cancers. It is involved in tumor growth and metastasis, for this reason it is one of the key enzyme to be inhibited, in an attempt to prevent tumor proliferation. However, it also plays an active role in cancer treatment, through its contribution in the conversion of the anti-cancer drug 5-fluorouracil (5-FU) to an irreversible inhibitor of thymidylate synthase (TS), responsible of the inhibition of the DNA synthesis. In this work, the intrinsic TP fluorescence has been investigated for the first time and exploited to study TP binding affinity for the unsubstituted 5-FU and for two 5-FU derivatives, designed to expose this molecule on liposomal membranes. These molecules were obtained by functionalizing the nitrogen atom with a chain consisting of six (1) or seven (2) units of glycol, linked to an alkyl moiety of 12 carbon atoms. Derivatives (1) and (2) exhibited an affinity for TP in the micromolar range, 10 times higher than the parent compound, irrespective of the length of the polyoxyethylenic spacer. This high affinity was maintained also when the compounds were anchored in liposomal membranes. Experimental results were supported by molecular dynamics simulations and docking calculations, supporting a feasible application of the designed supramolecular lipid structure in selective targeting of TP, to be potentially used as a drug delivery system or sensor device.

  2. Real-Time Discrimination and Versatile Profiling of Spontaneous Reactive Oxygen Species in Living Organisms with a Single Fluorescent Probe.

    Science.gov (United States)

    Zhang, Ruilong; Zhao, Jun; Han, Guangmei; Liu, Zhengjie; Liu, Cui; Zhang, Cheng; Liu, Bianhua; Jiang, Changlong; Liu, Renyong; Zhao, Tingting; Han, Ming-Yong; Zhang, Zhongping

    2016-03-23

    Fluorescent probes are powerful tools for the investigations of reactive oxygen species (ROS) in living organisms by visualization and imaging. However, the multiparallel assays of several ROS with multiple probes are often limited by the available number of spectrally nonoverlapping chromophores together with large invasive effects and discrepant biological locations. Meanwhile, the spontaneous ROS profilings in various living organs/tissues are also limited by the penetration capability of probes across different biological barriers and the stability in reactive in vivo environments. Here, we report a single fluorescent probe to achieve the effective discrimination and profiling of hydroxyl radicals (•OH) and hypochlorous acid (HClO) in living organisms. The probe is constructed by chemically grafting an additional five-membered heterocyclic ring and a lateral triethylene glycol chain to a fluorescein mother, which does not only turn off the fluorescence of fluorescein, but also create the dual reactive sites to ROS and the penetration capability in passing through various biological barriers. The reactions of probe with •OH and HClO simultaneously result in cyan and green emissions, respectively, providing the real-time discrimination and quantitative analysis of the two ROS in cellular mitochondria. Surprisingly, the accumulation of probes in the intestine and liver of a normal-state zebrafish and the transfer pathway from intestine-to-blood-to-organ/tissue-to-kidney-to-excretion clearly present the profiling of spontaneous •OH and HClO in these metabolic organs. In particular, the stress generation of •OH at the fresh wound of zebrafish is successfully visualized for the first time, in spite of its extremely short lifetime.

  3. In vivo imaging of membrane type-1 matrix metalloproteinase with a novel activatable near-infrared fluorescence probe.

    Science.gov (United States)

    Shimizu, Yoichi; Temma, Takashi; Hara, Isao; Makino, Akira; Kondo, Naoya; Ozeki, Ei-Ichi; Ono, Masahiro; Saji, Hideo

    2014-08-01

    Membrane type-1 matrix metalloproteinase (MT1-MMP) is a protease activating MMP-2 that mediates cleavage of extracellular matrix components and plays pivotal roles in tumor migration, invasion and metastasis. Because in vivo noninvasive imaging of MT1-MMP would be useful for tumor diagnosis, we developed a novel near-infrared (NIR) fluorescence probe that can be activated following interaction with MT1-MMP in vivo. MT1-hIC7L is an activatable fluorescence probe comprised of anti-MT1-MMP monoclonal antibodies conjugated to self-assembling polymer micelles that encapsulate NIR dyes (IC7-1, λem : 858 nm) at concentrations sufficient to cause fluorescence self-quenching. In aqueous buffer, MT1-hIC7L fluorescence was suppressed to background levels and increased approximately 35.5-fold in the presence of detergent. Cellular uptake experiments revealed that in MT1-MMP positive C6 glioma cells, MT1-hIC7L showed significantly higher fluorescence that increased with time as compared to hIC7L, a negative control probe lacking the anti-MT1-MMP monoclonal antibody. In MT1-MMP negative MCF-7 breast adenocarcinoma cells, both MT1-hIC7L and hIC7L showed no obvious fluorescence. In addition, the fluorescence intensity of C6 cells treated with MT1-hIC7L was suppressed by pre-treatment with an MT1-MMP endocytosis inhibitor (P imaging using probes intravenously administered to tumor-bearing mice showed that MT1-hIC7L specifically visualized C6 tumors (tumor-to-background ratios: 3.8 ± 0.3 [MT1-hIC7L] vs 3.1 ± 0.2 [hIC7L] 48 h after administration, P fluorescence in MCF-7 tumors. Together, these results show that MT1-hIC7L would be a potential activatable NIR probe for specifically detecting MT1-MMP-expressing tumors. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

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

    Science.gov (United States)

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

    2015-10-07

    Magnetic fluorescent nanoparticles (NPs) have great potential applications for diagnostics, imaging and therapy. We developed a facile polyol method to synthesize multifunctional Fe3O4@CeF3:Tb@CeF3 NPs with small size (CA) to obtain carboxyl-functionalized NPs (Fe3O4@CeF3:Tb@CeF3-COOH). Folic acid (FA) as an affinity ligand was then covalently conjugated onto NPs to yield Fe3O4@CeF3:Tb@CeF3-FA NPs. They were then applied as multimodal imaging agents for simultaneous in vitro targeted fluorescence imaging and magnetic resonance imaging (MRI) of HeLa cells with overexpressed folate receptors (FR). The results indicated that these NPs had strong luminescence and enhanced T2-weighted MR contrast and would be promising candidates as multimodal probes for both fluorescence and MRI imaging.

  5. Identification of Cannabis sativa L. using the 1-kbTHCA synthase-fluorescence in situ hybridization probe.

    Science.gov (United States)

    Jeangkhwoa, Pattraporn; Bandhaya, Achirapa; Umpunjun, Puangpaka; Chuenboonngarm, Ngarmnij; Panvisavas, Nathinee

    2017-03-01

    This study reports a successful application of fluorescence in situ hybridization (FISH) technique in the identification of Cannabis sativa L. cells recovered from fresh and dried powdered plant materials. Two biotin-16-dUTP-labeled FISH probes were designed from the Cannabis-specific tetrahydrocannabinolic acid synthase (THCAS) gene and the ITS region of the 45S rRNA gene. Specificity of probe-target hybridization was tested against the target and 4 non-target plant species, i.e., Humulus lupulus, Mitragyna speciosa, Papaver sp., and Nicotiana tabacum. The 1-kb THCA synthase hybridization probe gave Cannabis-specific hybridization signals, unlike the 700-bp Cannabis-ITS hybridization probe. Probe-target hybridization was also confirmed against 20 individual Cannabis plant samples. The 1-kb THCA synthase and 700-bp Cannabis-ITS hybridization probes clearly showed 2 hybridization signals per cell with reproducibility. The 1-kb THCA synthase probe did not give any FISH signal when tested against H. lupulus, its closely related member of the Canabaceae family. It was also showed that 1-kb THCA synthase FISH probe can be applied to identify small amount of dried powdered Cannabis material with an addition of rehydration step prior to the experimental process. This study provided an alternative identification method for Cannabis trace. Copyright © 2016. Published by Elsevier B.V.

  6. Long Range Polymer Chain Dynamics of Highly Flexible Polysiloxane in Solution Probed by Pyrene Excimer Fluorescence

    Directory of Open Access Journals (Sweden)

    Janine L. Thoma

    2018-03-01

    Full Text Available A poly(dimethylsiloxane-co-(3-aminopropylmethylsiloxane polymer (PDMS with 20.3 mol % of (3-aminopropylmethyl siloxane monomer has been labeled randomly with 1-pyreneacetyl groups to generate a series of polysiloxanes (Py-PDMS with pyrenyl contents ranging from 0.7 mol % to 5.2 mol % of the total number of structural units. The remainder of the amino groups were acetylated to avoid intra-chain quenching of the excited singlet states of pyrene via exciplex formation with free amino groups while allowing the formation of excimers to proceed. The fluorescence spectra and temporal decays of the Py-PDMS samples were acquired in tetrahydrofuran (THF, N,N-dimethylformamide (DMF, and dioxane. blob, the average rate constant for intra-chain pyrene excimer formation, was determined from the analysis of the fluorescence decays. blob was found to equal 1.16 (±0.13 × 109, 1.14 (±0.12 × 109, and 0.99 (±0.10 × 109 s−1 in THF, DMF, and dioxane, respectively, at room temperature. They are the largest values found to date for any polymeric backbone in these solvents. The qualitative relationship found here between blob and the chemical structures of the polymers indicates that the luminescence characteristics of randomly labeled polymers is a very useful method to probe the long range dynamics of chains of almost any polymer that is amenable to substitution by a lumophore.

  7. A novel fluorescent probe based on rhodamine hydrazone derivatives bearing a thiophene group for Al³⁺.

    Science.gov (United States)

    Li, Meng-xiao; Zhang, Xia; Fan, Yu-hua; Bi, Cai-feng

    2016-05-01

    In the present work, a novel 5-methyl-thiophene-carbaldehyde-functionalized rhodamine 6G Schiff base (RA) was designed and easily prepared as an Al(3+) fluorescent and colorimetric probe, which could selectively and sensitively detect Al(3+) by showing enhanced fluorescence emission. Meanwhile distinct color variation from colorless to pink also provided 'naked eye' detection of Al(3+), due to the ring spirolactam opening of the rhodamine derivative. Other metal ions (including K(+), Mg(2+), Na(+), Ba(2+), Mn(2+), Cd(2+), Fe(2+), Ni(2+), Pb(2+), Zn(2+), Hg(2+), Co(2+), Li(+), Sr(2+) and Cu(2+)) could only induce limited interference. The detection limit of the fluorescent probe was estimated to be 4.17 × 10(-6) M, the binding constant of the RA-Al(3+) complex was 1.4 × 10(6)  M(-1). Moreover, this fluorescent probe RA possessed high reversibility. As aluminum is a ubiquitous metal in nature and plays vital roles in many biological processes, this chemosensor could be explored for biological study applications. Copyright © 2015 John Wiley & Sons, Ltd.

  8. A spirobifluorene-based two-photon fluorescence probe for mercury ions and its applications in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Xiao, Haibo, E-mail: xiaohb@shnu.edu.cn; Zhang, Yanzhen; Zhang, Wu; Li, Shaozhi; Tan, Jingjing; Han, Zhongying

    2017-05-01

    A novel spirobifluorene derivative SPF-TMS, which containing dithioacetal groups and triphenylamine units, was synthesized. The probing behaviors toward various metal ions were investigated via UV/Vis absorption spectra as well as one-photon fluorescence changes. The results indicated that SPF-TMS exhibits high sensitivity and selectivity for mercury ions. The detection limit was at least 8.6 × 10{sup −8}M, which is excellent comparing with other optical sensors for Hg{sup 2+}. When measured by two-photon excited fluorescence technique in THF at 800 nm, the two-photon cross-section of SPF-TMS is 272 GM. Especially, upon reaction with mercury species, SPF-TMS yielded another two-photon dye SPF-DA. Both SPF-TMS and SPF-DA emit strong two-photon induced fluorescence and can be applied in cell imaging by two-photon microscopy. - Highlights: • We report a spirobifluorene-based molecule as two-photon fluorescent probe with large two-photon cross-section. • The molecule has exclusive selectivity and sensitivity for mercury species. • The molecule has large two-photon emission changes before and after addition of Hg{sup 2+}. • Both the probe and the mercury ion-promoted reaction product can be applied in cell imaging by two-photon microscopy.

  9. Turn-on fluorescence probes based on pyranine/viologen charge-transfer complexes for the determination of nucleotides

    Energy Technology Data Exchange (ETDEWEB)

    Schäferling, Michael, E-mail: Michael.schaeferling@utu.fi; Lang, Thomas; Schnettelker, Annette

    2014-10-15

    The formation of ground state charge-transfer complexes between pyranine (8-hydroxypyrene-1,3,6-trisulfonic acid) and viologen (paraquat) derivatives is utilized for the design of novel fluoroionophores for the determination of phosphate species, particularly of nucleotides. The strong quenching of the pyranine fluorescence by viologen-type charge transfer acceptors can be countermanded if these are functionalized with triethylammonium groups that serve as recognition elements for phosphate anions. We report on the fluorogenic responses of these water-soluble molecular probes in presence of different phosphates. Absorbance measurements give additional information on the charge transfer complex formation and the interaction with nucleotides. The experimental data show that these aggregates form attractive, simple and versatile fluorescence turn-on probes for nucleoside triphosphates. The reversibility of the fluorescence response is demonstrated by means of an enzymatic model assay using ATPase for the decomposition of adenosine triphosphate. - Highlights: • Pyranine/viologen charge-transfer complexes as molecular probe for ATP recognition. • Fluorescence turn on mechanism. • Selective compared to other nucleotides and phosphate anions. • Fast and reversible response applicable to monitor enzymatic reactions.

  10. Dual Mechanism of an Intramolecular Charge Transfer (ICT)-FRET-Based Fluorescent Probe for the Selective Detection of Hydrogen Peroxide.

    Science.gov (United States)

    Liang, Xiao; Xu, Xiaoyi; Qiao, Dan; Yin, Zheng; Shang, Luqing

    2017-12-14

    A dual-mechanism intramolecular charge transfer (ICT)-FRET fluorescent probe for the selective detection of H 2 O 2 in living cells has been designed and synthesized. This probe used a coumarin-naphthalimide hybrid as the FRET platform and a boronate moiety as the recognition group. Upon the addition of H 2 O 2 , the probe exhibited a redshifted (73 nm) fluorescence emission, and the ratio of fluorescence intensities at λ=558 and 485 nm (F 558 /F 485 ) shifted notably (up to 100-fold). Moreover, there was a good linearity (R 2 =0.9911) between the ratio and concentration of H 2 O 2 in the range of 0 to 60 μm, with a limit of detection of 0.28 μm (signal to noise ratio (S/N)=3). This probe could also detect enzymatically generated H 2 O 2 . Importantly, it could be used to visualize endogenous H 2 O 2 produced by stimulation from epidermal growth factor. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Raman probing of lipids, proteins, and mitochondria in skeletal myocytes: a case study on obesity

    DEFF Research Database (Denmark)

    Brazhe, Nadezda A.; Nikelshparg, Evelina I.; Prats, Clara

    2017-01-01

    We propose a novel approach to assess simultaneously lipid composition in lipid droplets, the redox state of cytochromes, and the relative amount of [Fe–S] clusters in the electron transport chain in the mitochondria of skeletal myocytes by means of near-infrared Raman spectroscopy. Mitochondria...... technique allows to estimate qualitatively the relative amount of cholesterol and unsaturated lipids, ordering of lipid phase in lipid droplets, changes in the redox state of c-type and b-type cytochromes, and the relative amount of [Fe–S] clusters in the mitochondria of intact myocytes. The proposed...

  12. Fluorescence-Guided Probes of Aptamer-Targeted Gold Nanoparticles with Computed Tomography Imaging Accesses for in Vivo Tumor Resection.

    Science.gov (United States)

    Li, Cheng-Hung; Kuo, Tsung-Rong; Su, Hsin-Jan; Lai, Wei-Yun; Yang, Pan-Chyr; Chen, Jinn-Shiun; Wang, Di-Yan; Wu, Yi-Chun; Chen, Chia-Chun

    2015-10-28

    Recent development of molecular imaging probes for fluorescence-guided surgery has shown great progresses for determining tumor margin to execute the tissue resection. Here we synthesize the fluorescent gold nanoparticles conjugated with diatrizoic acid and nucleolin-targeted AS1411 aptamer. The nanoparticle conjugates exhibit high water-solubility, good biocompatibility, visible fluorescence and strong X-ray attenuation for computed tomography (CT) contrast enhancement. The fluorescent nanoparticle conjugates are applied as a molecular contrast agent to reveal the tumor location in CL1-5 tumor-bearing mice by CT imaging. Furthermore, the orange-red fluorescence emitting from the conjugates in the CL1-5 tumor can be easily visualized by the naked eyes. After the resection, the IVIS measurements show that the fluorescence signal of the nanoparticle conjugates in the tumor is greatly enhanced in comparison to that in the controlled experiment. Our work has shown potential application of functionalized nanoparticles as a dual-function imaging agent in clinical fluorescence-guided surgery.

  13. Highly sensitive ratiometric detection of heparin and its oversulfated chondroitin sulfate contaminant by fluorescent peptidyl probe.

    Science.gov (United States)

    Mehta, Pramod Kumar; Lee, Hyeri; Lee, Keun-Hyeung

    2017-05-15

    The selective and sensitive detection of heparin, an anticoagulant in clinics as well as its contaminant oversulfated chondroitin sulfate (OSCS) is of great importance. We first reported a ratiometric sensing method for heparin as well as OSCS contaminants in heparin using a fluorescent peptidyl probe (Pep1, pyrene-GSRKR) and heparin-digestive enzyme. Pep1 exhibited a highly sensitive ratiometric response to nanomolar concentration of heparin in aqueous solution over a wide pH range (2~11) and showed highly selective ratiometric response to heparin among biological competitors such as hyaluronic acid and chondroitin sulfate. Pep1 showed a linear ratiometric response to nanomolar concentrations of heparin in aqueous solutions and in human serum samples. The detection limit for heparin was calculated to be 2.46nM (R 2 =0.99) in aqueous solutions, 2.98nM (R 2 =0.98) in 1% serum samples, and 3.43nM (R 2 =0.99) in 5% serum samples. Pep1 was applied to detect the contaminated OSCS in heparin with heparinase I, II, and III, respectively. The ratiometric sensing method using Pep1 and heparinase II was highly sensitive, fast, and efficient for the detection of OSCS contaminant in heparin. Pep1 with heparinase II could detect as low as 0.0001% (w/w) of OSCS in heparin by a ratiometric response. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Intermittent Fluorescence Oscillations in Lipid Droplets in a Live Normal and Lung Cancer Cell: Time-Resolved Confocal Microscopy.

    Science.gov (United States)

    Chowdhury, Rajdeep; Amin, Md Asif; Bhattacharyya, Kankan

    2015-08-27

    Intermittent structural oscillation in the lipid droplets of live lung cells is monitored using time-resolved confocal microscopy. Significant differences are observed between the lung cancer cell (A549) and normal (nonmalignant) lung cell (WI38). For this study, the lipid droplets are covalently labeled with a fluorescent dye, coumarin maleimide (7-diethylamino-3-(4-maleimido-phenyl)-4-methylcoumarin, CPM). The number of lipid droplets in the cancer cell is found to be ∼20-fold higher than that in the normal (nonmalignant) cell. The fluctuation in the fluorescence intensity of the dye (CPM) is attributed to the red-ox processes and periodic formation/rupture of the S-CPM bond. The amount of reactive oxygen species (ROS) is much higher in a cancer cell. This is manifested in faster oscillations (0.9 ± 0.3 s) in cancer cells compared to that in the normal cells (2.8 ± 0.7 s). Solvation dynamics in the lipid droplets of cancer cells is slower compared to that in the normal cell.

  15. Synthesis of strongly fluorescent carbon quantum dots modified with polyamidoamine and a triethoxysilane as quenchable fluorescent probes for mercury(II)

    International Nuclear Information System (INIS)

    Tang, Wenjie; Wang, Yan; Wang, Panpan; Di, Junwei; Wu, Ying; Yang, Jianping

    2016-01-01

    This article reports on the synthesis of water dispersible carbon quantum dots (CDs) by a one-step hydrothermal method using polyamidoamine (PAMAM) and (3-aminopropyl)triethoxysilane (APTES) as a platform and passivant. The resulting CDs are highly uniform and finely dispersed. The synergistic effect between PAMAM and APTES on the surface of the CDs results in a fluorescence that is much brighter than that of CDs modified with either APTES or PAMAM only. The fluorescence of the co-modified CDs is quenched by Hg(II) ions at fairly low concentrations. Under the optimum conditions, the intensity of quenched fluorescence drops with Hg(II) concentration in the range from 0.2 nM to 10 μM, and the detection limit is 87 fM. The effect of potentially interfering cations on the fluorescence revealed a high selectivity for Hg 2+ . The fluorescent probe was applied to the determination of Hg(II) in (spiked) waters and milk and gave recoveries between 95.6 and 107 %, with relative standard deviation between 4.4 and 6.0 %. (author)

  16. A BODIPY-Based Fluorescent Probe to Visually Detect Phosgene: Toward the Development of a Handheld Phosgene Detector.

    Science.gov (United States)

    Sayar, Melike; Karakuş, Erman; Güner, Tuğrul; Yildiz, Busra; Yildiz, Umit Hakan; Emrullahoğlu, Mustafa

    2018-03-02

    A boron-dipyrromethene (BODIPY)-based fluorescent probe with a phosgene-specific reactive motif shows remarkable selectivity toward phosgene, in the presence of which the nonfluorescent dye rapidly transforms into a new structure and induces a fluorescent response clearly observable to the naked eye under ultraviolet light. Given that dynamic, a prototypical handheld phosgene detector with a promising sensing capability that expedites the detection of gaseous phosgene without sophisticated instrumentation was developed. The proposed method using the handheld detector involves a rapid response period suitable for issuing early warnings during emergency situations. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. A Novel Water-soluble Ratiometric Fluorescent Probe Based on FRET for Sensing Lysosomal pH.

    Science.gov (United States)

    Song, Guang-Jie; Bai, Su-Yun; Luo, Jing; Cao, Xiao-Qun; Zhao, Bao-Xiang

    2016-11-01

    A new ratiometric fluorescent probe based on Förster resonance energy transfer (FRET) for sensing lysosomal pH has been developed. The probe (RMPM) was composed of imidazo[1,5-α]pyridine quaternary ammonium salt fluorophore as the FRET donor and the rhodamine moiety as the FRET acceptor. It's the first time to report that imidazo[1,5-α]pyridine quaternary ammonium salt acts as the FRET donor. The ratio of fluorescence intensity of the probe at two wavelengths (I 424 /I 581 ) changed significantly and responded linearly toward minor pH changes in the range of 5.4-6.6. It should be noted that it's rare to report that a ratiometric pH probe could detect so weak acidic pH with pKa = 6.31. In addition, probe RMPM exhibited excellent water-solubility, fast-response, all-right selectivity and brilliant reversibility. Moreover, RMPM has been successfully applied to sensing lysosomal pH in HeLa cells and has low cytotoxicity.

  18. Fluorescent Protein Voltage Probes Derived from ArcLight that Respond to Membrane Voltage Changes with Fast Kinetics

    Science.gov (United States)

    Han, Zhou; Jin, Lei; Platisa, Jelena; Cohen, Lawrence B.; Baker, Bradley J.; Pieribone, Vincent A.

    2013-01-01

    We previously reported the discovery of a fluorescent protein voltage probe, ArcLight, and its derivatives that exhibit large changes in fluorescence intensity in response to changes of plasma membrane voltage. ArcLight allows the reliable detection of single action potentials and sub-threshold activities in individual neurons and dendrites. The response kinetics of ArcLight (τ1-on ~10 ms, τ2-on ~ 50 ms) are comparable with most published genetically-encoded voltage probes. However, probes using voltage-sensing domains other than that from the Ciona intestinalis voltage sensitive phosphatase exhibit faster kinetics. Here we report new versions of ArcLight, in which the Ciona voltage-sensing domain was replaced with those from chicken, zebrafish, frog, mouse or human. We found that the chicken and zebrafish-based ArcLight exhibit faster kinetics, with a time constant (τ) less than 6ms for a 100 mV depolarization. Although the response amplitude of these two probes (8-9%) is not as large as the Ciona-based ArcLight (~35%), they are better at reporting action potentials from cultured neurons at higher frequency. In contrast, probes based on frog, mouse and human voltage sensing domains were either slower than the Ciona-based ArcLight or had very small signals. PMID:24312287

  19. Fluorescent protein voltage probes derived from ArcLight that respond to membrane voltage changes with fast kinetics.

    Directory of Open Access Journals (Sweden)

    Zhou Han

    Full Text Available We previously reported the discovery of a fluorescent protein voltage probe, ArcLight, and its derivatives that exhibit large changes in fluorescence intensity in response to changes of plasma membrane voltage. ArcLight allows the reliable detection of single action potentials and sub-threshold activities in individual neurons and dendrites. The response kinetics of ArcLight (τ1-on ~10 ms, τ2-on ~ 50 ms are comparable with most published genetically-encoded voltage probes. However, probes using voltage-sensing domains other than that from the Ciona intestinalis voltage sensitive phosphatase exhibit faster kinetics. Here we report new versions of ArcLight, in which the Ciona voltage-sensing domain was replaced with those from chicken, zebrafish, frog, mouse or human. We found that the chicken and zebrafish-based ArcLight exhibit faster kinetics, with a time constant (τ less than 6 ms for a 100 mV depolarization. Although the response amplitude of these two probes (8-9% is not as large as the Ciona-based ArcLight (~35%, they are better at reporting action potentials from cultured neurons at higher frequency. In contrast, probes based on frog, mouse and human voltage sensing domains were either slower than the Ciona-based ArcLight or had very small signals.

  20. Establishment of a novel immunoassay system for rapid detection of 2,4-dichlorophenoxyacetic acid residues based on magnetic-fluorescent probes

    Directory of Open Access Journals (Sweden)

    WANG Yuanfeng

    2014-12-01

    Full Text Available A novel immunoassay system based on magnetic-fluorescent probes was established to detect 2.4-dichlorophenoxyacetic acid (2,4-D residue in liquid system in food and agricultural products.The composites of anti-2,4-D antibody bound to Fe3O4@SiO2-NH2 was employed as the solid phase as well as magnetic probe.The composites composed of 2,4-D-OVA labeled with CdTe@SiO2-NH2 as the fluorescent probe was used to produce fluorescent signal.2,4-D and its fluorescent probe competed binding the antibody on the surface of the magnetic probe.The optimization of 2,4-D-OVA dosage,coupling PH and reaction time in preparing the fluorescent probe were investigated.It showed that in the synthesis of fluorescent probe 8.2 was the optimal pH,70 min was the optimal coupling time,500 μL amount of 2,4-D-OVA.The standard curve was obtained with the concentration of 2,4-D and the maximum fluorescence intensity.The detection limit of the assay was gotten and it was 3.55×10-8.One reaction step and one washing step were needed.The assay significantly shortened the testing time and amplified the detection signal compared with classic ELISA.

  1. Specificity and kinetics of alpha-synuclein binding to model membranes determined with fluorescent excited state intramolecular proton transfer (ESIPT) probe.

    Science.gov (United States)

    Shvadchak, Volodymyr V; Falomir-Lockhart, Lisandro J; Yushchenko, Dmytro A; Jovin, Thomas M

    2011-04-15

    Parkinson disease is characterized cytopathologically by the deposition in the midbrain of aggregates composed primarily of the presynaptic neuronal protein α-synuclein (AS). Neurotoxicity is currently attributed to oligomeric microaggregates subjected to oxidative modification and promoting mitochondrial and proteasomal dysfunction. Unphysiological binding to membranes of these and other organelles is presumably involved. In this study, we performed a systematic determination of the influence of charge, phase, curvature, defects, and lipid unsaturation on AS binding to model membranes using a new sensitive solvatochromic fluorescent probe. The interaction of AS with vesicular membranes is fast and reversible. The protein dissociates from neutral membranes upon thermal transition to the liquid disordered phase and transfers to vesicles with higher affinity. The binding of AS to neutral and negatively charged membranes occurs by apparently different mechanisms. Interaction with neutral bilayers requires the presence of membrane defects; binding increases with membrane curvature and rigidity and decreases in the presence of cholesterol. The association with negatively charged membranes is much stronger and much less sensitive to membrane curvature, phase, and cholesterol content. The presence of unsaturated lipids increases binding in all cases. These findings provide insight into the relation between membrane physical properties and AS binding affinity and dynamics that presumably define protein localization in vivo and, thereby, the role of AS in the physiopathology of Parkinson disease.

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

    Science.gov (United States)

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

    2001-01-01

    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. PMID:11239011

  3. Development and cytotoxicity of Schiff base derivative as a fluorescence probe for the detection of L-Arginine

    Science.gov (United States)

    Shang, Xuefang; Li, Jie; Guo, Kerong; Ti, Tongyu; Wang, Tianyun; Zhang, Jinlian

    2017-04-01

    Inspired from biological counter parts, chemical modification of Schiff base derivatives with function groups may provide a highly efficient method to detect amino acids. Therefore, a fluorescent probe involving Schiff base and hydroxyl group has been designed and prepared, which showed high response and specificity for Arginine (Arg) among normal eighteen standard kinds of amino acids (Alanine, Valine, Leucine, Isoleucine, Methionine, Asparticacid, Glutamicacid, Arginine, Glycine, Serine, Threonine, Asparagine, Phenylalanine, Histidine, Tryptophan, Proline, Lysine, Glutamine, Tyrosine and Cysteine). Furthermore, theoretical investigation further illustrated the possible binding mode in the host-guest interaction and the roles of molecular frontier orbitals in molecular interplay. In addition, the synthesized fluorescent probe exhibited high binding ability for Arg and low cytotoxicity to MCF-7 cells over a concentration range of 0-200 μg mL-1 which can be also used as a biosensor for the Arg detection in vivo.

