Molecules for Fluorescence Detection of Specific Chemicals

Science.gov (United States)

Fedor, Steve

2008-01-01

A family of fluorescent dye molecules has been developed for use in on-off fluorescence detection of specific chemicals. By themselves, these molecules do not fluoresce. However, when exposed to certain chemical analytes in liquid or vapor forms, they do fluoresce (see figure). These compounds are amenable to fixation on or in a variety of substrates for use in fluorescence-based detection devices: they can be chemically modified to anchor them to porous or non-porous solid supports or can be incorporated into polymer films. Potential applications for these compounds include detection of chemical warfare agents, sensing of acidity or alkalinity, and fluorescent tagging of proteins in pharmaceutical research and development. These molecules could also be exploited for use as two-photon materials for photodynamic therapy in the treatment of certain cancers and other diseases. A molecule in this family consists of a fluorescent core (such as an anthracene or pyrene) attached to two end groups that, when the dye is excited by absorption of light, transfer an electron to the core, thereby quenching the fluorescence. The end groups can be engineered so that they react chemically with certain analytes. Upon reaction, electrons on the end groups are no longer available for transfer to the core and, consequently, the fluorescence from the core is no longer quenched. The chemoselectivity of these molecules can be changed by changing the end groups. For example, aniline end groups afford a capability for sensing acids or acid halides (including those contained in chemical warfare agents). Pyridine or bipyridyl end groups would enable sensing of metal ions. Other chemicals that can be selectively detected through suitable choice of end groups include glucose and proteins. Moreover, the fluorescent cores can be changed to alter light-absorption and -emission characteristics: anthracene cores fluoresce at wavelengths around 500 nm, whereas perylene cores absorb and emit at

A brief introduction to single-molecule fluorescence methods

NARCIS (Netherlands)

Wildenberg, S.M.J.L.; Prevo, B.; Peterman, E.J.G.; Peterman, EJG; Wuite, GJL

2011-01-01

One of the more popular single-molecule approaches in biological science is single-molecule fluorescence microscopy, which is the subject of the following section of this volume. Fluorescence methods provide the sensitivity required to study biology on the single-molecule level, but they also allow

A brief introduction to single-molecule fluorescence methods

NARCIS (Netherlands)

van den Wildenberg, Siet M.J.L.; Prevo, Bram; Peterman, Erwin J.G.

2018-01-01

One of the more popular single-molecule approaches in biological science is single-molecule fluorescence microscopy, which will be the subject of the following section of this volume. Fluorescence methods provide the sensitivity required to study biology on the single-molecule level, but they also

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

International Nuclear Information System (INIS)

Jiang Dafeng; Liu Chunxia; Wang Lei; Jiang Wei

2010-01-01

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

Fluorescent scattering by molecules embedded in small particles

International Nuclear Information System (INIS)

1982-01-01

Studies are reported in these areas: double resonance in fluorescent and Raman scattering; surface enhanced Raman scattering; fluorescence by molecules embedded in small particles; fluorescence by a liquid droplet; and fluorescence by conical pits in surfaces

Fluorescence fluctuation spectroscopy (FFS)

CERN Document Server

Tetin, Sergey

2012-01-01

This new volume of Methods in Enzymology continues the legacy of this premier serial with quality chapters authored by leaders in the field. This volume covers fluorescence fluctuation spectroscopy and includes chapters on such topics as Förster resonance energy transfer (fret) with fluctuation algorithms, protein corona on nanoparticles by FCS, and FFS approaches to the study of receptors in live cells. Continues the legacy of this premier serial with quality chapters authored by leaders in the field Covers fluorescence fluctuation spectroscopy Contains chapters on such topics as Förster resonance energy transfer (fret) with fluctuation algorithms, protein corona on nanoparticles by FCS, and FFS approaches to the study of receptors in live cells.

Spectrally Resolved and Functional Super-resolution Microscopy via Ultrahigh-Throughput Single-Molecule Spectroscopy.

Science.gov (United States)

Yan, Rui; Moon, Seonah; Kenny, Samuel J; Xu, Ke

2018-03-20

As an elegant integration of the spatial and temporal dimensions of single-molecule fluorescence, single-molecule localization microscopy (SMLM) overcomes the diffraction-limited resolution barrier of optical microscopy by localizing single molecules that stochastically switch between fluorescent and dark states over time. While this type of super-resolution microscopy (SRM) technique readily achieves remarkable spatial resolutions of ∼10 nm, it typically provides no spectral information. Meanwhile, current scanning-based single-location approaches for mapping the positions and spectra of single molecules are limited by low throughput and are difficult to apply to densely labeled (bio)samples. In this Account, we summarize the rationale, design, and results of our recent efforts toward the integration of the spectral dimension of single-molecule fluorescence with SMLM to achieve spectrally resolved SMLM (SR-SMLM) and functional SRM ( f-SRM). By developing a wide-field scheme for spectral measurement and implementing single-molecule fluorescence on-off switching typical of SMLM, we first showed that in densely labeled (bio)samples it is possible to record the fluorescence spectra and positions of millions of single molecules synchronously within minutes, giving rise to ultrahigh-throughput single-molecule spectroscopy and SR-SMLM. This allowed us to first show statistically that for many dyes, single molecules of the same species exhibit near identical emission in fixed cells. This narrow distribution of emission wavelengths, which contrasts markedly with previous results at solid surfaces, allowed us to unambiguously identify single molecules of spectrally similar dyes. Crosstalk-free, multiplexed SRM was thus achieved for four dyes that were merely 10 nm apart in emission spectrum, with the three-dimensional SRM images of all four dyes being automatically aligned within one image channel. The ability to incorporate single-molecule fluorescence measurement with

Localizing internal friction along the reaction coordinate of protein folding by combining ensemble and single-molecule fluorescence spectroscopy

Science.gov (United States)

Borgia, Alessandro; Wensley, Beth G.; Soranno, Andrea; Nettels, Daniel; Borgia, Madeleine B.; Hoffmann, Armin; Pfeil, Shawn H.; Lipman, Everett A.; Clarke, Jane; Schuler, Benjamin

2012-01-01

Theory, simulations and experimental results have suggested an important role of internal friction in the kinetics of protein folding. Recent experiments on spectrin domains provided the first evidence for a pronounced contribution of internal friction in proteins that fold on the millisecond timescale. However, it has remained unclear how this contribution is distributed along the reaction and what influence it has on the folding dynamics. Here we use a combination of single-molecule Förster resonance energy transfer, nanosecond fluorescence correlation spectroscopy, microfluidic mixing and denaturant- and viscosity-dependent protein-folding kinetics to probe internal friction in the unfolded state and at the early and late transition states of slow- and fast-folding spectrin domains. We find that the internal friction affecting the folding rates of spectrin domains is highly localized to the early transition state, suggesting an important role of rather specific interactions in the rate-limiting conformational changes. PMID:23149740

Spectroscopy of Cold LiCa Molecules Formed on Helium Nanodroplets

Science.gov (United States)

2013-01-01

We report on the formation of mixed alkali–alkaline earth molecules (LiCa) on helium nanodroplets and present a comprehensive experimental and theoretical study of the ground and excited states of LiCa. Resonance enhanced multiphoton ionization time-of-flight (REMPI-TOF) spectroscopy and laser induced fluorescence (LIF) spectroscopy were used for the experimental investigation of LiCa from 15000 to 25500 cm–1. The 42Σ+ and 32Π states show a vibrational structure accompanied by distinct phonon wings, which allows us to determine molecular parameters as well as to study the interaction of the molecule with the helium droplet. Higher excited states (42Π, 52Σ+, 52Π, and 62Σ+) are not vibrationally resolved and vibronic transitions start to overlap. The experimental spectrum is well reproduced by high-level ab initio calculations. By using a multireference configuration interaction (MRCI) approach, we calculated the 19 lowest lying potential energy curves (PECs) of the LiCa molecule. On the basis of these calculations, we could identify previously unobserved transitions. Our results demonstrate that the helium droplet isolation approach is a powerful method for the characterization of tailor-made alkali–alkaline earth molecules. In this way, important contributions can be made to the search for optimal pathways toward the creation of ultracold alkali–alkaline earth ground state molecules from the corresponding atomic species. Furthermore, a test for PECs calculated by ab initio methods is provided. PMID:24028555

Study of the relaxation dynamics of Styryl 8 and of its solvent cage by sub-pico-second fluorescence laser spectroscopy

International Nuclear Information System (INIS)

Hebert, Philippe

1992-01-01

This research thesis addressed the study of the solvation dynamics of the fluorescent excited state of the styryl 8 molecule, and also the study of the photo-physical and photo-chemical properties, solvatochromism, fluorescence quantum efficiencies, non-radiative de-activation process, and photo-stability of this molecule. The development of a time-resolved (at a pico-second scale) fluorescence laser spectroscopy in a non linear crystal allowed the observation of styryl 8 short time fluorescence kinetics in different solvents, and the analysis of the evolution in time of its fluorescence spectra. Styryl rotation movements have also been studied with the same apparatus by performing time-resolved fluorescence anisotropy. The comparison between experimental results and those obtained with theoretical models highlights interactions between solute and solvent [fr

A Starting Point for Fluorescence-Based Single-Molecule Measurements in Biomolecular Research

Directory of Open Access Journals (Sweden)

Alexander Gust

2014-09-01

Full Text Available Single-molecule fluorescence techniques are ideally suited to provide information about the structure-function-dynamics relationship of a biomolecule as static and dynamic heterogeneity can be easily detected. However, what type of single-molecule fluorescence technique is suited for which kind of biological question and what are the obstacles on the way to a successful single-molecule microscopy experiment? In this review, we provide practical insights into fluorescence-based single-molecule experiments aiming for scientists who wish to take their experiments to the single-molecule level. We especially focus on fluorescence resonance energy transfer (FRET experiments as these are a widely employed tool for the investigation of biomolecular mechanisms. We will guide the reader through the most critical steps that determine the success and quality of diffusion-based confocal and immobilization-based total internal reflection fluorescence microscopy. We discuss the specific chemical and photophysical requirements that make fluorescent dyes suitable for single-molecule fluorescence experiments. Most importantly, we review recently emerged photoprotection systems as well as passivation and immobilization strategies that enable the observation of fluorescently labeled molecules under biocompatible conditions. Moreover, we discuss how the optical single-molecule toolkit has been extended in recent years to capture the physiological complexity of a cell making it even more relevant for biological research.

Fluorescence correlation spectroscopy to study antibody binding and stoichiometry of complexes

Science.gov (United States)

Swift, Kerry M.; Matayoshi, Edmund D.

2008-02-01

FCS (fluorescence correlation spectroscopy) was used to study the association at the single molecule level of tumor necrosis factor alpha (TNF-α) and two of its protein antagonists Humira (TM) (adalimumab), a fully humanized monoclonal antibody, and Enbrel (TM) (etanercept), a soluble form of the TNF receptor. Single molecule approaches potentially have the advantage not only of enhanced sensitivity, but also of observing at equilibrium the details that would otherwise be lost in classical ensemble experiments where heterogeneity is averaged. We prepared fluorescent conjugates of the protein drugs and their biological target, the trimeric soluble form of TNF-α. The bivalency of adalimumab and the trimeric nature of TNF-α potentially allow several forms of associative complexes that may differ in stoichiometry. Detailed knowledge of this reaction may be relevant to understanding adalimumab's pharmacological properties. Our FCS data showed that a single trimeric TNF-α can bind up to three adalimumab molecules. Under some conditions even larger complexes are formed, apparently the result of cross-linking of TNF-α trimers by adalimumab. In addition, distinct differences between Humira and Enbrel were observed in their association with TNF-α.

Time-resolved laser fluorescence spectroscopy of organic ligands by europium: Fluorescence quenching and lifetime properties

Science.gov (United States)

Nouhi, A.; Hajjoul, H.; Redon, R.; Gagné, J. P.; Mounier, S.

2018-03-01

Time-resolved Laser Fluorescence Spectroscopy (TRLFS) has proved its usefulness in the fields of biophysics, life science and geochemistry to characterize the fluorescence probe molecule with its chemical environment. The purpose of this study is to demonstrate the applicability of this powerful technique combined with Steady-State (S-S) measurements. A multi-mode factor analysis, in particular CP/PARAFAC, was used to analyze the interaction between Europium (Eu) and Humic substances (HSs) extracted from Saint Lawrence Estuary in Canada. The Saint Lawrence system is a semi-enclosed water stream with connections to the Atlantic Ocean and is an excellent natural laboratory. CP/PARAFAC applied to fluorescence S-S data allows introspecting ligands-metal interactions and the one-site 1:1 modeling gives information about the stability constants. From the spectral signatures and decay lifetimes data given by TRLFS, one can deduce the fluorescence quenching which modifies the fluorescence and discuss its mechanisms. Results indicated a relatively strong binding ability between europium and humic substances samples (Log K value varies from 3.38 to 5.08 at pH 7.00). Using the Stern-Volmer plot, it has been concluded that static and dynamic quenching takes places in the case of salicylic acid and europium interaction while for HSs interaction only a static quenching is observed.

Use of fluorescence spectroscopy to measure molecular autofluorescence in diabetic subjects

International Nuclear Information System (INIS)

Gomes, Cinthia Zanini

2011-01-01

Diabetes Mellitus (DM) comprises a complex metabolic syndrome, caused by reduced or absent secretion of insulin by pancreatic beta cells, leading to hyperglycemia. Hyperglycemia promotes glycation of proteins and, consequently, the appearance of advanced glycation end products (AGEs). Currently, diabetic patients are monitored by determining levels of glucose and glycated hemoglobin (HbA1c). The complications caused by hyperglycemia may be divided into micro and macrovascular complications, represented by retinopathy, nephropathy, neuropathy and cardiovascular disease. Albumin (HSA) is the most abundant serum protein in the human body and is subject to glycation. The Protoporphyrin IX (PpIX) is the precursor molecule of heme synthesis, structural component of hemoglobin. The in vitro and animals studies have indicated that hyperglycemia promotes a decrease in its concentration in erythrocytes. The fluorescence spectroscopy is a technique widely used in biomedical field. The autofluorescence corresponds to the intrinsic fluorescence present in some molecules, this being associated with the same structure. The aim of this study was to use fluorescence spectroscopy to measure levels of erythrocyte PpIX autofluorescence and AGE-HSA in diabetic and healthy subjects and compare them with levels of blood glucose and HbA1c. This study was conducted with 151 subjects (58 controls and 93 diabetics). Epidemiological data of patients and controls were obtained from medical records. For control subjects, blood glucose levels were obtained from medical records and levels of Hb1Ac obtained by using commercial kits. The determination of the PpIX autofluorescence was performed with excitation at 405 nm and emission at 632 nm. Determination of AGE-HSA was performed with excitation at 370 nm and emission at 455 nm. Approximately 50% of diabetic had micro and macrovascular lesions resulting from hyperglycemia. There were no significant differences in the PpIX emission intensity values

Fluorescence correlation spectroscopy analysis for accurate determination of proportion of doubly labeled DNA in fluorescent DNA pool for quantitative biochemical assays.

Science.gov (United States)

Hou, Sen; Sun, Lili; Wieczorek, Stefan A; Kalwarczyk, Tomasz; Kaminski, Tomasz S; Holyst, Robert

2014-01-15

Fluorescent double-stranded DNA (dsDNA) molecules labeled at both ends are commonly produced by annealing of complementary single-stranded DNA (ssDNA) molecules, labeled with fluorescent dyes at the same (3' or 5') end. Because the labeling efficiency of ssDNA is smaller than 100%, the resulting dsDNA have two, one or are without a dye. Existing methods are insufficient to measure the percentage of the doubly-labeled dsDNA component in the fluorescent DNA sample and it is even difficult to distinguish the doubly-labeled DNA component from the singly-labeled component. Accurate measurement of the percentage of such doubly labeled dsDNA component is a critical prerequisite for quantitative biochemical measurements, which has puzzled scientists for decades. We established a fluorescence correlation spectroscopy (FCS) system to measure the percentage of doubly labeled dsDNA (PDL) in the total fluorescent dsDNA pool. The method is based on comparative analysis of the given sample and a reference dsDNA sample prepared by adding certain amount of unlabeled ssDNA into the original ssDNA solution. From FCS autocorrelation functions, we obtain the number of fluorescent dsDNA molecules in the focal volume of the confocal microscope and PDL. We also calculate the labeling efficiency of ssDNA. The method requires minimal amount of material. The samples have the concentration of DNA in the nano-molar/L range and the volume of tens of microliters. We verify our method by using restriction enzyme Hind III to cleave the fluorescent dsDNA. The kinetics of the reaction depends strongly on PDL, a critical parameter for quantitative biochemical measurements. Copyright © 2013 Elsevier B.V. All rights reserved.

Photon-counting single-molecule spectroscopy for studying conformational dynamics and macromolecular interactions

Energy Technology Data Exchange (ETDEWEB)

Laurence, Ted Alfred [Univ. of California, Berkeley, CA (United States)

2002-01-01

Single-molecule methods have the potential to provide information about conformational dynamics and molecular interactions that cannot be obtained by other methods. Removal of ensemble averaging provides several benefits, including the ability to detect heterogeneous populations and the ability to observe asynchronous reactions. Single-molecule diffusion methodologies using fluorescence resonance energy transfer (FRET) are developed to monitor conformational dynamics while minimizing perturbations introduced by interactions between molecules and surfaces. These methods are used to perform studies of the folding of Chymotrypsin Inhibitor 2, a small, single-domain protein, and of single-stranded DNA (ssDNA) homopolymers. Confocal microscopy is used in combination with sensitive detectors to detect bursts of photons from fluorescently labeled biomolecules as they diffuse through the focal volume. These bursts are analyzed to extract fluorescence resonance energy transfer (FRET) efficiency. Advances in data acquisition and analysis techniques that are providing a more complete picture of the accessible molecular information are discussed. Photon Arrival-time Interval Distribution (PAID) analysis is a new method for monitoring macromolecular interactions by fluorescence detection with simultaneous determination of coincidence, brightness, diffusion time, and occupancy (proportional to concentration) of fluorescently-labeled molecules undergoing diffusion in a confocal detection volume. This method is based on recording the time of arrival of all detected photons, and then plotting the two-dimensional histogram of photon pairs, where one axis is the time interval between each pair of photons 1 and 2, and the second axis is the number of other photons detected in the time interval between photons 1 and 2. PAID is related to Fluorescence Correlation Spectroscopy (FCS) by a collapse of this histogram onto the time interval axis. PAID extends auto- and cross-correlation FCS

Photon-counting single-molecule spectroscopy for studying conformational dynamics and macromolecular interactions

International Nuclear Information System (INIS)

Laurence, Ted Alfred

2002-01-01

Single-molecule methods have the potential to provide information about conformational dynamics and molecular interactions that cannot be obtained by other methods. Removal of ensemble averaging provides several benefits, including the ability to detect heterogeneous populations and the ability to observe asynchronous reactions. Single-molecule diffusion methodologies using fluorescence resonance energy transfer (FRET) are developed to monitor conformational dynamics while minimizing perturbations introduced by interactions between molecules and surfaces. These methods are used to perform studies of the folding of Chymotrypsin Inhibitor 2, a small, single-domain protein, and of single-stranded DNA (ssDNA) homopolymers. Confocal microscopy is used in combination with sensitive detectors to detect bursts of photons from fluorescently labeled biomolecules as they diffuse through the focal volume. These bursts are analyzed to extract fluorescence resonance energy transfer (FRET) efficiency. Advances in data acquisition and analysis techniques that are providing a more complete picture of the accessible molecular information are discussed. Photon Arrival-time Interval Distribution (PAID) analysis is a new method for monitoring macromolecular interactions by fluorescence detection with simultaneous determination of coincidence, brightness, diffusion time, and occupancy (proportional to concentration) of fluorescently-labeled molecules undergoing diffusion in a confocal detection volume. This method is based on recording the time of arrival of all detected photons, and then plotting the two-dimensional histogram of photon pairs, where one axis is the time interval between each pair of photons 1 and 2, and the second axis is the number of other photons detected in the time interval between photons 1 and 2. PAID is related to Fluorescence Correlation Spectroscopy (FCS) by a collapse of this histogram onto the time interval axis. PAID extends auto- and cross-correlation FCS

Two-dimensional fluorescence lifetime correlation spectroscopy. 2. Application.

Science.gov (United States)

Ishii, Kunihiko; Tahara, Tahei

2013-10-03

In the preceding article, we introduced the theoretical framework of two-dimensional fluorescence lifetime correlation spectroscopy (2D FLCS). In this article, we report the experimental implementation of 2D FLCS. In this method, two-dimensional emission-delay correlation maps are constructed from the photon data obtained with the time-correlated single photon counting (TCSPC), and then they are converted to 2D lifetime correlation maps by the inverse Laplace transform. We develop a numerical method to realize reliable transformation, employing the maximum entropy method (MEM). We apply the developed actual 2D FLCS to two real systems, a dye mixture and a DNA hairpin. For the dye mixture, we show that 2D FLCS is experimentally feasible and that it can identify different species in an inhomogeneous sample without any prior knowledge. The application to the DNA hairpin demonstrates that 2D FLCS can disclose microsecond spontaneous dynamics of biological molecules in a visually comprehensible manner, through identifying species as unique lifetime distributions. A FRET pair is attached to the both ends of the DNA hairpin, and the different structures of the DNA hairpin are distinguished as different fluorescence lifetimes in 2D FLCS. By constructing the 2D correlation maps of the fluorescence lifetime of the FRET donor, the equilibrium dynamics between the open and the closed forms of the DNA hairpin is clearly observed as the appearance of the cross peaks between the corresponding fluorescence lifetimes. This equilibrium dynamics of the DNA hairpin is clearly separated from the acceptor-missing DNA that appears as an isolated diagonal peak in the 2D maps. The present study clearly shows that newly developed 2D FLCS can disclose spontaneous structural dynamics of biological molecules with microsecond time resolution.

Two dimensional laser induced fluorescence in the gas phase: a spectroscopic tool for studying molecular spectroscopy and dynamics

Science.gov (United States)

Gascooke, Jason R.; Lawrance, Warren D.

2017-11-01

Two dimensional laser induced fluorescence (2D-LIF) extends the usual laser induced fluorescence technique by adding a second dimension, the wavelength at which excited states emit, thereby significantly enhancing the information that can be extracted. It allows overlapping absorption features, whether they arise from within the same molecule or from different molecules in a mixture, to be associated with their appropriate "parent" state and/or molecule. While the first gas phase version of the technique was published a decade ago, the technique is in its infancy, having been exploited by only a few groups to date. However, its potential in gas phase spectroscopy and dynamics is significant. In this article we provide an overview of the technique and illustrate its potential with examples, with a focus on those utilising high resolution in the dispersed fluorescence dimension.

Nanosecond fluorescence spectroscopy

International Nuclear Information System (INIS)

Leskovar, B.

1985-03-01

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

Single-molecule fluorescence microscopy review: shedding new light on old problems.

Science.gov (United States)

Shashkova, Sviatlana; Leake, Mark C

2017-08-31

Fluorescence microscopy is an invaluable tool in the biosciences, a genuine workhorse technique offering exceptional contrast in conjunction with high specificity of labelling with relatively minimal perturbation to biological samples compared with many competing biophysical techniques. Improvements in detector and dye technologies coupled to advances in image analysis methods have fuelled recent development towards single-molecule fluorescence microscopy, which can utilize light microscopy tools to enable the faithful detection and analysis of single fluorescent molecules used as reporter tags in biological samples. For example, the discovery of GFP, initiating the so-called 'green revolution', has pushed experimental tools in the biosciences to a completely new level of functional imaging of living samples, culminating in single fluorescent protein molecule detection. Today, fluorescence microscopy is an indispensable tool in single-molecule investigations, providing a high signal-to-noise ratio for visualization while still retaining the key features in the physiological context of native biological systems. In this review, we discuss some of the recent discoveries in the life sciences which have been enabled using single-molecule fluorescence microscopy, paying particular attention to the so-called 'super-resolution' fluorescence microscopy techniques in live cells, which are at the cutting-edge of these methods. In particular, how these tools can reveal new insights into long-standing puzzles in biology: old problems, which have been impossible to tackle using other more traditional tools until the emergence of new single-molecule fluorescence microscopy techniques. © 2017 The Author(s).

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

Fluorescence Correlation Spectroscopy to Study Diffusion of Polymer Chains within Layered Hydrogen-Bonded Polymer Films

Science.gov (United States)

Pristinski, Denis; Kharlampieva, Evguenia; Sukhishvili, Svetlana

2002-03-01

Fluorescence Correlation Spectroscopy (FCS) has been used to probe molecular motions within polymer multilayers formed by hydrogen-bonding sequential self-assembly. Polyethylene glycol (PEG) molecules were end-labeled with the fluorescent tags, and self-assembled with polymethacrylic acid (PMAA) using layer-by-layer deposition. We have found that molecules included in the top adsorbed layer have significant mobility at the millisecond time scale, probably due to translational diffusion. However, their dynamics deviate from classical Brownian motion with a single diffusion time. Possible reasons for the deviation are discussed. We found that motions were significantly slowed with increasing depth within the PEG/PMAA multilayer. This phenomena occured in a narrow pH range around 4.0 in which intermolecular interactions were relatively weak.

Laser Induced Fluorescence Spectroscopy of Neutral and Ionized Polycyclic Aromatic Hydrocarbons in the Cosmic Simulation Chamber

Science.gov (United States)

Bejaoui, Salma; Salama, Farid; Contreras, Cesar; Sciamma O'Brien, Ella; Foing, Bernard; Pascale, Ehrenfreund

2015-01-01

Polycyclic aromatic hydrocarbon (PAH) molecules are considered the best carriers to account for the ubiquitous infrared emission bands. PAHs have also been proposed as candidates to explain the diffuse interstellar bands (DIBs), a series of absorption features seen on the interstellar extinction curve and are plausible carriers for the extended red emission (ERE), a photoluminescent process associated with a wide variety of interstellar environments. Extensive efforts have been devoted over the past two decades to characterize the physical and chemical properties of PAH molecules and ions in space. Absorption spectra of PAH molecules and ions trapped in solid matrices have been compared to the DIBs. Absorption spectra of several cold, isolated gas-phase PAHs have also been measured under experimental conditions that mimic the interstellar conditions. The purpose of this study is to provide a new dimension to the existing spectroscopic database of neutral and single ionized PAHs that is largely based on absorption spectra by adding emission spectroscopy data. The measurements are based on the laser induced fluorescence (LIF) technique and are performed with the Pulsed Discharge Nozzle (PDN) of the COSmIC laboratory facility at NASA Ames laboratory. The PDN generates a plasma in a free supersonic jet expansion to simulate the physical and the chemical conditions in interstellar environments. We focus, here, on the fluorescence spectra of large neutral PAHs and their cations where there is a lack of fluorescence spectroscopy data. The astronomical implications of the data (e.g., ERE) are examined.

Functionalization of gold and nanocrystalline diamond atomic force microscope tips for single molecule force spectroscopy

Science.gov (United States)

Drew, Michael E.

-ray photoelectron, infrared, and NEXAFS spectroscopy. The same photochemical method was used to attach molecules to NCD AFM tips. Selective fluorescent labeling and adhesion measurements indicated that the functionalization of undecylenic acid was successful. Studies towards applying NCD AFM tips for nanolithography on a polymer surface are also described.

Fluorescence blinking in MEH-PPV single molecules at low temperature

International Nuclear Information System (INIS)

Mirzov, O.; Cichos, F.; Borczyskowski, C. von; Scheblykin, I.

2005-01-01

Fluorescence intensity transients of single molecules of the conjugated polymer poly[2-methoxy,5-(2'-ethylhexyloxy)-p-phenylene-vinylene] (MEH-PPV) were studied at 15 K. Fluorescence blinking behavior was observed despite the expected low-temperature suppression of energy migration in such disordered molecular systems. Presence of the fluorescence blinking effect at 15 K indicates that the single molecules possess a collapsed conformation with characteristic size of not more than several nanometers, which corresponds to only a few exciton hops over a polymer chain

Improved Dye Stability in Single-Molecule Fluorescence Experiments

Science.gov (United States)

EcheverrÍa Aitken, Colin; Marshall, R. Andrew; Pugi, Joseph D.

Complex biological systems challenge existing single-molecule methods. In particular, dye stability limits observation time in singlemolecule fluorescence applications. Current approaches to improving dye performance involve the addition of enzymatic oxygen scavenging systems and small molecule additives. We present an enzymatic oxygen scavenging system that improves dye stability in single-molecule experiments. Compared to the currently-employed glucose-oxidase/catalase system, the protocatechuate-3,4-dioxygenase system achieves lower dissolved oxygen concentration and stabilizes single Cy3, Cy5, and Alexa488 fluorophores. Moreover, this system possesses none of the limitations associated with the glucose oxidase/catalase system. We also tested the effects of small molecule additives in this system. Biological reducing agents significantly destabilize the Cy5 fluorophore as a function of reducing potential. In contrast, anti-oxidants stabilize the Cy3 and Alexa488 fluorophores. We recommend use of the protocatechuate-3,4,-dioxygenase system with antioxidant additives, and in the absence of biological reducing agents. This system should have wide application to single-molecule fluorescence experiments.

Time-Resolved Fluorescence Spectroscopy and Fluorescence Lifetime Imaging Microscopy for Characterization of Dendritic Polymer Nanoparticles and Applications in Nanomedicine

Directory of Open Access Journals (Sweden)

Alexander Boreham

2016-12-01

Full Text Available The emerging field of nanomedicine provides new approaches for the diagnosis and treatment of diseases, for symptom relief and for monitoring of disease progression. One route of realizing this approach is through carefully constructed nanoparticles. Due to the small size inherent to the nanoparticles a proper characterization is not trivial. This review highlights the application of time-resolved fluorescence spectroscopy and fluorescence lifetime imaging microscopy (FLIM for the analysis of nanoparticles, covering aspects ranging from molecular properties to particle detection in tissue samples. The latter technique is particularly important as FLIM allows for distinguishing of target molecules from the autofluorescent background and, due to the environmental sensitivity of the fluorescence lifetime, also offers insights into the local environment of the nanoparticle or its interactions with other biomolecules. Thus, these techniques offer highly suitable tools in the fields of particle development, such as organic chemistry, and in the fields of particle application, such as in experimental dermatology or pharmaceutical research.

Time-Resolved Fluorescence Spectroscopy and Fluorescence Lifetime Imaging Microscopy for Characterization of Dendritic Polymer Nanoparticles and Applications in Nanomedicine.

Science.gov (United States)

Boreham, Alexander; Brodwolf, Robert; Walker, Karolina; Haag, Rainer; Alexiev, Ulrike

2016-12-24

The emerging field of nanomedicine provides new approaches for the diagnosis and treatment of diseases, for symptom relief and for monitoring of disease progression. One route of realizing this approach is through carefully constructed nanoparticles. Due to the small size inherent to the nanoparticles a proper characterization is not trivial. This review highlights the application of time-resolved fluorescence spectroscopy and fluorescence lifetime imaging microscopy (FLIM) for the analysis of nanoparticles, covering aspects ranging from molecular properties to particle detection in tissue samples. The latter technique is particularly important as FLIM allows for distinguishing of target molecules from the autofluorescent background and, due to the environmental sensitivity of the fluorescence lifetime, also offers insights into the local environment of the nanoparticle or its interactions with other biomolecules. Thus, these techniques offer highly suitable tools in the fields of particle development, such as organic chemistry, and in the fields of particle application, such as in experimental dermatology or pharmaceutical research.

Nano-sensing of the orientation of fluorescing molecules with active coated nano-particles

DEFF Research Database (Denmark)

Arslanagic, Samel; Ziolkowski, Richard W.

2015-01-01

The potential of using active coated nano-particles to determine the orientation of fluorescing molecules is reported. By treating each fluorescing molecule as an electric Hertzian dipole, single and multiple fluorescing molecules emitting coherently and incoherently in various orientations...... are considered in the presence of active coated nano-particles. It is demonstrated that in addition to offering a means to determine the orientation of a single molecule or the over-all orientation of the molecules surrounding it, the nature of the far-field response from the active coated nano...

Molecular spectroscopy

International Nuclear Information System (INIS)

Kokh, Eh.; Zonntag, B.

1981-01-01

The latest investigation results on molecular spectroscopy with application of synchrotron radiation in the region of vacuum ultraviolet are generalized. Some results on investigation of excited, superexcited and ionized molecule states with the use of adsorption spectroscopy, photoelectron spectroscopy, by fluorescent and mass-spectrometric methods are considered [ru

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

Science.gov (United States)

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

2014-05-01

performed in pellets (boric and humic acids mixture) using a portable system built by Embrapa Instrumentation. It comprises a diode laser (Coherent - CUBE) emitting at 405 nm (50 mW), and the detection of emission by a high sensitivity mini-spectrometer (USB4000 - Ocean Optics) using a range from 440 to 800 nm. In first step, the 3D tensors were then treated by the CP/PARAFAC algorithm to decompose the signal response after removing the diffusion signal : three components were extracted with a CORCONDIA over 60%. The first component can be associate an artefact of the measurement or boric acid fluorescence, the second and third component could the related to the two different fluorescence contributions of tryptophan molecule, one with central excitation/emission in 290/360 nm and other in 350/465 nm. The presence of a small quantity (i.e. few percent in mass) of humic acid (HA) is quenching drastically the TRP fluorescence. Complementary, measurements will be performed to understand this behaviour taking in account the absorption wavelength by the surface (colour) and by measuring the time life fluorescence of the samples. Humic acid fluorescence in pellets (BA and HA) cannot be observed using lamp + monochromator excitation due to low intensity of source. The same pellets were measure using LIFS system, and fluorescence intensity increased as a function of concentration of HA until occur the inner filter effect from 300 ppm, similar to the behaviour of HA in solution. Even whether solid surface measurements are easier, understanding is not yet clear. More investigation needs to be done. Moreover, it should be important to know if the use of CP/PARAFAC decomposition for such data is relevant with the trilinear model. References Milori, D.M.B.P., Galeti, H.V.A., Martin-Neto, L., Dieckow, J., González-Pérez, M., Bayer, C., Salton, J., 2006. Organic Matter Study of Whole Soil Samples Using Laser-Induced Fluorescence Spectroscopy. Soil Science Society of America Journal 70

Fluorescence Lifetime Correlation Spectroscopy (FLCS): Concepts, Applications and Outlook

Czech Academy of Sciences Publication Activity Database

Kapusta, Peter; Macháň, Radek; Benda, A.; Hof, Martin

2012-01-01

Roč. 13, č. 10 (2012), s. 12890-12910 E-ISSN 1422-0067 R&D Projects: GA ČR GBP208/12/G016 Institutional support: RVO:61388955 Keywords : fluorescence correlation spectroscopy (FCS) * time correlated single photon counting (TCSPC) * fluorescence cross-correlation spectroscopy (FCCS) Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.464, year: 2012

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

Energy Technology Data Exchange (ETDEWEB)

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

2014-01-15

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

Confined detection volume of fluorescence correlation spectroscopy by bare fiber probes.

Science.gov (United States)

Lu, Guowei; Lei, Franck H; Angiboust, Jean-François; Manfait, Michel

2010-04-01

A fiber-tip-based near-field fluorescence correlation spectroscopy (FCS) has been developed for confining the detection volume to sub-diffraction-limited dimensions. This near-field FCS is based on near-field illumination by coupling a scanning near-field optical microscope (SNOM) to a conventional confocal FCS. Single-molecule FCS analysis at 100 nM Rhodamine 6G has been achieved by using bare chemically etched, tapered fiber tips. The detection volume under control of the SNOM system has been reduced over one order of magnitude compared to that of the conventional confocal FCS. Related factors influencing the near-field FCS performance are investigated and discussed in detail. In this proof-of-principle study, the preliminary experimental results suggest that the fiber-tip-based near-field FCS might be a good alternative to realize localized analysis at the single-molecule level.

Surface Transient Binding-Based Fluorescence Correlation Spectroscopy (STB-FCS), a Simple and Easy-to-Implement Method to Extend the Upper Limit of the Time Window to Seconds.

Science.gov (United States)

Peng, Sijia; Wang, Wenjuan; Chen, Chunlai

2018-05-10

Fluorescence correlation spectroscopy is a powerful single-molecule tool that is able to capture kinetic processes occurring at the nanosecond time scale. However, the upper limit of its time window is restricted by the dwell time of the molecule of interest in the confocal detection volume, which is usually around submilliseconds for a freely diffusing biomolecule. Here, we present a simple and easy-to-implement method, named surface transient binding-based fluorescence correlation spectroscopy (STB-FCS), which extends the upper limit of the time window to seconds. We further demonstrated that STB-FCS enables capture of both intramolecular and intermolecular kinetic processes whose time scales cross several orders of magnitude.

Mechanisms of ultrafast fluorescence depletion spectroscopy and applications to measure slovation dynamics of coummarin 153 in methanol

International Nuclear Information System (INIS)

Yang Songqiu; Liu Jianyong; Zhou Panwang; Chen Junsheng; Han Keli; He Guozhong

2012-01-01

Subpicosecond fluorescence depletion spectroscopy (FDS) was used to measure the solvation dynamics of coumarin 153 (C153) in methanol. The FDS mechanisms were discussed. A quasi-continuous model was used to describe the solvational relaxation of excited states. The perturbations of the probe pulse on the excited sample system, including up-conversion and stimulated emission, were sufficiently discussed. For a probe molecule used in the FDS experiment, ensuring that the up-conversion perturbation can be negligible is important. FDS was found to be a good technique for measuring the solvation dynamics of C153 in methanol. - Highlights: ► Mechanisms of subpicosecond fluorescence depletion spectroscopy. ► Quasi-continuous model was used to describe the solvational relaxation. ► The solvation dynamics of coumarin 153 in methanol has been measured.

Fluorescence Spectroscopy, Exciton Dynamics and Photochemistry of Single Allophycocyanin Trimers

International Nuclear Information System (INIS)

Ying, Liming; Xie, Xiaoliang

1998-01-01

We report a study of the spectroscopy and exciton dynamics of the allophycocyanin trimer (APC), a light harvesting protein complex from cyanobacteria, by room-temperature single-molecule measurements of fluorescence spectra, lifetimes, intensity trajectories and polarization modulation. Emission spectra of individual APC trimers are found to be homogeneous on the time scale of seconds. In contrast, their emission lifetimes are found to be widely distributed, because of generation of exciton traps during the course of measurements. The intensity trajectories and polarization modulation experiments indicate reversible ixciton trap formation within the three quasi-independent pairs of strong interacting a84 and B84 chromophores in APC, as well a photobleaching of individual chromophores. Comparison experiments under continuous wave and pulsed excitation reveal a two-photon mechanism for generating exciton traps and/or photobleaching, which involves exciton-exciton annihilation. These single-molecule experiments provide new insights into exciton dynamics and photochemistry of light-harvesting complexes

Laser spectroscopy on organic molecules.

Science.gov (United States)

Imasaka, T

1996-06-01

Various laser spectrometric methods have been developed until now. Especially, laser fluorometry is most sensitive and is frequently combined with a separation technique such as capillary electrophoresis. For non-fluorescent compounds, photothermal spectrometry may be used instead. A diode laser is potentially useful for practical trace analysis, because of its low cost and long-term trouble-free operation. On the other hand, monochromaticity of the laser is essential in high-resolution spectrometry, e.g. in low temperature spectrometry providing a very sharp spectral feature. Closely-related compounds such as isomers can easily be differentiated, and information for assignment is obtained from the spectrum. Multiphoton ionization mass spectrometry is useful for soft ionization, providing additional information concerned with molecular weight and chemical structure. A short laser pulse with a sufficient energy is suitable for rapid heating of the solid surface. A matrix-assisted laser desorption/ion-ization technique is recently employed for introduction of a large biological molecule into a vacuum for mass analysis. In the future, laser spectrometry will be developed by a combination with state-of-the-art laser technology. In the 21st century, new laser spectrometry will be developed, which may be based on revolutionary ideas or unexpected discoveries. Such studies will open new frontiers in analytical laser spectroscopy.

Xanthines Studied via Femtosecond Fluorescence Spectroscopy

Directory of Open Access Journals (Sweden)

Pascale Changenet-Barret

2016-12-01

Full Text Available Xanthines represent a wide class of compounds closely related to the DNA bases adenine and guanine. Ubiquitous in the human body, they are capable of replacing natural bases in double helices and give rise to four-stranded structures. Although the use of their fluorescence for analytical purposes was proposed, their fluorescence properties have not been properly characterized so far. The present paper reports the first fluorescence study of xanthine solutions relying on femtosecond spectroscopy. Initially, we focus on 3-methylxanthine, showing that this compound exhibits non-exponential fluorescence decays with no significant dependence on the emission wavelength. The fluorescence quantum yield (3 × 10−4 and average decay time (0.9 ps are slightly larger than those found for the DNA bases. Subsequently, we compare the dynamical fluorescence properties of seven mono-, di- and tri-methylated derivatives. Both the fluorescence decays and fluorescence anisotropies vary only weakly with the site and the degree of methylation. These findings are in line with theoretical predictions suggesting the involvement of several conical intersections in the relaxation of the lowest singlet excited state.

Single-Molecule Fluorescence Studies of Membrane Transporters Using Total Internal Reflection Microscopy.

Science.gov (United States)

Goudsmits, Joris M H; van Oijen, Antoine M; Slotboom, Dirk J

2017-01-01

Cells are delineated by a lipid bilayer that physically separates the inside from the outer environment. Most polar, charged, or large molecules require proteins to reduce the energetic barrier for passage across the membrane and to achieve transport rates that are relevant for life. Here, we describe techniques to visualize the functioning of membrane transport proteins with fluorescent probes at the single-molecule level. First, we explain how to produce membrane-reconstituted transporters with fluorescent labels. Next, we detail the construction of a microfluidic flow cell to image immobilized proteoliposomes on a total internal reflection fluorescence microscope. We conclude by describing the methods that are needed to analyze fluorescence movies and obtain useful single-molecule data. © 2017 Elsevier Inc. All rights reserved.

Using Fluorescent Viruses for Detecting Bacteria in Water

Science.gov (United States)

Tabacco, Mary Beth; Qian, Xiaohua; Russo, Jaimie A.

2009-01-01

A method of detecting water-borne pathogenic bacteria is based partly on established molecular-recognition and fluorescent-labeling concepts, according to which bacteria of a species of interest are labeled with fluorescent reporter molecules and the bacteria can then be detected by fluorescence spectroscopy. The novelty of the present method lies in the use of bacteriophages (viruses that infect bacteria) to deliver the fluorescent reporter molecules to the bacteria of the species of interest.