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

    Science.gov (United States)

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

    2011-05-01

    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.

  5. A novel turn-on fluorescent strategy for sensing ascorbic acid using graphene quantum dots as fluorescent probe.

    Science.gov (United States)

    Liu, Hua; Na, Weidan; Liu, Ziping; Chen, Xueqian; Su, Xingguang

    2017-06-15

    In this paper, a facile and rapid fluorescence turn-on assay for fluorescent detection of ascorbic acid (AA) was developed by using the orange emission graphene quantum dots (GQDs). In the presence of horse radish peroxidase (HRP) and hydrogen peroxide (H 2 O 2 ), catechol can be oxidized by hydroxyl radicals and converted to o-benzoquinone, which can significantly quench the fluorescence of GQDs. However, when AA present in the system, it can consume part of H 2 O 2 and hydroxyl radicals to inhibit the generation of o-benzoquinone, resulting in fluorescence recovery. Under the optimized experimental conditions, the fluorescence intensity was linearly correlated with the concentration of H 2 O 2 in the range of 3.33-500µM with a detection limit of 1.2µM. The linear detection for AA was in the range from 1.11 to 300µM with a detection limit of 0.32µM. The proposed method was applied to the determination of AA in human serum samples with satisfactory results. Copyright © 2017. Published by Elsevier B.V.

  6. Synthesis and characterization of time-resolved fluorescence probes for evaluation of competitive binding to melanocortin receptors.

    Science.gov (United States)

    Alleti, Ramesh; Vagner, Josef; Dehigaspitiya, Dilani Chathurika; Moberg, Valerie E; Elshan, N G R D; Tafreshi, Narges K; Brabez, Nabila; Weber, Craig S; Lynch, Ronald M; Hruby, Victor J; Gillies, Robert J; Morse, David L; Mash, Eugene A

    2013-09-01

    Probes for use in time-resolved fluorescence competitive binding assays at melanocortin receptors based on the parental ligands MSH(4), MSH(7), and NDP-α-MSH were prepared by solid phase synthesis methods, purified, and characterized. The saturation binding of these probes was studied using HEK-293 cells engineered to overexpress the human melanocortin 4 receptor (hMC4R) as well as the human cholecystokinin 2 receptor (hCCK2R). The ratios of non-specific binding to total binding approached unity at high concentrations for each probe. At low probe concentrations, receptor-mediated binding and uptake was discernable, and so probe concentrations were kept as low as possible in determining Kd values. The Eu-DTPA-PEGO-MSH(4) probe exhibited low specific binding relative to non-specific binding, even at low nanomolar concentrations, and was deemed unsuitable for use in competition binding assays. The Eu-DTPA-PEGO probes based on MSH(7) and NDP-α-MSH exhibited Kd values of 27±3.9nM and 4.2±0.48nM, respectively, for binding with hMC4R. These probes were employed in competitive binding assays to characterize the interactions of hMC4R with monovalent and divalent MSH(4), MSH(7), and NDP-α-MSH constructs derived from squalene. Results from assays with both probes reflected only statistical enhancements, suggesting improper ligand spacing on the squalene scaffold for the divalent constructs. The Ki values from competitive binding assays that employed the MSH(7)-based probe were generally lower than the Ki values obtained when the probe based on NDP-α-MSH was employed, which is consistent with the greater potency of the latter probe. The probe based on MSH(7) was also competed with monovalent, divalent, and trivalent MSH(4) constructs that previously demonstrated multivalent binding in competitive binding assays against a variant of the probe based on NDP-α-MSH. Results from these assays confirm multivalent binding, but suggest a more modest increase in avidity for these

  7. Analysis of chemical equilibrium of silicon-substituted fluorescein and its application to develop a scaffold for red fluorescent probes.

    Science.gov (United States)

    Hirabayashi, Kazuhisa; Hanaoka, Kenjiro; Takayanagi, Toshio; Toki, Yuko; Egawa, Takahiro; Kamiya, Mako; Komatsu, Toru; Ueno, Tasuku; Terai, Takuya; Yoshida, Kengo; Uchiyama, Masanobu; Nagano, Tetsuo; Urano, Yasuteru

    2015-09-01

    Fluorescein is a representative green fluorophore that has been widely used as a scaffold of practically useful green fluorescent probes. Here, we report synthesis and characterization of a silicon-substituted fluorescein, i.e., 2-COOH TokyoMagenta (2-COOH TM), which is a fluorescein analogue in which the O atom at the 10' position of the xanthene moiety of fluorescein is replaced with a Si atom. This fluorescein analogue forms a spirolactone ring via intramolecular nucleophilic attack of the carboxylic group in a pH-dependent manner. Consequently, 2-COOH TM exhibits characteristic large pH-dependent absorption and fluorescence spectral changes: (1) 2-COOH TM is colorless at acidic pH, whereas fluorescein retains observable absorption and fluorescence even at acidic pH, and the absorption maximum is also shifted; (2) the absorption spectral change occurs above pH 7.0 for 2-COOH TM and below pH 7.0 for fluorescein; (3) 2-COOH TM shows a much sharper pH response than fluorescein because of its pKa inversion, i.e., pKa1 > pKa2. These features are also different from those of a compound without the carboxylic group, 2-Me TokyoMagenta (2-Me TM). Analysis of the chemical equilibrium between pH 3.0 and 11.0 disclosed that 2-COOH TM favors the colorless and nonfluorescent lactone form, compared with fluorescein. Substitution of Cl atoms at the 4' and 5' positions of the xanthene moiety of 2-COOH TM to obtain 2-COOH DCTM shifted the equilibrium so that the new derivative exists predominantly in the strongly fluorescent open form at physiological pH (pH 7.4). To demonstrate the practical utility of 2-COOH DCTM as a novel scaffold for red fluorescent probes, we employed it to develop a probe for β-galactosidase.

  8. Integrated femtosecond stimulated Raman scattering and two-photon fluorescence imaging of subcellular lipid and vesicular structures

    Science.gov (United States)

    Li, Xuesong; Lam, Wen Jiun; Cao, Zhe; Hao, Yan; Sun, Qiqi; He, Sicong; Mak, Ho Yi; Qu, Jianan Y.

    2015-11-01

    The primary goal of this study is to demonstrate that stimulated Raman scattering (SRS) as a new imaging modality can be integrated into a femtosecond (fs) nonlinear optical (NLO) microscope system. The fs sources of high pulse peak power are routinely used in multimodal nonlinear microscopy to enable efficient excitation of multiple NLO signals. However, with fs excitations, the SRS imaging of subcellular lipid and vesicular structures encounters significant interference from proteins due to poor spectral resolution and a lack of chemical specificity, respectively. We developed a unique NLO microscope of fs excitation that enables rapid acquisition of SRS and multiple two-photon excited fluorescence (TPEF) signals. In the in vivo imaging of transgenic C. elegans animals, we discovered that by cross-filtering false positive lipid signals based on the TPEF signals from tryptophan-bearing endogenous proteins and lysosome-related organelles, the imaging system produced highly accurate assignment of SRS signals to lipid. Furthermore, we demonstrated that the multimodal NLO microscope system could sequentially image lipid structure/content and organelles, such as mitochondria, lysosomes, and the endoplasmic reticulum, which are intricately linked to lipid metabolism.

  9. Methods of staining and visualization of sphingolipid enriched and non-enriched plasma membrane regions of Arabidopsis thaliana with fluorescent dyes and lipid analogues

    Directory of Open Access Journals (Sweden)

    Blachutzik Jörg O

    2012-08-01

    Full Text Available Abstract Background Sterols and Sphingolipids form lipid clusters in the plasma membranes of cell types throughout the animal and plant kingdoms. These lipid domains provide a medium for protein signaling complexes at the plasma membrane and are also observed to be principal regions of membrane contact at the inception of infection. We visualized different specific fluorescent lipophilic stains of the both sphingolipid enriched and non-sphingolipid enriched regions in the plasma membranes of live protoplasts of Arabidopsis thaliana. Results Lipid staining protocols for several fluorescent lipid analogues in plants are presented. The most emphasis was placed on successful protocols for the single and dual staining of sphingolipid enriched regions and exclusion of sphingolipid enriched regions on the plasma membrane of Arabidopsis thaliana protoplasts. A secondary focus was placed to ensure that these staining protocols presented still maintain cell viability. Furthermore, the protocols were successfully tested with the spectrally sensitive dye Laurdan. Conclusion Almost all existing staining procedures of the plasma membrane with fluorescent lipid analogues are specified for animal cells and tissues. In order to develop lipid staining protocols for plants, procedures were established with critical steps for the plasma membrane staining of Arabidopsis leaf tissue and protoplasts. The success of the plasma membrane staining protocols was additionally verified by measurements of lipid dynamics by the fluorescence recovery after photobleaching technique and by the observation of new phenomena such as time dependent lipid polarization events in living protoplasts, for which a putative physiological relevance is suggested.

  10. A highly selective turn-on fluorescent probe for Al3+ in aqueous solution based on quinoline Schiff-base

    Science.gov (United States)

    Huang, Peng-Cheng; Fang, Hao; Xiong, Jing-Jing; Wu, Fang-Ying

    2017-06-01

    A new Al3+-specific fluorescent probe NQ was designed and synthesized from 2-hydroxy-1-naphthaldehyde and 2-aminoquinoline. Upon the addition of Al3+, the fluorescent intensity of NQ was significantly enhanced compared with other examined metal ions in aqueous solution. The result of a Job’s plot indicated the formation of a 1:1 complex between the probe and Al3+, and the possible binding mode of the system between NQ and Al3+ was clarified by IR analysis and 1H NMR titration. Moreover, other metal ions examined had little effect on the detection of Al3+. The detection limit of NQ for Al3+ detection was 1.98 μM, which is lower than the level (7.4 μM) in drinking water defined by the World Health Organization. In addition, the fluorescent probe NQ could be recyclable simply through treatment with a proper reagent such as F-, and could also be used for the detection of Al3+ in real samples.

  11. Construction of an efficient two-photon fluorescent probe for imaging nitroreductase in live cells and tissues

    Science.gov (United States)

    Zhou, Liyi; Gong, Liang; Hu, Shunqin

    2018-06-01

    Compared with traditional confocal microscopy, two-photon fluorescence microscopy (TPFM), which excites a two-photon (TP) fluorophore by near-infrared light, provides improved three-dimensional image resolution with increased tissue-image depth (>500 μm) and an extended observation time. Therefore, the development of novel functional TP fluorophores has attracted great attention in recent years. Herein, a novel TP fluorophore CM-NH2, which have the donor-π-acceptor (D-π-A)-structure, was designed and synthesized. We further used this dye developed a new type of TP fluorescent probe CM-NO2 for detecting nitroreductase (NTR). Upon incubated with NTR for 15 min, CM-NO2 displayed a 90-fold fluorescence enhancement at 505 nm and the maximal TP action cross-section value after reaction was detected and calculated to be 200 GM at 760 nm. The probe exhibited excellent properties such as high sensitivity, high selectivity, low cytotoxicity, and high photostability. Moreover, the probe was utilized to image the tumor hypoxia in live HeLa cells. Finally, using the CM-NO2 to image NTR in tissues was demonstrated.

  12. Phenotypical characteristics of leukocyte membranes in Chernobyl clean-up workers from Latvia: use of the fluorescent probe ABM

    International Nuclear Information System (INIS)

    Kalnina, I.; Meirovics, I.; Garbuseva, N.; Bruvere, R.; Heisele, O.; Zvagule, T.; Volrate, A.; Feldmane, G.

    2001-01-01

    A fluorescent probe - aminoderivative of benzanthrone, AMB (developed at the Riga Technical University, Riga, Latvia) - has been previously shown to localise within the phospholipid bilayer of the cell membrane, and shown to affect the structural and functional properties of peripheral blood mononuclear cells (PBMC). The probe ABM was used to characterise the PBMC membranes of 97 Chernobyl clean-up workers from Latvia. The study was conducted in the years 1997-1998. After addition of the probe to PBMC, fluorescence intensity of ABM (F) was measured, the depolarisation value P was calculated, and emission spectra were recorded. Screening of all individuals showed 5 different patterns of fluorescence spectra. Four of the patterns had never been previously observed in healthy individuals or patients with tuberculosis, multiple sclerosis, oncologic patients, etc., examined by us. The spectral patterns of ABM suspensions were associated with ability of leukocytes to produce interferons, with the levels of immunoglobulins A, G, and M, the concentration of lead in peripheral blood, and with several neurologic diseases. The use of ABM allowed to show phenotypic differences in PBMC between Chernobyl clean-up workers and individuals who had never had professional contact with radioactivity. (author)

  13. Sets of RNA repeated tags and hybridization-sensitive fluorescent probes for distinct images of RNA in a living cell.

    Directory of Open Access Journals (Sweden)

    Takeshi Kubota

    Full Text Available BACKGROUND: Imaging the behavior of RNA in a living cell is a powerful means for understanding RNA functions and acquiring spatiotemporal information in a single cell. For more distinct RNA imaging in a living cell, a more effective chemical method to fluorescently label RNA is now required. In addition, development of the technology labeling with different colors for different RNA would make it easier to analyze plural RNA strands expressing in a cell. METHODOLOGY/PRINCIPAL FINDINGS: Tag technology for RNA imaging in a living cell has been developed based on the unique chemical functions of exciton-controlled hybridization-sensitive oligonucleotide (ECHO probes. Repetitions of selected 18-nucleotide RNA tags were incorporated into the mRNA 3'-UTR. Pairs with complementary ECHO probes exhibited hybridization-sensitive fluorescence emission for the mRNA expressed in a living cell. The mRNA in a nucleus was detected clearly as fluorescent puncta, and the images of the expression of two mRNAs were obtained independently and simultaneously with two orthogonal tag-probe pairs. CONCLUSIONS/SIGNIFICANCE: A compact and repeated label has been developed for RNA imaging in a living cell, based on the photochemistry of ECHO probes. The pairs of an 18-nt RNA tag and the complementary ECHO probes are highly thermostable, sequence-specifically emissive, and orthogonal to each other. The nucleotide length necessary for one tag sequence is much shorter compared with conventional tag technologies, resulting in easy preparation of the tag sequences with a larger number of repeats for more distinct RNA imaging.

  14. "Clickable" LNA/DNA probes for fluorescence sensing of nucleic acids and autoimmune antibodies

    DEFF Research Database (Denmark)

    Jørgensen, Anna S; Gupta, Pankaj; Wengel, Jesper

    2013-01-01

    Herein we describe fluorescent oligonucleotides prepared by click chemistry between novel alkyne-modified locked nucleic acid (LNA) strands and a series of fluorescent azides for homogeneous (all-in-solution) detection of nucleic acids and autoimmune antibodies.......Herein we describe fluorescent oligonucleotides prepared by click chemistry between novel alkyne-modified locked nucleic acid (LNA) strands and a series of fluorescent azides for homogeneous (all-in-solution) detection of nucleic acids and autoimmune antibodies....

  15. Dual PET and Near-Infrared Fluorescence Imaging Probes as Tools for Imaging in Oncology

    Science.gov (United States)

    An, Fei-Fei; Chan, Mark; Kommidi, Harikrishna; Ting, Richard

    2016-01-01

    OBJECTIVE The purpose of this article is to summarize advances in PET fluorescence resolution, agent design, and preclinical imaging that make a growing case for clinical PET fluorescence imaging. CONCLUSION Existing SPECT, PET, fluorescence, and MRI contrast imaging techniques are already deeply integrated into the management of cancer, from initial diagnosis to the observation and management of metastases. Combined positron-emitting fluorescent contrast agents can convey new or substantial benefits that improve on these proven clinical contrast agents. PMID:27223168

  16. Characterisation of corrosion processes of using electron micro-probe, scanning probe microscopy and synchrotron-generated x-ray fluorescence imaging

    International Nuclear Information System (INIS)

    Neufeld, A.K.; Cole, I.S.; Furman, S.A.; Isaacs, H.S.

    2002-01-01

    Full text: With recent advances in computerized technology, the study of chemical reactions can now be visualized as they occur in real time and has resulted in analytical techniques with orders of magnitude greater sensitivity and resolution. This ability offers the corrosion scientist a unique opportunity to study the processes relevant to degradation science which could only be theoretically considered. Neufeld el al (1,2) have attempted to explain in great detail the mechanism of corrosion initiation of zinc by using X-ray micro-probe, Scanning Kelvin probe, and more recently by using synchrotron-generated X-rays and X-ray fluorescence imaging. New results are presented from the synchrotron studies where the transport of ions in-situ has been investigated. The synthesis of information from the techniques will also be discussed in its relevance to atmospheric corrosion processes. Copyright (2002) Australian Society for Electron Microscopy Inc

  17. Probing Lipid Coating Dynamics of Quantum Dot Core Micelles via Forster Resonance Energy Transfer

    NARCIS (Netherlands)

    Zhao, Yiming; Schapotschnikow, Philipp; Skajaa, Torjus; Vlugt, Thijs J. H.; Mulder, Willem J. M.; de Mello Donegá, Celso; Meijerink, Andries

    2014-01-01

    Lipid coated nanocrystal assemblies are among the most extensively investigated nanoparticle platforms for biomedical imaging and therapeutic purposes. However, very few efforts have been addressed to the lipid coating exchange dynamics in such systems, which is key to our understanding of the

  18. Development of a Novel Fiber Optic Sensor Combined with a Fluorescence Turn-on Probe for Cu (II Detection

    Directory of Open Access Journals (Sweden)

    Ma J.

    2013-04-01

    Full Text Available Existing staining-based methodology for the detection of metal ions is not well suited for real-time or in situ use. This is a significant problem, given that these ions can have a considerable impact on both human health and the environment. Thus, there is growing interest and need for simple, rapid and in-situ monitoring techniques for the purpose of detecting various target analytes (e.g. heavy metals, which is of a significant importance in many fields ranging from environmental monitoring to the study of intracellular processes. Among various sensors developed, optical fiber-optic sensors (FOS, based on fluorescence, are one class of sensors that address this goal [1]. Optical fibers are ideal for environmental sensing applications because of their ability to transmit optical signals to and from the sensing region without the use of free-space optics. In this work, we present, for the first time, a simple FOS incorporating novel fluorescence turn-on mechanism [2] that could detect Cu (II as low as 10−4 M. Traditionally, fluorescence quenching or “turn-off” was used to detect Cu (II [3]. In recent years, fluorescence “turn-on” emerges as a preferable tool. The developed fiber-optic sensor has two fiber leads and one probe head. One fiber lead includes 6 fibers for He-Ne laser excitation light delivery (e-fibers. Another fiber lead has one receiving fiber (r-fiber connected to an Ocean Optics QE65000 scientific grade spectrometer, which is interrogated by a computer via USB connection. The SpectroSuite software is used to observe and to record all spectra. The probe head combines all fibers together to form a coaxial structure with the r-fiber placed in the center. The key component in the proposed fluorescent sensing system is a probe prepared by binding a receptor containing a zwitterionic chromophore (M1, through noncovalent interactions, to the fluorescent polymer (P1 resulting in quenching its emission. The sensing mechanism

  19. [Sentinel node detection using optonuclear probe (gamma and fluorescence) after green indocyanine and radio-isotope injections].

    Science.gov (United States)

    Poumellec, M-A; Dejode, M; Figl, A; Darcourt, J; Haudebourg, J; Sabah, Y; Voury, A; Martaens, A; Barranger, E

    2016-04-01

    Assess the biopsy's feasibility of the sentinel lymph node biopsy (SLNB) using optonuclear probe after of indocyanine green (ICG) and radio-isotope (RI) injections. Twenty-one patients with a localized breast cancer and unsuspicious axillary nodes underwent a SLNB after both injections of ICG and radio-isotope. One or more SLN were identified on the 21 patients (identification rate of 100%). The median number SLN was 2 (1-3). Twenty SLN were both radio-actives and fluorescents (54.1%), 11 fluorescent only (29.7%) and 6 were only radio-actives (16.2%). Seven patients had a metastatic SLN (8 SLN overall). Among them, only one had a micrometastasic SLN, 5 others had a macrometastatic SLN and one patient had two macrometastatic SLNs. Among the 8 metastatic SLN, 5 were both fluorescent and radioactive, 2 were only fluorescent and 1 was only radioactive. Detection SLN using optonuclear probe after indocyanine green and radio-isotope injections is effective and could be, after validation by randomized trial, a reliable alternative to the blue dye injection for teams who consider that combined detection as the reference. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  20. Kinetics of intercalation of fluorescent probes in magnesium–aluminium layered double hydroxide within a multiscale reaction–diffusion framework

    Science.gov (United States)

    Saliba, Daniel

    2016-01-01

    We report the synthesis of magnesium–aluminium layered double hydroxide (LDH) using a reaction–diffusion framework (RDF) that exploits the multiscale coupling of molecular diffusion with chemical reactions, nucleation and growth of crystals. In an RDF, the hydroxide anions are allowed to diffuse into an organic gel matrix containing the salt mixture needed for the precipitation of the LDH. The chemical structure and composition of the synthesized magnesium–aluminium LDHs are determined using powder X-ray diffraction (PXRD), thermo-gravimetric analysis, differential scanning calorimetry, solid-state nuclear magnetic resonance (SSNMR), Fourier transform infrared and energy dispersive X-ray spectroscopy. This novel technique also allows the investigation of the mechanism of intercalation of some fluorescent probes, such as the neutral three-dimensional rhodamine B (RhB) and the negatively charged two-dimensional 8-hydroxypyrene-1,3,6-trisulfonic acid (HPTS), using in situ steady-state fluorescence spectroscopy. The incorporation of these organic dyes inside the interlayer region of the LDH is confirmed via fluorescence microscopy, solid-state lifetime, SSNMR and PXRD. The activation energies of intercalation of the corresponding molecules (RhB and HPTS) are computed and exhibit dependence on the geometry of the involved probe (two or three dimensions), the charge of the fluorescent molecule (anionic, cationic or neutral) and the cationic ratio of the corresponding LDH. This article is part of the themed issue ‘Multiscale modelling at the physics–chemistry–biology interface’. PMID:27698034

  1. Kinetics of intercalation of fluorescent probes in magnesium-aluminium layered double hydroxide within a multiscale reaction-diffusion framework

    Science.gov (United States)

    Saliba, Daniel; Al-Ghoul, Mazen

    2016-11-01

    We report the synthesis of magnesium-aluminium layered double hydroxide (LDH) using a reaction-diffusion framework (RDF) that exploits the multiscale coupling of molecular diffusion with chemical reactions, nucleation and growth of crystals. In an RDF, the hydroxide anions are allowed to diffuse into an organic gel matrix containing the salt mixture needed for the precipitation of the LDH. The chemical structure and composition of the synthesized magnesium-aluminium LDHs are determined using powder X-ray diffraction (PXRD), thermo-gravimetric analysis, differential scanning calorimetry, solid-state nuclear magnetic resonance (SSNMR), Fourier transform infrared and energy dispersive X-ray spectroscopy. This novel technique also allows the investigation of the mechanism of intercalation of some fluorescent probes, such as the neutral three-dimensional rhodamine B (RhB) and the negatively charged two-dimensional 8-hydroxypyrene-1,3,6-trisulfonic acid (HPTS), using in situ steady-state fluorescence spectroscopy. The incorporation of these organic dyes inside the interlayer region of the LDH is confirmed via fluorescence microscopy, solid-state lifetime, SSNMR and PXRD. The activation energies of intercalation of the corresponding molecules (RhB and HPTS) are computed and exhibit dependence on the geometry of the involved probe (two or three dimensions), the charge of the fluorescent molecule (anionic, cationic or neutral) and the cationic ratio of the corresponding LDH. This article is part of the themed issue 'Multiscale modelling at the physics-chemistry-biology interface'.

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

    Science.gov (United States)

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

    1994-06-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1997-04-01

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

  4. Spectrofluorimetric quantification of bilirubin using yttrium-norfloxacin complex as a fluorescence probe in serum samples

    Energy Technology Data Exchange (ETDEWEB)

    Kamruzzaman, Mohammad; Alam, Al-Mahmnur [Department of Chemistry, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Hak Lee, Sang, E-mail: shlee@knu.ac.kr [Department of Chemistry, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Ho Kim, Young, E-mail: youngkim@knu.ac.kr [Research Institute of Advanced Energy Technology, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Kim, Gyu-Man [School of Mechanical Engineering, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Hyub Oh, Sang [Center for Gas Analysis, Korea Research Institute of Standards and Science, Daejeon 305-600 (Korea, Republic of)

    2012-11-15

    A simple and sensitive spectrofluorimetric method was developed to determine trace amounts of bilirubin (BR) using yttrium (Y{sup 3+})-norfloxacin (NFLX) complex as a fluorescence (FL) probe. NFLX can form a stable binary complex with Y{sup 3+} and markedly enhances the weak FL signal of the NFLX. The FL intensity of the Y{sup 3+}-NFLX complex decreased significantly in the presence of BR in a buffer solution at pH=7.2. Under optimal conditions, the FL intensity decreased according to the BR concentration and showed a good linear relationship in the range of 0.03-2.3 {mu}g mL{sup -1} of BR with a correlation coefficient of 0.9988. The limit of detection for the determination of BR was 2.8 ng mL{sup -1} with a relative standard deviation (RSD) of 1.55% for five replicate determination of 0.05 {mu}g mL{sup -1} BR. The presented method offers higher sensitivity with simple instrumentation and was applied successfully in detecting BR at low concentrations. Highlights: Black-Right-Pointing-Pointer Weak FL signal of NFLX was enhanced at 419 nm by forming binary complex with Y{sup 3+}. Black-Right-Pointing-Pointer The FL intensity of Y{sup 3+}-NFLX complex was quenched markedly in the presence of ATP. Black-Right-Pointing-Pointer NFLX can transfer energy to Y{sup 3+} and BR and form the Y{sup 3+}-NFLX-ATP ternary complex. Black-Right-Pointing-Pointer The reduced FL intensity of the system was correlated with the concentration of BR. Black-Right-Pointing-Pointer The method is applied to determine BR at low concentration (2.8 ng mL{sup -1}) in serum.

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

    Science.gov (United States)

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

    2011-01-01

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

  6. A convenient colorimetric and ratiometric fluorescent probe for detection of cyanide based on BODIPY derivative in aqueous media

    Directory of Open Access Journals (Sweden)

    Yanhua Yu

    2017-06-01

    Full Text Available A convenient colorimetric and ratiometric fluorescent probe based on BODIPY derivative for cyanide detection has been synthesized, whose structural contains a dicyanovinyl group used as a sensing unit. Among the tested analytes, such as CN−, F−, Cl−, Br−, I−, ClO4−, AcO−, NO3−, H2PO4− HSO4−, S2− and N3−, only CN− could react with dicyanovinyl moiety by nucleophilic addition, which disrupted the π-conjugation of the probe and hindered the intramolecular charge transfer (ICT, leading a blue shift of absorption and fluorescence spectrum and a concomitant color change from yellow to light pink. The detection limit of this probe was calculated to be 0.98 μM, which is lower than the maximum concentration in drinking water (1.9 μM permitted by the World Health Organization (WHO. Moreover, the probe showed excellent selectivity and anti-interference ability towards CN− over other anions. The reaction mechanism was fully supported by 1H NMR and MS spectrum.

  7. Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes for Rapid Identification of Candida albicans Directly from Blood Culture Bottles

    Science.gov (United States)

    Rigby, Susan; Procop, Gary W.; Haase, Gerhard; Wilson, Deborah; Hall, Geraldine; Kurtzman, Cletus; Oliveira, Kenneth; Von Oy, Sabina; Hyldig-Nielsen, Jens J.; Coull, James; Stender, Henrik

    2002-01-01

    A new fluorescence in situ hybridization (FISH) method that uses peptide nucleic acid (PNA) probes for identification of Candida albicans directly from positive-blood-culture bottles in which yeast was observed by Gram staining (herein referred to as yeast-positive blood culture bottles) is described. The test (the C. albicans PNA FISH method) is based on a fluorescein-labeled PNA probe that targets C. albicans 26S rRNA. The PNA probe is added to smears made directly from the contents of the blood culture bottle and hybridized for 90 min at 55°C. Unhybridized PNA probe is removed by washing of the mixture (30 min), and the smears are examined by fluorescence microscopy. The specificity of the method was confirmed with 23 reference strains representing phylogenetically related yeast species and 148 clinical isolates covering the clinically most significant yeast species, including C. albicans (n = 72), C. dubliniensis (n = 58), C. glabrata (n = 5), C. krusei (n = 2), C. parapsilosis (n = 4), and C. tropicalis (n = 3). The performance of the C. albicans PNA FISH method as a diagnostic test was evaluated with 33 routine and 25 simulated yeast-positive blood culture bottles and showed 100% sensitivity and 100% specificity. It is concluded that this 2.5-h method for the definitive identification of C. albicans directly from yeast-positive blood culture bottles provides important information for optimal antifungal therapy and patient management. PMID:12037084

  8. A NBD-based simple but effective fluorescent pH probe for imaging of lysosomes in living cells.

    Science.gov (United States)

    Cao, Xiang-Jian; Chen, Li-Na; Zhang, Xuan; Liu, Jin-Ting; Chen, Ming-Yu; Wu, Qiu-Rong; Miao, Jun-Ying; Zhao, Bao-Xiang

    2016-05-12

    NBDlyso with lysosome-locating morpholine moiety has been developed as a high selective and sensitive fluorescent pH probe. This probe can respond to acidic pH (2.0-7.0) in a short time (less than 1 min) and not almost change after continuously illuminated for an extended period by ultraviolet light. The fluorescence intensity of NBDlyso enhanced 100-fold in acidic solution, with very good linear relationship (R(2) = 0.996). The pKa of probe NBDlyso is 4.10. Therefore, NBDlyso was used to detect lysosomal pH changes successfully. Besides, X-ray crystallography was used to verify the structure of NBDlyso, and the recognition mechanism involving photo-induced electron transfer was interpreted theoretically by means of DFT and TDDFT calculations skillfully when NBDlyso comes into play under the acidic condition. This probe showed good ability to sense pH change in living cell image. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. New concepts of fluorescent probes for specific detection of DNA sequences: bis-modified oligonucleotides in excimer and exciplex detection.