Probing Protein Multidimensional Conformational Fluctuations by Single-Molecule Multiparameter Photon Stamping Spectroscopy

Science.gov (United States)

2015-01-01

Conformational motions of proteins are highly dynamic and intrinsically complex. To capture the temporal and spatial complexity of conformational motions and further to understand their roles in protein functions, an attempt is made to probe multidimensional conformational dynamics of proteins besides the typical one-dimensional FRET coordinate or the projected conformational motions on the one-dimensional FRET coordinate. T4 lysozyme hinge-bending motions between two domains along α-helix have been probed by single-molecule FRET. Nevertheless, the domain motions of T4 lysozyme are rather complex involving multiple coupled nuclear coordinates and most likely contain motions besides hinge-bending. It is highly likely that the multiple dimensional protein conformational motions beyond the typical enzymatic hinged-bending motions have profound impact on overall enzymatic functions. In this report, we have developed a single-molecule multiparameter photon stamping spectroscopy integrating fluorescence anisotropy, FRET, and fluorescence lifetime. This spectroscopic approach enables simultaneous observations of both FRET-related site-to-site conformational dynamics and molecular rotational (or orientational) motions of individual Cy3-Cy5 labeled T4 lysozyme molecules. We have further observed wide-distributed rotational flexibility along orientation coordinates by recording fluorescence anisotropy and simultaneously identified multiple intermediate conformational states along FRET coordinate by monitoring time-dependent donor lifetime, presenting a whole picture of multidimensional conformational dynamics in the process of T4 lysozyme open-close hinge-bending enzymatic turnover motions under enzymatic reaction conditions. By analyzing the autocorrelation functions of both lifetime and anisotropy trajectories, we have also observed the dynamic and static inhomogeneity of T4 lysozyme multidimensional conformational fluctuation dynamics, providing a fundamental

Quantitative frequency-domain fluorescence spectroscopy in tissues and tissue-like media

Science.gov (United States)

Cerussi, Albert Edward

1999-09-01

In the never-ending quest for improved medical technology at lower cost, modern near-infrared optical spectroscopy offers the possibility of inexpensive technology for quantitative and non-invasive diagnoses. Hemoglobin is the dominant chromophore in the 700-900 nm spectral region and as such it allows for the optical assessment of hemoglobin concentration and tissue oxygenation by absorption spectroscopy. However, there are many other important physiologically relevant compounds or physiological states that cannot be effectively sensed via optical methods because of poor optical contrast. In such cases, contrast enhancements are required. Fluorescence spectroscopy is an attractive component of optical tissue spectroscopy. Exogenous fluorophores, as well as some endogenous ones, may furnish the desperately needed sensitivity and specificity that is lacking in near-infrared optical tissue spectroscopy. The main focus of this thesis was to investigate the generation and propagation of fluorescence photons inside tissues and tissue-like media (i.e., scattering dominated media). The standard concepts of fluorescence spectroscopy have been incorporated into a diffusion-based picture that is sometimes referred to as photon migration. The novelty of this work lies in the successful quantitative recovery of fluorescence lifetimes, absolute fluorescence quantum yields, fluorophore concentrations, emission spectra, and both scattering and absorption coefficients at the emission wavelength from a tissue-like medium. All of these parameters are sensitive to the fluorophore local environment and hence are indicators of the tissue's physiological state. One application demonstrating the capabilities of frequency-domain lifetime spectroscopy in tissue-like media is a study of the binding of ethidium bromide to bovine leukocytes in fresh milk. Ethidium bromide is a fluorescent dye that is commonly used to label DNA, and hence visualize chromosomes in cells. The lifetime of

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

International Nuclear Information System (INIS)

Zhang Chuanfei; Lin Libin; Lou Fuhong; Peng Taiping

2001-01-01

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

Single molecule force spectroscopy: methods and applications in biology

International Nuclear Information System (INIS)

Shen Yi; Hu Jun

2012-01-01

Single molecule measurements have transformed our view of biomolecules. Owing to the ability of monitoring the activity of individual molecules, we now see them as uniquely structured, fluctuating molecules that stochastically transition between frequently many substrates, as two molecules do not follow precisely the same trajectory. Indeed, it is this discovery of critical yet short-lived substrates that were often missed in ensemble measurements that has perhaps contributed most to the better understanding of biomolecular functioning resulting from single molecule experiments. In this paper, we give a review on the three major techniques of single molecule force spectroscopy, and their applications especially in biology. The single molecular study of biotin-streptavidin interactions is introduced as a successful example. The problems and prospects of the single molecule force spectroscopy are discussed, too. (authors)

Seeing the electroporative uptake of cell-membrane impermeable fluorescent molecules and nanoparticles

Science.gov (United States)

Kim, Kisoo; Kim, Jeong Ah; Lee, Soon-Geul; Lee, Won Gu

2012-07-01

This paper presents direct visualization of uptake directionality for cell-membrane impermeant fluorescent molecules and fluorescence-doped nanoparticles at a single-cell level during electroporation. To observe directly the uptake direction, we used microchannel-type electroporation that can generate a relatively symmetric and uniform electric field. For all the image frames during electroporation, fluorescence intensities that occurred at cell membranes in both uptake directions toward the electrodes have been sequentially recorded and quantitatively analyzed pixel by pixel. In our experiments, we found that fluorescent molecules, even not labeled to target biomolecules, had their own uptake direction with different intensities. It is also observed that the uptake intensity toward the cell membrane had a maximal value at a certain electric voltage, not at the highest value of voltages applied. The results also imply that the uptake direction of fluorescence-doped nanoparticles can be determined by a net surface charge of uptake materials and sizes in the electroporative environments. In summary, we performed a quantitative screening and direct visualization of uptake directionality for a set of fluorescent molecules and fluorescence-doped nanoparticles using electric-pulsation. Taking a closer look at the uptake direction of exogenous materials will help researchers to understand an unknown uptake phenomenon in which way foreign materials are inclined to move, and furthermore to design functional nanoparticles for electroporative gene delivery.This paper presents direct visualization of uptake directionality for cell-membrane impermeant fluorescent molecules and fluorescence-doped nanoparticles at a single-cell level during electroporation. To observe directly the uptake direction, we used microchannel-type electroporation that can generate a relatively symmetric and uniform electric field. For all the image frames during electroporation, fluorescence intensities

Experimental station for gas phase fluorescence spectroscopy

International Nuclear Information System (INIS)

Stankiewicz, M.; Garcia, E. Melero; Ruiz, J. Alvarez; Erman, P.; Hatherly, P.A.; Kivimaeki, A.; Rachlew, E.; Rius i Riu, J.

2004-01-01

The details of an experimental setup for gas phase atomic and molecular fluorescence measurements using synchrotron radiation are described in this article. The most significant part of the apparatus is an optical arrangement, which allows for simultaneous measurements of dispersed as well as total fluorescence intensity using an effusive gas jet and an inbuilt gas cell assembled in a convenient plug and measure configuration. The first measurements concerning fluorescence of the N 2 molecule around the N 1s edge obtained with this setup are presented

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.

Photoionization of atoms and molecules

International Nuclear Information System (INIS)

Samson, J.A.R.

1976-01-01

A literature review on the present state of knowledge in photoionization is presented. Various experimental techniques that have been developed to study photoionization, such as fluorescence and photoelectron spectroscopy, mass spectroscopy, are examined. Various atoms and molecules were chosen to illustrate these techniques, specifically helium and xenon atoms and hydrogen molecules. Specialized photoionization such as in positive and negative ions, excited states, and free radicals is also treated. Absorption cross sections and ionization potentials are also discussed

A Project-Based Biochemistry Laboratory Promoting the Understanding and Uses of Fluorescence Spectroscopy in the Study of Biomolecular Structures and Interactions

Science.gov (United States)

Briese, Nicholas; Jakubowsk, Henry V.

2007-01-01

A laboratory project for a first semester biochemistry course is described, which integrates the traditional classroom study of the structure and function of biomolecules with the laboratory study of these molecules using fluorescence spectroscopy. Students are assigned a specific question addressing the stability/function of lipids, proteins, or…

Click strategies for single-molecule protein fluorescence.

Science.gov (United States)

Milles, Sigrid; Tyagi, Swati; Banterle, Niccolò; Koehler, Christine; VanDelinder, Virginia; Plass, Tilman; Neal, Adrian P; Lemke, Edward A

2012-03-21

Single-molecule methods have matured into central tools for studies in biology. Foerster resonance energy transfer (FRET) techniques, in particular, have been widely applied to study biomolecular structure and dynamics. The major bottleneck for a facile and general application of these studies arises from the need to label biological samples site-specifically with suitable fluorescent dyes. In this work, we present an optimized strategy combining click chemistry and the genetic encoding of unnatural amino acids (UAAs) to overcome this limitation for proteins. We performed a systematic study with a variety of clickable UAAs and explored their potential for high-resolution single-molecule FRET (smFRET). We determined all parameters that are essential for successful single-molecule studies, such as accessibility of the probes, expression yield of proteins, and quantitative labeling. Our multiparameter fluorescence analysis allowed us to gain new insights into the effects and photophysical properties of fluorescent dyes linked to various UAAs for smFRET measurements. This led us to determine that, from the extended tool set that we now present, genetically encoding propargyllysine has major advantages for state-of-the-art measurements compared to other UAAs. Using this optimized system, we present a biocompatible one-step dual-labeling strategy of the regulatory protein RanBP3 with full labeling position freedom. Our technique allowed us then to determine that the region encompassing two FxFG repeat sequences adopts a disordered but collapsed state. RanBP3 serves here as a prototypical protein that, due to its multiple cysteines, size, and partially disordered structure, is not readily accessible to any of the typical structure determination techniques such as smFRET, NMR, and X-ray crystallography.

Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy.

Science.gov (United States)

Stortz, Martin; Angiolini, Juan; Mocskos, Esteban; Wolosiuk, Alejandro; Pecci, Adali; Levi, Valeria

2018-05-01

The hierarchical organization of the cell nucleus into specialized open reservoirs and the nucleoplasm overcrowding impose restrictions to the mobility of biomolecules and their interactions with nuclear targets. These properties determine that many nuclear functions such as transcription, replication, splicing or DNA repair are regulated by complex, dynamical processes that do not follow simple rules. Advanced fluorescence microscopy tools and, in particular, fluorescence correlation spectroscopy (FCS) provide complementary and exquisite information on the dynamics of fluorescent labeled molecules moving through the nuclear space and are helping us to comprehend the complexity of the nuclear structure. Here, we describe how FCS methods can be applied to reveal the dynamical organization of the nucleus in live cells. Specifically, we provide instructions for the preparation of cellular samples with fluorescent tagged proteins and detail how FCS can be easily instrumented in commercial confocal microscopes. In addition, we describe general rules to set the parameters for one and two-color experiments and the required controls for these experiments. Finally, we review the statistical analysis of the FCS data and summarize the use of numerical simulations as a complementary approach that helps us to understand the complex matrix of molecular interactions network within the nucleus. Copyright © 2017 Elsevier Inc. All rights reserved.

Fluorescence excitation-emission matrix spectroscopy of vitiligo skin in vivo (Conference Presentation)

Science.gov (United States)

Zhao, Jianhua; Richer, Vincent; Al Jasser, Mohammed; Zandi, Soodabeh; Kollias, Nikiforos; Kalia, Sunil; Zeng, Haishan; Lui, Harvey

2016-02-01

Fluorescence signals depend on the intensity of the exciting light, the absorption properties of the constituent molecules, and the efficiency with which the absorbed photons are converted to fluorescence emission. The optical features and appearance of vitiligo have been explained primarily on the basis of reduced epidermal pigmentation, which results in abnormal white patches on the skin. The objective of this study is to explore the fluorescence properties of vitiligo and its adjacent normal skin using fluorescence excitation-emission matrix (EEM) spectroscopy. Thirty five (35) volunteers with vitiligo were acquired using a double-grating spectrofluorometer with excitation and emission wavelengths of 260-450 nm and 300-700 nm respectively. As expected, the most pronounced difference between the spectra obtained from vitiligo lesions compared to normally pigmented skin was that the overall fluorescence was much higher in vitiligo; these differences increased at shorter wavelengths, thus matching the characteristic spectral absorption of epidermal melanin. When comparing the fluorescence spectra from vitiligo to normal skin we detected three distinct spectral bands centered at 280nm, 310nm, and 335nm. The 280nm band may possibly be related to inflammation, whereas the 335 nm band may arise from collagen or keratin cross links. The source of the 310 nm band is uncertain; it is interesting to note its proximity to the 311 nm UV lamps used for vitiligo phototherapy. These differences are accounted for not only by changes in epidermal pigment content, but also by other optically active cutaneous biomolecules.

U(IV) fluorescence spectroscopy. A new speciation tool

Energy Technology Data Exchange (ETDEWEB)

Lehmann, Susanne; Brendler, Vinzenz [Helmholtz-Zentrum Dresden-Rossendorf e.V., Dresden (Germany). Surface Processes; Steudtner, Robin [Helmholtz-Zentrum Dresden-Rossendorf e.V., Dresden (Germany). Inst. of Resource Ecology

2017-06-01

We combined absorption and fluorescence spectroscopy to study the speciation of U(IV) in solution in concentrations down to 10{sup -6} M uranium. With our time-resolved laser-induced fluorescence setup we could determine the fluorescence decay time of U(IV) in perchloric as well as in chloric acid with 2.6 ± 0.3 ns at room temperature and 148.4 ± 6.5 ns at liquid nitrogen temperature. For the U(IV) sulfate system, we observed a bathochromic shift and a peak shape modification in the fluorescence spectra with increasing sulfate concentration in solution. Thus, the potential of U(IV) fluorescence for speciation analysis could be proven.

Precision analysis for standard deviation measurements of immobile single fluorescent molecule images.

Science.gov (United States)

DeSantis, Michael C; DeCenzo, Shawn H; Li, Je-Luen; Wang, Y M

2010-03-29

Standard deviation measurements of intensity profiles of stationary single fluorescent molecules are useful for studying axial localization, molecular orientation, and a fluorescence imaging system's spatial resolution. Here we report on the analysis of the precision of standard deviation measurements of intensity profiles of single fluorescent molecules imaged using an EMCCD camera.We have developed an analytical expression for the standard deviation measurement error of a single image which is a function of the total number of detected photons, the background photon noise, and the camera pixel size. The theoretical results agree well with the experimental, simulation, and numerical integration results. Using this expression, we show that single-molecule standard deviation measurements offer nanometer precision for a large range of experimental parameters.

Versatile single-molecule multi-color excitation and detection fluorescence setup for studying biomolecular dynamics

KAUST Repository

Sobhy, M. A.; Elshenawy, M. M.; Takahashi, Masateru; Whitman, B. H.; Walter, N. G.; Hamdan, S. M.

2011-01-01

Single-molecule fluorescence imaging is at the forefront of tools applied to study biomolecular dynamics both in vitro and in vivo. The ability of the single-molecule fluorescence microscope to conduct simultaneous multi-color excitation

Laser Spectroscopy of Ruthenium Containing Diatomic Molecules: RuH/D and RuP.

Science.gov (United States)

Adam, Allan G.; Konder, Ricarda M.; Nickerson, Nicole M.; Linton, Colan; Tokaryk, D. W.

2015-06-01

In the last few years, the Cheung group in Hong Kong and the Steimle group in Arizona have successfully studied several ruthenium containing diatomic molecules, RuX (X =C, O, N, B, using the laser-ablation molecular jet technique. Based on this success, the UNB spectroscopy group decided to try and find the optical signatures of other RuX molecules. Using CH_3OH and PH_3 as reactant gases, the RuH and RuP diatomic molecules have been detected in surveys of the 420 - 675 nm spectral region. RuD has also been made using fully deuterated methanol as a reactant. Dispersed fluorescence experiments have been performed to determine ground state vibrational frequencies and the presence of any low-lying electronic states. Rotationally resolved spectra for these molecules have also been taken and the analysis is proceeding. The most recent results will be presented. F. Wang et al., Journal of Chemical Physics 139, 174318 (2013). N. Wang et al., Journal of Physical Chemistry A 117, 13279 (2013). T. Steimle et al., Journal of Chemical Physics 119, 12965 (2003). N. Wang et al., Chemical Physics Letters 547, 21 (2012).

Detecting Kerogen as a Biosignature Using Colocated UV Time-Gated Raman and Fluorescence Spectroscopy.

Science.gov (United States)

Shkolyar, Svetlana; Eshelman, Evan J; Farmer, Jack D; Hamilton, David; Daly, Michael G; Youngbull, Cody

2018-04-01

The Mars 2020 mission will analyze samples in situ and identify any that could have preserved biosignatures in ancient habitable environments for later return to Earth. Highest priority targeted samples include aqueously formed sedimentary lithologies. On Earth, such lithologies can contain fossil biosignatures as aromatic carbon (kerogen). In this study, we analyzed nonextracted kerogen in a diverse suite of natural, complex samples using colocated UV excitation (266 nm) time-gated (UV-TG) Raman and laser-induced fluorescence spectroscopies. We interrogated kerogen and its host matrix in samples to (1) explore the capabilities of UV-TG Raman and fluorescence spectroscopies for detecting kerogen in high-priority targets in the search for possible biosignatures on Mars; (2) assess the effectiveness of time gating and UV laser wavelength in reducing fluorescence in Raman spectra; and (3) identify sample-specific issues that could challenge rover-based identifications of kerogen using UV-TG Raman spectroscopy. We found that ungated UV Raman spectroscopy is suited to identify diagnostic kerogen Raman bands without interfering fluorescence and that UV fluorescence spectroscopy is suited to identify kerogen. These results highlight the value of combining colocated Raman and fluorescence spectroscopies, similar to those obtainable by SHERLOC on Mars 2020, to strengthen the confidence of kerogen detection as a potential biosignature in complex natural samples. Key Words: Raman spectroscopy-Laser-induced fluorescence spectroscopy-Mars Sample Return-Mars 2020 mission-Kerogen-Biosignatures. Astrobiology 18, 431-453.

Portable fluorescence lifetime spectroscopy system for in-situ interrogation of biological tissues

Science.gov (United States)

Saito Nogueira, Marcelo; Cosci, Alessandro; Teixeira Rosa, Ramon Gabriel; Salvio, Ana Gabriela; Pratavieira, Sebastião; Kurachi, Cristina

2017-12-01

Fluorescence spectroscopy and lifetime techniques are potential methods for optical diagnosis and characterization of biological tissues with an in-situ, fast, and noninvasive interrogation. Several diseases may be diagnosed due to differences in the fluorescence spectra of targeted fluorophores, when, these spectra are similar, considering steady-state fluorescence, others may be detected by monitoring their fluorescence lifetime. Despite this complementarity, most of the current fluorescence lifetime systems are not robust and portable, and not being feasible for clinical applications. We describe the assembly of a fluorescence lifetime spectroscopy system in a suitcase, its characterization, and validation with clinical measurements of skin lesions. The assembled system is all encased and robust, maintaining its mechanical, electrical, and optical stability during transportation, and is feasible for clinical measurements. The instrument response function measured was about 300 ps, and the system is properly calibrated. At the clinical study, the system showed to be reliable, and the achieved spectroscopy results support its potential use as an auxiliary tool for skin diagnostics.

Visualizing Single-molecule DNA Replication with Fluorescence Microscopy

NARCIS (Netherlands)

Tanner, Nathan A.; Loparo, Joseph J.; Oijen, Antoine M. van

2009-01-01

We describe a simple fluorescence microscopy-based real-time method for observing DNA replication at the single-molecule level. A circular, forked DNA template is attached to a functionalized glass coverslip and replicated extensively after introduction of replication proteins and nucleotides. The

Detection of mechanical and disease stresses in citrus plants by fluorescence spectroscopy

Science.gov (United States)

Belasque, J., Jr.; Gasparoto, M. C. G.; Marcassa, L. G.

2008-04-01

We have investigated the detection of mechanical and disease stresses in citrus plants (Citrus limonia [L.] Osbeck) using laser-induced fluorescence spectroscopy. Due to its economic importance we have chosen to investigate the citrus canker disease, which is caused by the Xanthomonas axonopodis pv. citri bacteria. Mechanical stress was also studied because it plays an important role in the plant's infection by such bacteria. A laser-induced fluorescence spectroscopy system, composed of a spectrometer and a 532 nm10 mW excitation laser was used to perform fluorescence spectroscopy. The ratio of two chlorophyll fluorescence bands allows us to detect and discriminate between mechanical and disease stresses. This ability to discriminate may have an important application in the field to detect citrus canker infected trees.

Single Molecule Fluorescence: from Physical Fascination to Biological Relevance

OpenAIRE

Segers-Nolten, Gezina M.J.

2003-01-01

Confocal fluorescence microscopy is particularly well-known from the beautiful images that have been obtained with this technique from cells. Several cellular components could be nicely visualized simultaneously by staining them with different fluorophores. Not only for ensemble applications but also in single molecule research confocal fluorescence microscopy became a popular technique. In this thesis the possibilities are shown to study a complicated biological process, which is Nucleotide ...

Application of fluorescent and vibration spectroscopy for septic serum human albumin structure deformation during pathology

Science.gov (United States)

Zyubin, A.; Konstantinova, E.; Slezhkin, V.; Matveeva, K.; Samusev, I.; Bryukhanov, V.

2017-12-01

In this paper we perform results of conformational analysis of septic human serum albumin (HSA) carried out by Raman spectroscopy (RS), infrared (IR) spectroscopy and fluorescent spectroscopy. The main vibrational groups were identified and analyzed for septic HSA and its health control. Comparison between Raman and IR results were done. Fluorescent spectral changes of Trp-214 group were analyzed. Application of Raman, IR spectroscopy, fluorescent spectroscopy for conformational changes study of HSA during pathology were shown.

Assisted Interpretation of Laser-Induced Fluorescence Spectra of Egg-Based Binding Media Using Total Emission Fluorescence Spectroscopy

International Nuclear Information System (INIS)

Anglos, D.; Nevin, A.

2006-01-01

Laser-induced fluorescence (LIF) spectroscopy can provide nondestructive, qualitative analysis of protein-based binding media found in artworks. Fluorescence emissions from proteins in egg yolk and egg white are due to auto fluorescent aromatic amino acids as well as other native and age-related fluorophores, but the potential of fluorescence spectroscopy for the differentiation between binding media is dependent on the choice of a suitable excitation wavelength and limited by problems in interpretation. However, a better understanding of emission spectra associated with LIF can be achieved following comparisons with total emission fluorescence spectra where a series of consecutive emission spectra are recorded over a specific range. Results using nanosecond UV laser sources for LIF of egg-based binding media are presented which are rationalised following comparisons with total emission spectra. Specifically, fluorescence is assigned to tryptophan and oxidation products of amino acids; in the case of egg yolk, fatty-acid polymerisation and age-related degradation products account for the formation of fluorophores.

[Rapid identification of hogwash oil by using synchronous fluorescence spectroscopy].

Science.gov (United States)

Sun, Yan-Hui; An, Hai-Yang; Jia, Xiao-Li; Wang, Juan

2012-10-01

To identify hogwash oil quickly, the characteristic delta lambda of hogwash oil was analyzed by three dimensional fluorescence spectroscopy with parallel factor analysis, and the model was built up by using synchronous fluorescence spectroscopy with support vector machines (SVM). The results showed that the characteristic delta lambda of hogwash oil was 60 nm. Collecting original spectrum of different samples under the condition of characteristic delta lambda 60 nm, the best model was established while 5 principal components were selected from original spectrum and the radial basis function (RBF) was used as the kernel function, and the optimal penalty factor C and kernel function g were 512 and 0.5 respectively obtained by the grid searching and 6-fold cross validation. The discrimination rate of the model was 100% for both training sets and prediction sets. Thus, it is quick and accurate to apply synchronous fluorescence spectroscopy to identification of hogwash oil.

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

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.

Portable fluorescence lifetime spectroscopy system for in-situ interrogation of biological tissues.

Science.gov (United States)

Saito Nogueira, Marcelo; Cosci, Alessandro; Teixeira Rosa, Ramon Gabriel; Salvio, Ana Gabriela; Pratavieira, Sebastião; Kurachi, Cristina

2017-10-01

Fluorescence spectroscopy and lifetime techniques are potential methods for optical diagnosis and characterization of biological tissues with an in-situ, fast, and noninvasive interrogation. Several diseases may be diagnosed due to differences in the fluorescence spectra of targeted fluorophores, when, these spectra are similar, considering steady-state fluorescence, others may be detected by monitoring their fluorescence lifetime. Despite this complementarity, most of the current fluorescence lifetime systems are not robust and portable, and not being feasible for clinical applications. We describe the assembly of a fluorescence lifetime spectroscopy system in a suitcase, its characterization, and validation with clinical measurements of skin lesions. The assembled system is all encased and robust, maintaining its mechanical, electrical, and optical stability during transportation, and is feasible for clinical measurements. The instrument response function measured was about 300 ps, and the system is properly calibrated. At the clinical study, the system showed to be reliable, and the achieved spectroscopy results support its potential use as an auxiliary tool for skin diagnostics. (2017) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).

Fluorescent molecule incorporated metal-organic framework for fluoride sensing in aqueous solution

Energy Technology Data Exchange (ETDEWEB)

Zhao, Xudong, E-mail: zhaoxd_tykj@163.com; Wang, Yuanyang; Hao, Xiuli; Liu, Wen, E-mail: 13700509372@163.com

2017-04-30

Highlights: • Fluorescein sodium was successfully encapsulated in UiO-66 via in-situ synthesis. • FS@UiO-66 is one of the few Zr-MOF-based probes for fluoride so far. • FS@UiO-66 is a highly effective, fast-response and naked-eye sensor for fluoride. - Abstract: In this work, the fluorescent molecule (fluorescein sodium, FS) was successfully incorporated in the zirconium-based MOF (UiO-66) via in-situ synthesis method, which can be confirmed by FTIR spectra and fluorescence microscopic images. Based on this in-situ synthesis strategy, FS molecule can be immobilized tightly in the framework. Furthermore, the resulting FS@UiO-66 demonstrates to be a highly selective, real-time and naked-eye chemical sensor for fluoride in aqueous solution, which is mainly due to the release of FS molecule from FS@UiO-66 into the aqueous solution. Meanwhile, to the best of our knowledge, such Zr-MOF-based fluoride sensor is very rare so far. These results provide a promising approach to rationally design novel MOF-based fluorescent sensor for the target molecules.

Platinum plasmonic nanostructure arrays for massively parallel single-molecule detection based on enhanced fluorescence measurements

International Nuclear Information System (INIS)

Saito, Toshiro; Takahashi, Satoshi; Obara, Takayuki; Itabashi, Naoshi; Imai, Kazumichi

2011-01-01

We fabricated platinum bowtie nanostructure arrays producing fluorescence enhancement and evaluated their performance using two-photon photoluminescence and single-molecule fluorescence measurements. A comprehensive selection of suitable materials was explored by electromagnetic simulation and Pt was chosen as the plasmonic material for visible light excitation near 500 nm, which is preferable for multicolor dye-labeling applications like DNA sequencing. The observation of bright photoluminescence (λ = 500-600 nm) from each Pt nanostructure, induced by irradiation at 800 nm with a femtosecond laser pulse, clearly indicates that a highly enhanced local field is created near the Pt nanostructure. The attachment of a single dye molecule was attempted between the Pt triangles of each nanostructure by using selective immobilization chemistry. The fluorescence intensities of the single dye molecule localized on the nanostructures were measured. A highly enhanced fluorescence, which was increased by a factor of 30, was observed. The two-photon photoluminescence intensity and fluorescence intensity showed qualitatively consistent gap size dependence. However, the average fluorescence enhancement factor was rather repressed even in the nanostructure with the smallest gap size compared to the large growth of photoluminescence. The variation of the position of the dye molecule attached to the nanostructure may influence the wide distribution of the fluorescence enhancement factor and cause the rather small average value of the fluorescence enhancement factor.

Difference Raman spectroscopy of DNA molecules

International Nuclear Information System (INIS)

Anokhin, Andrey S; Yuzyuk, Yury I; Gorelik, Vladimir S; Dovbeshko, Galina I; Pyatyshev, Alexander Yu

2015-01-01

In this paper the micro-Raman spectra of calf DNA for different points of DNA sample have been recorded. The Raman spectra were made with help of difference Raman spectroscopy technique. Raman spectra were recorded with high spatial resolution from different points of the wet and dry samples in different spectral range (100÷4000cm −1 ) using two lasers: argon (514.5 nm) and helium -neon (632.8 nm). The significant differences in the Raman spectra for dry and wet DNA and for different points of DNA molecules were observed. The obtained data on difference Raman scattering spectra of DNA molecules may be used for identification of DNA types and for analysis of genetic information associated with the molecular structure of this molecule

Excitonic Coupling in Linear and Trefoil Trimer Perylenediimide Molecules Probed by Single-Molecule Spectroscopy

KAUST Repository

Yoo, Hyejin

2012-10-25

Perylenediimide (PDI) molecules are promising building blocks for photophysical studies of electronic interactions within multichromophore arrays. Such PDI arrays are important materials for fabrication of molecular nanodevices such as organic light-emitting diodes, organic semiconductors, and biosensors because of their high photostability, chemical and physical inertness, electron affinity, and high tinctorial strength over the entire visible spectrum. In this work, PDIs have been organized into linear (L3) and trefoil (T3) trimer molecules and investigated by single-molecule fluorescence microscopy to probe the relationship between molecular structures and interchromophoric electronic interactions. We found a broad distribution of coupling strengths in both L3 and T3 and hence strong/weak coupling between PDI units by monitoring spectral peak shifts in single-molecule fluorescence spectra upon sequential photobleaching of each constituent chromophore. In addition, we used a wide-field defocused imaging technique to resolve heterogeneities in molecular structures of L3 and T3 embedded in a PMMA polymer matrix. A systematic comparison between the two sets of experimental results allowed us to infer the correlation between intermolecular interactions and molecular structures. Our results show control of the PDI intermolecular interactions using suitable multichromophoric structures. © 2012 American Chemical Society.

Excitonic Coupling in Linear and Trefoil Trimer Perylenediimide Molecules Probed by Single-Molecule Spectroscopy

KAUST Repository

Yoo, Hyejin; Furumaki, Shu; Yang, Jaesung; Lee, Ji-Eun; Chung, Heejae; Oba, Tatsuya; Kobayashi, Hiroyuki; Rybtchinski, Boris; Wilson, Thea M.; Wasielewski, Michael R.; Vacha, Martin; Kim, Dongho

2012-01-01

Perylenediimide (PDI) molecules are promising building blocks for photophysical studies of electronic interactions within multichromophore arrays. Such PDI arrays are important materials for fabrication of molecular nanodevices such as organic light-emitting diodes, organic semiconductors, and biosensors because of their high photostability, chemical and physical inertness, electron affinity, and high tinctorial strength over the entire visible spectrum. In this work, PDIs have been organized into linear (L3) and trefoil (T3) trimer molecules and investigated by single-molecule fluorescence microscopy to probe the relationship between molecular structures and interchromophoric electronic interactions. We found a broad distribution of coupling strengths in both L3 and T3 and hence strong/weak coupling between PDI units by monitoring spectral peak shifts in single-molecule fluorescence spectra upon sequential photobleaching of each constituent chromophore. In addition, we used a wide-field defocused imaging technique to resolve heterogeneities in molecular structures of L3 and T3 embedded in a PMMA polymer matrix. A systematic comparison between the two sets of experimental results allowed us to infer the correlation between intermolecular interactions and molecular structures. Our results show control of the PDI intermolecular interactions using suitable multichromophoric structures. © 2012 American Chemical Society.

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

Fluorescence quenching of dye molecules near gold nanoparticles: radiative and nonradiative effects

NARCIS (Netherlands)

Dulkeith, E.; Morteani, A.C.; Niedereichholz, T.; Klar, T.A.; Feldman, J.; Levi, S.; van Veggel, F.C.J.M.; Reinhoudt, David; Möller, M.; Gittins, D.I.

2002-01-01

The radiative and nonradiative decay rates of lissamine dye molecules, chemically attached to differently sized gold nanoparticles, are investigated by means of time-resolved fluorescence experiments. A pronounced fluorescence quenching is observed already for the smallest nanoparticles of 1  nm

Single-Molecule Fluorescence Microscopy Reveals Local Diffusion Coefficients in the Pore Network of an Individual Catalyst Particle

NARCIS (Netherlands)

Hendriks, Frank|info:eu-repo/dai/nl/412642697; Meirer, Florian; Kubarev, Alexey V.; Ristanovic, Zoran|info:eu-repo/dai/nl/328233005; Roeffaers, Maarten B J; Vogt, Eelco T. C.|info:eu-repo/dai/nl/073717398; Bruijnincx, Pieter C. A.|info:eu-repo/dai/nl/33799529X; Weckhuysen, Bert M.|info:eu-repo/dai/nl/285484397

2017-01-01

We used single-molecule fluorescence microscopy to study self-diffusion of a feedstock-like probe molecule with nanometer accuracy in the macropores of a micrometer-sized, real-life fluid catalytic cracking (FCC) particle. Movies of single fluorescent molecules allowed their movement through the

Applying fluorescence correlation spectroscopy to investigate peptide-induced membrane disruption

DEFF Research Database (Denmark)

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

2017-01-01

to quantify leakage of fluorescent molecules of different sizes from large unilamellar lipid vesicles, thereby providing a tool for estimating the size of peptide-induced membrane disruptions. If fluorescently labeled lipids are incorporated into the membranes of the vesicles, FCS can also be used to obtain...

Emerging biomedical applications of time-resolved fluorescence spectroscopy

Science.gov (United States)

Lakowicz, Joseph R.; Szmacinski, Henryk; Koen, Peter A.

1994-07-01

Time-resolved fluorescence spectroscopy is presently regarded as a research tool in biochemistry, biophysics, and chemical physics. Advances in laser technology, the development of long-wavelength probes, and the use of lifetime-based methods are resulting in the rapid migration of time-resolved fluorescence to the clinical chemistry lab, to the patient's bedside, to flow cytometers, to the doctor's office, and even to home health care. Additionally, time-resolved imaging is now a reality in fluorescence microscopy, and will provide chemical imaging of a variety of intracellular analytes and/or cellular phenomena. In this overview paper we attempt to describe some of the opportunities available using chemical sensing based on fluorescence lifetimes, and to predict those applications of lifetime-based sensing which are most likely in the near future.

Fluorescence Correlation Spectroscopy to find the critical balance between extracellular association and intracellular dissociation of mRNA-complexes.

Science.gov (United States)

Zhang, Heyang; De Smedt, Stefaan C; Remaut, Katrien

2018-05-10

Fluorescence Correlation Spectroscopy (FCS) is a promising tool to study interactions on a single molecule level. The diffusion of fluorescent molecules in and out of the excitation volume of a confocal microscope leads to the fluorescence fluctuations that give information on the average number of fluorescent molecules present in the excitation volume and their diffusion coefficients. In this context, we complexed mRNA into lipoplexes and polyplexes and explored the association/dissociation degree of complexes by using gel electrophoresis and FCS. FCS enabled us to measure the association and dissociation degree of mRNA-based complexes both in buffer and protein-rich biological fluids such as human serum and ascitic fluid, which is a clear advantage over gel electrophoresis that was only applicable in protein-free buffer solutions. Furthermore, following the complex stability in buffer and biological fluids by FCS assisted to understand how complex characteristics, such as charge ratio and strength of mRNA binding, correlated to the transfection efficiency. We found that linear polyethyleneimine prevented efficient translation of mRNA, most likely due to a too strong mRNA binding, whereas the lipid based carrier Lipofectamine ® messengerMAX did succeed in efficient release and subsequent translation of mRNA in the cytoplasm of the cells. Overall, FCS is a reliable tool for the in depth characterization of mRNA complexes and can help us to find the critical balance keeping mRNA bound in complexes in the extracellular environment and efficient intracellular mRNA release leading to protein production. The delivery of messenger RNA (mRNA) to cells is promising to treat a variety of diseases. Therefore, the mRNA is typically packed in small lipid particles or polymer particles that help the mRNA to reach the cytoplasm of the cells. These particles should bind and carry the mRNA in the extracellular environment (e.g. blood, peritoneal fluid, ...), but should release

Fluorescence spectroscopy for medical and environmental diagnostics

International Nuclear Information System (INIS)

Johansson, Jonas.

1993-09-01

Fluorescence spectroscopy can be used for diagnostics in medical and environmental applications. The many aspects of fluorescence emission are utilized to enhance the accuracy of the diagnosis. A fluorescence detection system, based on nitrogen laser or dye laser excitation and optical multichannel detection, was constructed, and fluorescence spectra from human malignant tumours of various origins, were recorded. Tumour demarcation was observed using exogenous chromophores, as well as the endogenous tissue fluorescence. In particular, δ-amino levulinic acid was found to provide very good tumour demarcation. A multi-colour imaging system capable of simultaneous recording of four fluorescence images at selected wavelengths, was developed. Examples of processed images, based on the four sub-images, are shown for malignant tumours. In addition, data from photodynamic treatment of human malignant tumours are presented. Autofluorescence spectra from excised pieces of human atherosclerotic aorta and atherosclerotic coronary segment were found to be different from those of non-diseased vessels. Furthermore, fluorescence decay curves from atherosclerotic samples were found to differ from those of non-diseased samples. It is concluded that both spectral and temporal information should be utilized to enhance the demarcation. Methods for obtaining fluorescence data free from interference from blood, with applications to in vivo laser angioplasty of atherosclerosis, are discussed. The optical multichannel system and the multi-colour imaging system were integrated with a remote sensing system, originally used for environmental measurements, to obtain fluorescence spectra as well as fluorescence images of plants at a distance of up to 100 m. The fluorescence data from plants subject to environmental stress or senescent plants were found to differ from those obtained from healthy vegetation. 359 refs

Thousand-fold enhancement of single-molecule fluorescence near a single gold nanorod

NARCIS (Netherlands)

Yuan, H.; Khatua, S.; Zijlstra, P.; Yorulmaz, M.; Orrit, M.

2013-01-01

Single molecules: Large enhancements of single-molecule fluorescence up to 1100 times by using synthesized gold nanorods are reported (see picture). This high enhancement is achieved by selecting a dye with its adsorption and emission close to the surface plasmon resonance of the gold nanorods

Fluorescence spectroscopy as a tool for quality assessment of humic substances

Science.gov (United States)

Boguta, Patrycja

2016-04-01

*The studies were partly carried out within the framework of a research project. The project was financed from funds of National Science Center on the base of decision number DEC-2013/11/D/NZ9/02545. Fluorescence spectroscopy belongs to modern, non-destructive, rapid and relatively cheap methods, as well as for many years it was successfully used in studies of organic compounds in the fields of medicine, biology and chemistry. On the other hand, soil organic matter is a group of compounds with a complex spatial structure showing a large number of groups with different kinds of fluorophores. This could suggest the possibility of application of fluorescence spectroscopy in assessing the quality of humic substances as well as in monitoring of their chemical transformations. The aim of study was chemical description of humic and fulvic acids based on fluorescence spectra, as well as an attempt of evaluation of changes occurring under the influence of different pH and during interactions with various concentrations of metal. The humic and fulvic acids were isolated from chemically different soils. The measurements were carried out on Hitachi fluorescence spectrometer in solutions with a concentration of humic acids 40mg dm-3, at pH from 3 to 7, and for the evaluation of the metal impact: with increasing Zn concentrations (0-50mg dm-3). The fluorescence spectra were recorded in the form of synchronous and emission-excitation matrices (EEM). Studies have shown the presence of different groups of fluorophores. Synchronous spectra were characterized by a well-separated bands showing fluorescence in the area of low, medium and high wavelengths, suggesting the presence of structures, both weakly and strongly humified. EEM spectra revealed map of fluorophores within wide ranges of emission and excitation. Fluorophores differed in both position and intensity. The highest intensity was observed for compounds with the lowest humification degree which might be due to high amount

Emerging applications of fluorescence spectroscopy in medical microbiology field.

Science.gov (United States)

Shahzad, Aamir; Köhler, Gottfried; Knapp, Martin; Gaubitzer, Erwin; Puchinger, Martin; Edetsberger, Michael

2009-11-26

There are many diagnostic techniques and methods available for diagnosis of medically important microorganisms like bacteria, viruses, fungi and parasites. But, almost all these techniques and methods have some limitations or inconvenience. Most of these techniques are laborious, time consuming and with chances of false positive or false negative results. It warrants the need of a diagnostic technique which can overcome these limitations and problems. At present, there is emerging trend to use Fluorescence spectroscopy as a diagnostic as well as research tool in many fields of medical sciences. Here, we will critically discuss research studies which propose that Fluorescence spectroscopy may be an excellent diagnostic as well as excellent research tool in medical microbiology field with high sensitivity and specificity.

Emerging applications of fluorescence spectroscopy in medical microbiology field

Directory of Open Access Journals (Sweden)

Gaubitzer Erwin

2009-11-01

Full Text Available Abstract There are many diagnostic techniques and methods available for diagnosis of medically important microorganisms like bacteria, viruses, fungi and parasites. But, almost all these techniques and methods have some limitations or inconvenience. Most of these techniques are laborious, time consuming and with chances of false positive or false negative results. It warrants the need of a diagnostic technique which can overcome these limitations and problems. At present, there is emerging trend to use Fluorescence spectroscopy as a diagnostic as well as research tool in many fields of medical sciences. Here, we will critically discuss research studies which propose that Fluorescence spectroscopy may be an excellent diagnostic as well as excellent research tool in medical microbiology field with high sensitivity and specificity.

Optical Spectroscopy Of Charged Quantum Dot Molecules

Science.gov (United States)

Scheibner, M.; Bracker, A. S.; Stinaff, E. A.; Doty, M. F.; Gammon, D.; Ponomarev, I. V.; Reinecke, T. L.; Korenev, V. L.

2007-04-01

Coupling between two closely spaced quantum dots is observed by means of photoluminescence spectroscopy. Hole coupling is realized by rational crystal growth and heterostructure design. We identify molecular resonances of different excitonic charge states, including the important case of a doubly charged quantum dot molecule.

Single Fluorescent Molecules as Nano-Illuminators for Biological Structure and Function

Science.gov (United States)

Moerner, W. E.

2011-03-01

Since the first optical detection and spectroscopy of a single molecule in a solid (Phys. Rev. Lett. {62}, 2535 (1989)), much has been learned about the ability of single molecules to probe local nanoenvironments and individual behavior in biological and nonbiological materials in the absence of ensemble averaging that can obscure heterogeneity. Because each single fluorophore acts a light source roughly 1 nm in size, microscopic imaging of individual fluorophores leads naturally to superlocalization, or determination of the position of the molecule with precision beyond the optical diffraction limit, simply by digitization of the point-spread function from the single emitter. For example, the shape of single filaments in a living cell can be extracted simply by allowing a single molecule to move through the filament (PNAS {103}, 10929 (2006)). The addition of photoinduced control of single-molecule emission allows imaging beyond the diffraction limit (super-resolution) and a new array of acronyms (PALM, STORM, F-PALM etc.) and advances have appeared. We have used the native blinking and switching of a common yellow-emitting variant of green fluorescent protein (EYFP) reported more than a decade ago (Nature {388}, 355 (1997)) to achieve sub-40 nm super-resolution imaging of several protein structures in the bacterium Caulobacter crescentus: the quasi-helix of the actin-like protein MreB (Nat. Meth. {5}, 947 (2008)), the cellular distribution of the DNA binding protein HU (submitted), and the recently discovered division spindle composed of ParA filaments (Nat. Cell Biol. {12}, 791 (2010)). Even with these advances, better emitters would provide more photons and improved resolution, and a new photoactivatable small-molecule emitter has recently been synthesized and targeted to specific structures in living cells to provide super-resolution images (JACS {132}, 15099 (2010)). Finally, a new optical method for extracting three-dimensional position information based on

Optical absorption and fluorescence spectroscopy studies of Artepillin C, the major component of green propolis

Science.gov (United States)

Camuri, Isamara Julia; Costa, Adriano Batista; Ito, Amando Siuiti; Pazin, Wallance Moreira

2018-06-01

The bioactivity of propolis against several pathogens is well established, leading to the extensive consumption of that bee product to prevent diseases. Brazilian green propolis, collected by the species Apis mellifera, is one of the most consumed in the world. The chemical composition of green propolis is complex and it has been shown that it displays antioxidant, antimicrobial, anti-inflammatory and antitumor activities, especially due to the high content of Artepillin C. The molecule is a derivative of cinnamic acid with two prenylated groups, responsible for the improvement of the affinity of the compound for lipophilic environment. A carboxylic group (COOH) is also present in the molecule, making it a pH-sensitive compound and the pH-dependent structure of Artepillin C, may modulate its biological activity related to interactions with the cellular membrane of organisms and tissues. Molecular properties of Artepillin C on aqueous solution were examined by optical absorption, steady state and time-resolved fluorescence spectroscopies. Acid-base titration based on the spectral position of the near UV absorption band, resulted in the pKa value of 4.65 for the carboxylic group in Artepillin C. In acidic pH, below the pKa value, an absorption band raised around 350 nm at Artepillin C concentration above 50 μM, due to aggregation of the molecule. In neutral pH, with excitation at 310 nm, Artepillin C presents dual emission at 400 and 450 nm. In pH close to the pKa, the optical spectra show contribution from both protonated and deprotonated species. A three-exponential function was necessary to fit the intensity decays at the different pHs, dominated by a very short lifetime component, around 0.060 ns. The fast decay resulted in emission before fluorescence depolarization, and in values of fluorescence anisotropy higher than could be expected for monomeric forms of the compound. The results give fundamental knowledge about the protonation-deprotonation state of the

Single-molecule force spectroscopy: optical tweezers, magnetic tweezers and atomic force microscopy

Science.gov (United States)

Neuman, Keir C.; Nagy, Attila

2012-01-01

Single-molecule force spectroscopy has emerged as a powerful tool to investigate the forces and motions associated with biological molecules and enzymatic activity. The most common force spectroscopy techniques are optical tweezers, magnetic tweezers and atomic force microscopy. These techniques are described and illustrated with examples highlighting current capabilities and limitations. PMID:18511917

SISGR: Room Temperature Single-Molecule Detection and Imaging by Stimulated Emission Microscopy

Energy Technology Data Exchange (ETDEWEB)

Xie, Xiaoliang Sunney [Harvard Univ., Cambridge, MA (United States). Dept. of Chemistry and Chemical Biology

2017-03-13

Single-molecule spectroscopy has made considerable impact on many disciplines including chemistry, physics, and biology. To date, most single-molecule spectroscopy work is accomplished by detecting fluorescence. On the other hand, many naturally occurring chromophores, such as retinal, hemoglobin and cytochromes, do not have detectable fluorescence. There is an emerging need for single-molecule spectroscopy techniques that do not require fluorescence. In the last proposal period, we have successfully demonstrated stimulated emission microscopy, single molecule absorption, and stimulated Raman microscopy based on a high-frequency modulation transfer technique. These first-of-a- kind new spectroscopy/microscopy methods tremendously improved our ability to observe molecules that fluorescence weakly, even to the limit of single molecule detection for absorption measurement. All of these methods employ two laser beams: one (pump beam) excites a single molecule to a real or virtual excited state, and the other (probe beam) monitors the absorption/emission property of the single. We extract the intensity change of the probe beam with high sensitivity by implementing a high-frequency phase-sensitive detection scheme, which offers orders of magnitude improvement in detection sensitivity over direct absorption/emission measurement. However, single molecule detection based on fluorescence or absorption is fundamentally limited due to their broad spectral response. It is important to explore other avenues in single molecule detection and imaging which provides higher molecular specificity for studying a wide variety of heterogeneous chemical and biological systems. This proposal aimed to achieve single-molecule detection sensitivity with near resonance stimulated Raman scattering (SRS) microscopy. SRS microscopy was developed in our lab as a powerful technique for imaging heterogeneous samples based on their intrinsic vibrational contrasts, which provides much higher molecular

Dual time-resolved temperature-jump fluorescence and infrared spectroscopy for the study of fast protein dynamics.