    Science.gov (United States)

    Gbaj, A; Bichenkova, Ev; Walsh, L; Savage, He; Sardarian, Ar; Etchells, Ll; Gulati, A; Hawisa, S; Douglas, Kt

    2009-12-01

    The detection of single base mismatches in DNA is important for diagnostics, treatment of genetic diseases, and identification of single nucleotide polymorphisms. Highly sensitive, specific assays are needed to investigate genetic samples from patients. The use of a simple fluorescent nucleoside analogue in detection of DNA sequence and point mutations by hybridisation in solution is described in this study. The 5'-bispyrene and 3'-naphthalene oligonucleotide probes form an exciplex on hybridisation to target in water and the 5'-bispyrene oligonucleotide alone is an adequate probe to determine concentration of target present. It was also indicated that this system has a potential to identify mismatches and insertions. The aim of this work was to investigate experimental structures and conditions that permit strong exciplex emission for nucleic acid detectors, and show how such exciplexes can register the presence of mismatches as required in SNP analysis. This study revealed that the hybridisation of 5'-bispyrenyl fluorophore to a DNA target results in formation of a fluorescent probe with high signal intensity change and specificity for detecting a complementary target in a homogeneous system. Detection of SNP mutations using this split-probe system is a highly specific, simple, and accessible method to meet the rigorous requirements of pharmacogenomic studies. Thus, it is possible for the system to act as SNP detectors and it shows promise for future applications in genetic testing.

  10. Dansyl-8-aminoquinoline as a sensitive pH fluorescent probe with dual-responsive ranges in aqueous solutions.

    Science.gov (United States)

    Zhang, Min; Zheng, Shuyu; Ma, Liguo; Zhao, Meili; Deng, Lengfang; Yang, Liting; Ma, Li-Jun

    2014-04-24

    A sensitive pH fluorescent probe based on dansyl group, dansyl-8-aminoquinoline (DAQ), has been synthesized. The probe showed dual-responsive ranges to pH changes, one range from 2.00 to 7.95 and another one from 7.95 to 10.87 in aqueous solution, as it showed pKa values of 5.73 and 8.56 under acid and basic conditions, respectively. Furthermore, the pH response mechanism of the probe was explored successfully by using NMR spectra. The results indicated that the responses of DAQ to pH changes should attribute to the protonation of the nitrogen atom in the dimethylamino group and deprotonation of sulfonamide group. Copyright © 2014. Published by Elsevier B.V.

  11. Fluorescent probes for understanding soil water repellency: the novel application of a chemist's tool to soil science

    Science.gov (United States)

    Balshaw, Helen M.; Davies, Matthew L.; Doerr, Stefan H.; Douglas, Peter

    2015-04-01

    Food security and production is one of the key global issues faced by society. It has become essential to work the land efficiently, through better soil management and agronomy whilst protecting the environment from air and water pollution. The failure of soil to absorb water - soil water repellency can lead to major environmental problems such as increased overland flow and soil erosion, poor uptake of agricultural chemicals, and increased risk of groundwater pollution due to the rapid transfer of contaminants and nutrient leaching through uneven wetting and preferential flow pathways. Understanding the causes of soil hydrophobicity is essential for the development of effective methods for its amelioration, supporting environmental stability and food security. Organic compounds deposited on soil mineral or aggregate surfaces have long been recognised as a major factor in causing soil water repellency. It is widely accepted that the main groups of compounds responsible are long-chain acids, alkanes and other organic compounds with hydrophobic properties. However, when reapplied to sands and soils, the degree of water repellency induced by these compounds and mixtures varied widely with compound type, amount, and mixture, in a seemingly unpredictable way. Fluorescent and phosphorescent probes are widely used in chemistry and biochemistry due to their sensitive response to their physical and chemical environment, such as polarity, and viscosity. However, they have to-date not been used to study soil water repellency. Here we present preliminary work on the evaluation of fluorescent probes as tools to study two poorly understood features that determine the degree of wettability for water repellent soils: (i) the distribution of organics on soils; (ii) the changes in polarity at soil surfaces required for water drops to infiltrate. In our initial work we have examined probes adsorbed onto model soils, prepared by adsorption of specific organics onto acid washed sand

  12. A fast-response two-photon fluorescent probe for imaging endogenous H2O2 in living cells and tissues

    Science.gov (United States)

    Lu, Yanan; Shi, Xiaomin; Fan, Wenlong; Black, Cory A.; Lu, Zhengliang; Fan, Chunhua

    2018-02-01

    As a second messenger, hydrogen peroxide plays significant roles in numerous physiological and pathological processes and is related to various diseases including inflammatory disease, diabetes, neurodegenerative disorders, cardiovascular disease and Alzheimer's disease. Two-photon (TP) fluorescent probes reported for the detection of endogenous H2O2 are rare and most have drawbacks such as slow response and low sensitivity. In this report, we demonstrate a simple H2O2-specific TP fluorescent probe (TX-HP) containing a two-photon dye 6-hydroxy-2,3,4,4a-tetrahydro-1H-xanthen-1-one (TX) on the modulation of the ICT process. The probe exhibits a rapid fluorescent response to H2O2 in 9 min with both high sensitivity and selectivity. The probe can detect exogenous H2O2 in living cells. Furthermore, the probe is successfully utilized for imaging H2O2 in liver tissues.

  13. Dual-Modal Colorimetric/Fluorescence Molecular Probe for Ratiometric Sensing of pH and Its Application.

    Science.gov (United States)

    Wu, Luling; Li, Xiaolin; Huang, Chusen; Jia, Nengqin

    2016-08-16

    As traditional pH meters cannot work well for minute regions (such as subcellular organelles) and in harsh media, molecular pH-sensitive devices for monitoring pH changes in diverse local heterogeneous environments are urgently needed. Here, we report a new dual-modal colorimetric/fluorescence merocyanine-based molecular probe (CPH) for ratiometric sensing of pH. Compared with previously reported pH probes, CPH bearing the benzyl group at the nitrogen position of the indolium group and the phenol, which is used as the acceptor for proton, could respond to pH changes immediately through both the ratiometric fluorescence signal readout and naked-eye colorimetric observation. The sensing process was highly stable and reversible. Most importantly, the suitable pKa value (6.44) allows CPH to presumably accumulate in lysosomes and become a lysosome-target fluorescent probe. By using CPH, the intralysosomal pH fluctuation stimulated by antimalaria drug chloroquine was successfully tracked in live cells through the ratiometric fluorescence images. Additionally, CPH could be immobilized on test papers, which exhibited a rapid and reversible colorimetric response to acid/base vapor through the naked-eye colorimetric analysis. This proof-of-concept study presents the potential application of CPH as a molecular tool for monitoring intralysosomal pH fluctuation in live cells, as well as paves the way for developing the economic, reusable, and fast-response optical pH meters for colorimetric sensing acid/base vapor with direct naked-eye observation.

  14. Synthesis of water-soluble, ring-substituted squaraine dyes and their evaluation as fluorescent probes and labels

    Energy Technology Data Exchange (ETDEWEB)

    Tatarets, Anatoliy L. [SSI ' Institute for Single Crystals' of the National Academy of Sciences of Ukraine, 60 Lenin Ave., Kharkov 61001 (Ukraine); Fedyunyayeva, Irina A. [SSI ' Institute for Single Crystals' of the National Academy of Sciences of Ukraine, 60 Lenin Ave., Kharkov 61001 (Ukraine); Dyubko, Tatyana S. [SSI ' Institute for Single Crystals' of the National Academy of Sciences of Ukraine, 60 Lenin Ave., Kharkov 61001 (Ukraine); Povrozin, Yevgeniy A. [SSI ' Institute for Single Crystals' of the National Academy of Sciences of Ukraine, 60 Lenin Ave., Kharkov 61001 (Ukraine); Doroshenko, Andrey O. [Institute of Chemistry, V.N. Karazin National University, 4 Svobody Sq., Kharkov 61077 (Ukraine); Terpetschnig, Ewald A. [SETA BioMedicals, LLC, 2014 Silver Ct East, Urbana, IL 61801 (United States) and ISS, Inc., 1602 Newton Drive, Champaign, IL 61822 (United States)]. E-mail: ewaldte@juno.com; Patsenker, Leonid D. [SSI ' Institute for Single Crystals' of the National Academy of Sciences of Ukraine, 60 Lenin Ave., Kharkov 61001 (Ukraine); SETA BioMedicals, LLC, 2014 Silver Ct East, Urbana, IL 61801 (United States)

    2006-06-16

    A series of ring-substituted squaraines absorbing and emitting in the red and NIR spectral region was synthesized and their spectral and photophysical properties (quantum yields, fluorescence lifetimes) and photostabilities were measured and compared to Cy5, a commonly used fluorescent label. The absorption maxima in aqueous media were found to be between 628 and 667 nm and the emission maxima are between 642 and 685 nm. Squaraine dyes exhibit high extinction coefficients (163,000-265,000 M{sup -1} cm{sup -1}) and lower quantum yields (2-7%) in aqueous buffer but high quantum yields (up to 45%) and long fluorescence lifetimes (up to 3.3 ns) in presence of BSA. Dicyanomethylene- and thio-substituted squaraines exhibit an additional absorption around 400 nm with extinction coefficients between 21,500 and 44,500 M{sup -1} cm{sup -1}. These dyes are excitable not only with red but also with blue diode lasers or light emitting diodes. Due to the favourable spectral and photophysical properties these dyes can be used as fluorescent probes and labels for intensity- and fluorescence lifetime-based biomedical applications.

  15. Probing Contaminant-Induced Alterations in Chlorophyll Fluorescence by AC-Dielectrophoresis-Based 2D-Algal Array

    Directory of Open Access Journals (Sweden)

    Coralie Siebman

    2018-02-01

    Full Text Available The investigation of contaminant impact on algae requires rapid and reliable cell collection and optical detection. The capability of alternative current (AC dielectrophoresis (DEP collection of whole cell arrays with combined fluorescence microscopy detection to follow the alterations of chlorophyll fluorescence during environmental contaminant exposure was explored. The application of an AC-field of 100 V cm−1, 100 Hz for 30 min to capture and immobilize the cells of green alga Chlamydomonas reinhardtii in two-dimensional (2D arrays does not induce changes in chlorophyll fluorescence. The results demonstrate that DEP-based 2D-arrays allow non-invasive detection of chlorophyll fluorescence change upon exposure to high concentrations of copper oxide nanoparticles and ionic copper. These results were in agreement with data obtained by flow cytometry used as a comparative method. The tool was also applied to follow the effect of a number of ubiquitous contaminants such as inorganic mercury, methylmercury, and diuron. However, a statistically significant short-term effect was observed only for mercury. Overall, DEP-based 2D-arrays of algal cells with fluorescence detection appear to be suitable for stain-free probing the effects on the photosynthetic microorganisms in highly polluted environment.

  16. Combined fluorescence-Raman spectroscopy measurements with an optical fiber probe for the diagnosis of melanocytic lesions

    Science.gov (United States)

    Cosci, Alessandro; Cicchi, Riccardo; Rossari, Susanna; De Giorgi, Vincenzo; Massi, Daniela; Pavone, Francesco S.

    2012-02-01

    We have designed and developed an optical fiber-probe for spectroscopic measurements on human tissues. The experimental setup combines fluorescence spectroscopy and Raman spectroscopy in a multidimensional approach. Concerning fluorescence spectroscopy, the excitation is provided by two laser diodes, one emitting in the UV (378 nm) and the other emitting in the visible (445 nm). These two lasers are used to selectively excite fluorescence from NADH and FAD, which are among the brightest endogenous fluorophores in human tissues. For Raman and NIR spectroscopy, the excitation is provided by a third laser diode with 785 nm excitation wavelength. Laser light is delivered to the tissue through the central optical fiber of a fiber bundle. The surrounding 48 fibers of the bundle are used for collecting fluorescence and Raman and for delivering light to the spectrograph. Fluorescence and Raman spectra are acquired on a cooled CCD camera. The instrument has been tested on fresh human skin biopsies clinically diagnosed as malignant melanoma, melanocytic nevus, or healthy skin, finding an optimal correlation with the subsequent histological exam. In some cases our examination was not in agreement with the clinical observation, but it was with the histological exam, demonstrating that the system can potentially contribute to improve clinical diagnostic capabilities and hence reduce the number of unnecessary biopsies.

  17. Synthesis of water-soluble, ring-substituted squaraine dyes and their evaluation as fluorescent probes and labels

    International Nuclear Information System (INIS)

    Tatarets, Anatoliy L.; Fedyunyayeva, Irina A.; Dyubko, Tatyana S.; Povrozin, Yevgeniy A.; Doroshenko, Andrey O.; Terpetschnig, Ewald A.; Patsenker, Leonid D.

    2006-01-01

    A series of ring-substituted squaraines absorbing and emitting in the red and NIR spectral region was synthesized and their spectral and photophysical properties (quantum yields, fluorescence lifetimes) and photostabilities were measured and compared to Cy5, a commonly used fluorescent label. The absorption maxima in aqueous media were found to be between 628 and 667 nm and the emission maxima are between 642 and 685 nm. Squaraine dyes exhibit high extinction coefficients (163,000-265,000 M -1 cm -1 ) and lower quantum yields (2-7%) in aqueous buffer but high quantum yields (up to 45%) and long fluorescence lifetimes (up to 3.3 ns) in presence of BSA. Dicyanomethylene- and thio-substituted squaraines exhibit an additional absorption around 400 nm with extinction coefficients between 21,500 and 44,500 M -1 cm -1 . These dyes are excitable not only with red but also with blue diode lasers or light emitting diodes. Due to the favourable spectral and photophysical properties these dyes can be used as fluorescent probes and labels for intensity- and fluorescence lifetime-based biomedical applications

  18. A highly selective turn-on fluorescent probe for hypochlorous acid based on hypochlorous acid-induced oxidative intramolecular cyclization of boron dipyrromethene-hydrazone

    International Nuclear Information System (INIS)

    Chen, Wei-Chieh; Venkatesan, Parthiban; Wu, Shu-Pao

    2015-01-01

    Highlights: • A BODIPY-based fluorescent probe for sensing HOCl was developed. • The probe utilizes the HOCl-promoted cyclization in response to the amount of HOCl. • The probe might have application in the investigation of HOCl in biological systems. - Abstract: A BODIPY-based fluorescent probe, HBP, was developed for the detection of hypochlorous acid based on the specific hypochlorous acid-promoted oxidative intramolecular cyclization of heterocyclic hydrazone in response to the amount of HOCl. The reaction is accompanied by a 41-fold increase in the fluorescent quantum yield (from 0.004 to 0.164). The fluorescence intensity of the reaction between HOCl and HBP is linear in the HOCl concentration range of 1–8 μM with a detection limit of 2.4 nM (S/N = 3). Confocal fluorescence microscopy imaging using RAW264.7 cells showed that the new probe HBP could be used as an effective fluorescent probe for detecting HOCl in living cells

  19. A lysosome-locating and acidic pH-activatable fluorescent probe for visualizing endogenous H2O2 in lysosomes.

    Science.gov (United States)

    Liu, Jun; Zhou, Shunqing; Ren, Jing; Wu, Chuanliu; Zhao, Yibing

    2017-11-20

    There is increasing evidence indicating that lysosomal H 2 O 2 is closely related to autophagy and apoptotic pathways under both physiological and pathological conditions. Therefore, fluorescent probes that can be exploited to visualize H 2 O 2 in lysosomes are potential tools for exploring diverse roles of H 2 O 2 in cells. However, functional exploration of lysosomal H 2 O 2 is limited by the lack of fluorescent probes capable of compatibly sensing H 2 O 2 under weak acidic conditions (pH = 4.5) of lysosomes. Lower spatial resolution of the fluorescent visualization of lysosomal H 2 O 2 might be caused by the interference of signals from cytosolic and mitochondrial H 2 O 2 , as well as the non-specific distribution of the probes in cells. In this work, we developed a lysosome-locating and acidic-pH-activatable fluorescent probe for the detection and visualization of H 2 O 2 in lysosomes, which consists of a H 2 O 2 -responsive boronate unit, a lysosome-locating morpholine group, and a pH-activatable benzorhodol fluorophore. The response of the fluorescent probe to H 2 O 2 is significantly more pronounced under acidic pH conditions than that under neutral pH conditions. Notably, the present probe enables the fluorescence sensing of endogenous lysosomal H 2 O 2 in living cells without external stimulations, with signal interference from the cytoplasm and other intracellular organelles being negligible.

  20. A highly selective turn-on fluorescent probe for hypochlorous acid based on hypochlorous acid-induced oxidative intramolecular cyclization of boron dipyrromethene-hydrazone

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Wei-Chieh; Venkatesan, Parthiban; Wu, Shu-Pao, E-mail: spwu@mail.nctu.edu.tw

    2015-07-02

    Highlights: • A BODIPY-based fluorescent probe for sensing HOCl was developed. • The probe utilizes the HOCl-promoted cyclization in response to the amount of HOCl. • The probe might have application in the investigation of HOCl in biological systems. - Abstract: A BODIPY-based fluorescent probe, HBP, was developed for the detection of hypochlorous acid based on the specific hypochlorous acid-promoted oxidative intramolecular cyclization of heterocyclic hydrazone in response to the amount of HOCl. The reaction is accompanied by a 41-fold increase in the fluorescent quantum yield (from 0.004 to 0.164). The fluorescence intensity of the reaction between HOCl and HBP is linear in the HOCl concentration range of 1–8 μM with a detection limit of 2.4 nM (S/N = 3). Confocal fluorescence microscopy imaging using RAW264.7 cells showed that the new probe HBP could be used as an effective fluorescent probe for detecting HOCl in living cells.

  1. Investigation of molecular mechanisms of action of chelating drugs on protein-lipid model membranes by X-ray fluorescence

    International Nuclear Information System (INIS)

    Novikova, N. N.; Zheludeva, S. I.; Koval'chuk, M. V.; Stepina, N. D.; Erko, A. I.; Yur'eva, E. A.

    2009-01-01

    Protein-lipid films based on the enzyme alkaline phosphatase were subjected to the action of chelating drugs, which are used for accelerating the removal of heavy metals from the human body, and the elemental composition of the resulting films was investigated. Total-reflection X-ray fluorescence measurements were performed at the Berlin Electron Storage Ring Company for Synchrotron Radiation (BESSY) in Germany. A comparative estimation of the protective effect of four drugs (EDTA, succimer, xydiphone, and mediphon) on membrane-bound enzymes damaged by lead ions was made. The changes in the elemental composition of the protein-lipid films caused by high doses of chelating drugs were investigated. It was shown that state-of-the-art X-ray techniques can, in principle, be used to develop new methods for the in vitro evaluation of the efficiency of drugs, providing differential data on their actions.

  2. Synthesis and evaluation of [{sup 14}C]-Labelled and fluorescent-Tagged paclitaxel derivatives as new biological probes

    Energy Technology Data Exchange (ETDEWEB)

    Rao, C.S.; Chu, J.-J.; Lai, Y.-K. [Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan (China); Liu, R.-S. [Department of Chemistry, National Tsing Hua University, Hsinchu, Taiwan (China)

    1998-11-01

    Our present report deals with the preparation of hitherto unreported 7-([carbonyl-{sup 14}C]-acetyl)paclitaxel 4 and two new bioactive 7-substituted fluorescent taxoids (FITC 9 and rhodamine 11), as well as evaluation towards their applications as biological probes. The results in this report demonstrate that (a) the new paclitaxel derivatives 4, 9, 11 could be prepared with good yields starting from paclitaxel; (b) the [{sup 14}C]acetylation step was found to be better by using [{sup 14}C]acetic anhydride rather than [{sup 14}C]sodium acetate; (c) the radiochemical purity of 4 was 96% and its specific activity was 48 mCi/mmol; (d) the cytotoxicity of 4 was close to that of paclitaxel whereas 9, 11 were far less active than paclitaxel, but these cytotoxic levels were good enough for their biological applications; (e) the drug-quantitation by flow cytometric analysis using 9 and 11 was proved to be equally efficient with respect to the radioactivity-based determination employing 4; (f) the intracellular fluorescence mapping by 9 and 11 was found to be effective and the microtubule network pattern was visible in both the cases; (g) the overall fluorescence imaging efficiency was better with 11 while the intensity of fluorescence was higher with 9; (h) staining of nucleolus was observed in fluorescence studies of both 9 and 11. Based on these results, the newly prepared paclitaxel derivatives can be considered as efficient biological probes and should find further use in relevant applications. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

  3. Fluorescent Inhibitors as Tools To Characterize Enzymes: Case Study of the Lipid Kinase Phosphatidylinositol 4-Kinase IIIβ (PI4KB).

    Science.gov (United States)

    Humpolickova, Jana; Mejdrová, Ivana; Matousova, Marika; Nencka, Radim; Boura, Evzen

    2017-01-12

    The lipid kinase phosphatidylinositol 4-kinase IIIβ (PI4KB) is an essential host factor for many positive-sense single-stranded RNA (+RNA) viruses including human pathogens hepatitis C virus (HCV), Severe acute respiratory syndrome (SARS), coxsackie viruses, and rhinoviruses. Inhibitors of PI4KB are considered to be potential broad-spectrum virostatics, and it is therefore critical to develop a biochemical understanding of the kinase. Here, we present highly potent and selective fluorescent inhibitors that we show to be useful chemical biology tools especially in determination of dissociation constants. Moreover, we show that the coumarin-labeled inhibitor can be used to image PI4KB in cells using fluorescence-lifetime imaging microscopy (FLIM) microscopy.

  4. Chloroquine transport in Plasmodium falciparum. 1. Influx and efflux kinetics for live trophozoite parasites using a novel fluorescent chloroquine probe.

    Science.gov (United States)

    Cabrera, Mynthia; Natarajan, Jayakumar; Paguio, Michelle F; Wolf, Christian; Urbach, Jeffrey S; Roepe, Paul D

    2009-10-13

    Several models for how amino acid substitutions in the Plasmodium falciparum chloroquine resistance transporter (PfCRT) confer resistance to chloroquine (CQ) and other antimalarial drugs have been proposed. Distinguishing between these models requires detailed analysis of high-resolution CQ transport data that is unfortunately impossible to obtain with traditional radio-tracer methods. Thus, we have designed and synthesized fluorescent CQ analogues for drug transport studies. One probe places a NBD (6-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoic acid) group at the tertiary aliphatic N of CQ, via a flexible 6 C amide linker. This probe localizes to the malarial parasite digestive vacuole (DV) during initial perfusion under physiologic conditions and exhibits similar pharmacology relative to CQ, vs both CQ-sensitive (CQS) and CQ-resistant (CQR) parasites. Using live, synchronized intraerythrocytic parasites under continuous perfusion, we define NBD-CQ influx and efflux kinetics for CQS vs CQR parasites. Since this fluorescence approach provides data at much higher kinetic resolution relative to fast-filtration methods using (3)H-CQ, rate constants vs linear initial rates for CQ probe flux can be analyzed in detail. Importantly, we find that CQR parasites have a decreased rate constant for CQ influx into the DV and that this is due to mutation of PfCRT. Analysis of zero trans efflux for CQS and CQR parasites suggests that distinguishing between bound vs free pools of intra-DV drug probe is essential for proper kinetic analysis of efflux. The accompanying paper (DOI 10.1021/bi901035j ) further probes efflux kinetics for proteoliposomes containing purified, reconstituted PfCRT.

  5. Lateral mobility of plasma membrane lipids in dividing Xenopus eggs

    NARCIS (Netherlands)

    Laat, S.W. de; Tetteroo, P.A.T.; Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van

    1984-01-01

    The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xaopus Levis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein (“HEDAF”) and 5-(N-tetradecanoyl)aminofluorescein (“TEDAF”) as probes. The

  6. Investigations on therapeutic glucocerebrosidases through paired detection with fluorescent activity-based probes.

    Directory of Open Access Journals (Sweden)

    Wouter W Kallemeijn

    Full Text Available Deficiency of glucocerebrosidase (GBA causes Gaucher disease (GD. In the common non-neuronopathic GD type I variant, glucosylceramide accumulates primarily in the lysosomes of visceral macrophages. Supplementing storage cells with lacking enzyme is accomplished via chronic intravenous administration of recombinant GBA containing mannose-terminated N-linked glycans, mediating the selective uptake by macrophages expressing mannose-binding lectin(s. Two recombinant GBA preparations with distinct N-linked glycans are registered in Europe for treatment of type I GD: imiglucerase (Genzyme, contains predominantly Man(3 glycans, and velaglucerase (Shire PLC Man(9 glycans. Activity-based probes (ABPs enable fluorescent labeling of recombinant GBA preparations through their covalent attachment to the catalytic nucleophile E340 of GBA. We comparatively studied binding and uptake of ABP-labeled imiglucerase and velaglucerase in isolated dendritic cells, cultured human macrophages and living mice, through simultaneous detection of different GBAs by paired measurements. Uptake of ABP-labeled rGBAs by dendritic cells was comparable, as well as the bio-distribution following equimolar intravenous administration to mice. ABP-labeled rGBAs were recovered largely in liver, white-blood cells, bone marrow and spleen. Lungs, brain and skin, affected tissues in severe GD types II and III, were only poorly supplemented. Small, but significant differences were noted in binding and uptake of rGBAs in cultured human macrophages, in the absence and presence of mannan. Mannan-competed binding and uptake were largest for velaglucerase, when determined with single enzymes or as equimolar mixtures of both enzymes. Vice versa, imiglucerase showed more prominent binding and uptake not competed by mannan. Uptake of recombinant GBAs by cultured macrophages seems to involve multiple receptors, including several mannose-binding lectins. Differences among cells from different donors

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

    DEFF Research Database (Denmark)

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

    2014-01-01

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

  8. Curcumin-Based "Enhanced SNAr" Promoted Ultrafast Fluorescent Probe for Thiophenols Detection in Aqueous Solution and in Living Cells.

    Science.gov (United States)

    Yue, Yongkang; Huo, Fangjun; Zhang, Yongbin; Chao, Jianbin; Martínez-Máñez, Ramón; Yin, Caixia

    2016-11-01

    We report herein a highly selective and sensitive turn-on fluorescent probe (compound 1) with a fast response time (less than 2 min) for thiophenol detection based on an "enhanced S N Ar" reaction between thiophenols and a sulfonyl-ester moiety covalently attach to curcumin. Reaction of 1 in Hepes-MeOH (1:1, v/v, pH 7.4) in the presence of 4-methylthiophenol (MTP) resulted in a remarkable enhancement of the fluorescence. A linear response in the presence of MTP of the relative fluorescent intensity (F - F 0 ) of 1 at 536 nm in the 0-40 μM MTP concentration range was found. A limit of detection (LOD) for the detection of MTP of 26 nM, based on the definition by IUPAC (C DL = 3 Sb/m), was calculated. Probe 1 was applied to monitor and imaging exogenous MTP in live cells and to the detection of MTP in real water samples.

  9. A turn-on supramolecular fluorescent probe for sensing benzimidazole fungicides and its application in living cell imaging

    Science.gov (United States)

    Tang, Qing; Zhang, Jing; Sun, Tao; Wang, Cheng-Hui; Huang, Ying; Zhou, Qingdi; Wei, Gang

    2018-02-01

    A cucurbit[8]uril-based turn-on supramolecular fluorescent probe between cucurbit[8]uril (Q[8]) and pyronine Y (PyY) (designated 2PyY@Q[8]) in acidic aqueous solution showed a remarkable fluorescence 'turn-on' response to benzimidazole fungicides such as thiabendazole, fuberidazole and carbendazim. The 2PyY@Q[8] fluorescent probe can be used to detect benzimidazole fungicides with high sensitivity and selectivity with a detection limit of 10- 8 mol/L. A good linear relationship of emission intensity at 580 nm for benzimidazole fungicides at concentrations of 0.4-5.0 μmol/L was observed. The proposed sensing mechanism was investigated using 1H NMR spectroscopy combined with density functional theory calculations at the B3LYP/6-31G(d) level. The cell imaging study showed that the 2PyY@Q[8] complex could be used to image benzimidazole fungicide in prostate cancer (PC3) cells, which may help to elucidate relevant biological processes at the molecular level.