Science.gov (United States)

Davis, Caitlin M; Reddish, Michael J; Dyer, R Brian

2017-05-05

Time-resolved temperature-jump (T-jump) coupled with fluorescence and infrared (IR) spectroscopy is a powerful technique for monitoring protein dynamics. Although IR spectroscopy of the polypeptide amide I mode is more technically challenging, it offers complementary information because it directly probes changes in the protein backbone, whereas, fluorescence spectroscopy is sensitive to the environment of specific side chains. With the advent of widely tunable quantum cascade lasers (QCL) it is possible to efficiently probe multiple IR frequencies with high sensitivity and reproducibility. Here we describe a dual time-resolved T-jump fluorescence and IR spectrometer and its application to study protein folding dynamics. A Q-switched Ho:YAG laser provides the T-jump source for both time-resolved IR and fluorescence spectroscopy, which are probed by a QCL and Ti:Sapphire laser, respectively. The Ho:YAG laser simultaneously pumps the time-resolved IR and fluorescence spectrometers. The instrument has high sensitivity, with an IR absorbance detection limit of jump induced difference spectrum from 50ns to 0.5ms. This study demonstrates the power of the dual time-resolved T-jump fluorescence and IR spectroscopy to resolve complex folding mechanisms by complementary IR absorbance and fluorescence measurements of protein dynamics. Copyright © 2017 Elsevier B.V. All rights reserved.

Fluorescence lifetime spectroscopy in multiple-scattering environments: an application to biotechnology

Science.gov (United States)

Cerussi, Albert E.; Gratton, Enrico; Fantini, Sergio

1999-07-01

Over the past few years, there has been significant research activity devoted to the application of fluorescence spectroscopy to strongly scattering media, where photons propagate diffusely. Much of this activity focused on fluorescence as a source of contrast enhancement in optical tomography. Our efforts have emphasized the quantitative recovery of fluorescence parameters for spectroscopy. Using a frequency-domain diffusion-based model, we have successfully recovered the lifetime, the absolute quantum yield, the fluorophore concentration, and the emission spectrum of the fluorophore, as well as the absorption and the reduced scattering coefficients at the emission wavelength of the medium in different measurements. In this contribution, we present a sensitive monitor of the binding between ethidium bromide and bovine cells in fresh milk. The spectroscopic contrast was the approximately tenfold increase in the ethidium bromide lifetime upon binding to DNA. The measurement clearly demonstrated that we could quantitatively measure the density of cells in the milk, which is an application vital to the tremendous economic burden of bovine subclinical mastitis detection. Furthermore, we may in principle use the spirit of this technique as a quantitative monitor of the binding of fluorescent drugs inside tissues. This is a first step towards lifetime spectroscopy in tissues.

Spectroscopy of fractional Josephson vortex molecules

Energy Technology Data Exchange (ETDEWEB)

Goldobin, Edward; Gaber, Tobias; Buckenmaier, Kai; Kienzle, Uta; Sickinger, Hanna; Koelle, Dieter; Kleiner, Reinhold [Physikalisches Institut - Experimentalphysik II, Center for Collective Quantum Phenomena, Universitaet Tuebingen, Auf der Morgenstelle 14, D-72076 Tuebingen (Germany)

2010-07-01

Using tiny current injectors we create {kappa} discontinuities of the Josephson phase in a long Josephson junction. The junction reacts at the discontinuities by creating fractional Josephson vortices of size {lambda}{sub J} pinned at them. Such vortices carry the flux {phi}, which is a fraction of the magnetic flux quantum {phi}{sub 0}{approx}2.07 x 10{sup -15} Wb. Being pinned, a fractional vortex has an eigenfrequency (localized mode), which depends on {kappa} and applied bias current, and which lays within the plasma gap. If one considers a molecule consisting of several coupled fractional vortices, the eigenfrequency will split into several modes. We report on spectroscopy of a fractional vortex molecule performed in the thermal regime.

Using fluorescence correlation spectroscopy to study conformational changes in denatured proteins.

Science.gov (United States)

Sherman, Eilon; Itkin, Anna; Kuttner, Yosef Yehuda; Rhoades, Elizabeth; Amir, Dan; Haas, Elisha; Haran, Gilad

2008-06-01

Fluorescence correlation spectroscopy (FCS) is a sensitive analytical tool that allows dynamics and hydrodynamics of biomolecules to be studied under a broad range of experimental conditions. One application of FCS of current interest is the determination of the size of protein molecules in the various states they sample along their folding reaction coordinate, which can be accessed through the measurement of diffusion coefficients. It has been pointed out that the analysis of FCS curves is prone to artifacts that may lead to erroneous size determination. To set the stage for FCS studies of unfolded proteins, we first show that the diffusion coefficients of small molecules as well as proteins can be determined accurately even in the presence of high concentrations of co-solutes that change the solution refractive index significantly. Indeed, it is found that the Stokes-Einstein relation between the measured diffusion coefficient and solution viscosity holds even in highly concentrated glycerol or guanidinium hydrochloride (GuHCl) solutions. These measurements form the basis for an investigation of the structure of the denatured state of two proteins, the small protein L and the larger, three-domain protein adenylate kinase (AK). FCS is found useful for probing expansion in the denatured state beyond the unfolding transition. It is shown that the denatured state of protein L expands as the denaturant concentration increases, in a process akin to the transition from a globule to a coil in polymers. This process continues at least up to 5 M GuHCl. On the other hand, the denatured state of AK does not seem to expand much beyond 2 M GuHCl, a result that is in qualitative accord with single-molecule fluorescence histograms. Because both the unfolding transition and the coil-globule transition of AK occur at a much lower denaturant concentration than those of protein L, a possible correlation between the two phenomena is suggested.

Photoelectron spectroscopy of heavy atoms and molecules

International Nuclear Information System (INIS)

White, M.G.

1979-07-01

The importance of relativistic interactions in the photoionization of heavy atoms and molecules has been investigated by the technique of photoelectron spectroscopy. In particular, experiments are reported which illustrate the effects of the spin-orbit interaction in the neutral ground state, final ionic states and continuum states of the photoionization target

Fluorescence spectroscopy for the detection of potentially malignant disorders of the oral cavity: analysis of 30 cases

International Nuclear Information System (INIS)

Francisco, A L N; Correr, W R; Kurachi, C; Azevedo, L H; Galletta, V K; Pinto, C A L; Kowalski, L P

2014-01-01

Oral cancer is a major health problem worldwide and although early diagnosis of potentially malignant and malignant diseases is associated with better treatment results, a large number of cancers are initially misdiagnosed, with unfortunate consequences for long-term survival. Fluorescence spectroscopy is a noninvasive modality of diagnostic approach using induced fluorescence emission in tumors that can improve diagnostic accuracy. The objective of this study was to determine the ability to discriminate between normal oral mucosa and potentially malignant disorders by fluorescence spectroscopy. Fluorescence investigation under 408 and 532 nm excitation wavelengths was performed on 60 subjects, 30 with potentially malignant disorders and 30 volunteers with normal mucosa. Data was analyzed to correlate fluorescence patterns with clinical and histopathological diagnostics. Fluorescence spectroscopy used as a point measurement technique resulted in a great variety of spectral information. In a qualitative analysis of the fluorescence spectral characteristics of each type of injury evaluated, it was possible to discriminate between normal and abnormal oral mucosa. The results show the potential use of fluorescence spectroscopy for an improved discrimination of oral disorders. (paper)

Memory effects in single-molecule spectroscopy

International Nuclear Information System (INIS)

Schmitt, Daniel T.; Schulz, Michael; Reineker, Peter

2007-01-01

From the time series of LH2 optical single-molecule fluorescence excitation spectra of Rhodospirillum molischianum the memory function of the Mori-Zwanzig equation for the optical intensity is derived numerically. We show that the time dependence of the excited states is determined by at least three different non-Markovian stochastic processes with decay constants for the Mori-Zwanzig kernel on the order of 1-5min -1 . We suggest that this decay stems from the conformational motion of the protein scaffold of LH2

Laser-excited fluorescence spectroscopy of oxide glasses

International Nuclear Information System (INIS)

Weber, M.J.

1977-01-01

Laser-induced fluorescence line narrowing was applied to investigate the local fields and interactions of paramagnetic ions in oxide glasses. Studies included the site dependence of energy levels, radiative and nonradiative transition probabilities, homogeneous line broadening, and ion--ion energy transfer of rare earth ions. These results and the experimental techniques are reviewed briefly; the use of paramagnetic ions other than the rare earths is also considered. Recently, laser-excited fluorescence spectroscopy was used to investigate modifications in the local structure of lithium borate glass caused by compositional changes and phase separation and the site dependence of nonradiative relaxation of paramagnetic ions by multiphonon processes. These results and their implications are discussed. 6 figures

Blinking effect and the use of quantum dots in single molecule spectroscopy

International Nuclear Information System (INIS)

Rombach-Riegraf, Verena; Oswald, Peter; Bienert, Roland; Petersen, Jan; Domingo, M.P.; Pardo, Julian; Gräber, P.; Galvez, E.M.

2013-01-01

Highlights: ► It is possible to eliminate the blinking effect of a water-soluble QD. ► We provide a direct method to study protein function and dynamics at the single level. ► QD, potent tool for single molecule studies of biochemical and biological processes. -- Abstract: Luminescent semiconductor nanocrystals (quantum dots, QD) have unique photo-physical properties: high photostability, brightness and narrow size-tunable fluorescence spectra. Due to their unique properties, QD-based single molecule studies have become increasingly more popular during the last years. However QDs show a strong blinking effect (random and intermittent light emission), which may limit their use in single molecule fluorescence studies. QD blinking has been widely studied and some hypotheses have been done to explain this effect. Here we summarise what is known about the blinking effect in QDs, how this phenomenon may affect single molecule studies and, on the other hand, how the “on”/“off” states can be exploited in diverse experimental settings. In addition, we present results showing that site-directed binding of QD to cysteine residues of proteins reduces the blinking effect. This option opens a new possibility of using QDs to study protein–protein interactions and dynamics by single molecule fluorescence without modifying the chemical composition of the solution or the QD surface.

Blinking effect and the use of quantum dots in single molecule spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Rombach-Riegraf, Verena; Oswald, Peter; Bienert, Roland; Petersen, Jan [Albert-Ludwigs-Universitaet Freiburg, Institut fuer Physikalische Chemie, Albertstrasse 23a, 79104 Freiburg (Germany); Domingo, M.P. [Instituto de Carboquimica (CSIC), Miguel Luesma 4, 50018 Zaragoza (Spain); Pardo, Julian [Grupo Apoptosis, Inmunidad y Cancer, Departamento Bioquimica y Biologia Molecular y Celular, Fac. Ciencias, Universidad de Zaragoza, Zaragoza (Spain); Fundacion Aragon I-D (ARAID), Gobierno de Aragon, Zaragoza (Spain); Immune Effector Cells Group, Aragon Health Research Institute (IIS Aragon), Biomedical Research Centre of Aragon (CIBA) Fundacion Aragon I-D - ARAID, Gobierno de Aragon, Zaragoza (Spain); Graeber, P. [Albert-Ludwigs-Universitaet Freiburg, Institut fuer Physikalische Chemie, Albertstrasse 23a, 79104 Freiburg (Germany); Galvez, E.M., E-mail: eva@icb.csic.es [Instituto de Carboquimica (CSIC), Miguel Luesma 4, 50018 Zaragoza (Spain); Immune Effector Cells Group, Aragon Health Research Institute (IIS Aragon), Biomedical Research Centre of Aragon (CIBA) Fundacion Aragon I-D - ARAID, Gobierno de Aragon, Zaragoza (Spain)

2013-01-04

Highlights: Black-Right-Pointing-Pointer It is possible to eliminate the blinking effect of a water-soluble QD. Black-Right-Pointing-Pointer We provide a direct method to study protein function and dynamics at the single level. Black-Right-Pointing-Pointer QD, potent tool for single molecule studies of biochemical and biological processes. -- Abstract: Luminescent semiconductor nanocrystals (quantum dots, QD) have unique photo-physical properties: high photostability, brightness and narrow size-tunable fluorescence spectra. Due to their unique properties, QD-based single molecule studies have become increasingly more popular during the last years. However QDs show a strong blinking effect (random and intermittent light emission), which may limit their use in single molecule fluorescence studies. QD blinking has been widely studied and some hypotheses have been done to explain this effect. Here we summarise what is known about the blinking effect in QDs, how this phenomenon may affect single molecule studies and, on the other hand, how the 'on'/'off' states can be exploited in diverse experimental settings. In addition, we present results showing that site-directed binding of QD to cysteine residues of proteins reduces the blinking effect. This option opens a new possibility of using QDs to study protein-protein interactions and dynamics by single molecule fluorescence without modifying the chemical composition of the solution or the QD surface.

Mapping vortex-like hydrodynamic flow in microfluidic networks using fluorescence correlation spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Ke Liu; Yu Tian; Burrows, Sean M.; Reif, Randall D. [Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX 79409-1061 (United States); Pappas, Dimitri, E-mail: d.pappas@ttu.edu [Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX 79409-1061 (United States)

2009-09-28

The ability to quickly measure flow parameters in microfluidic devices is critical for micro total analysis system ({mu}TAS) applications. Macrofluidic methods to assess flow suffer from limitations that have made conventional methods unsuitable for the flow behavior profiling. Single molecule fluorescence correlation spectroscopy (FCS) has been employed in our study to characterize the fluidic vortex generating at a T-shape junction of microscale channels. Due to its high spatial and temporal resolution, the corresponding magnitudes relative to different flow rates in the main channel can be quantitatively differentiated using flow time ({tau}{sub F}) measurements of dye molecules traversing the detection volume in buffer solution. Despite the parabolic flow in the channel upstream, a heterogeneous distribution of flow has been detected across the channel intersection. In addition, our current observations also confirmed the aspect of vortex-shaped flow in low-shear design that was developed previously for cell culture. This approach not only overcomes many technical barriers for examining hydrodynamic vortices and movements in miniature structures without physically integrating any probes, but it is also especially useful for the hydrodynamic studies in polymer-glass based micro -reactor and -mixer.

Investigating single molecule adhesion by atomic force spectroscopy.

Science.gov (United States)

Stetter, Frank W S; Kienle, Sandra; Krysiak, Stefanie; Hugel, Thorsten

2015-02-27

Atomic force spectroscopy is an ideal tool to study molecules at surfaces and interfaces. An experimental protocol to couple a large variety of single molecules covalently onto an AFM tip is presented. At the same time the AFM tip is passivated to prevent unspecific interactions between the tip and the substrate, which is a prerequisite to study single molecules attached to the AFM tip. Analyses to determine the adhesion force, the adhesion length, and the free energy of these molecules on solid surfaces and bio-interfaces are shortly presented and external references for further reading are provided. Example molecules are the poly(amino acid) polytyrosine, the graft polymer PI-g-PS and the phospholipid POPE (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine). These molecules are desorbed from different surfaces like CH3-SAMs, hydrogen terminated diamond and supported lipid bilayers under various solvent conditions. Finally, the advantages of force spectroscopic single molecule experiments are discussed including means to decide if truly a single molecule has been studied in the experiment.

Revealing the Raft Domain Organization in the Plasma Membrane by Single-Molecule Imaging of Fluorescent Ganglioside Analogs.

Science.gov (United States)

Suzuki, Kenichi G N; Ando, Hiromune; Komura, Naoko; Konishi, Miku; Imamura, Akihiro; Ishida, Hideharu; Kiso, Makoto; Fujiwara, Takahiro K; Kusumi, Akihiro

2018-01-01

Gangliosides have been implicated in a variety of physiological processes, particularly in the formation and function of raft domains in the plasma membrane. However, the scarcity of suitable fluorescent ganglioside analogs had long prevented us from determining exactly how gangliosides perform their functions in the live-cell plasma membrane. With the development of new fluorescent ganglioside analogs, as described by Komura et al. (2017), this barrier has been broken. We can now address the dynamic behaviors of gangliosides in the live-cell plasma membrane, using fluorescence microscopy, particularly by single-fluorescent molecule imaging and tracking. Single-molecule tracking of fluorescent GM1 and GM3 revealed that these molecules are transiently and dynamically recruited to monomers (monomer-associated rafts) and homodimer rafts of the raftophilic GPI-anchored protein CD59 in quiescent cells, with exponential residency times of 12 and 40ms, respectively, in a manner dependent on raft-lipid interactions. Upon CD59 stimulation, which induces CD59-cluster signaling rafts, the fluorescent GM1 and GM3 analogs were recruited to the signaling rafts, with a lifetime of 48ms. These results represent the first direct evidence that GPI-anchored receptors and gangliosides interact in a cholesterol-dependent manner. Furthermore, they show that gangliosides continually move in and out of rafts that contain CD59 in an extremely dynamic manner, with much higher frequency than expected previously. Such studies would not have been possible without fluorescent ganglioside probes, which exhibit native-like behavior and single-molecule tracking. In this chapter, we review the methods for single-molecule tracking of fluorescent ganglioside analogs and the results obtained by applying these methods. © 2018 Elsevier Inc. All rights reserved.

Endogenous synchronous fluorescence spectroscopy (SFS) of basal cell carcinoma-initial study

Science.gov (United States)

Borisova, E.; Zhelyazkova, Al.; Keremedchiev, M.; Penkov, N.; Semyachkina-Glushkovskaya, O.; Avramov, L.

2016-01-01

The human skin is a complex, multilayered and inhomogeneous organ with spatially varying optical properties. Analysis of cutaneous fluorescence spectra could be a very complicated task; therefore researchers apply complex mathematical tools for data evaluation, or try to find some specific approaches, that would simplify the spectral analysis. Synchronous fluorescence spectroscopy (SFS) allows improving the spectral resolution, which could be useful for the biological tissue fluorescence characterization and could increase the tumour detection diagnostic accuracy.

Single-Molecule Chemistry with Surface- and Tip-Enhanced Raman Spectroscopy.

Science.gov (United States)

Zrimsek, Alyssa B; Chiang, Naihao; Mattei, Michael; Zaleski, Stephanie; McAnally, Michael O; Chapman, Craig T; Henry, Anne-Isabelle; Schatz, George C; Van Duyne, Richard P

2017-06-14

Single-molecule (SM) surface-enhanced Raman spectroscopy (SERS) and tip-enhanced Raman spectroscopy (TERS) have emerged as analytical techniques for characterizing molecular systems in nanoscale environments. SERS and TERS use plasmonically enhanced Raman scattering to characterize the chemical information on single molecules. Additionally, TERS can image single molecules with subnanometer spatial resolution. In this review, we cover the development and history of SERS and TERS, including the concept of SERS hot spots and the plasmonic nanostructures necessary for SM detection, the past and current methodologies for verifying SMSERS, and investigations into understanding the signal heterogeneities observed with SMSERS. Moving on to TERS, we cover tip fabrication and the physical origins of the subnanometer spatial resolution. Then, we highlight recent advances of SMSERS and TERS in fields such as electrochemistry, catalysis, and SM electronics, which all benefit from the vibrational characterization of single molecules. SMSERS and TERS provide new insights on molecular behavior that would otherwise be obscured in an ensemble-averaged measurement.

Two-focus fluorescence correlation spectroscopy

International Nuclear Information System (INIS)

Dertinger, T.

2007-05-01

Fluorescence Correlation Spectroscopy (FCS) has been invented more than 30 years ago and experienced a renaissance after stable and affordable laser sources and low-noise single-photon detectors have become available. Its ability to measure diffusion coefficients at nanomolar concentrations of analyte made it a widely used tool in biophysics. However, in recent years it has been shown by many authors that aberrational (e.g. astigmatism) and photophysical effects (e.g. optical saturation) may influence the result of an FCS experiment dramatically, so that a precise and reliable estimation of the diffusion coefficient is no longer possible. In this thesis, we report on the development, implementation, and application of a new and robust modification of FCS that we termed two-focus FCS (2fFCS) and which fulfils two requirements: (i) It introduces an external ruler into the measurement by generating two overlapping laser foci of precisely known and fixed distance. (ii) These two foci and corresponding detection regions are generated in such a way that the corresponding molecule detection functions (MDFs) are sufficiently well described by a simple two-parameter model yielding accurate diffusion coefficients when applied to 2fFCS data analysis. Both these properties enable us to measure absolute values of the diffusion coefficient with an accuracy of a few percent. Moreover, it will turn out that the new technique is robust against refractive index mismatch, coverslide thickness deviations, and optical saturation effects, which so often trouble conventional FCS measurements. This thesis deals mainly with the introduction of the new measurement scheme, 2fFCS, but also presents several applications with far-reaching importance. (orig.)

Two-focus fluorescence correlation spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Dertinger, T.

2007-05-15

Fluorescence Correlation Spectroscopy (FCS) has been invented more than 30 years ago and experienced a renaissance after stable and affordable laser sources and low-noise single-photon detectors have become available. Its ability to measure diffusion coefficients at nanomolar concentrations of analyte made it a widely used tool in biophysics. However, in recent years it has been shown by many authors that aberrational (e.g. astigmatism) and photophysical effects (e.g. optical saturation) may influence the result of an FCS experiment dramatically, so that a precise and reliable estimation of the diffusion coefficient is no longer possible. In this thesis, we report on the development, implementation, and application of a new and robust modification of FCS that we termed two-focus FCS (2fFCS) and which fulfils two requirements: (i) It introduces an external ruler into the measurement by generating two overlapping laser foci of precisely known and fixed distance. (ii) These two foci and corresponding detection regions are generated in such a way that the corresponding molecule detection functions (MDFs) are sufficiently well described by a simple two-parameter model yielding accurate diffusion coefficients when applied to 2fFCS data analysis. Both these properties enable us to measure absolute values of the diffusion coefficient with an accuracy of a few percent. Moreover, it will turn out that the new technique is robust against refractive index mismatch, coverslide thickness deviations, and optical saturation effects, which so often trouble conventional FCS measurements. This thesis deals mainly with the introduction of the new measurement scheme, 2fFCS, but also presents several applications with far-reaching importance. (orig.)

Application of laser fluorescence spectroscopy by two-photon excitation into atomic hydrogen density measurement in reactive plasmas

International Nuclear Information System (INIS)

Kajiwara, Toshinori; Takeda, Kazuyuki; Kim, Hee Je; Park, Won Zoo; Muraoka, Katsunori; Akazaki, Masanori; Okada, Tatsuo; Maeda, Mitsuo.

1990-01-01

Density profiles of hydrogen atoms in reactive plasmas of hydrogen and methane gases were measured, for the first time, using the laser fluorescence spectroscopy by two-photon excitation of Lyman beta transition and observation at the Balmer alpha radiation. Absolute density determinations showed atomic densities of around 3 x 10 17 m -3 , or the degree of dissociation to be 10 -4 . Densities along the axis perpendicular to the RF electrode showed peaked profiles, which were due to the balance of atomic hydrogen production by electron impact on molecules against diffusion loss to the walls. (author)

Synthesis and Application of Polymeric Fluorescent Compounds Based on Coumarin

Directory of Open Access Journals (Sweden)

Guojun Liu

2015-06-01

Full Text Available In this work, a multifunctional yellowing inhibitor was synthesized by the Pechmann method. In order to obtain the target compound, 7-hydroxy-4-methyl coumarin was prepared by using the raw materials of resorcinol and ethyl acetoacetate, with toluene-p-sulfonic acid as the catalyst. New polymeric fluorescent compounds were synthesized by connecting the 7-hydroxy-4-methyl coumarin, the hindered amine light stabilizer 4-amion-2,2,6,6-tetramentylniperidine, and a series of polyethylene glycol segments into the same molecule with cyanuric chloride as a bridge. The structures of the synthesized molecules were confirmed by FT-IR, 1H NMR, and elemental analysis. The luminescent properties of the fluorescent compounds were studied by UV-vis spectroscopy and fluorescence spectroscopy. The integration effect between the fluorescent compounds and paper was tested by a scanning electron microscope. The light stability effect on the paper sheet was tested using an ultraviolet aging apparatus. The results indicate that the polymeric fluorescent compounds had a positive effect on the light stability of the high-yield pulp.

In-vivo optical detection of cancer using chlorin e6 – polyvinylpyrrolidone induced fluorescence imaging and spectroscopy

International Nuclear Information System (INIS)

Chin, William WL; Thong, Patricia SP; Bhuvaneswari, Ramaswamy; Soo, Khee Chee; Heng, Paul WS; Olivo, Malini

2009-01-01

Photosensitizer based fluorescence imaging and spectroscopy is fast becoming a promising approach for cancer detection. The purpose of this study was to examine the use of the photosensitizer chlorin e6 (Ce6) formulated in polyvinylpyrrolidone (PVP) as a potential exogenous fluorophore for fluorescence imaging and spectroscopic detection of human cancer tissue xenografted in preclinical models as well as in a patient. Fluorescence imaging was performed on MGH human bladder tumor xenografted on both the chick chorioallantoic membrane (CAM) and the murine model using a fluorescence endoscopy imaging system. In addition, fiber optic based fluorescence spectroscopy was performed on tumors and various normal organs in the same mice to validate the macroscopic images. In one patient, fluorescence imaging was performed on angiosarcoma lesions and normal skin in conjunction with fluorescence spectroscopy to validate Ce6-PVP induced fluorescence visual assessment of the lesions. Margins of tumor xenografts in the CAM model were clearly outlined under fluorescence imaging. Ce6-PVP-induced fluorescence imaging yielded a specificity of 83% on the CAM model. In mice, fluorescence intensity of Ce6-PVP was higher in bladder tumor compared to adjacent muscle and normal bladder. Clinical results confirmed that fluorescence imaging clearly captured the fluorescence of Ce6-PVP in angiosarcoma lesions and good correlation was found between fluorescence imaging and spectral measurement in the patient. Combination of Ce6-PVP induced fluorescence imaging and spectroscopy could allow for optical detection and discrimination between cancer and the surrounding normal tissues. Ce6-PVP seems to be a promising fluorophore for fluorescence diagnosis of cancer

Raman and fluorescent scattering by molecules embedded in small particles

International Nuclear Information System (INIS)

Chew, H.W.; McNulty, P.J.

1983-01-01

We have formulated a model for fluorescent and Raman scattering by molecules embedded in or in the vicinity of small particles. The model takes into account the size, shape, refractive index, and morphology of the host particles. Analytic and numerical results have been obtained for spherical (one and more layers, including magnetic dipole transitions) cylindrical and spheroidal particles. Particular attention has been given to the spherical case with fluorescent/Raman scatterers uniformly distributed in the particles radiating both coherently and incohorently. Depolarization effects have been studied with suitable averaging process, and good agreement with experiment has been obtained. Analytic and numerical results have been obtained for the elastic scattering of evanescent waves; these results are useful for the study of fluorescent under excitation by evanescent waves

Laser induced fluorescence spectroscopy for FTU

International Nuclear Information System (INIS)

Hughes, T.P.

1995-07-01

Laser induced fluorescence spectroscopy (LIFS) is based on the absorption of a short pulse of tuned laser light by a group of atoms and the observation of the resulting fluorescence radiation from the excited state. Because the excitation is resonant it is very efficient, and the fluorescence can be many times brighter than the normal spontaneous emission, so low number densities of the selected atoms can be detected and measured. Good spatial resolution can be achieved by using a narrow laser beam. If the laser is sufficiently monochromatic, and it can be tuned over the absorption line profile of the selected atoms, information can also be obtained about the velocities of the atoms from the Doppler effect which can broaden and shift the line. In this report two topics are examined in detail. The first is the effect of high laser irradiance, which can cause 'power broadening' of the apparent absorption line profile. The second is the effect of the high magnetic field in FTU. Detailed calculations are given for LIFS of neutral iron and molybdenum atoms, including the Zeeman effect, and the implementation of LIFS for these atoms on FTU is discussed

"FluSpec": A Simulated Experiment in Fluorescence Spectroscopy

Science.gov (United States)

Bigger, Stephen W.; Bigger, Andrew S.; Ghiggino, Kenneth P.

2014-01-01

The "FluSpec" educational software package is a fully contained tutorial on the technique of fluorescence spectroscopy as well as a simulator on which experiments can be performed. The procedure for each of the experiments is also contained within the package along with example analyses of results that are obtained using the software.

Spectroscopy and Chemistry of Cold Molecules

Science.gov (United States)

Momose, Takamasa

2012-06-01

Molecules at low temperatures are expected to behave quite differently from those at high temperatures because pronounced quantum effects emerge from thermal averages. Even at 10 K, a significant enhancement of reaction cross section is expected due to tunneling and resonance effects. Chemistry at this temperature is very important in order to understand chemical reactions in interstellar molecular clouds. At temperatures lower than 1 K, collisions and intermolecular interactions become qualitatively different from those at high temperatures because of the large thermal de Broglie wavelength of molecules. Collisions at these temperatures must be treated as the interference of molecular matter waves, but not as hard sphere collisions. A Bose-Einstein condensate is a significant state of matter as a result of coherent matter wave interaction. Especially, dense para-H_2 molecules are predicted to become a condensate even around 1 K. A convenient method to investigate molecules around 1 K is to dope molecules in cold matrices. Among various matrices, quantum hosts such as solid para-H_2 and superfluid He nano-droplets have been proven to be an excellent host for high-resolution spectroscopy. Rovibrational motion of molecules in these quantum hosts is well quantized on account of the weak interactions and the softness of quantum environment. The linewidths of infrared spectra of molecules in the quantum hosts are extremely narrow compared with those in other matrices. The sharp linewidths allow us to resolve fine spectral structures originated in subtle interactions between guest and host molecules. In this talk, I will describe how the splitting and lineshape of high-resolution spectra of molecules in quantum hosts give us new information on the static and dynamical interactions of molecules in quantum medium. The topics include dynamical response of superfluid environment upon rotational excitation, and possible superfluid phase of para-H_2 clusters. I will also

Time-Gated Raman Spectroscopy for Quantitative Determination of Solid-State Forms of Fluorescent Pharmaceuticals.

Science.gov (United States)

Lipiäinen, Tiina; Pessi, Jenni; Movahedi, Parisa; Koivistoinen, Juha; Kurki, Lauri; Tenhunen, Mari; Yliruusi, Jouko; Juppo, Anne M; Heikkonen, Jukka; Pahikkala, Tapio; Strachan, Clare J

2018-04-03

Raman spectroscopy is widely used for quantitative pharmaceutical analysis, but a common obstacle to its use is sample fluorescence masking the Raman signal. Time-gating provides an instrument-based method for rejecting fluorescence through temporal resolution of the spectral signal and allows Raman spectra of fluorescent materials to be obtained. An additional practical advantage is that analysis is possible in ambient lighting. This study assesses the efficacy of time-gated Raman spectroscopy for the quantitative measurement of fluorescent pharmaceuticals. Time-gated Raman spectroscopy with a 128 × (2) × 4 CMOS SPAD detector was applied for quantitative analysis of ternary mixtures of solid-state forms of the model drug, piroxicam (PRX). Partial least-squares (PLS) regression allowed quantification, with Raman-active time domain selection (based on visual inspection) improving performance. Model performance was further improved by using kernel-based regularized least-squares (RLS) regression with greedy feature selection in which the data use in both the Raman shift and time dimensions was statistically optimized. Overall, time-gated Raman spectroscopy, especially with optimized data analysis in both the spectral and time dimensions, shows potential for sensitive and relatively routine quantitative analysis of photoluminescent pharmaceuticals during drug development and manufacturing.

Video-rate confocal microscopy for single-molecule imaging in live cells and superresolution fluorescence imaging.

Science.gov (United States)

Lee, Jinwoo; Miyanaga, Yukihiro; Ueda, Masahiro; Hohng, Sungchul

2012-10-17

There is no confocal microscope optimized for single-molecule imaging in live cells and superresolution fluorescence imaging. By combining the swiftness of the line-scanning method and the high sensitivity of wide-field detection, we have developed a, to our knowledge, novel confocal fluorescence microscope with a good optical-sectioning capability (1.0 μm), fast frame rates (fluorescence detection efficiency. Full compatibility of the microscope with conventional cell-imaging techniques allowed us to do single-molecule imaging with a great ease at arbitrary depths of live cells. With the new microscope, we monitored diffusion motion of fluorescently labeled cAMP receptors of Dictyostelium discoideum at both the basal and apical surfaces and obtained superresolution fluorescence images of microtubules of COS-7 cells at depths in the range 0-85 μm from the surface of a coverglass. Copyright © 2012 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Micro-hole array fluorescent sensor based on AC-Dielectrophoresis (DEP) for simultaneous analysis of nano-molecules

Science.gov (United States)

Kim, Hye Jin; Kang, Dong-Hoon; Lee, Eunji; Hwang, Kyo Seon; Shin, Hyun-Joon; Kim, Jinsik

2018-02-01

We propose a simple fluorescent bio-chip based on two types of alternative current-dielectrophoretic (AC-DEP) force, attractive (positive DEP) and repulsive (negative DEP) force, for simultaneous nano-molecules analysis. Various radius of micro-holes on the bio-chip are designed to apply the different AC-DEP forces, and the nano-molecules are concentrated inside the micro-hole arrays according to the intensity of the DEP force. The bio-chip was fabricated by Micro Electro Mechanical system (MEMS) technique, and was composed of two layers; a SiO2 layer and Ta/Pt layer were accomplished for an insulation layer and a top electrode with micro-hole arrays to apply electric fields for DEP force, respectively. Each SiO2 and Ta/Pt layers were deposited by thermal oxidation and sputtering, and micro-hole arrays were fabricated with Inductively Coupled Plasma (ICP) etching process. For generation of each positive and negative DEP at micro-holes, we applied two types of sine-wave AC voltage with different frequency range alternately. The intensity of the DEP force was controlled by the radius of the micro-hole and size of nano-molecule, and calculated with COMSOL multi-physics. Three types of nano-molecules labelled with different fluorescent dye were used and the intensity of nano-molecules was examined by the fluorescent optical analysis after applying the DEP force. By analyzing the fluorescent intensities of the nano-molecules, we verify the various nano-molecules in analyte are located successfully inside corresponding micro-holes with different radius according to their size.

Fluorescent scattering by molecules embedded in small particles. Progress report, May 1, 1977--October 31, 1978

International Nuclear Information System (INIS)

Chew, H.; McNulty, P.J.

1978-01-01

A model for the fluorescence and Raman scattering by molecules that comprise or are embedded in small particles was developed and numerical calculations performed. The emphasis during this first year of the contract was on writing and testing the computer programs necessary for numerical calculations and on demonstrating the extent of the potential effects that the geometrical and optical properties of the particle would have on the Raman and fluorescent emissions. For the purpose of demonstrating effects emphasis was focused upon the case of isotropically polarizable molecules that fluoresce or Raman scatter through electric dipole transitions. Some preliminary results are described. One result of these investigations that is of particular significance for remote sensing of pollutants is that it would be a serious mistake to use inelastic scattering techniques such as Raman and fluorescent scattering for quantitative assay of specific molecules in aerosols containing particulates without taking into account the size, structure and refractive index of the particles. A list of publications is included

APD detectors for biological fluorescence spectroscopy

International Nuclear Information System (INIS)

Mazeres, S.; Borrel, V.; Magenc, C.; Courrech, J.L.; Bazer-Bachi, R.

2006-01-01

Fluorescence spectroscopy is a very convenient and widely used method for studying the molecular background of biological processes [L. Salome, J.L. Cazeil, A. Lopez, J.F. Tocanne, Eur. Biophys. J. 27 (1998) 391-402]. Chromophores are included in the structure under study and a flash of laser light induces fluorescence (Fluorescence Recovery After Photo-bleaching), the decay of which yields information on the polarity, the speed of rotation, and the speed of diffusion as well as on the temporal and spatial evolution of interactions between molecular species. The method can even be used to study living cells [J.F. Tocanne, L. Cezanne, A. Lopez, Prog. Lipid Res. 33 (1994) 203-237, L. Cezanne, A. Lopez, F. Loste, G. Parnaud, O. Saurel, P. Demange, J.F. Tocanne, Biochemistry 38 (1999) 2779-2786]. This is classically performed with a PM-based system. For biological reasons a decrease of the excitation of the cells is highly desirable. Because the fluorescence response then becomes fainter a significant improvement in detector capability would be welcome. We present here results obtained with an Avalanche Photo Diode (APD)-based system. The small sensitive area of detection allows a very significant improvement in signal/noise ratio, improvement in gain, and the opening-up of a new parameter space. With these new detectors we can begin the study of information transmission between cells through morphine receptors. This work involves both electronics engineers and biophysicists, so results and techniques in both fields will be presented here

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.

Spectroscopy of Molecules in Extreme Rotational States Using AN Optical Centrifuge

Science.gov (United States)

Mullin, Amy S.; Toro, Carlos; Echibiri, Geraldine; Liu, Qingnan

2012-06-01

Our lab has developed a high-power optical centrifuge that is capable of trapping and spinning large number densities of molecules into extreme rotational states. By coupling this device with high resolution transient IR absorption spectroscopy, we measure the time-resolved behavior and energy profiles of individual ro-vibrational states of molecules in very high rotational states. Recent results will be discussed on the spectroscopy of new rotational states, collisional dynamics in the optical centrifuge, spatially-dependent energy profiles and possibilities for new chemistry induced by centrifugal forces.

Efficient encapsulation of chloroform with cryptophane-M and the formation of exciplex studied by fluorescence spectroscopy.

Science.gov (United States)

Shi, Yanqi; Li, Xueming; Yang, Jianchun; Gao, Fang; Tao, Chuanyi

2011-03-01

Efficient encapsulation of small molecules with supermolecules is one of significantly important subjects due to strong application potentials. This article presents the interaction between cryptophane-M and chloroform by fluorescence spectroscopy. The sonicated cryptophane-M solution exhibits light green color in chloroform, and the solid obtained from the evaporation of chloroform also has different color from that of cryptophane-M. In contrast, the sonicated cryptophane-M solutions in other solvents are colorless, and the solid obtained from the evaporation of these solvents has the same color as that of cryptophane-M. Furthermore, the freshly prepared cryptophane-M solution in different solvents is almost colorless, and the solid obtained from the evaporation of these solvents displays the same color as that of cryptophane-M. Although the sonicated cryptophane-M solutions in different solvents have very similar absorption spectra, they exhibit quite different emission spectra in chloroform. In contrast, the freshly-prepared cryptophane-M solutions show similar absorption and emission spectroscopy in various solvents. The variation of the fluorescence spectroscopy in binary solvents with the increasing chloroform ratio suggests that cryptophane-M and chloroform form a 1:1 exciplex, and the binding constant is estimated to be 292.95 M(-1). Although all solvents are able to enter into the cavity of cryptophane-M, only chloroform can stay in the cavity of cryptophane-M for a while, which is mostly due to the strong intermolecular interaction between cryptophane-M and chloroform, and this results in the formation of the exciplex between them. © Springer Science+Business Media, LLC 2010

Limitations of superfluid helium droplets as host system revealed by electronic spectroscopy of embedded molecules

Energy Technology Data Exchange (ETDEWEB)

Premke, Tobias

2016-02-19

Superfluid helium nanodroplets serve a unique cryogenic host system ideal to prepare cold molecules and clusters. Structures as well as dynamic processes can be examined by means of high resolution spectroscopy. Dopant spectra are accompanied by helium-induced spectroscopic features which reveal information on the dopant to helium interaction. For this reason the experimental research focuses on the investigation of such helium-induced effects in order to provide new information on the microsolvation inside the droplets. Since the quantitative understanding of helium-induced spectral features is essential to interpret molecular spectra recorded in helium droplets, this study contributes further experimental details on microsolvation in superfluid helium droplets. For this purpose two contrary systems were examined by means of high resolution electronic spectroscopy. The first one, phthalocyanine (Pc), is a planar organic molecule offering a huge and planar surface to the helium atoms and thus, the non-superfluid helium solvation layer can form different structures. The second system is iodine and in contrast to Pc it is of simple molecular shape. That means that in this case different complex structures of the non-superfluid helium solvation layer and the dopant can be expected to be avoided. Thus, both molecules should show clear differences in their microsolvation behavior. In this work a detailed examination of different spectroscopic properties of phthalocyanine is given by means of fluorescence excitation and dispersed emission spectroscopy. It raises legitimate doubts about the assignment of experimentally observed signals to features predicted by the model of the microsolvation. Even though there are no experimental observations which disprove the empirical model for the solvation in helium droplets, an unambiguous assignment of the helium-induced spectroscopic structures is often not possible. In the second part of this work, the investigation of the

Developing DNA nanotechnology using single-molecule fluorescence.

Science.gov (United States)

Tsukanov, Roman; Tomov, Toma E; Liber, Miran; Berger, Yaron; Nir, Eyal

2014-06-17

CONSPECTUS: An important effort in the DNA nanotechnology field is focused on the rational design and manufacture of molecular structures and dynamic devices made of DNA. As is the case for other technologies that deal with manipulation of matter, rational development requires high quality and informative feedback on the building blocks and final products. For DNA nanotechnology such feedback is typically provided by gel electrophoresis, atomic force microscopy (AFM), and transmission electron microscopy (TEM). These analytical tools provide excellent structural information; however, usually they do not provide high-resolution dynamic information. For the development of DNA-made dynamic devices such as machines, motors, robots, and computers this constitutes a major problem. Bulk-fluorescence techniques are capable of providing dynamic information, but because only ensemble averaged information is obtained, the technique may not adequately describe the dynamics in the context of complex DNA devices. The single-molecule fluorescence (SMF) technique offers a unique combination of capabilities that make it an excellent tool for guiding the development of DNA-made devices. The technique has been increasingly used in DNA nanotechnology, especially for the analysis of structure, dynamics, integrity, and operation of DNA-made devices; however, its capabilities are not yet sufficiently familiar to the community. The purpose of this Account is to demonstrate how different SMF tools can be utilized for the development of DNA devices and for structural dynamic investigation of biomolecules in general and DNA molecules in particular. Single-molecule diffusion-based Förster resonance energy transfer and alternating laser excitation (sm-FRET/ALEX) and immobilization-based total internal reflection fluorescence (TIRF) techniques are briefly described and demonstrated. To illustrate the many applications of SMF to DNA nanotechnology, examples of SMF studies of DNA hairpins and

Assessment of the unidentified organic matter fraction in fogwater using fluorescence spectroscopy

Science.gov (United States)

Valsaraj, K.; Birdwell, J.

2010-07-01

Dissolved organic matter (DOM) in fogwaters from southeastern Louisiana and central-eastern China has been characterized using excitation-emission matrix (EEM) fluorescence spectroscopy. The results demonstrate that fluorescence spectroscopy can be used to obtain a qualitative assessment of the large fraction of fogwater organic carbon (~40 - 80% by weight) that cannot be identified in terms of specific chemical compounds. The method has the principle advantage that it can be applied at natural abundance concentrations, thus eliminating the need for large sample volumes required to isolate DOM for characterization by other spectroscopic (NMR, FTIR) and chemical (elemental) analyses. It was anticipated that the fogwater organic matter fluorescence spectra would resemble those of surface and rain waters, containing peaks indicative of both humic substances and fluorescent amino acids. Humic- and protein-like fluorophores were observed in the fogwater spectra and fluorescence-derived indices had values comparable to other natural waters. Biological character (intensity of tyrosine and tryptophan peaks) was found to increase with organic carbon concentration. Fogwater organic matter appears to contain a mixture of terrestrially- and microbially-derived material. The fluorescence results show that most of the unidentified fogwater organic carbon can be represented by humic-like and biologically-derived substances similar to those present in other aquatic systems.