  10. A dual-emission and large Stokes shift fluorescence probe for real-time discrimination of ROS/RNS and imaging in live cells.

    Science.gov (United States)

    Guo, Ting; Cui, Lei; Shen, Jiaoning; Wang, Rui; Zhu, Weiping; Xu, Yufang; Qian, Xuhong

    2013-03-04

    A novel dual-emission fluorescence probe has been developed for specific and sensitive detection of hypochlorite (ClO(-)). Upon addition of ClO(-), significant changes in fluorescence emission intensity at two discrete wavelengths were observed. Meanwhile OONO(-) led to only a single-channel fluorescence enhancement. This feature makes it a clear advantage in distinguishing ClO(-), RNS from other ROS.

  11. TaqMan MGB probe fluorescence real-time quantitative PCR for rapid detection of Chinese Sacbrood virus.

    Directory of Open Access Journals (Sweden)

    Ma Mingxiao

    Full Text Available Sacbrood virus (SBV is a picorna-like virus that affects honey bees (Apis mellifera and results in the death of the larvae. Several procedures are available to detect Chinese SBV (CSBV in clinical samples, but not to estimate the level of CSBV infection. The aim of this study was develop an assay for rapid detection and quantification of this virus. Primers and probes were designed that were specific for CSBV structural protein genes. A TaqMan minor groove binder (MGB probe-based, fluorescence real-time quantitative PCR was established. The specificity, sensitivity and stability of the assay were assessed; specificity was high and there were no cross-reactivity with healthy larvae or other bee viruses. The assay was applied to detect CSBV in 37 clinical samples and its efficiency was compared with clinical diagnosis, electron microscopy observation, and conventional RT-PCR. The TaqMan MGB-based probe fluorescence real-time quantitative PCR for CSBV was more sensitive than other methods tested. This assay was a reliable, fast, and sensitive method that was used successfully to detect CSBV in clinical samples. The technology can provide a useful tool for rapid detection of CSBV. This study has established a useful protocol for CSBV testing, epidemiological investigation, and development of animal models.

  12. Rational design of reversible fluorescent probes for live-cell imaging and quantification of fast glutathione dynamics

    Science.gov (United States)

    Umezawa, Keitaro; Yoshida, Masafumi; Kamiya, Mako; Yamasoba, Tatsuya; Urano, Yasuteru

    2017-03-01

    Alterations in glutathione (GSH) homeostasis are associated with a variety of diseases and cellular functions, and therefore, real-time live-cell imaging and quantification of GSH dynamics are important for understanding pathophysiological processes. However, existing fluorescent probes are unsuitable for these purposes due to their irreversible fluorogenic mechanisms or slow reaction rates. In this work, we have successfully overcome these problems by establishing a design strategy inspired by Mayr's work on nucleophilic reaction kinetics. The synthesized probes exhibit concentration-dependent, reversible and rapid absorption/fluorescence changes (t1/2 = 620 ms at [GSH] = 1 mM), as well as appropriate Kd values (1-10 mM: within the range of intracellular GSH concentrations). We also developed FRET-based ratiometric probes, and demonstrated that they are useful for quantifying GSH concentration in various cell types and also for real-time live-cell imaging of GSH dynamics with temporal resolution of seconds.

  13. New fluorescent probes of the hydroxyl radical: characterisation and modelization of the reactivity of coumarin derivatives with HO

    International Nuclear Information System (INIS)

    Louit, G.

    2005-10-01

    The hydroxyl radical is involved in a wide range of different fields, from oxidative stress to atmospheric chemistry. In addition to the study of oxidative damage in biological media, the hydroxyl radical detection allows to perform a dosimetry when it is produced by ionising radiation. The aims of this work have been double: - to improve the detection of the hydroxyl radical by the design of new probes - to improve knowledge on the reactive pathways in which the hydroxyl radical is involved. We have studied the coumarin molecule, as well as 6 derivatives that we have synthesised, as fluorescent probes of the hydroxyl radical. Firstly, fluorescence spectroscopy and HPLC chromatography have allowed the evaluation of the sensibility and selectivity of detection of the probes. Consequently to this study, two applications have been developed, concerning the determination of rate constants by competition kinetics and bidimensional dosimetry. Secondly, we have studied the reactivity of the hydroxyl radical through the regioselectivity of its addition on the aromatic cycle. This problem was addressed by the combined use of experimental methods such as time resolved kinetics and HPLC along with interpretation from classical and ab initio modelization. (author)

  14. Hydrogel Micro-/Nanosphere Coated by a Lipid Bilayer: Preparation and Microscopic Probing

    Directory of Open Access Journals (Sweden)

    Sarah Rahni

    2017-02-01

    Full Text Available The result of polymeric nanogels and lipid vesicles interaction—lipobeads—can be considered as multipurpose containers for future therapeutic applications, such as targeted anticancer chemotherapy with superior tumor response and minimum side effects. In this work, micrometer sized lipobeads were synthesized by two methods: (i mixing separately prepared microgels made of poly(N-isopropylacrylamide (PNIPA and phospholipid vesicles of micrometer or nanometer size and (ii polymerization within the lipid vesicles. For the first time, a high vacuum scanning electron microscopy was shown to be suitable for a quick validation of the structural organization of wet lipobeads and their constituents without special sample preparation. In particular, the structural difference of microgels prepared by thermal and UV-polymerization in different solvents was revealed and three types of giant liposomes were recognized under high vacuum in conjunction with their size, composition, and method of preparation. Importantly, the substructure of the hydrogel core and multi- and unilamellar constructions of the peripheral lipid part were explicitly distinguished on the SEM images of lipobeads, justifying the spontaneous formation of a lipid bilayer on the surface of microgels and evidencing an energetically favorable structural organization of the hydrogel/lipid bilayer assembly. This key property can facilitate lipobeads’ preparation and decrease technological expenses on their scaled production. The comparison of the SEM imaging with the scanning confocal and atomic force microscopies data are also presented in the discussion.

  15. Preparation of a Two-Photon Fluorescent Probe for Imaging H2O2 in Lysosomes in Living Cells and Tissues.

    Science.gov (United States)

    Ren, Mingguang; Deng, Beibei; Kong, Xiuqi; Tang, Yonghe; Lin, Weiying

    2017-01-01

    Hydrogen peroxide (H 2 O 2 ) plays important roles in many physiological and pathological processes. At the cellular organelle level, the abnormal concentrations of H 2 O 2 in the lysosomes may cause redox imbalance and the loss of the critical functions of the lysosomes. Herein, we describe the preparation of a potent lysosome-targeted two-photon fluorescent probe (Lyso-HP) for the detection of H 2 O 2 in the lysosomes in the living cells. This unique fluorescent probe can also be employed to effectively detect H 2 O 2 in the living tissues using two-photon fluorescence microscopy.

  16. Subunits of highly Fluorescent Protein R-Phycoerythrin as Probes for Cell Imaging and Single-Molecule Detection

    Energy Technology Data Exchange (ETDEWEB)

    Isailovic, Dragan [Iowa State Univ., Ames, IA (United States)

    2005-01-01

    The purposes of our research were: (1) To characterize subunits of highly fluorescent protein R-Phycoerythrin (R-PE) and check their suitability for single-molecule detection (SMD) and cell imaging, (2) To extend the use of R-PE subunits through design of similar proteins that will be used as probes for microscopy and spectral imaging in a single cell, and (3) To demonstrate a high-throughput spectral imaging method that will rival spectral flow cytometry in the analysis of individual cells. We first demonstrated that R-PE subunits have spectroscopic and structural characteristics that make them suitable for SMD. Subunits were isolated from R-PE by high-performance liquid chromatography (HPLC) and detected as single molecules by total internal reflection fluorescence microscopy (TIRFM). In addition, R-PE subunits and their enzymatic digests were characterized by several separation and detection methods including HPLC, capillary electrophoresis, sodium dodecyl sulfate-polyacrilamide gel electrophoresis (SDS-PAGE) and HPLC-electrospray ionization mass spectrometry (ESI-MS). Favorable absorption and fluorescence of the R-PE subunits and digest peptides originate from phycoerythrobilin (PEB) and phycourobilin (PUB) chromophores that are covalently attached to cysteine residues. High absorption coefficients and strong fluorescence (even under denaturing conditions), broad excitation and emission fluorescence spectra in the visible region of electromagnetic spectrum, and relatively low molecular weights make these molecules suitable for use as fluorescence labels of biomolecules and cells. We further designed fluorescent proteins both in vitro and in vivo (in Escherichia coli) based on the highly specific attachment of PEB chromophore to genetically expressed apo-subunits of R-PE. In one example, apo-alpha and apo-beta R-PE subunits were cloned from red algae Polisiphonia boldii (P. boldii), and expressed in E. coli. Although expressed apo-subunits formed inclusion

  17. A novel “Turn-On” fluorescent probe for F− detection in aqueous solution and its application in live-cell imaging

    International Nuclear Information System (INIS)

    Xu, Jian; Sun, Shaobo; Li, Qian; Yue, Ying; Li, Yingdong; Shao, Shijun

    2014-01-01

    Highlights: • A novel BODIPY-based “Turn-On” fluorescent probe was synthesized. • Highly selective detection of fluoride ions in 100% aqueous solution. • Study of sensing mechanism using density functional theory (DFT) calculations. • Fluorescent bioimaging of F − ion in A549 and ATII cells. - Abstract: A novel probe incorporating quaternized 4-pyridinium group into a BODIPY molecule was synthesized and studied for the selective detection of fluoride ions (F − ) in aqueous solution. The design was based on a fluoride-specific desilylation reaction and the “Turn-On” fluorescent response of probe 1 to F − was ascribed to the inhibition of photoinduced electron transfer (PET) process. The probe displayed many desired properties such as high specificity, appreciable solubility, desirable response time and low toxicity to mammalian cells. There was a good linearity between the fluorescence intensity and the concentrations of F − in the range of 0.1–1 mM with a detection limit of 0.02 mM. The sensing mechanism was confirmed by the NMR, electrospray ionization mass spectrum, optical spectroscopy and the mechanism of “Turn-On” fluorescent response was also determinated by a density functional theory (DFT) calculation using Gaussian 03 program. Moreover, the probe was successfully applied for the fluorescence imaging of F − in human epithelial lung cancer (A549) cells and alveolar type II (ATII) cells under physiological conditions

  18. A novel “Turn-On” fluorescent probe for F{sup −} detection in aqueous solution and its application in live-cell imaging

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Jian [Key Laboratory of Chemistry of Northwestern Plant Resources and Key Laboratory for Natural Medicine of Gansu Province, Lanzhou institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); University of Chinese Academy of Sciences, Beijing 100039 (China); Sun, Shaobo [Institute of Integrated Traditional and Western Medicine, Gansu University of Chinese Medicine, Lanzhou 730000 (China); Li, Qian; Yue, Ying [Key Laboratory of Chemistry of Northwestern Plant Resources and Key Laboratory for Natural Medicine of Gansu Province, Lanzhou institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Li, Yingdong [Institute of Integrated Traditional and Western Medicine, Gansu University of Chinese Medicine, Lanzhou 730000 (China); Shao, Shijun, E-mail: sjshao@licp.cas.cn [Key Laboratory of Chemistry of Northwestern Plant Resources and Key Laboratory for Natural Medicine of Gansu Province, Lanzhou institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China)

    2014-11-07

    Highlights: • A novel BODIPY-based “Turn-On” fluorescent probe was synthesized. • Highly selective detection of fluoride ions in 100% aqueous solution. • Study of sensing mechanism using density functional theory (DFT) calculations. • Fluorescent bioimaging of F{sup −} ion in A549 and ATII cells. - Abstract: A novel probe incorporating quaternized 4-pyridinium group into a BODIPY molecule was synthesized and studied for the selective detection of fluoride ions (F{sup −}) in aqueous solution. The design was based on a fluoride-specific desilylation reaction and the “Turn-On” fluorescent response of probe 1 to F{sup −} was ascribed to the inhibition of photoinduced electron transfer (PET) process. The probe displayed many desired properties such as high specificity, appreciable solubility, desirable response time and low toxicity to mammalian cells. There was a good linearity between the fluorescence intensity and the concentrations of F{sup −} in the range of 0.1–1 mM with a detection limit of 0.02 mM. The sensing mechanism was confirmed by the NMR, electrospray ionization mass spectrum, optical spectroscopy and the mechanism of “Turn-On” fluorescent response was also determinated by a density functional theory (DFT) calculation using Gaussian 03 program. Moreover, the probe was successfully applied for the fluorescence imaging of F{sup −} in human epithelial lung cancer (A549) cells and alveolar type II (ATII) cells under physiological conditions.

  19. Sensitive and rapid detection of endogenous hydrogen sulfide distributing in different mouse viscera via a two-photon fluorescent probe

    International Nuclear Information System (INIS)

    Chen, Qian; Yang, Jinfeng; Li, Yinhui; Zheng, Jing; Yang, Ronghua

    2015-01-01

    Development of efficient methods for detection of endogenous H 2 S in living cells and tissues is of considerable significance for better understanding the biological and pathological functions of H 2 S. Two-photon (TP) fluorescent probes are favorable as powerful molecular tools for studying physiological process due to its non-invasiveness, high spatiotemporal resolution and deep-tissues imaging. Up to date, several TP probes for intracellular H 2 S imaging have been designed, but real-time imaging of endogenous H 2 S-related biological processes in tissues is hampered due to low sensitivity, long response time and interference from other biothiols. To address this issue, we herein report a novel two-photon fluorescent probe (TPP-H 2 S) for highly sensitive and fast monitoring and imaging H 2 S levels in living cells and tissues. In the presence of H 2 S, it exhibits obviously improved sensitivity (LOD: 0.12 μM) and fast response time (about 2 min) compared with the reported two-photon H 2 S probes. With two-photon excitation, TPP-H 2 S displays high signal-to-noise ratio and sensitivity even no interference in cell growth media. As further application, TPP-H 2 S is applied for fast imaging of H 2 S in living cells and different fresh tissues by two-photon confocal microscope. Most importantly we first measured the endogenous H 2 S level in different viscera by vivisection and found that the distribution of endogenous H 2 S mostly in brain, liver and lung. The excellent sensing properties of TPP-H 2 S make it a practically useful tool for further studying biological roles of H 2 S. - Highlights: • This two-photon probe exhibits an improved sensitivity and response time to H 2 S. • This probe shows excellent membrane permeability and fast visualization of H 2 S in living cells and tissues. • This probe is successfully applied to measure the endogenously produced H 2 S levels in different viscera of mouse.

  20. Carbon dots as fluorescent probes for "off-on" detection of Cu2+ and L-cysteine in aqueous solution.

    Science.gov (United States)

    Zong, Jie; Yang, Xiaoling; Trinchi, Adrian; Hardin, Simon; Cole, Ivan; Zhu, Yihua; Li, Chunzhong; Muster, Tim; Wei, Gang

    2014-01-15

    Copper ion (Cu(2+)) and L-cysteine (L-Cys) detection is critically important since an abnormal level of Cu(2+) or L-Cys is an indicator for many diseases. In this paper, we demonstrate an "off-on" approach for highly sensitive and selective detection of Cu(2+) and L-Cys using carbon dots (CDs) as fluorescent probes. CDs were prepared by using mesoporous silica (MS) spheres as nanoreactors. The binding ability of CDs towards metal ions was examined by comparing the fluorescence intensities of CDs before and after the addition of the metal ions. The addition of Cu(2+) cations leads to their absorption on the surface of CDs and the significant fluorescence quench of CDs (turn-off). The resulting in CDs-Cu(2+) system was found to be sensitive to L-Cys. The addition of L-Cys not only serves to shelter the CDs effectively from being quenched, but also to reverse the quenching and restore the fluorescence (turn-on) due to its ability to remove Cu(2+) from the surface of CDs. This method is facile, rapid, low cost, and environment-friendly. A detection limit as low as 2.3×10(-8) M for Cu(2+) and 3.4×10(-10) M for L-Cys is obtained, which is promising for biological applications. © 2013 Elsevier B.V. All rights reserved.

  1. Spectral-fluorescent study of the interaction of the polymethine dye probe Cyan 2 with chondroitin-4-sulfate

    Science.gov (United States)

    Tatikolov, Alexander S.; Akimkin, Timofey M.; Panova, Ina G.; Yarmoluk, Sergiy M.

    2017-04-01

    The noncovalent interaction of the polymethine dye probe 3,3‧,9-trimethylthiacarbocyanine iodide (Cyan 2) with chondroitin-4-sulfate (C4S) in buffer solutions with different pH and in water in the absence of buffers has been studied by spectral-fluorescent methods. It has been shown that in all media studied, at relatively high concentrations, the dye is bound to C4S mainly as a monomer, which is accompanied by a steep rise of fluorescence (the intermediate formation of dye aggregates on the biopolymer is also observed). From the dependence of the fluorescence quantum yield on the concentration of C4S, the parameters of binding of the dye monomer to C4S were obtained: the effective binding constant K, the number of the monomeric C4S units n per one dye monomer bound to C4S, and the fluorescence quantum yield of the bound dye monomer Φfb. The dependence of Φfb (and K) on pH of the medium is not monotonic: it has a minimum in the region of neutral pH and a growth in the regions of acid and basic pH. This can be explained by changing the charge of a C4S macromolecule as a function of pH and related conformational alterations in the biopolymer, which can affect the rigidity of a dye molecule and the energy of its interaction with the biopolymer.

  2. Ultrafast Dynamics of Sb-Corroles: A Combined Vis-Pump Supercontinuum Probe and Broadband Fluorescence Up-Conversion Study

    Directory of Open Access Journals (Sweden)

    Clark Zahn

    2017-07-01

    Full Text Available Corroles are a developing class of tetrapyrrole-based molecules with significant chemical potential and relatively unexplored photophysical properties. We combined femtosecond broadband fluorescence up-conversion and fs broadband Vis-pump Vis-probe spectroscopy to comprehensively characterize the photoreaction of 5,10,15-tris-pentafluorophenyl-corrolato-antimony(V-trans-difluoride (Sb-tpfc-F2. Upon fs Soret band excitation at ~400 nm, the energy relaxed almost completely to Q band electronic excited states with a time constant of 500 ± 100 fs; this is evident from the decay of Soret band fluorescence at around 430 nm and the rise time of Q band fluorescence, as well as from Q band stimulated emission signals at 600 and 650 nm with the same time constant. Relaxation processes on a time scale of 10 and 20 ps were observed in the fluorescence and absorption signals. Triplet formation showed a time constant of 400 ps, with an intersystem crossing yield from the Q band to the triplet manifold of between 95% and 99%. This efficient triplet formation is due to the spin-orbit coupling of the antimony ion.

  3. Spectrofluorimetric determination of trace amount of coenzyme II using ciprofloxacin-terbium complex as a fluorescent probe

    International Nuclear Information System (INIS)

    Bian Weiwei; Wang Yusheng; Zhu Xiaojing; Jiang Chongqiu

    2006-01-01

    A new spectrofluorimetric method was developed for the determination of trace amount of nicotinamide adenine dinucleotide phosphate (NADP). Using terbium ion (Tb 3+ )-ciprofloxacin (CIP) complex as a fluorescent probe, in the buffer solution of pH=9.00, NADP can remarkably enhance the fluorescence intensity of the Tb 3+ -CIP complex at λ=545nm and the enhanced fluorescence intensity of Tb 3+ ion is in proportion to the concentration of NADP. Optimum conditions for the determination of NADP were also investigated. The dynamic range for the determination of NADP is 4.9x10 -7 -3.7x10 -6 molL -1 with detection limit of 1.3x10 -7 molL -1 . This method is simple, practical and relatively free interference from coexisting substances and can be successfully applied to determination of NADP in synthetic water samples. Moreover, the enhancement mechanisms of the fluorescence intensity in the Tb 3+ -CIP system and the Tb 3+ -CIP-NADP system have been also discussed

  4. Novel Fluorescent Microemulsion: Probing Properties, Investigating Mechanism, and Unveiling Potential Application.

    Science.gov (United States)

    Hou, Mengna; Dang, Leping; Liu, Tiankuo; Guo, Yun; Wang, Zhanzhong

    2017-08-09

    Nanoscale microemulsions have been utilized as delivery carriers for nutraceuticals and active biological drugs. Herein, we designed and synthesized a novel oil in water (O/W) fluorescent microemulsion based on isoamyl acetate, polyoxyethylene castor oil EL (CrEL), and water. The microemulsion emitted bright blue fluorescence, thus exhibiting its potential for active drug detection with label-free strategy. The microemulsion exhibited excitation-dependent emission and distinct red shift with longer excitation wavelengths. Lifetime and quantum yield of fluorescent microemulsion were 2.831 ns and 5.0%, respectively. An excellent fluorescent stability of the microemulsion was confirmed by altering pH, ionic strength, temperature, and time. Moreover, we proposed a probable mechanism of fluorochromic phenomenon, in connection with the aromatic ring structure of polyoxyethylene ether substituent in CrEL. Based on our findings, we concluded that this new fluorescent microemulsion is a promising drug carrier that can facilitate active drug detection with a label-free strategy. Although further research is required to understand the exact mechanism behind its fluorescence property, this work provided valuable guidance to develop new biosensors based on fluorescent microemulsion.

  5. Correlative Light-Electron Microscopy of Lipid-Encapsulated Fluorescent Nanodiamonds for Nanometric Localization of Cell Surface Antigens.

    Science.gov (United States)

    Hsieh, Feng-Jen; Chen, Yen-Wei; Huang, Yao-Kuan; Lee, Hsien-Ming; Lin, Chun-Hung; Chang, Huan-Cheng

    2018-02-06

    Containing an ensemble of nitrogen-vacancy centers in crystal matrices, fluorescent nanodiamonds (FNDs) are a new type of photostable markers that have found wide applications in light microscopy. The nanomaterial also has a dense carbon core, making it visible to electron microscopy. Here, we show that FNDs encapsulated in biotinylated lipids (bLs) are useful for subdiffraction imaging of antigens on cell surface with correlative light-electron microscopy (CLEM). The lipid encapsulation enables not only good dispersion of the particles in biological buffers but also high specific labeling of live cells. By employing the bL-encapsulated FNDs to target CD44 on HeLa cell surface through biotin-mediated immunostaining, we obtained the spatial distribution of these antigens by CLEM with a localization accuracy of ∼50 nm in routine operations. A comparative study with dual-color imaging, in which CD44 was labeled with FND and MICA/MICB was labeled with Alexa Fluor 488, demonstrated the superior performance of FNDs as fluorescent fiducial markers for CLEM of cell surface antigens.

  6. An Optimized Set of Fluorescence In Situ Hybridization Probes for Detection of Pancreatobiliary Tract Cancer in Cytology Brush Samples.

    Science.gov (United States)

    Barr Fritcher, Emily G; Voss, Jesse S; Brankley, Shannon M; Campion, Michael B; Jenkins, Sarah M; Keeney, Matthew E; Henry, Michael R; Kerr, Sarah M; Chaiteerakij, Roongruedee; Pestova, Ekaterina V; Clayton, Amy C; Zhang, Jun; Roberts, Lewis R; Gores, Gregory J; Halling, Kevin C; Kipp, Benjamin R

    2015-12-01

    Pancreatobiliary cancer is detected by fluorescence in situ hybridization (FISH) of pancreatobiliary brush samples with UroVysion probes, originally designed to detect bladder cancer. We designed a set of new probes to detect pancreatobiliary cancer and compared its performance with that of UroVysion and routine cytology analysis. We tested a set of FISH probes on tumor tissues (cholangiocarcinoma or pancreatic carcinoma) and non-tumor tissues from 29 patients. We identified 4 probes that had high specificity for tumor vs non-tumor tissues; we called this set of probes pancreatobiliary FISH. We performed a retrospective analysis of brush samples from 272 patients who underwent endoscopic retrograde cholangiopancreatography for evaluation of malignancy at the Mayo Clinic; results were available from routine cytology and FISH with UroVysion probes. Archived residual specimens were retrieved and used to evaluate the pancreatobiliary FISH probes. Cutoff values for FISH with the pancreatobiliary probes were determined using 89 samples and validated in the remaining 183 samples. Clinical and pathologic evidence of malignancy in the pancreatobiliary tract within 2 years of brush sample collection was used as the standard; samples from patients without malignancies were used as negative controls. The validation cohort included 85 patients with malignancies (46.4%) and 114 patients with primary sclerosing cholangitis (62.3%). Samples containing cells above the cutoff for polysomy (copy number gain of ≥2 probes) were classified as positive in FISH with the UroVysion and pancreatobiliary probes. Multivariable logistic regression was used to estimate associations between clinical and pathology findings and results from FISH. The combination of FISH probes 1q21, 7p12, 8q24, and 9p21 identified cancer cells with 93% sensitivity and 100% specificity in pancreatobiliary tissue samples and were therefore included in the pancreatobiliary probe set. In the validation cohort of

  7. Towards Fluorescence In Vivo Hybridization (FIVH) Detection of H. pylori in Gastric Mucosa Using Advanced LNA Probes

    DEFF Research Database (Denmark)

    Fontenete, Sílvia; Leite, Marina; Guimarães, Nuno

    2015-01-01

    acid (LNA)/ 2' O-methyl RNA (2'OMe) probe using standard phosphoramidite chemistry and FISH hybridization was then successfully performed both on adhered and suspended bacteria at 37°C. In this work we simplified, shortened and adapted FISH to work at gastric pH values, meaning that the hybridization...... step now takes only 30 minutes and, in addition to the buffer, uses only urea and probe at non-toxic concentrations. Importantly, the sensitivity and specificity of the FISH method was maintained in the range of conditions tested, even at low stringency conditions (e.g., low pH). In conclusion......In recent years, there have been several attempts to improve the diagnosis of infection caused by Helicobacter pylori. Fluorescence in situ hybridization (FISH) is a commonly used technique to detect H. pylori infection but it requires biopsies from the stomach. Thus, the development of an in vivo...

  8. A fluorescent probe distinguishes between inhibition of early and late steps of lipopolysaccharide biogenesis in whole cells

    Science.gov (United States)

    Moison, Eileen; Xie, Ran; Zhang, Ge; Lebar, Matthew D.; Meredith, Timothy C.; Kahne, Daniel

    2017-01-01

    Lipopolysaccharide (LPS) biogenesis in Gram-negative organisms involves its biosynthesis in the cytoplasm and subsequent transport across three cellular compartments to the cell surface. We developed a fluorescent probe that allows us to determine the spatial distribution of LPS in whole cells. We show that polymyxin B nonapeptide (PMBN) containing a dansyl fluorophore specifically binds to LPS in membranes. We show that this probe detects decreases in LPS levels on the cell surface when LPS biosynthesis is inhibited at an early step. We also can detect accumulation of LPS in particular subcellular locations when LPS assembly is blocked during transport, allowing us to differentiate inhibitors targeting early and late stages of LPS biogenesis. PMID:28248483

  9. Y2O3: Eu,Zn nanocrystals as a fluorescent probe for the detection of biotin

    International Nuclear Information System (INIS)

    Ganjkhanlou, Y.; Hosseinnia, A.; Kazemzad, M.; Khanlarkhani, A.; Moghaddam, A.B.

    2012-01-01

    We report on the application of nanocrystals (NCs) of the type Y 2 O 3 : Eu,Zn as a probe for the fluorescent detection of biotin in aqueous solution. The NCs were dispersed in water in the presence of various surface modifiers including mercaptoethanol (ME), monoethanolamine and ethylene glycol. Both the absorbance of surfactant and the stability of the suspensions were investigated in order to optimize the experimental conditions. ME is found to be the most suitable surfactant for stabilization of the suspended NCs. Their photoluminescence intensity is found to be quenched by biotin. The Stern-Volmer constant for the quenching process is 7.6 x 10 3 M -1 . This NC probe can be applied to the detection of biotin in the 1-60 μM concentration range with detection limit of 1.89 μM. The possible mechanisms of quenching also are discussed. (author)

  10. Texture of lipid bilayer domains

    DEFF Research Database (Denmark)

    Jensen, Uffe Bernchou; Brewer, Jonathan R.; Midtiby, Henrik Skov

    2009-01-01

    We investigate the texture of gel (g) domains in binary lipid membranes composed of the phospholipids DPPC and DOPC. Lateral organization of lipid bilayer membranes is a topic of fundamental and biological importance. Whereas questions related to size and composition of fluid membrane domain...... are well studied, the possibility of texture in gel domains has so far not been examined. When using polarized light for two-photon excitation of the fluorescent lipid probe Laurdan, the emission intensity is highly sensitive to the angle between the polarization and the tilt orientation of lipid acyl...... chains. By imaging the intensity variations as a function of the polarization angle, we map the lateral variations of the lipid tilt within domains. Results reveal that gel domains are composed of subdomains with different lipid tilt directions. We have applied a Fourier decomposition method...