On-Line Monitoring of Fermentation Processes by Near Infrared and Fluorescence Spectroscopy

DEFF Research Database (Denmark)

Svendsen, Carina

Monitoring and control of fermentation processes is important to ensure high product yield, product quality and product consistency. More knowledge on on-line analytical techniques such as near infrared and fluorescence spectroscopy is desired in the fermentation industry to increase the efficiency...... of on-line monitoring systems. The primary aim of this thesis is to elucidate and explore the dynamics in fermentation processes by spectroscopy. Though a number of successful on-line lab-scale monitoring systems have been reported, it seems that several challenges are still met, which limits the number...... of full-scale systems implemented in industrial fermentation processes. This thesis seeks to achieve a better understanding of the techniques near infrared and fluorescence spectroscopy and thereby to solve some of the challenges that are encountered. The thesis shows the advantages of applying real...

Application of fluorescence spectroscopy and imaging in the detection of a photosensitizer in photodynamic therapy

Science.gov (United States)

Zang, Lixin; Zhao, Huimin; Zhang, Zhiguo; Cao, Wenwu

2017-02-01

Photodynamic therapy (PDT) is currently an advanced optical technology in medical applications. However, the application of PDT is limited by the detection of photosensitizers. This work focuses on the application of fluorescence spectroscopy and imaging in the detection of an effective photosenzitizer, hematoporphyrin monomethyl ether (HMME). Optical properties of HMME were measured and analyzed based on its absorption and fluorescence spectra. The production mechanism of its fluorescence emission was analyzed. The detection device for HMME based on fluorescence spectroscopy was designed. Ratiometric method was applied to eliminate the influence of intensity change of excitation sources, fluctuates of excitation sources and photo detectors, and background emissions. The detection limit of this device is 6 μg/L, and it was successfully applied to the diagnosis of the metabolism of HMME in the esophageal cancer cells. To overcome the limitation of the point measurement using fluorescence spectroscopy, a two-dimensional (2D) fluorescence imaging system was established. The algorithm of the 2D fluorescence imaging system is deduced according to the fluorescence ratiometric method using bandpass filters. The method of multiple pixel point addition (MPPA) was used to eliminate fluctuates of signals. Using the method of MPPA, SNR was improved by about 30 times. The detection limit of this imaging system is 1.9 μg/L. Our systems can be used in the detection of porphyrins to improve the PDT effect.

Membrane mobility and microdomain association of the dopaminetransporter studied with fluorescence correlation spectroscopy and fluorescence recovery after photobleaching

DEFF Research Database (Denmark)

Adkins, Erika; Samuvel, Devadoss; Fog, Jacob

2007-01-01

To investigate microdomain association of the dopamine transporter (DAT), we employed FCS (fluorescence correlation spectroscopy) and FRAP (fluorescence recovery after photobleaching). In non-neuronal cells (HEK293), FCS measurements revealed for the YFP-DAT (DAT tagged with yellow fluorescent...... protein) a diffusion coefficient (D) of ~3.6 × 10-9 cm2/s, consistent with a relatively freely diffusible protein. In neuronally derived cells (N2a), we were unable to perform FCS measurements on plasma membrane-associated protein due to photobleaching, suggesting partial immobilization...

Fluorescence detection of single molecules using pulsed near-field optical excitation and time correlated photon counting

International Nuclear Information System (INIS)

Ambrose, W.P.; Goodwin, P.M.; Martin, J.C.; Keller, R.A.

1994-01-01

Pulsed excitation, time correlated single photon counting and time gated detection are used in near-field optical microscopy to enhance fluorescence images and measure the fluorescence lifetimes of single molecules of Rhodamine 6G on silica surfaces. Time gated detection is used to reject prompt scattered background and to improve the image signal to noise ratio. The excited state lifetime of a single Rhodamine 6G molecule is found to depend on the position of the near-field probe. We attribute the lifetime variations to spontaneous emission rate alterations by the fluorescence reflected from and quenching by the aluminum coated probe

Time gated fluorescence lifetime imaging and micro-volume spectroscopy using two-photon excitation

NARCIS (Netherlands)

Sytsma, J.; Vroom, J.M.; de Grauw, C.J.; Gerritsen, H.C.

A scanning microscope utilizing two-photon excitation in combination with fluorescence lifetime contrast is presented. The microscope makes use of a tunable femtosecond titanium:sapphire laser enabling the two-photon excitation of a broad range of fluorescent molecules, including UV probes.

Transition paths in single-molecule force spectroscopy.

Science.gov (United States)

Cossio, Pilar; Hummer, Gerhard; Szabo, Attila

2018-03-28

In a typical single-molecule force spectroscopy experiment, the ends of the molecule of interest are connected by long polymer linkers to a pair of mesoscopic beads trapped in the focus of two laser beams. At constant force load, the total extension, i.e., the end-to-end distance of the molecule plus linkers, is measured as a function of time. In the simplest systems, the measured extension fluctuates about two values characteristic of folded and unfolded states, with occasional transitions between them. We have recently shown that molecular (un)folding rates can be recovered from such trajectories, with a small linker correction, as long as the characteristic time of the bead fluctuations is shorter than the residence time in the unfolded (folded) state. Here, we show that accurate measurements of the molecular transition path times require an even faster apparatus response. Transition paths, the trajectory segments in which the molecule (un)folds, are properly resolved only if the beads fluctuate more rapidly than the end-to-end distance of the molecule. Therefore, over a wide regime, the measured rates may be meaningful but not the transition path times. Analytic expressions for the measured mean transition path times are obtained for systems diffusing anisotropically on a two-dimensional free energy surface. The transition path times depend on the properties both of the molecule and of the pulling device.

The photodissociation and reaction dynamics of vibrationally excited molecules

Energy Technology Data Exchange (ETDEWEB)

Crim, F.F. [Univ. of Wisconsin, Madison (United States)

1993-12-01

This research determines the nature of highly vibrationally excited molecules, their unimolecular reactions, and their photodissociation dynamics. The goal is to characterize vibrationally excited molecules and to exploit that understanding to discover and control their chemical pathways. Most recently the author has used a combination of vibrational overtone excitation and laser induced fluorescence both to characterize vibrationally excited molecules and to study their photodissociation dynamics. The author has also begun laser induced grating spectroscopy experiments designed to obtain the electronic absorption spectra of highly vibrationally excited molecules.

Quantification of protein based on single-molecule counting by total internal reflection fluorescence microscopy with adsorption equilibrium

International Nuclear Information System (INIS)

Wang Lei; Xu Guang; Shi Zhikun; Jiang Wei; Jin Wenrui

2007-01-01

We developed a sensitive single-molecule imaging method for quantification of protein by total internal reflection fluorescence microscopy with adsorption equilibrium. In this method, the adsorption equilibrium of protein was achieved between solution and glass substrate. Then, fluorescence images of protein molecules in a evanescent wave field were taken by a highly sensitive electron multiplying charge coupled device. Finally, the number of fluorescent spots corresponding to the protein molecules in the images was counted. Alexa Fluor 488-labeled goat anti-rat IgG(H + L) was chosen as the model protein. The spot number showed an excellent linear relationship with protein concentration. The concentration linear range was 5.4 x 10 -11 to 8.1 x 10 -10 mol L -1

Quantitative analysis of essential oils of Thymus daenensis using laser-induced fluorescence and Raman spectroscopy.

Science.gov (United States)

Khoshroo, H; Khadem, H; Bahreini, M; Tavassoli, S H; Hadian, J

2015-11-10

Laser-induced fluorescence and Raman spectroscopy are used for the investigation of different genotypes of Thymus daenensis native to the Ilam province of Iran. Different genotypes of T. daenensis essential oils, labeled T1 through T7, possess slight differences with regard to the composition of the thymol. The gas chromatography-mass spectrometry (GC-MS) method is performed to determine the concentration of each constituent as a reference method. The Raman spectra of different concentrations of pure thymol dissolved in hexane as standard samples are obtained via a laboratory prototype Raman spectroscopy setup for the calculation of the calibration curve. The regression coefficient and limit of detection are calculated. The possibility of the differentiation of different genotypes of T. daenensis is also examined by laser-induced fluorescence spectroscopy, although we do not know the exact amounts of their components. All the fluorescence spectral information is used jointly by cluster analysis to differentiate between 7 genotypes. Our results demonstrate the acceptable precision of Raman spectroscopy with GC-MS and corroborate the capacity of Raman spectroscopy in applications in the quantitative analysis field. Furthermore, the cluster analysis results show that laser-induced fluorescence spectroscopy is an acceptable technique for the rapid classification of different genotypes of T. daenensis without having any previous information of their exact amount of constituents. So, the ability to rapidly and nondestructively differentiate between genotypes makes it possible to efficiently select high-quality herbs from many samples.

Terahertz-Radiation-Enhanced Emission of Fluorescence from Gas Plasma

International Nuclear Information System (INIS)

Liu Jingle; Zhang, X.-C.

2009-01-01

We report the study of femtosecond laser-induced air plasma fluorescence under the illumination of terahertz (THz) pulses. Semiclassical modeling and experimental verification indicate that time-resolved THz radiation-enhanced emission of fluorescence is dominated by the electron kinetics and the electron-impact excitation of gas molecules or ions. We demonstrate that the temporal waveform of the THz field could be retrieved from the transient enhanced fluorescence, making omnidirectional, coherent detection available for THz time-domain spectroscopy.

Single-Molecule Interfacial Electron Transfer

Energy Technology Data Exchange (ETDEWEB)

Ho, Wilson [Univ. of California, Irvine, CA (United States)

2018-02-03

Interfacial electron transfer (ET) plays an important role in many chemical and biological processes. Specifically, interfacial ET in TiO₂-based systems is important to solar energy technology, catalysis, and environmental remediation technology. However, the microscopic mechanism of interfacial ET is not well understood with regard to atomic surface structure, molecular structure, bonding, orientation, and motion. In this project, we used two complementary methodologies; single-molecule fluorescence spectroscopy, and scanning-tunneling microscopy and spectroscopy (STM and STS) to address this scientific need. The goal of this project was to integrate these techniques and measure the molecular dependence of ET between adsorbed molecules and TiO₂ semiconductor surfaces and the ET induced reactions such as the splitting of water. The scanning probe techniques, STM and STS, are capable of providing the highest spatial resolution but not easily time-resolved data. Single-molecule fluorescence spectroscopy is capable of good time resolution but requires further development to match the spatial resolution of the STM. The integrated approach involving Peter Lu at Bowling Green State University (BGSU) and Wilson Ho at the University of California, Irvine (UC Irvine) produced methods for time and spatially resolved chemical imaging of interfacial electron transfer dynamics and photocatalytic reactions. An integral aspect of the joint research was a significant exchange of graduate students to work at the two institutions. This project bridged complementary approaches to investigate a set of common problems by working with the same molecules on a variety of solid surfaces, but using appropriate techniques to probe under ambient (BGSU) and ultrahigh vacuum (UCI) conditions. The molecular level understanding of the fundamental interfacial electron transfer processes obtained in this joint project will be important for developing efficient light harvesting

UV-visible and resonance Raman spectroscopy of halogen molecules in clathrate hydrates

Energy Technology Data Exchange (ETDEWEB)

Janda, K.C.; Kerenskaya, G.; Goldsheleger, I.U.; Apkarian, V.A.; Fleischer, E.B. [California Univ., Irvine, CA (United States). Dept. of Chemistry

2008-07-01

Resonance Raman spectroscopy was used to study halogen clathrate hydrate solids. In particular, this paper presented an ultraviolet-visible spectra for a polycrystalline sample of chlorine clathrate hydrate and two single crystal samples of bromine clathrate hydrate. UV-visible spectroscopy was used to study the interactions between the halogen guest molecule and the host water lattice. The spectrum for chlorine hydrate had a strong temperature dependence, while the spectra for bromine clathrate hydrate single crystals had a stable cubic type 2 structure as well as a tetragonal structure. A metastable cubic type 1 structure was also observed. Resonance Raman spectroscopy showed how the molecules fit into the host cages. 25 refs., 2 tabs., 7 figs.

Cytosine Radical Cations: A Gas-Phase Study Combining IRMPD Spectroscopy, UVPD Spectroscopy, Ion-Molecule Reactions, and Theoretical Calculations

Czech Academy of Sciences Publication Activity Database

Lesslie, M.; Lawler, J. T.; Dang, A.; Korn, J. A.; Bím, Daniel; Steinmetz, V.; Maitre, P.; Tureček, F.; Ryzhov, V.

2017-01-01

Roč. 18, č. 10 (2017), s. 1293-1301 ISSN 1439-4235 Institutional support: RVO:61388963 Keywords : ion-molecule reactions * IRMPD spectroscopy * nucleobases * radical ions * UVPD spectroscopy Subject RIV: CF - Physical ; Theoretical Chemistry OBOR OECD: Physical chemistry Impact factor: 3.075, year: 2016

Membrane mobility and microdomain association of the dopamine transporter studied with fluorescence correlation spectroscopy and fluorescence recovery after photobleaching

DEFF Research Database (Denmark)

Adkins, Erika M; Samuvel, Devadoss J; Fog, Jacob U

2007-01-01

To investigate microdomain association of the dopamine transporter (DAT), we employed FCS (fluorescence correlation spectroscopy) and FRAP (fluorescence recovery after photobleaching). In non-neuronal cells (HEK293), FCS measurements revealed for the YFP-DAT (DAT tagged with yellow fluorescent...... protein) a diffusion coefficient (D) of approximately 3.6 x 10(-9) cm2/s, consistent with a relatively freely diffusible protein. In neuronally derived cells (N2a), we were unable to perform FCS measurements on plasma membrane-associated protein due to photobleaching, suggesting partial immobilization...

Single molecule microscopy and spectroscopy: concluding remarks.

Science.gov (United States)

van Hulst, Niek F

2015-01-01

Chemistry is all about molecules: control, synthesis, interaction and reaction of molecules. All too easily on a blackboard, one draws molecules, their structures and dynamics, to create an insightful picture. The dream is to see these molecules in reality. This is exactly what "Single Molecule Detection" provides: a look at molecules in action at ambient conditions; a breakthrough technology in chemistry, physics and biology. Within the realms of the Royal Society of Chemistry, the Faraday Discussion on "Single Molecule Microscopy and Spectroscopy" was a very appropriate topic for presentation, deliberation and debate. Undoubtedly, the Faraday Discussions have a splendid reputation in stimulating scientific debates along the traditions set by Michael Faraday. Interestingly, back in the 1830's, Faraday himself pursued an experiment that led to the idea that atoms in a compound were joined by an electrical component. He placed two opposite electrodes in a solution of water containing a dissolved compound, and observed that one of the elements of the compound accumulated on one electrode, while the other was deposited on the opposite electrode. Although Faraday was deeply opposed to atomism, he had to recognize that electrical forces were responsible for the joining of atoms. Probably a direct view on the atoms or molecules in his experiment would have convinced him. As such, Michael Faraday might have liked the gathering at Burlington House in September 2015 (). Surely, with the questioning eyes of his bust on the 1st floor corridor, the non-believer Michael Faraday has incited each passer-by to enter into discussion and search for deeper answers at the level of single molecules. In these concluding remarks, highlights of the presented papers and discussions are summarized, complemented by a conclusion on future perspectives.

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.

Laser Spectroscopy and AB Initio Calculations on the TaF Molecule

Science.gov (United States)

Ng, Kiu Fung; Zou, Wenli; Liu, Wenjian; Cheung, Allan S. C.

2016-06-01

Electronic transition spectrum of the tantalum monoflouride (TaF) molecule in the spectral region between 448 and 520 nm has been studied using the technique of laser-ablation/reaction free jet expansion and laser induced fluorescence spectroscopy. TaF molecule was produced by reacting laser-ablated tantalum atoms with sulfur hexafluoride gas seeded in argon. Sixteen vibrational bands with resolved rotational structure have been recorded and analyzed, which were organized into six electronic transition systems and the ground state has been identified to be the X3Σ-(0+) state with bond length, ro, and equilibrium vibrational frequency, ωe, determined to be 1.8209 Å and 700.1 wn respectively. In addition, four vibrational bands belong to another transition system involving lower state with Ω = 2 component has also been analyzed. All observed transitions are with ΔΩ = 0. Least-squares fit of the measured line positions yielded molecular constants for the electronic states involved. The Λ-S and Ω states of TaF were calculated at the state-averaged complete active space self-consistent field (SA-CASSCF) and the subsequent internally contracted multi-reference configuration interaction with singles and doubles and Davidson's cluster correction (MRCISD+Q) levels of theory with the active space of 4 electrons in 6 orbitals, that is, the molecular orbitals corresponding to Ta 5d6s are active. The spin-orbit coupling (SOC) is calculated by the state-interaction approach at the SA-CASSCF level via the relativistic effective core potentials (RECPs) spin-orbit operator, where the diagonal elements of the spin-orbit matrix are replaced by the above MRCISD+Q energies. The spectroscopic properties of the ground and many low-lying electronic states of the TaF molecule will be reported. With respect to the observed electronic states in this work, the calculated results are in good agreement with our experimental determinations. This work represents the first experimental

Versatile single-molecule multi-color excitation and detection fluorescence setup for studying biomolecular dynamics

KAUST Repository

Sobhy, M. A.

2011-11-07

Single-molecule fluorescence imaging is at the forefront of tools applied to study biomolecular dynamics both in vitro and in vivo. The ability of the single-molecule fluorescence microscope to conduct simultaneous multi-color excitation and detection is a key experimental feature that is under continuous development. In this paper, we describe in detail the design and the construction of a sophisticated and versatile multi-color excitation and emission fluorescence instrument for studying biomolecular dynamics at the single-molecule level. The setup is novel, economical and compact, where two inverted microscopes share a laser combiner module with six individual laser sources that extend from 400 to 640 nm. Nonetheless, each microscope can independently and in a flexible manner select the combinations, sequences, and intensities of the excitation wavelengths. This high flexibility is achieved by the replacement of conventional mechanical shutters with acousto-optic tunable filter (AOTF). The use of AOTF provides major advancement by controlling the intensities, duration, and selection of up to eight different wavelengths with microsecond alternation time in a transparent and easy manner for the end user. To our knowledge this is the first time AOTF is applied to wide-field total internal reflection fluorescence (TIRF) microscopy even though it has been commonly used in multi-wavelength confocal microscopy. The laser outputs from the combiner module are coupled to the microscopes by two sets of four single-mode optic fibers in order to allow for the optimization of the TIRF angle for each wavelength independently. The emission is split into two or four spectral channels to allow for the simultaneous detection of up to four different fluorophores of wide selection and using many possible excitation and photoactivation schemes. We demonstrate the performance of this new setup by conducting two-color alternating excitation single-molecule fluorescence resonance energy

Solid-phase single molecule biosensing using dual-color colocalization of fluorescent quantum dot nanoprobes

Science.gov (United States)

Liu, Jianbo; Yang, Xiaohai; Wang, Kemin; Wang, Qing; Liu, Wei; Wang, Dong

2013-10-01

The development of solid-phase surface-based single molecule imaging technology has attracted significant interest during the past decades. Here we demonstrate a sandwich hybridization method for highly sensitive detection of a single thrombin protein at a solid-phase surface based on the use of dual-color colocalization of fluorescent quantum dot (QD) nanoprobes. Green QD560-modified thrombin binding aptamer I (QD560-TBA I) were deposited on a positive poly(l-lysine) assembled layer, followed by bovine serum albumin blocking. It allowed the thrombin protein to mediate the binding of the easily detectable red QD650-modified thrombin binding aptamer II (QD650-TBA II) to the QD560-TBA I substrate. Thus, the presence of the target thrombin can be determined based on fluorescent colocalization measurements of the nanoassemblies, without target amplification or probe separation. The detection limit of this assay reached 0.8 pM. This fluorescent colocalization assay has enabled single molecule recognition in a separation-free detection format, and can serve as a sensitive biosensing platform that greatly suppresses the nonspecific adsorption false-positive signal. This method can be extended to other areas such as multiplexed immunoassay, single cell analysis, and real time biomolecule interaction studies.The development of solid-phase surface-based single molecule imaging technology has attracted significant interest during the past decades. Here we demonstrate a sandwich hybridization method for highly sensitive detection of a single thrombin protein at a solid-phase surface based on the use of dual-color colocalization of fluorescent quantum dot (QD) nanoprobes. Green QD560-modified thrombin binding aptamer I (QD560-TBA I) were deposited on a positive poly(l-lysine) assembled layer, followed by bovine serum albumin blocking. It allowed the thrombin protein to mediate the binding of the easily detectable red QD650-modified thrombin binding aptamer II (QD650-TBA II) to

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

Detecting aromatic compounds on planetary surfaces using ultraviolet time-resolved fluorescence spectroscopy

Science.gov (United States)

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

2018-02-01

Many aromatic organic molecules exhibit strong and characteristic fluorescence when excited with ultraviolet radiation. As laser excitation in the ultraviolet generates both fluorescence and resonantly enhanced Raman scattering of aromatic vibrational modes, combined Raman and fluorescence instruments have been proposed to search for organic compounds on Mars. In this work the time-resolved fluorescence of a suite of 24 compounds composed of 2-5 ringed alternant, non-alternant, and heterocyclic PAHs was measured. Fluorescence instrumentation with similar specifications to a putative flight instrument was capable of observing the fluorescence decay of these compounds with a sub-ns resolution. Incorporating time-resolved capabilities was also found to increase the ability to discriminate between individual PAHs. Incorporating time-resolved fluorescence capabilities into an ultraviolet gated Raman system intended for a rover or lander can increase the ability to detect and characterize PAHs on planetary surfaces.

Radio frequency scanning tunneling spectroscopy for single-molecule spin resonance.