  11. A rhodamine-labeled citalopram analogue as a high-affinity fluorescent probe for the serotonin transporter

    DEFF Research Database (Denmark)

    Zhang, Peng; Jørgensen, Trine Nygaard; Løland, Claus Juul

    2013-01-01

    A novel fluorescent ligand was synthesized as a high-affinity, high specificity probe for visualizing the serotonin transporter (SERT). The rhodamine fluorophore was extended from an aniline substitution on the 5-position of the dihydroisobenzofuran ring of citalopram (2, 1-(3-(dimethylamino......)propyl)-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile), using an ethylamino linker. The resulting rhodamine-labeled ligand 8 inhibited [3H]5-HT uptake in COS-7 cells (Ki = 225 nM) with similar potency to the tropane-based JHC 1-064 (1), but with higher specificity towards the SERT relative...

  12. Anti-VEGF antibody conjugated CdHgTe quantum dots as a fluorescent probe for imaging in living mouse

    Energy Technology Data Exchange (ETDEWEB)

    Pang, Lili; Cui, Hongjing [Department of Analytical Chemistry, China Pharmaceutical University, Nanjing (China); Liu, Yu [Department of Biochemistry, School of Life Science and Technology, China Pharmaceutical University, Nanjing (China); Zhong, Wenying, E-mail: wyzhong@cpu.edu.cn [Department of Analytical Chemistry, China Pharmaceutical University, Nanjing (China); Key laboratory of Biomedical Functional Materials, China Pharmaceutical University, Nanjing (China)

    2016-05-15

    The dual-function anti-VEGF antibody conjugated CdHgTe quantum dots with good targeting property was successfully prepared. In this system, anti-VEGF antibody is not only a target agent but also a therapeutic drug. The anti-VEGF antibody conjugated near-infrared quantum dots can achieve the purposes of detection and treatment at the same time. As-prepared dual-function fluorescent probe in this work has been successfully applied for in vivo and in vitro imaging, which is promising in rapid tumor detection.

  13. Hydration of poly( N-isopropylacrylamide) brushes on micro-silica beads measured by a fluorescent probe

    Science.gov (United States)

    Hattori, Yusuke; Nagase, Kenichi; Kobayashi, Jun; Kikuchi, Akihiko; Akiyama, Yoshikatsu; Kanazawa, Hideko; Okano, Teruo

    2010-05-01

    Hydration of poly( N-isopropylacrylamide) brushes on silica micro-beads was investigated using a fluorescent probe method. The free ends, the bottom, and the random of brushes were labeled with dansyl group. The emission spectra at the thin brushes were reduced with increasing temperature regardless of their labeling locations. At the free ends of thick brushes, the emission intensity was enhanced at 500 nm and reduced at 455 nm by heating, which was corresponding to the local micro-environmental change around the free ends. The spectral shift was speculated to be due to the enhancement of the flexibility and the hydration of thick brushes.

  14. Hydrophobic analogues of rhodamine B and rhodamine 101: potent fluorescent probes of mitochondria in living C. elegans

    Directory of Open Access Journals (Sweden)

    Laurie F. Mottram

    2012-12-01

    Full Text Available Mitochondria undergo dynamic fusion and fission events that affect the structure and function of these critical energy-producing cellular organelles. Defects in these dynamic processes have been implicated in a wide range of human diseases including ischemia, neurodegeneration, metabolic disease, and cancer. To provide new tools for imaging of mitochondria in vivo, we synthesized novel hydrophobic analogues of the red fluorescent dyes rhodamine B and rhodamine 101 that replace the carboxylate with a methyl group. Compared to the parent compounds, methyl analogues termed HRB and HR101 exhibit slightly red-shifted absorbance and emission spectra (5–9 nm, modest reductions in molar extinction coefficent and quantum yield, and enhanced partitioning into octanol compared with aqueous buffer of 10-fold or more. Comparison of living C. elegans (nematode roundworm animals treated with the classic fluorescent mitochondrial stains rhodamine 123, rhodamine 6G, and rhodamine B, as well as the structurally related fluorophores rhodamine 101, and basic violet 11, revealed that HRB and HR101 are the most potent mitochondrial probes, enabling imaging of mitochondrial motility, fusion, and fission in the germline and other tissues by confocal laser scanning microscopy after treatment for 2 h at concentrations as low as 100 picomolar. Because transgenes are poorly expressed in the germline of these animals, these small molecules represent superior tools for labeling dynamic mitochondria in this tissue compared with the expression of mitochondria-targeted fluorescent proteins. The high bioavailabilty of these novel fluorescent probes may facilitate the identification of agents and factors that affect diverse aspects of mitochondrial biology in vivo.

  15. A Thiazole Coumarin (TC) Turn-On Fluorescence Probe for AT-Base Pair Detection and Multipurpose Applications in Different Biological Systems

    Science.gov (United States)

    Narayanaswamy, Nagarjun; Kumar, Manoj; Das, Sadhan; Sharma, Rahul; Samanta, Pralok K.; Pati, Swapan K.; Dhar, Suman K.; Kundu, Tapas K.; Govindaraju, T.

    2014-01-01

    Sequence-specific recognition of DNA by small turn-on fluorescence probes is a promising tool for bioimaging, bioanalytical and biomedical applications. Here, the authors report a novel cell-permeable and red fluorescent hemicyanine-based thiazole coumarin (TC) probe for DNA recognition, nuclear staining and cell cycle analysis. TC exhibited strong fluorescence enhancement in the presence of DNA containing AT-base pairs, but did not fluoresce with GC sequences, single-stranded DNA, RNA and proteins. The fluorescence staining of HeLa S3 and HEK 293 cells by TC followed by DNase and RNase digestion studies depicted the selective staining of DNA in the nucleus over the cytoplasmic region. Fluorescence-activated cell sorting (FACS) analysis by flow cytometry demonstrated the potential application of TC in cell cycle analysis in HEK 293 cells. Metaphase chromosome and malaria parasite DNA imaging studies further confirmed the in vivo diagnostic and therapeutic applications of probe TC. Probe TC may find multiple applications in fluorescence spectroscopy, diagnostics, bioimaging and molecular and cell biology. PMID:25252596

  16. New Concepts of Fluorescent Probes for Specific Detection of DNA Sequences: Bis-Modified Oligonucleotides in Excimer and Exciplex Detection

    Directory of Open Access Journals (Sweden)

    Gbaj A

    2009-01-01

    Full Text Available The detection of single base mismatches in DNA is important for diagnostics, treatment of genetic diseases, and identification of single nucleotide polymorphisms. Highly sensitive, specific assays are needed to investigate genetic samples from patients. The use of a simple fluorescent nucleoside analogue in detection of DNA sequence and point mutations by hybridisation in solution is described in this study. The 5’-bispyrene and 3’-naphthalene oligonucleotide probes form an exciplex on hybridisation to target in water and the 5’-bispyrene oligonucleotide alone is an adequate probe to determine concentration of target present. It was also indicated that this system has a potential to identify mismatches and insertions. The aim of this work was to investigate experimental structures and conditions that permit strong exciplex emission for nucleic acid detectors, and show how such exciplexes can register the presence of mismatches as required in SNP analysis. This study revealed that the hybridisation of 5'-bispyrenyl fluorophore to a DNA target results in formation of a fluorescent probe with high signal intensity change and specificity for detecting a complementary target in a homogeneous system. Detection of SNP mutations using this split-probe system is a highly specific, simple, and accessible method to meet the rigorous requirements of pharmacogenomic studies. Thus, it is possible for the system to act as SNP detectors and it shows promise for future applications in genetic testing.

  17. Fluorescence studies on gamma irradiated egg lecithin liposomal membrane

    International Nuclear Information System (INIS)

    Pandey, B.N.; Mishra, K.P.

    1998-01-01

    Alterations in structure and organization of sonicated EYL liposomal vesicular membrane after irradiation was investigated by DPH fluorescence probe which is a well known reporter for the environment of hydrophobic interior of membrane. Results of present study have demonstrated that loss of DPH fluorescence in liposomal membrane is linked to free radical mediated structural alterations possibly rigidization in the lipid bilayer

  18. Fluorescent probe for turn-on sensing of L-cysteine by ensemble of AuNCs and polymer protected AuNPs.

    Science.gov (United States)

    Xu, Xiaozhe; Qiao, Juan; Li, Nan; Qi, Li; Zhang, Shufeng

    2015-06-16

    A new fluorescent probe based on ensemble of gold nanoclusters (AuNCs) and polymer protected gold nanoparticles (AuNPs) for turn-on sensing of L-cysteine was designed and prepared. The AuNCs were protected by bovine serum albumin and had strong fluorescence. The polymer protected AuNPs were synthesized by a facile in situ strategy at room temperature and could quench the fluorescence of AuNCs due to the Förster resonance energy transfer. Interestingly, it has been observed that the quenched fluorescence of AuNCs was recovered by L-cysteine, which could induce the aggregation of polymer protected AuNPs by sulfur group. Then the prepared fluorescent probe was successfully used for determination of L-Cys in human urines, which would have an evolving aspect and promote the subsequent exploration. Copyright © 2015 Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

    Yerramilli, V Siddartha; Kim, Kyung Hyuk

    2018-03-16

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

  20. A lysosome-targetable turn-on fluorescent probe for the detection of thiols in living cells based on a 1,8-naphthalimide derivative

    Science.gov (United States)

    Liang, Beibei; Wang, Baiyan; Ma, Qiujuan; Xie, Caixia; Li, Xian; Wang, Suiping

    2018-03-01

    Biological thiols, like cysteine (Cys), homocysteine (Hcy) and glutathione (GSH), play crucial roles in biological systems and in lysosomal processes. Highly selective probes for detecting biological thiols in lysomes of living cells are rare. In this work, a lysosome-targetable turn-on fluorescent probe for the detection of thiols in living cells was designed and synthesized based on a 1,8-naphthalimide derivative. The probe has a 4-(2-aminoethyl)morpholine unit as a lysosome-targetable group and an acrylate group as the thiol recognition unit as well as a fluorescence quencher. In the absence of biothiols, the probe displayed weak fluorescence due to the photoinduced electron transfer (PET) process. Upon the addition of biothiols, the probe exhibited an enhanced fluorescence emission centered at 550 nm due to cleavage of the acrylate moiety. The probe had high selectivity toward biothiols. Moreover, the probe features fast response time, excitation in the visible region and ability of working in a wide pH range. The linear response range covers a concentration range of Cys from 1.5 × 10- 7 to 1.0 × 10- 5 mol·L- 1 and the detection limit is 6.9 × 10- 8 mol·L- 1 for Cys. The probe has been successfully applied to the confocal imaging of biothiols in lysosomes of A549 cells with low cell toxicity. Furthermore, the method was successfully applied to the determination of thiols in a complex multicomponent mixture such as human serum, which suggests our proposed method has great potential for diagnostic purposes. All of such good properties prove it can be used to monitor biothiols in lysosomes of living cells and to be a good fluorescent probe for the selective detection of thiols.

  1. Irregular bilayer structure in vesicles prepared from Halobacterium cutirubrum lipids

    Science.gov (United States)

    Lanyi, J. K.

    1974-01-01

    Fluorescent probes were used to study the structure of the cell envelope of Halobacterium cutirubrum, and, in particular, to explore the effect of the heterogeneity of the lipids in this organism on the structure of the bilayers. The fluorescence polarization of perylene was followed in vesicles of unfractionated lipids and polar lipids as a function of temperature in 3.4 M solutions of NaCl, NaNO3, and KSCN, and it was found that vesicles of unfractionated lipids were more perturbed by chaotropic agents than polar lipids. The dependence of the relaxation times of perylene on temperature was studied in cell envelopes and in vesicles prepared from polar lipids, unfractionated lipids, and mixtures of polar and neutral lipids.

  2. A dihydrazone based fluorescent probe for selective determination of Al{sup 3+} ions

    Energy Technology Data Exchange (ETDEWEB)

    Pratap Singh, Divya; Singh, Vinod P., E-mail: singvp@yahoo.co.in

    2014-11-15

    A highly selective fluorescent sensor N,N′-bis((2-hydroxynaphthalen-1-yl)methylene) oxalohydrazide (H{sub 2}ohn) for the determination of Al{sup 3+} ions was synthesized and characterized by different physico-chemical and spectroscopic techniques. The single crystal structure of H{sub 2}ohn receptor has also been reported. The H{sub 2}ohn shows an enhanced fluorescence in the presence of Al{sup 3+} ions in ethanol–water (2:3 v/v) solution. Other cations viz. Na{sup +}, K{sup +}, Mg{sup 2+}, Ca{sup 2+}, Mn{sup 2+}, Fe{sup 3+}, Co{sup 2+}, Ni{sup 2+}, Cu{sup 2+}, Zn{sup 2+}, Pb{sup 2+}, Cd{sup 2+} and Hg{sup 2+} show no appreciable change in fluorescence intensity. The binding mode of H{sub 2}ohn receptor with Al{sup 3+} was studied by UV–visible, fluorescence and {sup 1}H NMR titrations. The receptor acts as a dibasic hexadentate ligand and interacts with two Al{sup 3+} ions with a high binding constant K{sub B}=2.62×10{sup 11} M{sup −2}. The lowest detection limit for Al{sup 3+} complex of H{sub 2}ohn was determined to be 8.56×10{sup −10} M. The structures of H{sub 2}ohn and its Al(III) complex were also optimized by DFT calculations. - Highlights: • A dihydrazone based fluorescent sensor is synthesized. • Characterized by IR, NMR, UV−visible and mass spectral analysis. • The receptor serves as a selective fluorescent sensor for Al{sup 3+} over other cations. • The sensing property is monitored by UV−visible, fluorescence and NMR spectroscopy. • A high binding constant of the receptor with Al{sup 3+} is reported here.

  3. A light-up fluorescent probe for citrate detection based on bispyridinum amides with aggregation-induced emission feature.

    Science.gov (United States)

    Liu, Chenchen; Hang, Yandi; Jiang, Tao; Yang, Ji; Zhang, Xiao; Hua, Jianli

    2018-02-01

    Citrate is an important intermediate in the citric acid cycle, a vital metabolic pathway for animals, plants and bacteria. It is of great significance to detect its levels in human beings because several diseases may cause the abnormal of citrate. In this paper, a new turn-on fluorescent sensor (TPE-Py) using the classic tetraphenylethylene (TPE) as the aggregation-induced emission (AIE) fluorophore and bipyridinium-based amides as the recognition receptor has been synthesized for the detection of citrate. The probe exhibits good selectivity and sensitivity to citrate with a relatively low detection limit (1.0 × 10 -7 M). The enhancement of the fluorescence is relevant with the AIE property based on the complexation of TPE-Py with citrate caused by the hydrogen bonding and electrostatic interactions between the bipyridinium diamides and citrate, which has been proved by 1 H NMR and mass spectra titration, scanning electronic microscope and dynamic light scattering analyses. More importantly, the quantification of citrate in artificial urine may develop TPE-Py fluorometric probe for the citrate detection in real biosystems. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Fluorescent pH-Sensing Probe Based on Biorefinery Wood Lignosulfonate and Its Application in Human Cancer Cell Bioimaging.

    Science.gov (United States)

    Xue, Yuyuan; Liang, Wanshan; Li, Yuan; Wu, Ying; Peng, Xinwen; Qiu, Xueqing; Liu, Jinbin; Sun, Runcang

    2016-12-28

    A water-soluble, ratiometric fluorescent pH probe, L-SRhB, was synthesized via grafting spirolactam Rhodamine B (SRhB) to lignosulfonate (LS). As the ring-opening product of L-SRhB, FL-SRhB was also prepared. The pH-response experiment indicated that L-SRhB showed a rapid response to pH changes from 4.60 to 6.20 with a pK a of 5.35, which indicated that L-SRhB has the potential for pH detection of acidic organelle. In addition, the two probes were internalized successfully by living cells through the endocytosis pathway and could distinguish normal cells from cancer cells by different cell staining rates. In addition, L-SRhB showed obvious cytotoxicity to cancer cells, whereas it was nontoxic to normal cells in the same condition. L-SRhB might have potential in cancer therapy. L-SRhB might be a promising ratiometric fluorescent pH sensor and bioimaging dye for the recognition of cancer cells. The results also provided a new perspective to the high-value utilization of lignin.

  5. Amino acid detection using fluoroquinolone–Cu2+ complex as a switch-on fluorescent probe by competitive complexation without derivatization

    International Nuclear Information System (INIS)

    Farokhcheh, Alireza; Alizadeh, Naader

    2014-01-01

    In this work, we describe the use of fluoroquinolone–Cu 2+ complex as a competitive switch-on fluorescence probe for amino acid determination without derivatization. The fluorescence intensity of this probe, which has been reduced due to effective quenching by Cu 2+ ion, increases drastically by an addition of amino acid (glycine, phenylalanine, sarcosine, aspargine, alanine, proline, arginine, aspartic acid, glutamic acid, lysine, leucine and isoleucine). The overall stability constants of Cu 2+ ion complexes with amino acids were determined by fluorometric titration of fluoroquinolone-Cu 2+ complex with the amino acid solution. Furthermore, the probe shows high calibration sensitivity toward aspartic acid. The fluorescence signal depends linearly on the amino acid concentration within the range of concentration from 1.2×10 −7 to 1.1×10 −5 mol L −1 for aspartic acid. The detection limit was found 2.7×10 −8 mol L −1 with the relative standard deviation (RSD%) about 2.1% (five replicate). -- Highlights: • Amino acids are detected by using fluoroquinolone–Cu 2+ complex as fluorescent probe. • Amino acids were detected based on a competitive complexation reaction. • Probe has been able to recognize amino acids through switch-on fluorescence behavior. • Ultra-trace level of aspartic and glutamic acid is determined without derivatization

  6. Amino acid detection using fluoroquinolone–Cu{sup 2+} complex as a switch-on fluorescent probe by competitive complexation without derivatization

    Energy Technology Data Exchange (ETDEWEB)

    Farokhcheh, Alireza; Alizadeh, Naader, E-mail: alizaden@modares.ac.ir

    2014-01-15

    In this work, we describe the use of fluoroquinolone–Cu{sup 2+} complex as a competitive switch-on fluorescence probe for amino acid determination without derivatization. The fluorescence intensity of this probe, which has been reduced due to effective quenching by Cu{sup 2+} ion, increases drastically by an addition of amino acid (glycine, phenylalanine, sarcosine, aspargine, alanine, proline, arginine, aspartic acid, glutamic acid, lysine, leucine and isoleucine). The overall stability constants of Cu{sup 2+} ion complexes with amino acids were determined by fluorometric titration of fluoroquinolone-Cu{sup 2+} complex with the amino acid solution. Furthermore, the probe shows high calibration sensitivity toward aspartic acid. The fluorescence signal depends linearly on the amino acid concentration within the range of concentration from 1.2×10{sup −7} to 1.1×10{sup −5} mol L{sup −1} for aspartic acid. The detection limit was found 2.7×10{sup −8} mol L{sup −1} with the relative standard deviation (RSD%) about 2.1% (five replicate). -- Highlights: • Amino acids are detected by using fluoroquinolone–Cu{sup 2+} complex as fluorescent probe. • Amino acids were detected based on a competitive complexation reaction. • Probe has been able to recognize amino acids through switch-on fluorescence behavior. • Ultra-trace level of aspartic and glutamic acid is determined without derivatization.

  7. Dansyl-Galactoside, a Fluorescent Probe of Active Transport in Bacterial Membrane Vesicles*

    Science.gov (United States)

    Reeves, John P.; Shechter, Emanuel; Weil, Rudolf; Kaback, H. R.

    1973-01-01

    A fluorescent galactoside, 2-(N-dansyl)-aminoethyl β-D-thiogalactoside (dansyl-galactoside), competitively inhibits lactose transport by membrane vesicles of Escherichia coli, but is not actively transported. An increase in dansyl-galactoside fluorescence is observed upon addition of D-lactate. The fluorescence increase is not observed in membrane vesicles lacking the β-galactoside transport system, and is blocked or rapidly reversed by addition of β-galactosides, sulfhydryl reagents, inhibitors of D-lactate oxidation, or uncoupling agents. The fluorescence increase exhibits an emission maximum at 500 nm and excitation maxima at 345 nm and at 292 nm. The latter excitation maximum is absent unless D-lactate is added, indicating that the bound dansyl-galactoside molecules are excited by energy transfer from the membrane proteins. Titration of vesicles with dansyl-galactoside in the presence of D-lactate demonstrates that the β-galactoside carrier protein represents about 3.3% of the total membrane protein. The data indicate that D-lactate oxidation leads to binding of the fluorescent galactoside to the β-galactoside carrier protein in such a manner that the dansyl group is transferred to a hydrophobic environment within the membrane. PMID:4583021

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

    Science.gov (United States)

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

    2000-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Fei-Fei An

    2017-06-01

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

  10. Turn-on Fluorescent Probe for Exogenous and Endogenous Imaging of Hypochlorous Acid in Living Cells and Quantitative Application in Flow Cytometry.

    Science.gov (United States)

    Zhan, Zixuan; Liu, Rui; Chai, Li; Li, Qiuyan; Zhang, Kexin; Lv, Yi

    2017-09-05

    Hypochlorous acid (HClO) acts as a dominant microbicidal mediator in the natural immune system, and the excess production of hypochlorites is related to a series of diseases. Thus, it is vitally important and necessary to develop a highly sensitive and selective method for HClO detection in living systems, and most of fluorescent probes are mainly focused on cells imaging. Besides, accurate HClO quantitative information about individual cells in a large cell population is extremely important for understanding inflammation and cellular apoptosis as well. In our work, a turn-on fluorescent probe has been synthesized, which can selectively and sensitively detect HClO with fast response time. The probe is almost nonfluorescent possibly due to both the spirolactam form of fluorescein and unbridged C═N bonds which can undergo a nonradiative decay process in the excited state. Upon the addition of ClO - , the probe was oxidized to ring-opened fluorescent form and the fluorescence intensity was greatly enhanced. In live cell experiments, the probe was successfully applied to image exogenous ClO - in HeLa cells and endogenous HClO in RAW 264.7 macrophage cells. In particular, the quantitative information on exogenous and endogenous HClO can also be acquired in flow cytometry. Therefore, the probe not only can image exogenous and endogenous HClO but also provides a new and promising platform to quantitatively detect HClO in flow cytometry.

  11. A novel pH-sensitive polymeric fluorescent probe: Synthesis, characterization and optical properties

    Energy Technology Data Exchange (ETDEWEB)

    Chen Dongyun; Xu Qingfeng; Xia Xuewei; Ge Jianfeng [Key Laboratory of Organic Synthesis of Jiangsu Province, College of Chemistry, Chemical Engineering and Material Science, Soochow University, Suzhou, Jiangsu, 215123 (China); Lu Jianmei, E-mail: lujm@suda.edu.cn [Key Laboratory of Organic Synthesis of Jiangsu Province, College of Chemistry, Chemical Engineering and Material Science, Soochow University, Suzhou, Jiangsu, 215123 (China); Li Najun, E-mail: linajun@sina.com [Key Laboratory of Organic Synthesis of Jiangsu Province, College of Chemistry, Chemical Engineering and Material Science, Soochow University, Suzhou, Jiangsu, 215123 (China)

    2010-04-15

    A novel initiator containing proflavine derivative moiety, 3,6-dibromo-isobutyramide acridine (DIA), was synthesized and initiated the atom transfer radical polymerization (ATRP) of methyl methacrylate (MMA). A water-soluble monomer, N,N-dimethylacrylamide (DMAA) was also initiated by DIA for comparison. The obtained fluorescent polymers, PMMA-DIA and PDMAA-DIA, were characterized by {sup 1}H NMR, GPC and TGA. The emission spectra of the fluorescent polymers exhibit obvious changes in color and fluorescence intensity along with pH varied in range of 3.0-9.0. In addition, the obtained polymers present good film-forming capacity and the films also have a high quantum yield and pH response. Both oil-soluble PMMA-DIA and water-soluble PDMAA-DIA have steady optical and chemical properties by containing proflavine moiety in the main chain.

  12. A KIND OF FLUORESCENCE PROBE TO STUDY THE KINETICS OF POLYMERIZATION PROCESS

    Institute of Scientific and Technical Information of China (English)

    YANG Guoqiang; WU Shikang

    1994-01-01

    Fluorescence properties of 1-phenyl-3-(4'-nitrophenyl) pyrazoline (PNP) were studied in bulk polymerization process of methylmethacrylate (MMA). The fluorescence intensity of PNP was enhanced and the emission maximum was blue shifted with the polymerization progress. In the period of auto-acceleration of the polymerization the enhancement of fluorescence intensity and blue shift of peak wavelength in spectra could be observed evidently. This means that the solvatochromic properties of PNP are influenced not only by the solvent polarity but also by the viscosity of the medium(especially by the phase transition). In solid state PNP emits from the charge transfer excited state without solvent relaxation. The transient emission spectra and the results from Bakhshiev model of solvent relaxation coincide with that from the polymerization experiment.

  13. Quantum dots (QDs) based fluorescence probe for the sensitive determination of kaempferol

    Science.gov (United States)

    Tan, Xuanping; Liu, Shaopu; Shen, Yizhong; He, Youqiu; Yang, Jidong

    2014-12-01

    In this work, using the quenching of fluorescence of thioglycollic acid (TGA)-capped CdTe quantum dots (QDs), a novel method for the determination of kaempferol (KAE) has been developed. Under optimum conditions, a linear calibration plot of the quenched fluorescence intensity at 552 nm against the concentration of KAE was observed in the range of 4-44 μg mL-1 with a detection limit (3σ/K) of 0.79 μg mL-1. In addition, the detailed reaction mechanism has also been proposed on the basis of electron transfer supported by ultraviolet-visible (UV-vis) absorption and fluorescence (FL) spectroscopy. The method has been applied for the determination of KAE in pharmaceutical preparations with satisfactory results. The proposed method manifested several advantages such as high sensitivity, short analysis time, low cost and ease of operation.

  14. Thiophene-derivatized Fluorescent Benzamides as Possible Probes for Histone Deacetylases

    International Nuclear Information System (INIS)

    Seo, Young Jun

    2013-01-01

    We have synthesized a series of novel fluorescent benzamides inhibitors possessing intrinsic fluorescence properties. Most of these benzamide fluorophores exhibit high quantum yields, making them suitable for use in imaging studies, with colors ranging from blue to green; a couple of them were also water-soluble. Notably, TB1 and TB2 display a high quantum yield and TB1 exhibits high binding affinity to HDAC enzymes. We believe that these new fluorescent benzamide inhibitors might be useful diagnostic tools for in vitro studies of HDACs. Histone deacetylases (HDACs) are crucial gene regulating enzymes that control the expression of histones-epigenetic targets in research related to developing new therapies for cancer, central nervous system disorders, and heart disease. The deacetylation of histones is a vital repression process in transcriptional gene expression; it also affects apoptosis, cell-cycle arrest, and angiogenesis

  15. Thiophene-derivatized Fluorescent Benzamides as Possible Probes for Histone Deacetylases

    Energy Technology Data Exchange (ETDEWEB)

    Seo, Young Jun [Chonbuk National Univ., Jeonju (Korea, Republic of)

    2013-08-15

    We have synthesized a series of novel fluorescent benzamides inhibitors possessing intrinsic fluorescence properties. Most of these benzamide fluorophores exhibit high quantum yields, making them suitable for use in imaging studies, with colors ranging from blue to green; a couple of them were also water-soluble. Notably, TB1 and TB2 display a high quantum yield and TB1 exhibits high binding affinity to HDAC enzymes. We believe that these new fluorescent benzamide inhibitors might be useful diagnostic tools for in vitro studies of HDACs. Histone deacetylases (HDACs) are crucial gene regulating enzymes that control the expression of histones-epigenetic targets in research related to developing new therapies for cancer, central nervous system disorders, and heart disease. The deacetylation of histones is a vital repression process in transcriptional gene expression; it also affects apoptosis, cell-cycle arrest, and angiogenesis.

  16. Fluorescent carbon nanoparticles derived from natural materials of mango fruit for bio-imaging probes

    Science.gov (United States)

    Jeong, Chan Jin; Roy, Arup Kumer; Kim, Sung Han; Lee, Jung-Eun; Jeong, Ji Hoon; Insik; Park, Sung Young

    2014-11-01

    Water soluble fluorescent carbon nanoparticles (FCP) obtained from a single natural source, mango fruit, were developed as unique materials for non-toxic bio-imaging with different colors and particle sizes. The prepared FCPs showed blue (FCP-B), green (FCP-G) and yellow (FCP-Y) fluorescence, derived by the controlled carbonization method. The FCPs demonstrated hydrodynamic diameters of 5-15 nm, holding great promise for clinical applications. The biocompatible FCPs demonstrated great potential in biological fields through the results of in vitro imaging and in vivo biodistribution. Using intravenously administered FCPs with different colored particles, we precisely defined the clearance and biodistribution, showing rapid and efficient urinary excretion for safe elimination from the body. These findings therefore suggest the promising possibility of using natural sources for producing fluorescent materials.Water soluble fluorescent carbon nanoparticles (FCP) obtained from a single natural source, mango fruit, were developed as unique materials for non-toxic bio-imaging with different colors and particle sizes. The prepared FCPs showed blue (FCP-B), green (FCP-G) and yellow (FCP-Y) fluorescence, derived by the controlled carbonization method. The FCPs demonstrated hydrodynamic diameters of 5-15 nm, holding great promise for clinical applications. The biocompatible FCPs demonstrated great potential in biological fields through the results of in vitro imaging and in vivo biodistribution. Using intravenously administered FCPs with different colored particles, we precisely defined the clearance and biodistribution, showing rapid and efficient urinary excretion for safe elimination from the body. These findings therefore suggest the promising possibility of using natural sources for producing fluorescent materials. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr04805a

  17. Enhanced fluorescence imaging of live cells by effective cytosolic delivery of probes.

    Directory of Open Access Journals (Sweden)

    Marzia Massignani

    Full Text Available BACKGROUND: Microscopic techniques enable real-space imaging of complex biological events and processes. They have become an essential tool to confirm and complement hypotheses made by biomedical scientists and also allow the re-examination of existing models, hence influencing future investigations. Particularly imaging live cells is crucial for an improved understanding of dynamic biological processes, however hitherto live cell imaging has been limited by the necessity to introduce probes within a cell without altering its physiological and structural integrity. We demonstrate herein that this hurdle can be overcome by effective cytosolic delivery. PRINCIPAL FINDINGS: We show the delivery within several types of mammalian cells using nanometre-sized biomimetic polymer vesicles (a.k.a. polymersomes that offer both highly efficient cellular uptake and endolysomal escape capability without any effect on the cellular metabolic activity. Such biocompatible polymersomes can encapsulate various types of probes including cell membrane probes and nucleic acid probes as well as labelled nucleic acids, antibodies and quantum dots. SIGNIFICANCE: We show the delivery of sufficient quantities of probes to the cytosol, allowing sustained functional imaging of live cells over time periods of days to weeks. Finally the combination of such effective staining with three-dimensional imaging by confocal laser scanning microscopy allows cell imaging in complex three-dimensional environments under both mono-culture and co-culture conditions. Thus cell migration and proliferation can be studied in models that are much closer to the in vivo situation.

  18. The fluorescence lifetime of BRI1-GFP as probe for the noninvasive determination of the membrane potential in living cells

    Science.gov (United States)

    Elgass, K.; Caesar, K.; Schleifenbaum, F.; Meixner, A. J.; Harter, K.

    2010-02-01

    As the excited state lifetime of a fluorescent molecule depends on its environment, it is possible to use it as a probe for physico-chemical parameters of the surrounding medium. Whereas this is well known for many solid guest/host systems, only few reports of quantitative, temporal resolved in vivo studies to monitor the nano-environment for a protein-coupled chromophore such as GFP are known from literature. Here we present a novel approach to determine the membrane potential of living (plant) cells based on the fluorescence lifetime (FLT) analysis of membrane-located GFP. By using confocal sample scanning microscopy (CSSM) combined with fluorescence lifetime imaging microscopy, we recently showed that the phytohormone brassinolide (BL) induces cell wall expansion and a decrease in the FLT of the BRI1-GFP in living cells of Arabidopsis thaliana seedlings. BRI1 is the dominant functional receptor for BL in Arabidopsis and locates to the plasma membrane. Although the dependence of the FLT of GFP on its physico-chemical environment such as pH-value, refractive index and pressure has been reported, the observed FLT decrease of BRI1-GFP in response to BL application could not be explained by these parameters. However, our in vivo FLT and CSSM analyses indicate that the BLinduced change in the FLT of BRI1-GFP is caused by hyperpolarisation of the plasma membrane (Em). Thus, our results indicate that BRI1-GFP serves as sensitive and non-invasive probe for recording the Em of the plasma membrane in living plant cells with high spatio-temporal resolution.

  19. Highly Sensitive Fluorescence Probe Based on Functional SBA-15 for Selective Detection of Hg2+

    Directory of Open Access Journals (Sweden)

    Wang Xiaoyu

    2010-01-01

    Full Text Available Abstract An inorganic–organic hybrid fluorescence chemosensor (DA/SBA-15 was prepared by covalent immobilization of a dansylamide derivative into the channels of mesoporous silica material SBA-15 via (3-aminopropyltriethoxysilane (APTES groups. The primary hexagonally ordered mesoporous structure of SBA-15 was preserved after the grafting procedure. Fluorescence characterization shows that the obtained inorganic–organic hybrid composite is highly selective and sensitive to Hg2+ detection, suggesting the possibility for real-time qualitative or quantitative detection of Hg2+ and the convenience for potential application in toxicology and environmental science.

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

    International Nuclear Information System (INIS)

    Urano, Yasuteru

    2008-01-01

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

  1. A Conjugated Aptamer-Gold Nanoparticle Fluorescent Probe for Highly Sensitive Detection of rHuEPO-α

    Directory of Open Access Journals (Sweden)

    Zhaoyang Zhang

    2011-11-01

    Full Text Available We present here a novel conjugated aptamer-gold nanoparticle (Apt-AuNPs fluorescent probe and its application for specific detection of recombinant human erythropoietin-α (rHuEPO-α. In this nanobiosensor, 12 nm AuNPs function as both a nano-scaffold and a nano-quencher (fluorescent energy acceptor, on the surface of which the complementary sequences are linked (as cODN-AuNPs and pre-hybridized with carboxymethylfluorescein (FAM-labeled anti-rHuEPO-α aptamers. Upon target protein binding, the aptamers can be released from the AuNP surface and the fluorescence signal is restored. Key variables such as the length of linker, the hybridization site and length have been designed and optimized. Full performance evaluation including sensitivity, linear range and interference substances are also described. This nanobiosensor provides a promising approach for a simple and direct quantification of rHuEPO-α concentrations as low as 0.92 nM within a few hours.

  2. Concise and Efficient Fluorescent Probe via an Intromolecular Charge Transfer for the Chemical Warfare Agent Mimic Diethylchlorophosphate Vapor Detection.

    Science.gov (United States)

    Yao, Junjun; Fu, Yanyan; Xu, Wei; Fan, Tianchi; Gao, Yixun; He, Qingguo; Zhu, Defeng; Cao, Huimin; Cheng, Jiangong

    2016-02-16

    Sarin, used as chemical warfare agents (CWAs) for terrorist attacks, can induce a number of virulent effects. Therefore, countermeasures which could realize robust and convenient detection of sarin are in exigent need. A concise charge-transfer colorimetric and fluorescent probe (4-(6-(tert-butyl)pyridine-2-yl)-N,N-diphenylaniline, TBPY-TPA) that could be capable of real-time and on-site monitoring of DCP vapor was reported in this contribution. Upon contact with DCP, the emission band red-shifted from 410 to 522 nm upon exposure to DCP vapor. And the quenching rate of TBPY-TPA reached up to 98% within 25 s. Chemical substances such as acetic acid (HAc), dimethyl methylphosphonate (DMMP), pinacolyl methylphosphonate (PAMP), and triethyl phosphate (TEP) do not interfere with the detection. A detection limit for DCP down to 2.6 ppb level is remarkably achieved which is below the Immediately Dangerous to Life or Health concentration. NMR data suggested that a transformation of the pyridine group into pyridinium salt via a cascade reaction is responsible for the sensing process which induced the dramatic fluorescent red shift. All of these data suggest TBPY-TPA is a promising fluorescent sensor for a rapid, simple, and low-cost method for DCP detection, which could be easy to prepare as a portable chemosensor kit for its practical application in real-time and on-site monitoring.

  3. Design and synthesis of new fluorescent probe for rapid and highly sensitive detection of proteins via electrophoretic gel stain.

    Science.gov (United States)

    Suzuki, Yoshio; Takagi, Nobuyuki; Chimuro, Tomoyuki; Shinohara, Atsushi; Sakaguchi, Nao; Hiratsuka, Atsunori; Yokoyama, Kenji

    2011-06-01

    A new fluorescent molecular probe, 2,2'-(1E,1'E)-2,2'-(4-(dicyanomethylene)-4H-pyrane-2,6-diyl)bis(ethene-2,1-diyl)bis(sodium benzenesulfonate) salt (1), possessing the cyanopyranyl moieties and two benzene sulfonic acid groups was designed and synthesized to detect proteins in solution and for high-throughput SDS-PAGE. Compound 1 exhibited no fluorescence in the absence of proteins; however, it exhibited strong fluorescence on the addition of bovine serum albumin as a result of intramolecular charge transfer. Compared with the conventional protocols for in-gel protein staining, such as SYPRO Ruby and silver staining, 1 achieves higher sensitivity, even though it offers a simplified, higher throughput protocol. In fact, the total time required for protein staining was 60-90 min under optimum conditions much shorter than that required by the less-sensitive silver staining or SYPRO Ruby staining protocols. Moreover, 1 was successfully applied to protein identification by mass spectrometry via in-gel tryptic digestion, Western blotting, and native PAGE together with protein staining by 1, which is a modified protocol of blue native PAGE (BN-PAGE). Thus, 1 may facilitate high-sensitivity protein detection, and it may be widely applicable as a convenient tool in various scientific and medical fields. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. A Bright Future for Precision Medicine: Advances in Fluorescent Chemical Probe Design and Their Clinical Application.

    Science.gov (United States)

    Garland, Megan; Yim, Joshua J; Bogyo, Matthew

    2016-01-21

    The Precision Medicine Initiative aims to use advances in basic and clinical research to develop therapeutics that selectively target and kill cancer cells. Under the same doctrine of precision medicine, there is an equally important need to visualize these diseased cells to enable diagnosis, facilitate surgical resection, and monitor therapeutic response. Therefore, there is a great opportunity for chemists to develop chemically tractable probes that can image cancer in vivo. This review focuses on recent advances in the development of optical probes, as well as their current and future applications in the clinical management of cancer. The progress in probe development described here suggests that optical imaging is an important and rapidly developing field of study that encourages continued collaboration among chemists, biologists, and clinicians to further refine these tools for interventional surgical imaging, as well as for diagnostic and therapeutic applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. A novel ''donor-π-acceptor'' type fluorescence probe for sensing pH: mechanism and application in vivo.

    Science.gov (United States)

    Chao, Jianbin; Wang, Huijuan; Zhang, Yongbin; Yin, Caixia; Huo, Fangjun; Song, Kailun; Li, Zhiqing; Zhang, Ting; Zhao, Yaqin

    2017-11-01

    A novel pH fluorescent probe 1-(pyren-1-yl)-3-(6-methoxypridin-3-yl)-acrylketone, (PMPA), which had a pyrene structure attached to methoxypyridine, was synthesized for monitoring extremely acidic and alkaline pH. The pH titrations indicated that PMPA displayed a remarkable emission enhancement with a pK a of 2.70 and responded linearly to minor pH fluctuations within the extremely acidic range of 1.26-3.97. Interestingly, PMPA also exhibited strong pH-dependent characteristics with pK a 9.32 and linear response to extreme-alkalinity range of 8.54-10.36. In addition, PMPA displayed a good selectivity, excellent photostability and large Stokes shift (167nm). Furthermore, the probe PMPA had excellent cell membrane permeability and was applied successfully to rapidly detect pH in living cells. pH value in these organs was closely related to many diseases, so these findings suggested that the probe had potential application in pH detecting for disease diagnosis. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Circular dichroism probed by two-photon fluorescence microscopy in enantiopure chiral polyfluorene thin films

    NARCIS (Netherlands)

    Savoini, M.; Wu, Xiaofei; Celebrano, M.; Ziegler, J.; Biagioni, P.; Meskers, S.C.J.; Duo, L.; Hecht, B.; Finazzi, M.

    2012-01-01

    Two-photon fluorescence scanning confocal microscopy sensitive to circular dichroism with a diffraction-limited resolution well below 500 nm is demonstrated. With this method, the spatial variation of the circular dichroism of thermally annealed chiral polyfluorene thin films has been imaged. We

  7. A fluorescent probe which allows highly specific thiol labeling at low pH

    DEFF Research Database (Denmark)

    Nielsen, Jonas W.; Jensen, Kristine Steen; Hansen, Rosa E.

    2012-01-01

    and properties of a thiol-specific reagent, fluorescent cyclic activated disulfide (FCAD), which includes the fluorescein moiety as fluorophore and utilizes a variation of thiol-disulfide exchange chemistry. The leaving-group character of FCAD makes it reactive at pH 3, allowing modification at low pH, limiting...

  8. ABC transporters affect the detection of intracellular oxidants by fluorescent probes

    Czech Academy of Sciences Publication Activity Database

    Procházková, J.; Kubala, Lukáš; Kotasová, H.; Gudernová, I.; Šrámková, Z.; Pekarová, Michaela; Sarkadi, B.; Pachernik, J.

    2011-01-01

    Roč. 45, č. 7 (2011), s. 779-787 ISSN 1071-5762 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Institutional support: RVO:68081707 Keywords : Fluorescence * redox status * mitochondria Subject RIV: BO - Biophysics Impact factor: 2.878, year: 2011

  9. Preparation and use of Cu nanoclusters as fluorescent probes to determine Au(III) ions

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jinshui, E-mail: jsliu@sina.com; Wang, Bin; Xu, Meijiao; Wang, Lizhen; Zhou, Zihan

    2017-05-15

    Polyethyleneimine-capped Cu nanoclusters (PEI-CuNCs) with a mean diameter of 2.5 nm and that emitted blue light at 485 nm were synthesized. The fluorescence of PEI-CuNCs was strongly quenched when Au(III) ions were present. Because of this specific response, a PEI-CuNC-based fluorescent turn-off sensor was developed. Under optimum conditions, a good linear relationship (R{sup 2}=0.993) was found between the relative fluorescence intensity (F{sub 0}/F) and the Au(III) ion concentration between 0.5 and 15 µM. The detection limit for Au(III) ions was 0.06 µM. The quenching mechanism was assessed by performing X-ray photoelectron spectroscopy spectra studies. The fluorescence quenching was ascribed to photo-induced electron transfer from the PEI-CuNCs to the AuCl{sub 4}{sup −} occur. Using The method described is a new way of quantifying Au(III) ions.

  10. Fluorescence spectral correlation spectroscopy (FSCS) for probes with highly overlapping emission spectra

    Czech Academy of Sciences Publication Activity Database

    Benda, A.; Kapusta, Peter; Hof, Martin; Gaus, K.

    2014-01-01

    Roč. 22, č. 3 (2014), s. 2973-2988 ISSN 1094-4087 R&D Projects: GA AV ČR KJB400400904; GA ČR GBP208/12/G016 Institutional support: RVO:61388955 Keywords : spectroscopy * fluorescence and luminiscence * confocal microscopy Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.488, year: 2014

  11. Probe transfer with and without membrane fusion in a fluorescence fusion assay

    NARCIS (Netherlands)

    Ohki, S; Flanagan, TD; Hoekstra, D

    1998-01-01

    An analysis of the R(18) fusion assay was made during the fusion of the Sendai virus with erythrocyte ghosts. The increase in R(18) fluorescence, reflecting the interaction process, was evaluated in terms of the different processes that in principle may contribute to this increase, that is,

  12. Quantitative imaging of glutathione in live cells using a reversible reaction-based ratiometric fluorescent probe

    Science.gov (United States)

    Glutathione (GSH) plays an important role in maintaining redox homeostasis inside cells. Currently, there are no methods available to quantitatively assess the GSH concentration in live cells. Live cell fluorescence imaging revolutionized the understanding of cell biology and has become an indispens...

  13. Fluorescence correlation spectroscopy on electron transfer reactions : probing inter- and intramolecular redox processes

    NARCIS (Netherlands)

    Sen, S.

    2016-01-01

    We developed a new FRET-based technique, “Fluredox”, which allows fluorescence readout of the redox state of oxido-reductases at single molecule level. Commercially available red-absorbing fluorophore ATTO655 was selected for labeling Azurin, a small blue mononuclear copper protein. Single molecule

  14. Structural properties of oligonucleotide monolayers on gold surfaces probed by fluorescence investigations.

    Science.gov (United States)

    Rant, Ulrich; Arinaga, Kenji; Fujita, Shozo; Yokoyama, Naoki; Abstreiter, Gerhard; Tornow, Marc

    2004-11-09

    We present optical investigations on the conformation of oligonucleotide layers on Au surfaces. Our studies concentrate on the effect of varying surface coverage densities on the structural properties of layers of 12- and 24mer single-stranded DNA, tethered to the Au surface at one end while being labeled with a fluorescent marker at the opposing end. The distance-dependent energy transfer from the marker dye to the metal surface, which causes quenching of the observed fluorescence, is used to provide information on the orientation of the DNA strands relative to the surface. Variations in the oligonucleotide coverage density, as determined from electrochemical quantification, over 2 orders of magnitude are achieved by employing different preparation conditions. The observed enhancement in fluorescence intensity with increasing DNA coverage can be related to a model involving mutual steric interactions of oligonucleotides on the surface, as well as fluorescence quenching theory. Finally, the applicability of the presented concepts for investigations of heterogeneous monolayers is demonstrated by means of studying the coadsorption of mercaptohexanol onto DNA-modified Au surfaces.

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

    International Nuclear Information System (INIS)

    Jaillon, Franck; Zheng Wei; Huang Zhiwei

    2008-01-01

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

  16. Monitoring of Au(iii) species in plants using a selective fluorescent probe.

    Science.gov (United States)

    Li, Zhen; Xu, Yuqing; Fu, Jie; Zhu, Hailiang; Qian, Yong

    2018-01-23

    A colorimetric and ratiometric probe with a push-pull chromophore dicyanoisophorone system, AuP, has been developed for the detection of Au(iii) species with highly sensitive and selective response to real-water samples and living tissues of Arabidopsis thaliana.

  17. A novel water-soluble 1,8-naphthalimide as a fluorescent pH-probe and a molecular logic circuit

    International Nuclear Information System (INIS)

    Georgiev, Nikolai I.; Dimitrova, Margarita D.; Krasteva, Paoleta V.; Bojinov, Vladimir B.

    2017-01-01

    A novel highly water-soluble fluorescence sensing 1,8-naphthalimide is synthesized and investigated. The novel compound is designed on the “fluorophore-receptor 1 -spacer-receptor 2 ” model as a molecular fluorescence probe for determination of ions in 100% aqueous media. The novel probe comprising hydrazide and N-methylpiperazine substituents is capable of operating simultaneously via ICT and PET signaling mechanism and of recognizing selectively protons and hydroxyl anions over the representative metal ions and anions. Due to the remarkable fluorescence changes as a function of pH the system is able to act as a three output combinatorial logic circuit with two chemical inputs. Two INHIBIT gates in fluorescence and absorption mode as well as an IMPLICATION logic gate are obtained. Because of the parallel action of both INHIBIT gates a magnitude digital comparator is achieved for the first time in this way.

  18. Biophysical characterization of the fluorescent protein voltage probe VSFP2.3 based on the voltage-sensing domain of Ci-VSP

    DEFF Research Database (Denmark)

    Lundby, Alicia; Akemann, Walther; Knöpfel, Thomas

    2010-01-01

    A voltage sensitive phosphatase was discovered in the ascidian Ciona intestinalis. The phosphatase, Ci-VSP, contains a voltage-sensing domain homologous to those known from voltage-gated ion channels, but unlike ion channels, the voltage-sensing domain of Ci-VSP can reside in the cell membrane...... as a monomer. We fused the voltage-sensing domain of Ci-VSP to a pair of fluorescent reporter proteins to generate a genetically encodable voltage-sensing fluorescent probe, VSFP2.3. VSFP2.3 is a fluorescent voltage probe that reports changes in membrane potential as a FRET (fluorescence resonance energy....... Neutralization of an arginine in S4, previously suggested to be a sensing charge, and measuring associated sensing currents indicate that this charge is likely to reside at the membrane-aqueous interface rather than within the membrane electric field. The data presented give us insights into the voltage-sensing...

  19. Recent Developments in Fluorescence Correlation Spectroscopy for Diffusion Measurements in Planar Lipid Membranes

    Czech Academy of Sciences Publication Activity Database

    Macháň, Radek; Hof, Martin

    2010-01-01

    Roč. 11, č. 2 (2010), s. 427-457 E-ISSN 1422-0067 R&D Projects: GA ČR GA203/08/0114; GA AV ČR GEMEM/09/E006 Institutional research plan: CEZ:AV0Z40400503 Keywords : lateral diffusion * fluorescence fluctuation spectroscopy * confocal microscopy Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.279, year: 2010

  20. Identification of squid species by melting temperature shifts on fluorescence melting curve analysis (FMCA) using single dual-labeled probe

    Science.gov (United States)

    Koh, Eunjung; Song, Ha Jeong; Kwon, Na Young; Kim, Gi Won; Lee, Kwang Ho; Jo, Soyeon; Park, Sujin; Park, Jihyun; Park, Eun Kyeong; Hwang, Seung Yong

    2017-06-01

    Real time PCR is a standard method for identification of species. One of limitations of the qPCR is that there would be false-positive result due to mismatched hybridization between target sequence and probe depending on the annealing temperature in the PCR condition. As an alternative, fluorescence melting curve analysis (FMCA) could be applied for species identification. FMCA is based on a dual-labeled probe. Even with subtle difference of target sequence, there are visible melting temperature (Tm) shift. One of FMCA applications is distinguishing organisms distributed and consumed globally as popular food ingredients. Their prices are set by species or country of origin. However, counterfeiting or distributing them without any verification procedure are becoming social problems and threatening food safety. Besides distinguishing them in naked eye is very difficult and almost impossible in any processed form. Therefore, it is necessary to identify species in molecular level. In this research three species of squids which have 1-2 base pair differences each are selected as samples since they have the same issue. We designed a probe which perfectly matches with one species and the others mismatches 2 and 1 base pair respectively and labeled with fluorophore and quencher. In an experiment with a single probe, we successfully distinguished them by Tm shift depending on the difference of base pair. By combining FMCA and qPCR chip, smaller-scale assay with higher sensitivity and resolution could be possible, andc furthermore, enabling results analysis with smart phone would realize point-of-care testing (POCT).

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

    Czech Academy of Sciences Publication Activity Database

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

    2016-01-01

    Roč. 55, č. 1 (2016), s. 174-178 ISSN 1433-7851 R&D Projects: GA ČR GBP206/12/G151; GA ČR(CZ) GC14-03141J Institutional support: RVO:61388963 ; RVO:61388955 Keywords : DNA * fluorescence spectroscopy * fluorescent probes * nucleosides * time-resolved spectroscopy Subject RIV: CC - Organic Chemistry ; BO - Biophysics (UFCH-W) Impact factor: 11.994, year: 2016

  2. A novel fluorescence probe based on triphenylamine Schiff base for bioimaging and responding to pH and Fe{sup 3+}

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lei; Yang, Xiaodong; Chen, Xiuli [School of Chemistry and Chemical Engineering, South China University of Technology, Guangzhou 510640 (China); Zhou, Yuping [Guangdong Provincial key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China); Lu, Xiaodan; Yan, Chenggong; Xu, Yikai [Medical Imaging Center, Nanfang Hospital, Southern Medical University, Guangzhou 510515 (China); Liu, Ruiyuan, E-mail: ruiyliu@smu.edu.cn [Guangdong Provincial key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China); Qu, Jinqing, E-mail: cejqqu@scut.edu.cn [School of Chemistry and Chemical Engineering, South China University of Technology, Guangzhou 510640 (China)

    2017-03-01

    A novel fluorescence probe 1 based on triphenylamine was synthesized and characterized by NMR, IR, high resolution mass spectrometry and elemental analysis. Its fluorescence was quenched when pH below 2. There was a linear relationship between the fluorescence intensity and pH value ranged from 2 to 7. And its fluorescence emission was reversibility in acidic and alkaline solution. Furthermore, it exhibited remarkable selectivity and high sensitivity to Fe{sup 3+} and was able to detect Fe{sup 3+} in aqueous solution with low detection limit of 0.511 μM. Job plot showed that the binding stoichiometry of 1 with Fe{sup 3+} was 1:1. Further observations of {sup 1}H NMR titration suggested that coordination interaction between Fe{sup 3+} and nitrogen atom on C =N bond promoted the intramolecular charge transfer (ICT) or energy transfer process causing fluorescence quenching. Additionally, 1 was also able to be applied for detecting Fe{sup 3+} in living cell and bioimaging. - Graphical abstract: Triphenylamine based fluorescence probe can detect pH and Fe{sup 3+} simultaneously in aqueous solution and be applied for detecting Fe{sup 3+} in living cell and bioimaging. - Highlights: • The fluorescence probe is sensitive to pH in strong acid conditions. • The fluorescence chemosensor can detect pH and Fe{sup 3+} simultaneously. • The recognition is able to carry out in aqueous solution. • The probe can also be applied for detecting Fe{sup 3+} in living cell and bioimaging. • The sensor is synthesized easily with one step.

  3. Does fluoride disrupt hydrogen bond network in cationic lipid bilayer? Time-dependent fluorescence shift of Laurdan and molecular dynamics simulations

    Czech Academy of Sciences Publication Activity Database

    Pokorná, Šárka; Jurkiewicz, Piotr; Vazdar, M.; Cwiklik, Lukasz; Jungwirth, Pavel; Hof, Martin

    2014-01-01

    Roč. 141, č. 22 (2014), 22D516 ISSN 0021-9606 R&D Projects: GA ČR GBP208/12/G016 Institutional support: RVO:61388955 ; RVO:61388963 Keywords : fluorescence sfifts * Cationic lipids * Hydrogen bond networks Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.952, year: 2014

  4. Green fluorescent protein changes the conductance of connexin 43 (Cx43) hemichannels reconstituted in planar lipid bilayers.

    Science.gov (United States)

    Carnarius, Christian; Kreir, Mohamed; Krick, Marcel; Methfessel, Christoph; Moehrle, Volker; Valerius, Oliver; Brüggemann, Andrea; Steinem, Claudia; Fertig, Niels

    2012-01-20

    In mammalian tissues, connexin 43 (Cx43) is the most prominent member of the connexin family. In a single lipid bilayer, six connexin subunits assemble into a hemichannel (connexon). Direct communication of apposing cells is realized by two adjacent hemichannels, which can form gap junction channels. Here, we established an expression system in Pichia pastoris to recombinantly produce and purify Cx43 as well as Cx43 fused to green fluorescent protein (GFP). Proteins were isolated from crude cell membrane fractions via affinity chromatography. Cx43 and Cx43-GFP hemichannels were reconstituted in giant unilamellar vesicles as proven by fluorescence microscopy, and their electrophysiological behavior was analyzed on the single channel level by planar patch clamping. Cx43 and Cx43-GFP both showed an ohmic behavior and a voltage-dependent open probability. Cx43 hemichannels exhibited one major mean conductance of 224 ± 26 picosiemens (pS). In addition, a subconductance state at 124 ± 5 pS was identified. In contrast, the analysis of Cx43-GFP single channels revealed 10 distinct conductance states in the range of 15 to 250 pS, with a larger open probability at 0 mV as compared with Cx43, which suggests that intermolecular interactions between the GFP molecules alter the electrophysiology of the protein.

  5. Green Fluorescent Protein Changes the Conductance of Connexin 43 (Cx43) Hemichannels Reconstituted in Planar Lipid Bilayers*

    Science.gov (United States)

    Carnarius, Christian; Kreir, Mohamed; Krick, Marcel; Methfessel, Christoph; Moehrle, Volker; Valerius, Oliver; Brüggemann, Andrea; Steinem, Claudia; Fertig, Niels

    2012-01-01

    In mammalian tissues, connexin 43 (Cx43) is the most prominent member of the connexin family. In a single lipid bilayer, six connexin subunits assemble into a hemichannel (connexon). Direct communication of apposing cells is realized by two adjacent hemichannels, which can form gap junction channels. Here, we established an expression system in Pichia pastoris to recombinantly produce and purify Cx43 as well as Cx43 fused to green fluorescent protein (GFP). Proteins were isolated from crude cell membrane fractions via affinity chromatography. Cx43 and Cx43-GFP hemichannels were reconstituted in giant unilamellar vesicles as proven by fluorescence microscopy, and their electrophysiological behavior was analyzed on the single channel level by planar patch clamping. Cx43 and Cx43-GFP both showed an ohmic behavior and a voltage-dependent open probability. Cx43 hemichannels exhibited one major mean conductance of 224 ± 26 picosiemens (pS). In addition, a subconductance state at 124 ± 5 pS was identified. In contrast, the analysis of Cx43-GFP single channels revealed 10 distinct conductance states in the range of 15 to 250 pS, with a larger open probability at 0 mV as compared with Cx43, which suggests that intermolecular interactions between the GFP molecules alter the electrophysiology of the protein. PMID:22139870

  6. Single-cell Raman and fluorescence microscopy reveal the association of lipid bodies with phagosomes in leukocytes

    Science.gov (United States)

    van Manen, Henk-Jan; Kraan, Yvonne M.; Roos, Dirk; Otto, Cees

    2005-01-01

    Cellular imaging techniques based on vibrational spectroscopy have become powerful tools in cell biology because the molecular composition of subcellular compartments can be visualized without the need for labeling. Using high-resolution, nonresonant confocal Raman microscopy on individual cells, we demonstrate here that lipid bodies (LBs) rich in arachidonate as revealed by their Raman spectra associate with latex bead-containing phagosomes in neutrophilic granulocytes. This finding was corroborated in macrophages and in PLB-985 cells, which can be induced to differentiate into neutrophil-like cells, by selective staining of LBs and visualization by confocal fluorescence microscopy. We further show that the accumulation of LBs near phagosomes is mediated at least in part by the flavohemoprotein gp91phox (in which “phox” is phagocyte oxidase), because different LB distributions around phagocytosed latex beads were observed in WT and gp91phox-deficient PLB-985 cells. gp91phox, which accumulates in the phagosomal membrane, is the catalytic subunit of the leukocyte NADPH oxidase, a critical enzyme in the innate immune response. Finally, time-lapse fluorescence microscopy experiments on neutrophils revealed that the LB-phagosome association is transient, similar to the “kiss-and-run” behavior displayed by endosomes involved in phagosome maturation. Because arachidonic acid (AA) has been shown to be involved in NADPH oxidase activation and phagosome maturation in neutrophils and macrophages, respectively, the findings reported here suggest that LBs may provide a reservoir of AA for local activation of these essential leukocyte functions. PMID:16002471

  7. Probing into the Interaction of Nicotine and Bovine Submaxillary Mucin: NMR, Fluorescence, and FTIR Approaches

    Directory of Open Access Journals (Sweden)

    Xiaoxiang Liao

    2016-01-01

    Full Text Available Nicotine, the important component of cigarette products, may have an impact on the oral environment after inhalation. The research of interaction between nicotine and bovine submaxillary mucin (BSM contributes to understand the binding mechanism of nicotine and BSM, and the effects of nicotine on the structure and function of the mucin. NMR data demonstrated that the interaction between nicotine and BSM did exist, and it was pyrrolidyl ring of nicotine playing the major role in the binding. The quenching mechanisms of nicotine and BSM in different pH were different: for acidic environment, the quenching was dynamic; while it became static in the alkaline circumstance. Synchronous fluorescence spectra indicated that nicotine had effect on the microenvironment of the Trp rather than Tyr residue. Meanwhile, the impact of nicotine on the conformation of BSM was also confirmed by 3D fluorescence and FTIR spectra.

  8. Characterization of microenvironment polarity and solvent accessibility of polysilsesquioxane xerogels by the fluorescent probe technique

    Energy Technology Data Exchange (ETDEWEB)

    Shea, K.J.; Zhu, H.D. [Univ., of California, Irvine, CA (United States). Dept. of Chemistry; Loy, D.A. [Sandia National Labs., Albuquerque, NM (United States)

    1995-05-01

    Poly (1, 4 bis(triethoxysilyl)benzene) (PTESB), a representative of a new type of organic-inorganic hybrid polysilsesquioxane material, was characterized by fluorescence spectroscopy for both microenvironmental polarity and solvent accessibility. A dansyl fluorescent molecule was incorporated into the bulk as well as onto the surface of both PTESB and silica materials. Information about the microenvironment polarity and accessibility of PTESB to various organic solvents was determined and compared to that of silica gel. This study found that both the bulk and surface of PTESB are less polar than that of the silica material. The silica material is accessible to polar solvents and water, while YMB is accessible to polar solvents but not to water. The hydrophobicity of PTESB differentiates these new materials from silica gel.

  9. Carbon dots prepared from citric acid and urea as fluorescent probes for hypochlorite and peroxynitrite

    International Nuclear Information System (INIS)

    Simões, Eliana F. C.; Leitão, João M. M.; Esteves da Silva, Joaquim C. G.

    2016-01-01

    Carbon dots (CDs) were prepared from citric acid and urea, and their fluorescence was found to be quenched by hypochlorite and peroxynitrite. Microwave based synthesis gives CDs with excitation/emission wavelength-dependent quantum yields (8 % at 400/520 nm; 10 % at 360/451 nm; 12 % at 350/420 nm). Quenching of fluorescence depends on pH values, and response is most selective and sensitive to hypochlorite at pH 4, and to peroxynitrite at pH 9. The lower detection limits are 0.5 and 1.5 μM, respectively. The method was successfully applied to quantify hypochlorite and peroxynitrite in standard solutions and in spiked dilute serum samples. (author)

  10. Selective Labeling of Proteins on Living Cell Membranes Using Fluorescent Nanodiamond Probes

    Directory of Open Access Journals (Sweden)

    Shingo Sotoma

    2016-03-01

    Full Text Available The impeccable photostability of fluorescent nanodiamonds (FNDs is an ideal property for use in fluorescence imaging of proteins in living cells. However, such an application requires highly specific labeling of the target proteins with FNDs. Furthermore, the surface of unmodified FNDs tends to adsorb biomolecules nonspecifically, which hinders the reliable targeting of proteins with FNDs. Here, we combined hyperbranched polyglycerol modification of FNDs with the β-lactamase-tag system to develop a strategy for selective imaging of the protein of interest in cells. The combination of these techniques enabled site-specific labeling of Interleukin-18 receptor alpha chain, a membrane receptor, with FNDs, which eventually enabled tracking of the diffusion trajectory of FND-labeled proteins on the membrane surface.

  11. Enzyme-Free Detection of Mutations in Cancer DNA Using Synthetic Oligonucleotide Probes and Fluorescence Microscopy

    DEFF Research Database (Denmark)

    Miotke, Laura; Maity, Arindam; Ji, Hanlee

    2015-01-01

    BACKGROUND: Rapid reliable diagnostics of DNA mutations are highly desirable in research and clinical assays. Current development in this field goes simultaneously in two directions: 1) high-throughput methods, and 2) portable assays. Non-enzymatic approaches are attractive for both types...... 1000-fold above the potential detection limit. CONCLUSION: Overall, the novel assay we describe could become a new approach to rapid, reliable and enzyme-free diagnostics of cancer or other associated DNA targets. Importantly, stoichiometry of wild type and mutant targets is conserved in our assay...... of methods since they would allow rapid and relatively inexpensive detection of nucleic acids. Modern fluorescence microscopy is having a huge impact on detection of biomolecules at previously unachievable resolution. However, no straightforward methods to detect DNA in a non-enzymatic way using fluorescence...

  12. Development of species-specific rDNA probes for Giardia by multiple fluorescent in situ hybridization combined with immunocytochemical identification of cyst wall antigens.

    Science.gov (United States)

    Erlandsen, Stanley L; Jarroll, Edward; Wallis, Peter; van Keulen, Harry

    2005-08-01

    In this study, we describe the development of fluorescent oligonucleotide probes to variable regions in the small subunit of 16S rRNA in three distinct Giardia species. Sense and antisense probes (17-22 mer) to variable regions 1, 3, and 8 were labeled with digoxygenin or selected fluorochomes (FluorX, Cy3, or Cy5). Optimal results were obtained with fluorochome-labeled oligonucleotides for detection of rRNA in Giardia cysts. Specificity of fluorescent in situ hybridization (FISH) was shown using RNase digestion and high stringency to diminish the hybridization signal, and oligonucleotide probes for rRNA in Giardia lamblia, Giardia muris, and Giardia ardeae were shown to specifically stain rRNA only within cysts or trophozoites of those species. The fluorescent oligonucleotide specific for rRNA in human isolates of Giardia was positive for ten different strains. A method for simultaneous FISH detection of cysts using fluorescent antibody (genotype marker) and two oligonucleotide probes (species marker) permitted visualization of G. lamblia and G. muris cysts in the same preparation. Testing of an environmental water sample revealed the presence of FISH-positive G. lamblia cysts with a specific rDNA probe for rRNA, while negative cysts were presumed to be of animal or bird origin.

  13. Photoluminescence studies of a Terbium(III) complex as a fluorescent probe for DNA detection

    Energy Technology Data Exchange (ETDEWEB)

    Khorasani-Motlagh, Mozhgan, E-mail: mkhorasani@chem.usb.ac.ir; Noroozifar, Meissam; Niroomand, Sona; Moodi, Asieh

    2013-11-15

    The photoluminescence properties of a Tb(III) complex of the form [Tb(phen){sub 2}Cl{sub 3}·OH{sub 2}] (phen=1,10-phenanthroline) in different solvents are presented. It shows the characteristic luminescence of the corresponding Ln{sup 3+} ion in the visible region. The emission intensity of this complex in coordinating solvent is higher than non-coordinating one. The suggested mechanism for the energy transfer between the ligand and Tb{sup 3+} ion is the intramolecular energy transfer mechanism. The interactions of the Tb(III) complex with fish salmon DNA are studied by fluorescence spectroscopy, circular dichroism study and viscosity measurements. The results of fluorescence titration reveal that DNA strongly quenches the intrinsic fluorescence of the complex through a static quenching procedure. The binding constant (K{sub b}) of the above metal complex at 25 °C is determined by the fluorescence titration method and it is found to be (8.06±0.01)×10{sup 3} M{sup −1}. The thermodynamic parameters (ΔH{sup 0}>0, ΔS{sup 0}>0 and ΔG{sup 0}<0) indicate that the hydrophobic interactions play a major role in DNA–Tb complex association. The results support the claim that the title complex bonds to FS-DNA by a groove mode. -- Highlights: • Photoluminescence of [Tb(phen){sub 2}Cl{sub 3}·OH{sub 2}] in different solvents are studied. • Tb(III) complex shows good binding affinity to FS DNA with K{sub b}=(8.06±0.01)×10{sup 3} M{sup −1}. • Viscosity of DNA almost unchanged by increasing amount of Tb complex. • CD spectrum of DNA has a little change with increasing amount of Tb complex. • Thermodynamic parameters indicate that the binding reaction is entropically driven.

  14. Determination of sparfloxacin with CdSe/CdS quantum dots as fluorescent probes

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Ming, E-mail: gxglzws@foxmail.com; Yan, Xiaoya; Xiong, Ling

    2015-01-15

    Water-soluble CdSe/CdS quantum dots (QDs) modified with thioglycolic acid (TGA) were synthesized. A novel method for determination of sparfloxacin (SPFX) has been developed based on quenching of the fluorescence of QDs at 556 nm wavelength. The optimum fluorescence intensity was found in 0.067 mol L{sup −1} KH{sub 2}PO{sub 4}–Na{sub 2}HPO{sub 4} buffer solution at pH 6.47 of 3.0×10{sup −5} mol L{sup −1} QDs. When the concentration of quantum dots is 3.0×10{sup −5} mol L{sup −1} the fluorescence quenching intensity of QDs is linearly proportional to the concentration of SPFX from 0.5 μg mL{sup −1} to 30 μg mL{sup −1}, with correlation coefficient R=0.9983. The detection limit for SPFX was 0.1391 μg mL{sup −1}. The method was used for determination of SPFX in tablets, and the results agreed with the claimed value. Trace amounts of SPFX in milk were also determined with the recovery of 95.3–106.8%. - Highlights: • Water-soluble CdSe/CdS quantum dots modified with thioglycolic acid were synthesized. • Determination of sparfloxacin was based on quenching of the fluorescence of QDs. • The detection limit for sparfloxacin was 0.1391 μg mL{sup −1}. • The method has been used successfully to determine SPFX in tablets and milk.

  15. mathFISH, a Web Tool That Uses Thermodynamics-Based Mathematical Models for In Silico Evaluation of Oligonucleotide Probes for Fluorescence In Situ Hybridization▿ †

    OpenAIRE

    Yilmaz, L. Safak; Parnerkar, Shreyas; Noguera, Daniel R.

    2010-01-01

    Mathematical models of RNA-targeted fluorescence in situ hybridization (FISH) for perfectly matched and mismatched probe/target pairs are organized and automated in web-based mathFISH (http://mathfish.cee.wisc.edu). Offering the users up-to-date knowledge of hybridization thermodynamics within a theoretical framework, mathFISH is expected to maximize the probability of success during oligonucleotide probe design.

  16. mathFISH, a web tool that uses thermodynamics-based mathematical models for in silico evaluation of oligonucleotide probes for fluorescence in situ hybridization.

    Science.gov (United States)

    Yilmaz, L Safak; Parnerkar, Shreyas; Noguera, Daniel R

    2011-02-01

    Mathematical models of RNA-targeted fluorescence in situ hybridization (FISH) for perfectly matched and mismatched probe/target pairs are organized and automated in web-based mathFISH (http://mathfish.cee.wisc.edu). Offering the users up-to-date knowledge of hybridization thermodynamics within a theoretical framework, mathFISH is expected to maximize the probability of success during oligonucleotide probe design.

  17. A Pyrene- and Phosphonate-Containing Fluorescent Probe as Guest Molecule in a Host Polymer Matrix

    Directory of Open Access Journals (Sweden)

    Jacqueline Marchand-Brynaert

    2013-02-01

    Full Text Available New host-guest materials have been prepared by incorporation of a home-made organic probe displaying a pyrene motif and a phosphonate function into a regular amphiphilic copolymer. Using powder X-Ray diffraction, photoluminescence and FT-IR spectroscopy, we have been able to study the non-covalent interactions between the host matrix and the guest molecule in the solid state. Interestingly, we have shown that the matrix directs the guest spatial localization and alters its properties. Thanks to the comparison of pyrene vs. N-pyrenylmaleimide derivatives, the influence of the chemical nature of the guest molecules on the non-covalent interactions with the host have been studied. In addition, using polyethylene glycol as a reference host, we have been able to evidence a true matrix effect within our new insertion materials. The phosphonated guest molecule appears to be a novel probe targeting the hydrophilic domain of the host copolymer.

  18. Dual imaging probes for magnetic resonance imaging and fluorescence microscopy based on perovskite manganite nanoparticles

    Czech Academy of Sciences Publication Activity Database

    Kačenka, M.; Kaman, Ondřej; Kotek, J.; Falteisek, L.; Černý, J.; Jirák, D.; Herynek, V.; Zacharovová, K.; Berková, A.; Jendelová, Pavla; Kupčík, Jaroslav; Pollert, Emil; Veverka, Pavel; Lukeš, I.

    2011-01-01

    Roč. 21, č. 1 (2011), s. 157-164 ISSN 0959-9428 R&D Projects: GA AV ČR KAN200200651 Institutional research plan: CEZ:AV0Z10100521; CEZ:AV0Z50390703; CEZ:AV0Z40720504 Keywords : cellular labelling * dual probe * magnetic nanoparticles * MRI * silica coating Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 5.968, year: 2011

  19. Semiconductor quantum dots as fluorescent probes for in vitro and in vivo bio-molecular and cellular imaging

    Directory of Open Access Journals (Sweden)

    Sarwat B. Rizvi

    2010-08-01

    Full Text Available Over the years, biological imaging has seen many advances, allowing scientists to unfold many of the mysteries surrounding biological processes. The ideal imaging resolution would be in nanometres, as most biological processes occur at this scale. Nanotechnology has made this possible with functionalised nanoparticles that can bind to specific targets and trace processes at the cellular and molecular level. Quantum dots (QDs or semiconductor nanocrystals are luminescent particles that have the potential to be the next generation fluorophores. This paper is an overview of the basics of QDs and their role as fluorescent probes for various biological imaging applications. Their potential clinical applications and the limitations that need to be overcome have also been discussed.

  20. Determination of the ratio between mercaptalbumin and nonmercaptalbumin by HPLC with fluorescence probe specifically binding to albumin.

    Science.gov (United States)

    Ohyama, Kaname; Kishikawa, Naoya; Matsuo, Aya; Imazato, Takahiro; Ueki, Yukitaka; Wada, Mitsuhiro; Nakashima, Kenichiro; Kuroda, Naotaka

    2014-01-01

    A simple and selective HPLC-fluorescence (FL) method with FL probe, 4-[4-(4-dimethylaminophenyl)-5-phenyl-1H-imidazol-2-yl]benzoic acid methyl ester (DAPIM), for simultaneous determination of mercaptalbumin (HMA) and nonmercaptalbumin (HNA1) was developed. After HMA and HNA1 were separated on an ion-exchange column, they were on-line and post-column mixed with DAPIM. The DAPIM-albumin complex produces FL (λex 370nm and λem 510nm); however, DAPIM solution never gives the FL. Based on this mechanism, selective determination of HMA and HNA1 were achieved without any pretreatment and interfering peak. The proposed method was applied to the measurement of HMA and HNA1 in human serum of healthy volunteers and diabetes mellitus patients. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. Use of fluorescent probes for ROS to tease apart Type I and Type II photochemical pathways in photodynamic therapy

    DEFF Research Database (Denmark)

    Garcia-Diaz, Maria; Huang, Ying-Ying; Hamblin, Michael R

    2016-01-01

    ) include superoxide, hydrogen peroxide and hydroxyl radical (HO), while singlet oxygen ((1)O2) is produced by energy transfer. Diverse methods exist to distinguish between these two pathways, some of which are more specific or more sensitive than others. In this review we cover the use of two fluorescence...... probes: singlet oxygen sensor green (SOSG) detects (1)O2; and 4-hydroxyphenyl-fluorescein (HPF) that detects HO. Interesting data was collected concerning the photochemical pathways of functionalized fullerenes compared to tetrapyrroles, stable synthetic bacteriochlorins with and without central metals......Photodynamic therapy involves the excitation of a non-toxic dye by harmless visible light to produce a long-lived triplet state that can interact with molecular oxygen to produce reactive oxygen species (ROS), which can damage biomolecules and kill cells. ROS produced by electron transfer (Type 1...

  2. Utilization of fluorescent probe association for simultaneous assessment of plasmatic, acrosomal, and mitochondrial membranes of rooster spermatozoa

    Directory of Open Access Journals (Sweden)

    ECC Celeghini

    2007-09-01

    Full Text Available This experiment was designed with the objective of developing a simple, practical, and high repeatability technique for the simultaneous evaluation of the integrity of the plasmatic and acrosomal membranes, as well as funcional mitochondria of domestic fowl spermatozoa using an association of fluorescent probes. Four ejaculates (motility > 80% and abnormal morphology < 10% from each of six Ross male broiler breeder (n=24 were diluted in TALP sperm medium (25x10(6 spermatozoa/mL and split into two aliquots, and one of these aliquots was flash frozen in liquid nitrogen and thawed to damage all cellular membranes. Three treatments were prepared from these aliquots, with the following ratios of Fresh semen:Flash frozen semen: 100:0 (T100, 50:50 (T50, and 0:100 (T0. A 150-µL aliquot of diluted semen was placed in a microcentrifuge tube with the addition of 2-µL PI, 2-µL MITO, and 50-µL FITC-PSA, and incubated at 38.5º C/8 min in the dark. An 8-µL sample was placed on a slide, coverslipped, and examined by epifluorescence microscopy. Each sample had 200 cells counted and classified based on the fluorescence emitted by each probe. By regression analysis, plasma membrane integrity, as detected by PI, was determined as: v=4.17+0.82X (R²=0.95. Acrosome integrity, as detected by FITC-PSA, generated the equation: v=4.19+0.84X (R²=0.96. Functional mitochondria was estimated by the equation v=3.20+0.83X (R²=0.96. This is an efficient technique to simultaneously evaluate plasmatic, acrosomal, and mitochondrial membranes in fowl sperm. It is suggested that its application in flow cytometry systems allows this methodology to be applied in large scale.

  3. Novel lanthanide pH fluorescent probes based on multiple emissions and its visible-light-sensitized feature

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Jintai [School of Chemistry and Environment, South China Normal University, Guangzhou 510006 (China); Zheng, Yuhui, E-mail: yhzheng78@scnu.edu.cn [School of Chemistry and Environment, South China Normal University, Guangzhou 510006 (China); Wang, Qianming, E-mail: qmwang@scnu.edu.cn [Key Laboratory of Theoretical Chemistry of Environment, Ministry of Education, School of Chemistry and Environment, South China Normal University, Guangzhou 510006 (China); School of Chemistry and Environment, South China Normal University, Guangzhou 510006 (China); Guangzhou Key Laboratory of Materials for Energy Conversion and Storage, Guangzhou 510006 (China); State Key Laboratory of Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082 (China); Zeng, Zhi [School of Chemistry and Environment, South China Normal University, Guangzhou 510006 (China); Zhang, Cheng Cheng [Departments of Physiology and Developmental Biology, University of Texas Southwestern Medical Center, Dallas (United States)

    2014-08-11

    Graphical abstract: A new type of Eu(III) ofloxacin complex as the fluorescent pH indicator has been reported. Compared to pure ligand, the complex offers more distinguished color changes (green–red–blue) derived from both lanthanide line emissions and the secondary ionization steps of ofloxacin. - Highlights: • The pH probe offers a very wide working range in water (pH 1–14). • The emission changes have multiple colors. • Long-lived excited state lifetimes of Eu(III) has been used. • Two types of pH sensitive hydrogels were fabricated. - Abstract: A new type of Eu(III) ofloxacin complex as the fluorescent pH indicator has been presented. Compared to pure ligand, the complex offers more distinguished color changes (green–red–blue) derived from both lanthanide line emissions and the secondary ionization steps of ofloxacin. During the concentration dependence experiments, the photoluminescence studies on the complex showed that the excitation of this pH probe can occur at a very long wavelength which extends to visible range (Ex = 427 nm). Furthermore, the functional complex was successfully incorporated into soft networks and two novel luminescent hydrogels (rod and film) were fabricated. The soft materials also exhibited specific responses towards the pH variation. Finally, the onion cell-stain experiments were carried out to further confirm the validity of pH dependence and the results support the idea that the material will be suitable for monitoring biological samples in the future.

  4. Probing endocytosis from the enterocyte brush border using fluorescent lipophilic dyes

    DEFF Research Database (Denmark)

    Danielsen, E Michael

    2015-01-01

    The small intestinal brush border is a specialized cell membrane that needs to withstand the solubilizing effect of bile salts during assimilation of dietary nutrients and to achieve detergent resistance; it is highly enriched in glycolipids organized in lipid raft microdomains. In the present work......-toluenesulfonate), and CellMask Orange plasma membrane stain were used to study endocytosis from the enterocyte brush border of organ-cultured porcine mucosal explants. All the dyes readily incorporated into the brush border but were not detectably endocytosed by 5 min, indicating a slow uptake compared with other cell types...

  5. Multimodality Imaging Probe for Positron Emission Tomography and Fluorescence Imaging Studies

    Directory of Open Access Journals (Sweden)

    Suresh K. Pandey

    2014-05-01

    Full Text Available Our goal is to develop multimodality imaging agents for use in cell tracking studies by positron emission tomography (PET and optical imaging (OI. For this purpose, bovine serum albumin (BSA was complexed with biotin (histologic studies, 5(6- carboxyfluorescein, succinimidyl ester (FAM SE (OI studies, and diethylenetriamine pentaacetic acid (DTPA for chelating gallium 68 (PET studies. For synthesis of BSA-biotin-FAM-DTPA, BSA was coupled to (+-biotin N-hydroxysuccinimide ester (biotin-NHSI. BSA- biotin was treated with DTPA-anhydride and biotin-BSA-DTPA was reacted with FAM. The biotin-BSA-DTPA-FAM was reacted with gallium chloride 3 to 5 mCi eluted from the generator using 0.1 N HCl and was passed through basic resin (AG 11 A8 and 150 mCi (100 μL, pH 7–8 was incubated with 0.1 mg of FAM conjugate (100 μL at room temperature for 15 minutes to give 66Ga-BSA-biotin-DTPA-FAM. A shaved C57 black mouse was injected with FAM conjugate (50 μL at one flank and FAM-68Ga (50 μL, 30 mCi at the other. Immediately after injection, the mouse was placed in a fluorescence imaging system (Kodak In-Vivo F, Bruker Biospin Co., Woodbridge, CT and imaged (Λex: 465 nm, Λem: 535 nm, time: 8 seconds, Xenon Light Source, Kodak. The same mouse was then placed under an Inveon microPET scanner (Siemens Medical Solutions, Knoxville, TN injected (intravenously with 25 μCi of 18F and after a half-hour (to allow sufficient bone uptake was imaged for 30 minutes. Molecular weight determined using matrix-associated laser desorption ionization (MALDI for the BSA sample was 66,485 Da and for biotin-BSA was 67,116 Da, indicating two biotin moieties per BSA molecule; for biotin-BSA-DTPA was 81,584 Da, indicating an average of 30 DTPA moieties per BSA molecule; and for FAM conjugate was 82,383 Da, indicating an average of 1.7 fluorescent moieties per BSA molecule. Fluorescence imaging clearly showed localization of FAM conjugate and FAM-68Ga at respective flanks of the mouse

  6. InP/ZnS Nanocrystals as Fluorescent Probes for the Detection of ATP

    OpenAIRE

    Massadeh, Salam; Nann, Thomas

    2014-01-01

    This article reports on a study on fluorescence adenosine triphosphate (ATP) detection by InP/ZnS quantum dots (QDs). We present a spectroscopic analysis displaying the effect of enzymatic reactions of glucose oxidase (GOX) and hexokinase (HEX) on the InP/ZnS quantum dots at physiological pH. The InP/ZnS quantum dots act as glucose sensors in the presence of GOX, Glu and ATP, and their luminescence quenches during the release of hydrogen peroxide from the reaction. However, in the presence of...

  7. Fluorescently tagged iminoalditol glycosidase inhibitors as novel biological probes and diagnostics

    DEFF Research Database (Denmark)

    Lundt, Inge; Steiner, Andreas J.; Stütz, Arnold E.

    2006-01-01

    1,5-Dideoxy-1,5-imino-D-glucitol, the corresponding D-manno and L-ido epimers as well as the powerful b-glucosidase inhibitor isofagomine were N-alkylated with di-, tri-, as well as tetraethylene glycol derived straight chain spacer arms by a set of simple standard procedures. The terminal functi...... functional groups of the spacer arms, primary amines, were employed to introduce fluorescent dansyl moieties. Resulting derivatives showed glycosidase inhibitory activities comparable to those of the parent compounds....

  8. Determination of sertraline in pharmaceutical and biological samples using 1, 10-phenanthroline-terbium probe and silver nanoparticles enhanced fluorescence

    Energy Technology Data Exchange (ETDEWEB)

    Lotfi, Ali, E-mail: alilotfi67@gmail.com [Young Researchers and Elite Club, Tabriz Branch, Islamic Azad University, Tabriz (Iran, Islamic Republic of); Manzoori, Jamshid L. [Department of Chemistry, Tabriz Branch, Islamic Azad University, Tabriz (Iran, Islamic Republic of); Mohagheghi, Arash [Clinical Psychiatry Research Center, Tabriz University of Medical Sciences, Tabriz (Iran, Islamic Republic of)

    2017-05-15

    Sertraline is an antidepressant widely prescribed for major depressive disorders. In this contribution we report a novel, rapid and sensitive spectrofluorimetric technique, developed and validated for the determination of sertraline in pharmaceutical, human urine and human plasma samples, based on the fluorescence enhancement of the sertraline by 1, 10-phenanthroline-terbium probe with Ag nanoparticles (AgNPs). The effect of pH, buffer concentration, the order of addition of reagents, terbium and 1, 10-phenanthroline concentrations, and concentration of Ag nanoparticles (AgNPs) as well as reaction time on the fluorescence intensity were investigated and the optimum conditions were determined. The linear range for determination of sertraline was obtained as 0.001–3 mg L{sup −1}. The limit of detection (b+3s) and the limit of quantification was calculated as 2.9×10{sup −4} mg L{sup −1} and 9.8×10{sup −4} mg L{sup −1}, respectively. The interference effects of common excipients found in pharmaceutical preparations were studied. The presented technique was used to determine the sertraline in pharmaceutical samples, human urine and plasma as real samples. The presented method was indicated a comparable results with the standard analytical techniques for sertraline. Good linearity, reproducibility, recovery and limit of detection have made this method suitable for determination of sertraline in various types of samples.

  9. Asymmetric rotor-like probes to polarized fluorescence study of the macroscopically oriented uniaxial media: Model parameters recognition

    International Nuclear Information System (INIS)

    Buczkowski, M.; Fisz, J.J.

    2008-01-01

    In this paper the possibility of the numerical data modelling in the case of angle- and time-resolved fluorescence spectroscopy is investigated. The asymmetric fluorescence probes are assumed to undergo the restricted rotational diffusion in a hosting medium. This process is described quantitatively by the diffusion tensor and the aligning potential. The evolution of the system is expressed in terms of the Smoluchowski equation with an appropriate time-developing operator. A matrix representation of this operator is calculated, then symmetrized and diagonalized. The resulting propagator is used to generate the synthetic noisy data set that imitates results of experimental measurements. The data set serves as a groundwork to the χ 2 optimization, performed by the genetic algorithm followed by the gradient search, in order to recover model parameters, which are diagonal elements of the diffusion tensor, aligning potential expansion coefficients and directions of the electronic dipole moments. This whole procedure properly identifies model parameters, showing that the outlined formalism should be taken in the account in the case of analysing real experimental data

  10. Efficient and Scalable Synthesis of 4-Carboxy-Pennsylvania Green Methyl Ester: A Hydrophobic Building Block for Fluorescent Molecular Probes.

    Science.gov (United States)

    Woydziak, Zachary R; Fu, Liqiang; Peterson, Blake R

    2014-01-01

    Fluorinated fluorophores are valuable tools for studies of biological systems. However, amine-reactive single-isomer derivatives of these compounds are often very expensive. To provide an inexpensive alternative, we report a practical synthesis of 4-carboxy-Pennsylvania Green methyl ester. Derivatives of this hydrophobic fluorinated fluorophore, a hybrid of the dyes Oregon Green and Tokyo Green, are often cell permeable, enabling labeling of intracellular targets and components. Moreover, the low pKa of Pennsylvania Green (4.8) confers bright fluorescence in acidic cellular compartments such as endosomes, enhancing its utility for chemical biology investigations. To improve access to the key intermediate 2,7-difluoro-3,6-dihydroxyxanthen-9-one, we subjected bis-(2,4,5-trifluorophenyl)methanone to iterative nucleophilic aromatic substitution by hydroxide on scales of > 40 g. This intermediate was used to prepare over 15 grams of pure 4-carboxy-Pennsylvania Green methyl ester in 28% overall yield without requiring chromatography. This compound can be converted into the amine reactive N -hydroxysuccinimidyl ester in essentially quantitative yield for the synthesis of a wide variety of fluorescent molecular probes.

  11. Determination of sertraline in pharmaceutical and biological samples using 1, 10-phenanthroline-terbium probe and silver nanoparticles enhanced fluorescence

    International Nuclear Information System (INIS)

    Lotfi, Ali; Manzoori, Jamshid L.; Mohagheghi, Arash

    2017-01-01

    Sertraline is an antidepressant widely prescribed for major depressive disorders. In this contribution we report a novel, rapid and sensitive spectrofluorimetric technique, developed and validated for the determination of sertraline in pharmaceutical, human urine and human plasma samples, based on the fluorescence enhancement of the sertraline by 1, 10-phenanthroline-terbium probe with Ag nanoparticles (AgNPs). The effect of pH, buffer concentration, the order of addition of reagents, terbium and 1, 10-phenanthroline concentrations, and concentration of Ag nanoparticles (AgNPs) as well as reaction time on the fluorescence intensity were investigated and the optimum conditions were determined. The linear range for determination of sertraline was obtained as 0.001–3 mg L −1 . The limit of detection (b+3s) and the limit of quantification was calculated as 2.9×10 −4 mg L −1 and 9.8×10 −4 mg L −1 , respectively. The interference effects of common excipients found in pharmaceutical preparations were studied. The presented technique was used to determine the sertraline in pharmaceutical samples, human urine and plasma as real samples. The presented method was indicated a comparable results with the standard analytical techniques for sertraline. Good linearity, reproducibility, recovery and limit of detection have made this method suitable for determination of sertraline in various types of samples.

  12. Curcumin as fluorescent probe for directly monitoring in vitro uptake of curcumin combined paclitaxel loaded PLA-TPGS nanoparticles

    Science.gov (United States)

    Nguyen, Hoai Nam; Thu Ha, Phuong; Sao Nguyen, Anh; Nguyen, Dac Tu; Doan Do, Hai; Nguyen Thi, Quy; Nhung Hoang Thi, My

    2016-06-01

    Theranostics, which is the combination of both therapeutic and diagnostic capacities in one dose, is a promising tool for both clinical application and research. Although there are many chromophores available for optical imaging, their applications are limited due to the photobleaching property or intrinsic toxicity. Curcumin, a natural compound extracted from the rhizome of curcuma longa, is well known thanks to its bio-pharmaceutical activities and strong fluorescence as biocompatible probe for bio-imaging. In this study, we aimed to fabricate a system with dual functions: diagnostic and therapeutic, based on poly(lactide)-tocopheryl polyethylene glycol succinate (PLA-TPGS) micelles co-loaded curcumin (Cur) and paclitaxel (PTX). Two kinds of curcumin nanoparticle (NP) were fabricated and characterized by Fourier transform infrared spectroscopy, field emission scanning electron microscopy and dynamic light scattering methods. The cellular uptake and fluorescent activities of curcumin in these systems were also tested by bioassay studies, and were compared with paclitaxe-oregon. The results showed that (Cur + PTX)-PLA-TPGS NPs is a potential system for cancer theranostics.

  13. Curcumin as fluorescent probe for directly monitoring in vitro uptake of curcumin combined paclitaxel loaded PLA-TPGS nanoparticles

    International Nuclear Information System (INIS)

    Nguyen, Hoai Nam; Ha, Phuong Thu; Do, Hai Doan; Nguyen, Anh Sao; Nguyen, Dac Tu; Thi, Quy Nguyen; Thi, My Nhung Hoang

    2016-01-01

    Theranostics, which is the combination of both therapeutic and diagnostic capacities in one dose, is a promising tool for both clinical application and research. Although there are many chromophores available for optical imaging, their applications are limited due to the photobleaching property or intrinsic toxicity. Curcumin, a natural compound extracted from the rhizome of curcuma longa, is well known thanks to its bio-pharmaceutical activities and strong fluorescence as biocompatible probe for bio-imaging. In this study, we aimed to fabricate a system with dual functions: diagnostic and therapeutic, based on poly(lactide)-tocopheryl polyethylene glycol succinate (PLA-TPGS) micelles co-loaded curcumin (Cur) and paclitaxel (PTX). Two kinds of curcumin nanoparticle (NP) were fabricated and characterized by Fourier transform infrared spectroscopy, field emission scanning electron microscopy and dynamic light scattering methods. The cellular uptake and fluorescent activities of curcumin in these systems were also tested by bioassay studies, and were compared with paclitaxe-oregon. The results showed that (Cur + PTX)-PLA-TPGS NPs is a potential system for cancer theranostics. (paper)

  14. Isostructural fluorescent and radioactive probes for monitoring neural stem and progenitor cell transplants

    Energy Technology Data Exchange (ETDEWEB)

    Schaffer, Paul [McMaster Nuclear Reactor, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada); Gleave, Jacqueline A. [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, L8N 3Z5 (Canada); Lemon, Jennifer A. [Department of Medical Physics and Applied Radiation Sciences, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada); Reid, Leslie C. [Department of Chemistry, McMaster University, Hamilton, Ontario, L8S 4M1 (Canada); Pacey, Laura K.K. [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, L8N 3Z5 (Canada); Farncombe, Troy H. [Department of Nuclear Medicine, Hamilton Health Sciences, Hamilton, Ontario, L8N 3Z5 (Canada); Boreham, Douglas R. [Department of Medical Physics and Applied Radiation Sciences, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada); Zubieta, Jon [Department of Chemistry, Syracuse University, Syracuse, NY 13244-4100 (United States); Babich, John W. [Molecular Insight Pharmaceuticals Inc., Cambridge, MA 02142 (United States); Doering, Laurie C. [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, L8N 3Z5 (Canada); Valliant, John F. [Department of Medical Physics and Applied Radiation Sciences, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada); Department of Chemistry, McMaster University, Hamilton, Ontario, L8S 4M1 (Canada)], E-mail: valliant@mcmaster.ca

    2008-02-15

    A construct for tagging neurospheres and monitoring cell transplantations was developed using a new technology for producing luminescent and radiolabeled probes that have identical structures. The HIV1-Tat basic domain derivatives NAcGRKKRRQRRR(SAACQ)G (SAACQ-1) and [NAcGRKKRRQRRR(Re(CO){sub 3}SAACQ)G]{sup +} (ReSAACQ-1) were prepared in excellent yields using the single amino acid chelate-quinoline (SAACQ) ligand and its Re(I) complex and conventional automated peptide synthesis methods. The distribution of the luminescent Re probe, using epifluorescence microscopy, showed that it localized primarily in the cell nucleus with a significant degree of association on the nuclear envelope. A smaller amount was found to be dispersed in the cytoplasm. The {sup 99m}Tc analogue was then prepared in 43{+-}7% (n=12) yield and very high effective specific activity. Following incubation, average uptake of the probe in neurospheres ranged between 10 and 20 Bq/cell. As determined by colorimetric assays, viability for cells labeled with high effective specific activity {sup 99m}TcSAACQ-1 was 97{+-}4% at 2 h postlabeling and 85{+-}25% at 24 h postlabeling for incubation activities ranging from 245 to 8900 Bq/cell. DNA analysis showed that at these levels, there was no significant difference between the extent of DNA damage in the treated cells versus control cells. A series of preliminary SPECT/CT studies of transplants in mice were performed, which showed that the strategy is convenient and feasible and that it is possible to routinely assess procedures noninvasively and determine the number of cells transplanted.

  15. Towards Fluorescence In Vivo Hybridization (FIVH Detection of H. pylori in Gastric Mucosa Using Advanced LNA Probes.

    Directory of Open Access Journals (Sweden)

    Sílvia Fontenete

    Full Text Available In recent years, there have been several attempts to improve the diagnosis of infection caused by Helicobacter pylori. Fluorescence in situ hybridization (FISH is a commonly used technique to detect H. pylori infection but it requires biopsies from the stomach. Thus, the development of an in vivo FISH-based method (FIVH that directly detects and allows the visualization of the bacterium within the human body would significantly reduce the time of analysis, allowing the diagnosis to be performed during endoscopy. In a previous study we designed and synthesized a phosphorothioate locked nucleic acid (LNA/ 2' O-methyl RNA (2'OMe probe using standard phosphoramidite chemistry and FISH hybridization was then successfully performed both on adhered and suspended bacteria at 37°C. In this work we simplified, shortened and adapted FISH to work at gastric pH values, meaning that the hybridization step now takes only 30 minutes and, in addition to the buffer, uses only urea and probe at non-toxic concentrations. Importantly, the sensitivity and specificity of the FISH method was maintained in the range of conditions tested, even at low stringency conditions (e.g., low pH. In conclusion, this methodology is a promising approach that might be used in vivo in the future in combination with a confocal laser endomicroscope for H. pylori visualization.

  16. Towards Fluorescence In Vivo Hybridization (FIVH) Detection of H. pylori in Gastric Mucosa Using Advanced LNA Probes

    Science.gov (United States)

    Fontenete, Sílvia; Leite, Marina; Guimarães, Nuno; Madureira, Pedro; Ferreira, Rui Manuel; Figueiredo, Céu; Wengel, Jesper; Azevedo, Nuno Filipe

    2015-01-01

    In recent years, there have been several attempts to improve the diagnosis of infection caused by Helicobacter pylori. Fluorescence in situ hybridization (FISH) is a commonly used technique to detect H. pylori infection but it requires biopsies from the stomach. Thus, the development of an in vivo FISH-based method (FIVH) that directly detects and allows the visualization of the bacterium within the human body would significantly reduce the time of analysis, allowing the diagnosis to be performed during endoscopy. In a previous study we designed and synthesized a phosphorothioate locked nucleic acid (LNA)/ 2’ O-methyl RNA (2’OMe) probe using standard phosphoramidite chemistry and FISH hybridization was then successfully performed both on adhered and suspended bacteria at 37°C. In this work we simplified, shortened and adapted FISH to work at gastric pH values, meaning that the hybridization step now takes only 30 minutes and, in addition to the buffer, uses only urea and probe at non-toxic concentrations. Importantly, the sensitivity and specificity of the FISH method was maintained in the range of conditions tested, even at low stringency conditions (e.g., low pH). In conclusion, this methodology is a promising approach that might be used in vivo in the future in combination with a confocal laser endomicroscope for H. pylori visualization. PMID:25915865

  17. Rapid Identification of Staphylococcus aureus Directly from Blood Cultures by Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes

    Science.gov (United States)

    Oliveira, Kenneth; Procop, Gary W.; Wilson, Deborah; Coull, James; Stender, Henrik

    2002-01-01

    A new fluorescence in situ hybridization (FISH) method with peptide nucleic acid (PNA) probes for identification of Staphylococcus aureus directly from positive blood culture bottles that contain gram-positive cocci in clusters (GPCC) is described. The test (the S. aureus PNA FISH assay) is based on a fluorescein-labeled PNA probe that targets a species-specific sequence of the 16S rRNA of S. aureus. Evaluations with 17 reference strains and 48 clinical isolates, including methicillin-resistant and methicillin-susceptible S. aureus species, coagulase-negative Staphylococcus species, and other clinically relevant and phylogenetically related bacteria and yeast species, showed that the assay had 100% sensitivity and 96% specificity. Clinical trials with 87 blood cultures positive for GPCC correctly identified 36 of 37 (97%) of the S. aureus-positive cultures identified by standard microbiological methods. The positive and negative predictive values were 100 and 98%, respectively. It is concluded that this rapid method (2.5 h) for identification of S. aureus directly from blood culture bottles that contain GPCC offers important information for optimal antibiotic therapy. PMID:11773123

  18. Naphthalene-based fluorescent probes for glutathione and their applications in living cells and patients with sepsis

    Science.gov (United States)

    Li, Jun; Kwon, Younghee; Chung, Kyung Soo; Lim, Chang Su; Lee, Dayoung; Yue, Yongkang; Yoon, Jisoo; Kim, Gyoungmi; Nam, Sang-Jip; Chung, Youn Wook; Kim, Hwan Myung; Yin, Caixia; Ryu, Ji-Hwan; Yoon, Juyoung

    2018-01-01

    Rationale: Among the biothiols-related diseases, sepsis is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection and can result in severe oxidative stress and damage to multiple organs. In this study, we aimed to develop a fluorescence chemosensor that can both detect GSH and further predict sepsis. Methods: In this study, two new naphthalene dialdehyde compounds containing different functional groups were synthesized, and the sensing abilities of these compounds towards biothiols and its applications for prediction of sepsis were investigated. Results: Our study revealed that the newly developed probe 6-methoxynaphthalene-2, 3-dicarbaldehyde (MNDA) has two-photon is capable of detecting GSH in live cells with two-photon microscopy (TPM) under the excitation at a wavelength of 900 nm. Furthermore, two GSH detection probes naphthalene-2,3-dicarboxaldehyde (NDA) and 6-fluoronaphthalene-2,3-dicarbaldehyde (FNDA) not only can detect GSH in living cells, but also showed clinical significance for the diagnosis and prediction of mortality in patients with sepsis. Conclusions: These results open up a promising direction for further medical diagnostic techniques. PMID:29507630

  19. Small molecule probes finely differentiate between various ds- and ss-DNA and RNA by fluorescence, CD and NMR response

    Energy Technology Data Exchange (ETDEWEB)

    Crnolatac, Ivo; Rogan, Iva; Majić, Boris; Tomić, Sanja [Division of Organic Chemistry and Biochemistry, Division of Physical Chemistry, Ruđer Bošković Institute, P.O. Box 180, 10002 Zagreb (Croatia); Deligeorgiev, Todor [Faculty of Chemistry and Pharmacy, University of Sofia (Bulgaria); Horvat, Gordan [Department of Physical Chemistry, Faculty of Science/Chemistry, Horvatovac 102A, HR-10000 Zagreb (Croatia); Makuc, Damjan; Plavec, Janez [Slovenian NMR Centre, National Institute of Chemistry, Hajdrihova 19, Ljubljana (Slovenia); EN-FIST Centre of Excellence, Trg Osvobodilne Fronte 13, Ljubljana (Slovenia); Pescitelli, Gennaro [Department of Chemistry, University of Pisa, Via Moruzzi 13, Pisa (Italy); Piantanida, Ivo, E-mail: pianta@irb.hr [Division of Organic Chemistry and Biochemistry, Division of Physical Chemistry, Ruđer Bošković Institute, P.O. Box 180, 10002 Zagreb (Croatia)

    2016-10-12

    Two small molecules showed intriguing properties of analytical multipurpose probes, whereby one chromophore gives different signal for many different DNA/RNA by application of several highly sensitive spectroscopic methods. Dyes revealed pronounced fluorescence ratiomeric differentiation between ds-AU-RNA, AT-DNA and GC-DNA in approximate order 10:8:1. Particularly interesting, dyes showed specific fluorimetric response for poly rA even at 10-fold excess of any other ss-RNA, and moreover such emission selectivity is preserved in multicomponent ss-RNA mixtures. The dyes also showed specific chiral recognition of poly rU in respect to the other ss-RNA by induced CD (ICD) pattern in visible range (400–500 nm), which was attributed to the dye-side-chain contribution to binding (confirmed by absence of any ICD band for reference compound lacking side-chain). Most intriguingly, minor difference in the side-chain attached to dye chromophore resulted in opposite sign of dye-ICD pattern, whereby differences in NMR NOESY contacts and proton chemical shifts between two dye/oligo rU complexes combined with MD simulations and CD calculations attributed observed bisignate ICD to the dimeric dye aggregate within oligo rU. - Highlights: • Novel dyes emit fluorescence only for poly rA even at high excess of all other ss-RNA. • Fluorescence response for AT-DNA is 8 times stronger than for GC-DNA. • Florescence induced by ds-RNA is 20% stronger that emission induced by ds-DNA. • Intrinsically non-chiral, dyes show strong and characteristic ICD response for poly rU.

  20. Zwitterionic Silane Copolymer for Ultra-Stable and Bright Biomolecular Probes Based on Fluorescent Quantum Dot Nanoclusters.

    Science.gov (United States)

    Dembele, Fatimata; Tasso, Mariana; Trapiella-Alfonso, Laura; Xu, Xiangzhen; Hanafi, Mohamed; Lequeux, Nicolas; Pons, Thomas

    2017-05-31

    Fluorescent semiconductor quantum dots (QDs) exhibit several unique properties that make them suitable candidates for biomolecular sensing, including high brightness, photostability, broad excitation, and narrow emission spectra. Assembling these QDs into robust and functionalizable nanosized clusters (QD-NSCs) can provide fluorescent probes that are several orders of magnitude brighter than individual QDs, thus allowing an even greater sensitivity of detection with simplified instrumentation. However, the formation of compact, antifouling, functionalizable, and stable QD-NSCs remains a challenging task, especially for a use at ultralow concentrations for single-molecule detection. Here, we describe the development of fluorescent QD-NSCs envisioned as a tool for fast and sensitive biomolecular recognition. First, QDs were assembled into very compact 100-150 nm diameter spherical aggregates; the final QD-NSCs were obtained by growing a cross-linked silica shell around these aggregates. Hydrolytic stability in several concentration and pH conditions is a key requirement for a potential and efficient single-molecule detection tool. However, the hydrolysis of Si-O-Si bonds leads to desorption of monosilane-based surface groups at very low silica concentrations or in a slightly basic medium. Thus, we designed a novel multidentate copolymer composed of multiple silane as well as zwitterionic monomers. Coating silica beads with this multidentate copolymer provided a robust surface chemistry that was demonstrated to be stable against hydrolysis, even at low concentrations. Copolymer-coated silica beads also showed low fouling properties and high colloidal stability in saline solutions. Furthermore, incorporation of additional azido-monomers enabled easy functionalization of QD-NSCs using copper-free bio-orthogonal cyclooctyne-azide click chemistry, as demonstrated by a biotin-streptavidin affinity test.

  1. Fluorescent probe studies of polarity and solvation within room temperature ionic liquids: a review.

    Science.gov (United States)

    Pandey, Shubha; Baker, Sheila N; Pandey, Siddharth; Baker, Gary A

    2012-09-01

    Ionic liquids display an array of useful and sometimes unconventional, solvent features and have attracted considerable interest in the field of green chemistry for the potential they hold to significantly reduce environmental emissions. Some of these points have a bearing on the chemical reactivity of these systems and have also generated interest in the physical and theoretical aspects of solvation in ionic liquids. This review presents an introduction to the field of ionic liquids, followed by discussion of investigations into the solvation properties of neat ionic liquids or mixed systems including ionic liquids as a major or minor component. The ionic liquid based multicomponent systems discussed are composed of other solvents, other ionic liquids, carbon dioxide, surfactants or surfactant solutions. Although we clearly focus on fluorescence spectroscopy as a tool to illuminate ionic liquid systems, the issues discussed herein are of general relevance to discussions of polarity and solvent effects in ionic liquids. Transient solvation measurements carried out by means of time-resolved fluorescence measurements are particularly powerful for their ability to parameterize the kinetics of the solvation process in ionic liquids and are discussed as well.

  2. Design, synthesis and evaluation of a new fluorescent probe for measuring polymyxin-lipopolysaccharide binding interactions

    Science.gov (United States)

    Soon, Rachel L.; Velkov, Tony; Chiu, Francis; Thompson, Philip E.; Kancharla, Rashmi; Roberts, Kade; Larson, Ian; Nation, Roger L.; Li, Jian

    2011-01-01

    Fluorescence assays employing semi-synthetic or commercial dansyl-polymyxin B, have been widely employed to assess the affinity of polycations, including polymyxins, for bacterial cells and lipopolysaccharide (LPS). The five primary γ-amines on diaminobutyric-acid residues of polymyxin B are potentially derivatized with dansyl-cholride. Mass spectrometric analysis of the commercial product revealed a complex mixture of di- or tetra- dansyl-substituted polymyxin B. We synthesized a mono-substituted fluorescent derivative, dansyl[Lys]1polymyxinB3. The affinity of polymyxin for purified Gram-negative LPS, and whole bacterial cells was investigated. The affinity of dansyl[Lys]1polymyxinB3 for LPS was comparable to polymyxin B and colistin, and considerably greater (kd dansyl[Lys]1polymyxinB3 to LPS, attributed to electrostatic interactions. The hydrophobic dansyl moiety imparted a greater entropic contribution to the dansyl[Lys]1polymyxinB3-LPS reaction. Molecular modeling revealed a loss of electrostatic contact within the dansyl[Lys]1polymyxinB3-LPS complex due to steric hindrance from the dansyl[Lys]1 fluorophore; this corresponded with diminished antibacterial activity (MIC ≥ 16 μg/mL). Dansyl[Lys]1polymyxinB3 may prove useful as a screening tool for drug development. PMID:21050838

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

    Science.gov (United States)

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

    2015-08-01

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

  4. Photostable bipolar fluorescent probe for video tracking plasma membranes related cellular processes.

    Science.gov (United States)

    Zhang, Xinfu; Wang, Chao; Jin, Liji; Han, Zhuo; Xiao, Yi

    2014-08-13

    Plasma membranes can sense the stimulations and transmit the signals from extracellular environment and then make further responses through changes in locations, shapes or morphologies. Common fluorescent membrane markers are not well suited for long time tracking due to their shorter retention time inside plasma membranes and/or their lower photostability. To this end, we develop a new bipolar marker, Mem-SQAC, which can stably insert into plasma membranes of different cells and exhibits a long retention time over 30 min. Mem-SQAC also inherits excellent photostability from the BODIPY dye family. Large two-photon absorption cross sections and long wavelength fluorescence emissions further enhance the competitiveness of Mem-SQAC as a membrane marker. By using Mem-SQAC, significant morphological changes of plasma membranes have been monitored during heavy metal poisoning and drug induced apoptosis of MCF-7 cells; the change tendencies are so distinctly different from each other that they can be used as indicators to distinguish different cell injuries. Further on, the complete processes of endocytosis toward Staphylococcus aureus and Escherichia coli by RAW 264.7 cells have been dynamically tracked. It is discovered that plasma membranes take quite different actions in response to the two bacteria, information unavailable in previous research reports.

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

    Science.gov (United States)

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

    2014-11-01

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

  6. Development and use of fluorescent 16S rRNA-targeted probes for the specific detection of Methylophaga species by in situ hybridization in marine sediments.

    Science.gov (United States)

    Janvier, Monique; Regnault, Béatrice; Grimont, Patrick

    2003-09-01

    Methylotrophic bacteria are widespread in nature. They may play an important role in the cycling of carbon and in the metabolism of dimethylsulfide in a marine environment. Bacteria belonging to the genus Methylophaga are a unique group of aerobic, halophilic, non-methane-utilizing methylotrophs. Two 16S rRNA-targeted oligonucleotide probes were developed for the specific detection of Methylophaga species, marine methylobacteria, by fluorescence in situ hybridization. Probe MPH-730 was highly specific for all members of the genus Methylophaga while probe MPHm-994 targeted exclusively M. marina. The application of these probes were demonstrated by the detection of Methylophaga species in enrichment cultures from various marine sediments. All isolates recovered were visualized by using the genus specific probe MPH-730. The results were confirmed by 16S rDNA sequencing which demonstrated that all selected isolates belong to Methylophaga. Five isolates could be detected by the M. marina-specific probe MPHm-994 and were confirmed by rRNA gene restriction pattern (ribotyping). With the development of these specific probes, fluorescence in situ hybridization shows that the genus Methylophaga is widespread in marine samples.

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

    Science.gov (United States)

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

    2017-10-02

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

  8. Dual lanthanide-doped complexes: the development of a time-resolved ratiometric fluorescent probe for anthrax biomarker and a paper-based visual sensor.

    Science.gov (United States)

    Wang, Qi-Xian; Xue, Shi-Fan; Chen, Zi-Han; Ma, Shi-Hui; Zhang, Shengqiang; Shi, Guoyue; Zhang, Min

    2017-08-15

    In this work, a novel time-resolved ratiometric fluorescent probe based on dual lanthanide (Tb: terbium, and Eu: europium)-doped complexes (Tb/DPA@SiO 2 -Eu/GMP) has been designed for detecting anthrax biomarker (dipicolinic acid, DPA), a unique and major component of anthrax spores. In such complexes-based probe, Tb/DPA@SiO 2 can serve as a stable reference signal with green fluorescence and Eu/GMP act as a sensitive response signal with red fluorescence for ratiometric fluorescent sensing DPA. Additionally, the probe exhibits long fluorescence lifetime, which can significantly reduce the autofluorescence interferences from biological samples by using time-resolved fluorescence measurement. More significantly, a paper-based visual sensor for DPA has been devised by using filter paper embedded with Tb/DPA@SiO 2 -Eu/GMP, and we have proved its utility for fluorescent detection of DPA, in which only a handheld UV lamp is used. In the presence of DPA, the paper-based visual sensor, illuminated by a handheld UV lamp, would result in an obvious fluorescence color change from green to red, which can be easily observed with naked eyes. The paper-based visual sensor is stable, portable, disposable, cost-effective and easy-to-use. The feasibility of using a smartphone with easy-to-access color-scanning APP as the detection platform for quantitative scanometric assays has been also demonstrated by coupled with our proposed paper-based visual sensor. This work unveils an effective method for accurate, sensitive and selective monitoring anthrax biomarker with backgroud-free and self-calibrating properties. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Study of the Combined Effect of Ibuprofen and Cholesterol on the Microviscosity and Ordering of Model Lipid Membranes by Timeresolved Measurement of Fluorescence Anisotropy Decay

    Science.gov (United States)

    Yefimova, S. L.; Tkacheva, T. N.; Kasian, N. A.

    2017-05-01

    The timeresolved fluorescence anisotropy decay of perylene incorporated into the lipid Ladipalmitoylphosphatidylch oline (DPPC) membrane has been studied to evaluate the membranotropic action of the nonsteroidal antiinflammatory drug, ibuprofen, and the combined effect of ibuprofen and cholesterol. The rotation correlation times (φ) and limiting anisotropy (r∞ ) permit an independent estimation of the effects of these additives on the microviscosity and ordering of model lipid membranes in different phase states. Ibuprofen was shown to cause a significant decrease in the DPPC membrane microviscosity in the gel phase with hardly any effect on the liquidcrystal phase. However, in both phases, ibuprofen diminishes the ordering of the lipid hydrophobic chains. A marked additive effect is noted when ibuprofen is embedded in the liquid membrane enriched with cholesterol, which manifests itself in substantial fluidization and disordering or the liquid membrane by the action of the components on the lipid membrane. Ibuprofen in the liquidcrystal phase causes leveling of the fluidizing and ordering effects of cholesterol.

  10. Study of the Conformational State of Non-Cross-Linked and Cross-Linked Poly(alkylmethyldiallylammonium chlorides) in Aqueous Solution by Fluorescence Probing

    NARCIS (Netherlands)

    Wang, Guang-Jia; Engberts, Jan B.F.N.

    The aggregation behaviour of novel non-cross-linked and cross-linked poly(alkylmethyldiallylammonium chlorides) in aqueous solutions has been investigated by fluorescence spectroscopy using pyrene as a probe. These copolymers were found to exhibit similar aggregate properties as the corresponding

  11. Reaction-based small-molecule fluorescent probes for dynamic detection of ROS and transient redox changes in living cells and small animals.

    Science.gov (United States)

    Lü, Rui

    2017-09-01

    Dynamic detection of transient redox changes in living cells and animals has broad implications for human health and disease diagnosis, because intracellular redox homeostasis regulated by reactive oxygen species (ROS) plays important role in cell functions, normal physiological functions and some serious human diseases (e.g., cancer, Alzheimer's disease, diabetes, etc.) usually have close relationship with the intracellular redox status. Small-molecule ROS-responsive fluorescent probes can act as powerful tools for dynamic detection of ROS and redox changes in living cells and animals through fluorescence imaging techniques; and great advances have been achieved recently in the design and synthesis of small-molecule ROS-responsive fluorescent probes. This article highlights up-to-date achievements in designing and using the reaction-based small-molecule fluorescent probes (with high sensitivity and selectivity to ROS and redox cycles) in the dynamic detection of ROS and transient redox changes in living cells and animals through fluorescence imaging. Copyright © 2017. Published by Elsevier Ltd.

  12. Real-Time Tracking the Synthesis and Degradation of Albumin in Complex Biological Systems with a near-Infrared Fluorescent Probe.

    Science.gov (United States)

    Jin, Qiang; Feng, Lei; Zhang, Shui-Jun; Wang, Dan-Dan; Wang, Fang-Jun; Zhang, Yi; Cui, Jing-Nan; Guo, Wen-Zhi; Ge, Guang-Bo; Yang, Ling

    2017-09-19

    In this study, a novel fluorescent detection system for biological sensing of human albumin (HA) was developed on the basis of the pseudoesterase activity and substrate preference of HA. The designed near-infrared (NIR) fluorescent probe (DDAP) could be effectively hydrolyzed by HA, accompanied by significant changes in both color and