Science.gov (United States)

Müllegger, Stefan; Tebi, Stefano; Das, Amal K; Schöfberger, Wolfgang; Faschinger, Felix; Koch, Reinhold

2014-09-26

We probe nuclear and electron spins in a single molecule even beyond the electromagnetic dipole selection rules, at readily accessible magnetic fields (few mT) and temperatures (5 K) by resonant radio-frequency current from a scanning tunneling microscope. We achieve subnanometer spatial resolution combined with single-spin sensitivity, representing a 10 orders of magnitude improvement compared to existing magnetic resonance techniques. We demonstrate the successful resonant spectroscopy of the complete manifold of nuclear and electronic magnetic transitions of up to ΔI(z)=±3 and ΔJ(z)=±12 of single quantum spins in a single molecule. Our method of resonant radio-frequency scanning tunneling spectroscopy offers, atom-by-atom, unprecedented analytical power and spin control with an impact on diverse fields of nanoscience and nanotechnology.

Rapid measurement of meat spoilage using fluorescence spectroscopy

Science.gov (United States)

Wu, Binlin; Dahlberg, Kevin; Gao, Xin; Smith, Jason; Bailin, Jacob

2017-02-01

Food spoilage is mainly caused by microorganisms, such as bacteria. In this study, we measure the autofluorescence in meat samples longitudinally over a week in an attempt to develop a method to rapidly detect meat spoilage using fluorescence spectroscopy. Meat food is a biological tissue, which contains intrinsic fluorophores, such as tryptophan, collagen, nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) etc. As meat spoils, it undergoes various morphological and chemical changes. The concentrations of the native fluorophores present in a sample may change. In particular, the changes in NADH and FAD are associated with microbial metabolism, which is the most important process of the bacteria in food spoilage. Such changes may be revealed by fluorescence spectroscopy and used to indicate the status of meat spoilage. Therefore, such native fluorophores may be unique, reliable and nonsubjective indicators for detection of spoiled meat. The results of the study show that the relative concentrations of all above fluorophores change as the meat samples kept in room temperature ( 19° C) spoil. The changes become more rapidly after about two days. For the meat samples kept in a freezer ( -12° C), the changes are much less or even unnoticeable over a-week-long storage.

Direct solid surface fluorescence spectroscopy of standard chemicals and humic acid in ternary system.

Science.gov (United States)

Mounier, S; Nicolodelli, G; Redon, R; Milori, D M B P

2017-04-15

The front face fluorescence spectroscopy is often used to quantify chemicals in well-known matrices as it is a rapid and powerful technique, with no sample preparation. However it was not used to investigate extracted organic matter like humic substances. This work aims to fully investigate for the first time front face fluorescence spectroscopy response of a ternary system including boric acid, tryptophan and humic substances, and two binaries system containing quinine sulfate or humic substance in boric acid. Pure chemicals, boric acid, tryptophan, quinine sulfate and humic acid were mixed together in solid pellet at different contents from 0 to 100% in mass. The measurement of excitation emission matrix of fluorescence (3D fluorescence) and laser induced fluorescence were then done in the front face mode. Fluorescence matrices were decomposed using the CP/PARAFAC tools after scattering treatments. Results show that for 3D fluorescence there is no specific component for tryptophan and quinine sulfate, and that humic substances lead to a strong extinction effect for mixture containing quinine sulfate. Laser induced fluorescence gives a very good but non-specific related response for both quinine sulfate and tryptophan. No humic substances fluorescence response was found, but extinction effect is observed as for 3D fluorescence. This effect is stronger for quinine sulfate than for tryptophan. These responses were modeled using a simple absorbance versus emission model. Copyright © 2017 Elsevier B.V. All rights reserved.

Numerical simulation of dynamic quenching of dual-split fluorescence of molecules with intramolecular hydrogen bonds

International Nuclear Information System (INIS)

Morozov, V.A.; Chuvulkin, N.D.; Smolenskij, E.A.; Dubina, Yu.M.

2014-01-01

The dynamic quenching of intensity pulses of the dual-split fluorescence (DSF) has been simulated using numerical solutions of the equations for the population matrix of five states of the model fluorescent molecule (FM). The state with the highest energy is considered as resonantly excited by irradiation, and two other excited states populated by subsequent relaxation processes are taken as initial states for the FM transitions with emission of the DSF photons. The FM model parameters are selected to fit typical parameters of the molecules with intramolecular proton photo transfer. Quenching is considered as a consequence of non-radiative decay of the FM excited states due to collisions with the quencher molecules. Examples of two types of the DSF quenching of the FM are given. The first type leads to an intramolecular radiationless decay of particular excited states of the FM, and the second one results in radiationless transitions from the same states to the quencher molecule states. (authors)

Fluorescence Spectroscopy and Chemometric Modeling for Bioprocess Monitoring

Directory of Open Access Journals (Sweden)

Saskia M. Faassen

2015-04-01

Full Text Available On-line sensors for the detection of crucial process parameters are desirable for the monitoring, control and automation of processes in the biotechnology, food and pharma industry. Fluorescence spectroscopy as a highly developed and non-invasive technique that enables the on-line measurements of substrate and product concentrations or the identification of characteristic process states. During a cultivation process significant changes occur in the fluorescence spectra. By means of chemometric modeling, prediction models can be calculated and applied for process supervision and control to provide increased quality and the productivity of bioprocesses. A range of applications for different microorganisms and analytes has been proposed during the last years. This contribution provides an overview of different analysis methods for the measured fluorescence spectra and the model-building chemometric methods used for various microbial cultivations. Most of these processes are observed using the BioView® Sensor, thanks to its robustness and insensitivity to adverse process conditions. Beyond that, the PLS-method is the most frequently used chemometric method for the calculation of process models and prediction of process variables.

Fluorescence Spectroscopy and Chemometric Modeling for Bioprocess Monitoring

Science.gov (United States)

Faassen, Saskia M.; Hitzmann, Bernd

2015-01-01

On-line sensors for the detection of crucial process parameters are desirable for the monitoring, control and automation of processes in the biotechnology, food and pharma industry. Fluorescence spectroscopy as a highly developed and non-invasive technique that enables the on-line measurements of substrate and product concentrations or the identification of characteristic process states. During a cultivation process significant changes occur in the fluorescence spectra. By means of chemometric modeling, prediction models can be calculated and applied for process supervision and control to provide increased quality and the productivity of bioprocesses. A range of applications for different microorganisms and analytes has been proposed during the last years. This contribution provides an overview of different analysis methods for the measured fluorescence spectra and the model-building chemometric methods used for various microbial cultivations. Most of these processes are observed using the BioView® Sensor, thanks to its robustness and insensitivity to adverse process conditions. Beyond that, the PLS-method is the most frequently used chemometric method for the calculation of process models and prediction of process variables. PMID:25942644

Simultaneous membrane interaction of amphipathic peptide monomers, self-aggregates and cargo complexes detected by fluorescence correlation spectroscopy.

Science.gov (United States)

Vasconcelos, Luís; Lehto, Tõnis; Madani, Fatemeh; Radoi, Vlad; Hällbrink, Mattias; Vukojević, Vladana; Langel, Ülo

2018-02-01

Peptides able to translocate cell membranes while carrying macromolecular cargo, as cell-penetrating peptides (CPPs), can contribute to the field of drug delivery by enabling the transport of otherwise membrane impermeable molecules. Formation of non-covalent complexes between amphipathic peptides and oligonucleotides is driven by electrostatic and hydrophobic interactions. Here we investigate and quantify the coexistence of distinct molecular species in multiple equilibria, namely peptide monomer, peptide self-aggregates and peptide/oligonucleotide complexes. As a model for the complexes, we used a stearylated peptide from the PepFect family, PF14 and siRNA. PF14 has a cationic part and a lipid part, resembling some characteristics of cationic lipids. Fluorescence correlation spectroscopy (FCS) and fluorescence cross-correlation spectroscopy (FCCS) were used to detect distinct molecular entities in solution and at the plasma membrane of live cells. For that, we labeled the peptide with carboxyrhodamine 6G and the siRNA with Cyanine 5. We were able to detect fluorescent entities with diffusional properties characteristic of the peptide monomer as well as of peptide aggregates and peptide/oligonucleotide complexes. Strategies to avoid peptide adsorption to solid surfaces and self-aggregation were developed and allowed successful FCS measurements in solution and at the plasma membrane. The ratio between the detected molecular species was found to vary with pH, peptide concentration and the proximity to the plasma membrane. The present results suggest that the diverse cellular uptake mechanisms, often reported for amphipathic CPPs, might result from the synergistic effect of peptide monomers, self-aggregates and cargo complexes, distributed unevenly at the plasma membrane. Copyright © 2017 Elsevier B.V. All rights reserved.

Use of fluorescence spectroscopy to measure molecular autofluorescence in diabetic subjects; Utilizacao da espectroscopia de fluorescencia para mensuramento de moleculas autofluorescentes em individuos diabeticos

Energy Technology Data Exchange (ETDEWEB)

Gomes, Cinthia Zanini

2011-07-01

Diabetes Mellitus (DM) comprises a complex metabolic syndrome, caused by reduced or absent secretion of insulin by pancreatic beta cells, leading to hyperglycemia. Hyperglycemia promotes glycation of proteins and, consequently, the appearance of advanced glycation end products (AGEs). Currently, diabetic patients are monitored by determining levels of glucose and glycated hemoglobin (HbA1c). The complications caused by hyperglycemia may be divided into micro and macrovascular complications, represented by retinopathy, nephropathy, neuropathy and cardiovascular disease. Albumin (HSA) is the most abundant serum protein in the human body and is subject to glycation. The Protoporphyrin IX (PpIX) is the precursor molecule of heme synthesis, structural component of hemoglobin. The in vitro and animals studies have indicated that hyperglycemia promotes a decrease in its concentration in erythrocytes. The fluorescence spectroscopy is a technique widely used in biomedical field. The autofluorescence corresponds to the intrinsic fluorescence present in some molecules, this being associated with the same structure. The aim of this study was to use fluorescence spectroscopy to measure levels of erythrocyte PpIX autofluorescence and AGE-HSA in diabetic and healthy subjects and compare them with levels of blood glucose and HbA1c. This study was conducted with 151 subjects (58 controls and 93 diabetics). Epidemiological data of patients and controls were obtained from medical records. For control subjects, blood glucose levels were obtained from medical records and levels of Hb1Ac obtained by using commercial kits. The determination of the PpIX autofluorescence was performed with excitation at 405 nm and emission at 632 nm. Determination of AGE-HSA was performed with excitation at 370 nm and emission at 455 nm. Approximately 50% of diabetic had micro and macrovascular lesions resulting from hyperglycemia. There were no significant differences in the PpIX emission intensity values

Single-Molecule Analysis of Pre-mRNA Splicing with Colocalization Single-Molecule Spectroscopy (CoSMoS).

Science.gov (United States)

Braun, Joerg E; Serebrov, Victor

2017-01-01

Recent development of single-molecule techniques to study pre-mRNA splicing has provided insights into the dynamic nature of the spliceosome. Colocalization single-molecule spectroscopy (CoSMoS) allows following spliceosome assembly in real time at single-molecule resolution in the full complexity of cellular extracts. A detailed protocol of CoSMoS has been published previously (Anderson and Hoskins, Methods Mol Biol 1126:217-241, 2014). Here, we provide an update on the technical advances since the first CoSMoS studies including slide surface treatment, data processing, and representation. We describe various labeling strategies to generate RNA reporters with multiple dyes (or other moieties) at specific locations.

A Nanofluidic Mixing Device for High-throughput Fluorescence Sensing of Single Molecules

NARCIS (Netherlands)

Mathwig, Klaus; Fijen, C.; Fontana, M.; Lemay, S.G.; Hohlbein, J.C.

2017-01-01

We introduce a nanofluidic mixing device entirely fabricated in glass for the fluorescence detection of single molecules. The design consists of a nanochannel T-junction and allows the continuous monitoring of chemical or enzymatic reactions of analytes as they arrive from two independent inlets.

Distribution of diffusion times determined by fluorescence (lifetime) correlation spectroscopy

Czech Academy of Sciences Publication Activity Database

Pánek, Jiří; Loukotová, Lenka; Hrubý, Martin; Štěpánek, Petr

2018-01-01

Roč. 51, č. 8 (2018), s. 2796-2804 ISSN 0024-9297 R&D Projects: GA MŠk(CZ) LO1507 Institutional support: RVO:61389013 Keywords : polymer solution * fluorescence correlation spectroscopy * diffusion time distribution Subject RIV: CD - Macromolecular Chemistry OBOR OECD: Polymer science Impact factor: 5.835, year: 2016

Comparison of fluorescence rejection methods of baseline correction and shifted excitation Raman difference spectroscopy

Science.gov (United States)

Cai, Zhijian; Zou, Wenlong; Wu, Jianhong

2017-10-01

Raman spectroscopy has been extensively used in biochemical tests, explosive detection, food additive and environmental pollutants. However, fluorescence disturbance brings a big trouble to the applications of portable Raman spectrometer. Currently, baseline correction and shifted-excitation Raman difference spectroscopy (SERDS) methods are the most prevailing fluorescence suppressing methods. In this paper, we compared the performances of baseline correction and SERDS methods, experimentally and simulatively. Through the comparison, it demonstrates that the baseline correction can get acceptable fluorescence-removed Raman spectrum if the original Raman signal has good signal-to-noise ratio, but it cannot recover the small Raman signals out of large noise background. By using SERDS method, the Raman signals, even very weak compared to fluorescence intensity and noise level, can be clearly extracted, and the fluorescence background can be completely rejected. The Raman spectrum recovered by SERDS has good signal to noise ratio. It's proved that baseline correction is more suitable for large bench-top Raman system with better quality or signal-to-noise ratio, while the SERDS method is more suitable for noisy devices, especially the portable Raman spectrometers.

Optical fluorescence spectroscopy to detect hepatic necrosis after normothermic ischemia: animal model

Science.gov (United States)

Romano, Renan A.; Vollet-Filho, Jose D.; Pratavieira, Sebastião.; Fernandez, Jorge L.; Kurachi, Cristina; Bagnato, Vanderlei S.; Castro-e-Silva, Orlando; Sankarankutty, Ajith K.

2015-06-01

Liver transplantation is a well-established treatment for liver failure. However, the success of the transplantation procedure depends on liver graft conditions. The tissue function evaluation during the several transplantation stages is relevant, in particular during the organ harvesting, when a decision is made concerning the viability of the graft. Optical fluorescence spectroscopy is a good option because it is a noninvasive and fast technique. A partial normothermic hepatic ischemia was performed in rat livers, with a vascular occlusion of both median and left lateral lobes, allowing circulation only for the right lateral lobe and the caudate lobe. Fluorescence spectra under excitation at 532 nm (doubled frequency Nd:YAG laser) were collected using a portable spectrometer (USB2000, Ocean Optics, USA). The fluorescence emission was collected before vascular occlusion, after ischemia, and 24 hours after reperfusion. A morphometric histology analysis was performed as the gold standard evaluation - liver samples were analyzed, and the percentage of necrotic tissue was obtained. The results showed that changes in the fluorescence emission after ischemia can be correlated with the amount of necrosis evaluated by a morphometric analysis, the Pearson correlation coefficient of the generated model was 0.90 and the root mean square error was around 20%. In this context, the laser-induced fluorescence spectroscopy technique after normothermic ischemia showed to be a fast and efficient method to differentiate ischemic injury from viable tissues.

Quantitative structural modeling on the wavelength interval (Δλ) in synchronous fluorescence spectroscopy

Science.gov (United States)

Samari, Fayezeh; Yousefinejad, Saeed

2017-11-01

Emission fluorescence spectroscopy has an extremely restricted scope of application to analyze of complex mixtures since its selectivity is reduced by the extensive spectral overlap. Synchronous fluorescence spectroscopy (SFS) is a technique enables us to analyze complex mixtures with overlapped emission and/or excitation spectra. The difference of excitation and emission wavelength of compounds (interval wavelength or Δλ) is an important characteristic in SFS. Thus a multi-parameter model was constructed to predict Δλ in 63 fluorescent compounds and the regression coefficient in training set, cross validation and test set were 0.88, 0.85 and 0.91 respectively. Furthermore, the applicability and validity of model were evaluated using different statistical methods such as y-scrambling and applicability domain. It was concluded that increasing average valence connectivity, number of Al2-NH functional group and Geary autocorrelation (lag 4) with electronegative weights can lead to increasing Δλ in the fluorescent compounds. The current study obtained an insight into the structural properties of compounds effective on their Δλ as an important parameter in SFS.

Some fluorescence properties of dimethylaminochalcone and its novel cyclic analogues

Science.gov (United States)

Tomečková, Vladimíra; Poškrobová, Martina; Štefanišinová, Miroslava; Perjési, Pál

2009-12-01

This paper demonstrates the basic character (polarity, solubility, colour, absorption and fluorescence quantum yield) of synthetic dimethylaminochalcone ( 1) and its cyclic analogues measured in toluene, chloroform, dimethylsulfoxide and ethanol, which have been studied by absorption and fluorescence spectroscopy. The biologically active dye 4'-dimethylaminochalcone ( 1b) and its less flexible analogues 4-dimethylaminoindanone ( 2b), -tetralone ( 3b), and -benzosuberone ( 4b) are lipophilic molecules that displayed the best solubility in toluene and chloroform. The highest fluorescence and quantum yields of compounds 1 and 2 have been obtained in DMSO and chloroform. Quenching effect of fluorescence compounds ( 1- 4) has been studied in the mixture of the most polar organic solvents DMSO and water. In the presence of water, fluorescence of compound 1 has been quenched the best from all studied chalcones and emission maxima of molecules 1- 4 have been shifted to the longer wavelengths. Quenching effect of fluorescence by water was in order 1 > 2 > 3 > 4.

Single Molecule Spectroscopy in Chemistry, Physics and Biology Nobel Symposium

CERN Document Server

Gräslund, Astrid; Widengren, Jerker

2010-01-01

Written by the leading experts in the field, this book describes the development and current state-of-the-art in single molecule spectroscopy. The application of this technique, which started 1989, in physics, chemistry and biosciences is displayed.

Rationale for single molecule detection by means of Raman spectroscopy

International Nuclear Information System (INIS)

Gaponenko, S.V.; Guzatov, D.V.

2009-01-01

A consistent quantum electrodynamical description is proposed of Raman scattering of light by a molecule in a medium with a modified photon density of states. Enhanced local density of states near a metal nanobody is shown to increase a scattering rate by several orders of magnitude, thus providing a rationale for experimental detection of single molecules by means of Raman spectroscopy. For an ellipsoidal particle 10 14 -fold enhancement of the Raman scattering cross-section is obtained. (authors)

Damping kinetics of annihilation slowed fluorescence of aromatic molecules in Langmuir films

International Nuclear Information System (INIS)

Ibraev, N.Kh.; Latonin, V.A.; Satybaldina, D.Zh.

1996-01-01

The paper is devoted to study of decay mechanism of phosphors molecules triplet states in the Langmuir-Blogett (L-B) films. The L-B-films have being prepared on the base of anthracene, pyrene, 3,4-benzopyrene and 1,2-benzanthracene molecules. Spectral and kinetic investigations were carried out in photons counting regime. Long-lived luminescence was observed under excitation of the LB-film by nitrogen laser radiation (λ=337 nm) .Spectrum of the long-lived luminescence coincides with spectrum of fast fluorescence of same film. Received spectral data indicate on similarity of LB-films structure of aromatic molecules and its crystals. It was defined, that deactivation of triplet states of aromatic molecules in LB-films has being carried out mainly in the result of annihilation of triplet excitation. 14 refs., 2 tabs., 5 figs

Time-resolved spectroscopy defines perturbation in molecules

International Nuclear Information System (INIS)

Ahmed, K.

1998-01-01

Time-resolved LIF spectroscopy is employed in order to investigate perturbations in different excited electronic state of alkali molecules. Dunham Coefficients are used to search the selected excited ro-vibrational level, which is overlap with the other nearby excited states. Lifetime measurement has been performed of more than 50 ro-vibrational levels. Out of these 25 levels were observed drastically different lifetimes from the other unperturbed levels. In this report, influence of different perturbations on this anomalous behavior is investigated and discussed. (author)

Laser resonant ionization spectroscopy and laser-induced resonant fluorescence spectra of samarium atom

International Nuclear Information System (INIS)

Jin, Changtai

1995-01-01

We have measured new high-lying levels of Sm atom by two-colour resonant photoionisation spectroscopy; we have observed the isotope shifts of Sm atom by laser-induced resonant fluorescence spectroscopy; the lifetime of eight low-lying levels of Sm atom were measured by using pulsed laser-Boxcar technique in atomic beam.

2D fluorescence spectroscopy for monitoring ion-exchange membrane based technologies - Reverse electrodialysis (RED).

Science.gov (United States)

Pawlowski, Sylwin; Galinha, Claudia F; Crespo, João G; Velizarov, Svetlozar

2016-01-01

Reverse electrodialysis (RED) is one of the emerging, membrane-based technologies for harvesting salinity gradient energy. In RED process, fouling is an undesirable operation constraint since it leads to a decrease of the obtainable net power density due to increasing stack electric resistance and pressure drop. Therefore, early fouling detection is one of the main challenges for successful RED technology implementation. In the present study, two-dimensional (2D) fluorescence spectroscopy was used, for the first time, as a tool for fouling monitoring in RED. Fluorescence excitation-emission matrices (EEMs) of ion-exchange membrane surfaces and of natural aqueous streams were acquired during one month of a RED stack operation. Fouling evolvement on the ion-exchange membrane surfaces was successfully followed by 2D fluorescence spectroscopy and quantified using principal components analysis (PCA). Additionally, the efficiency of cleaning strategy was assessed by measuring the membrane fluorescence emission intensity before and after cleaning. The anion-exchange membrane (AEM) surface in contact with river water showed to be significantly affected due to fouling by humic compounds, which were found to cross through the membrane from the lower salinity (river water) to higher salinity (sea water) stream. The results obtained show that the combined approach of using 2D fluorescence spectroscopy and PCA has a high potential for studying fouling development and membrane cleaning efficiency in ion exchange membrane processes. Copyright © 2015 Elsevier Ltd. All rights reserved.

Spectral characterization of crude oil using fluorescence (synchronous and time-resolved) and NIR (Near Infrared Spectroscopy); Caracterizacao espectral do petroleo utilizando fluorescencia (sincronizada e resolvida no tempo) e NIR (Near Infrared Spectroscopy)

Energy Technology Data Exchange (ETDEWEB)

Falla Sotelo, F.; Araujo Pantoja, P.; Lopez-Gejo, J.; Le Roux, G.A.C.; Nascimento, C.A.O. [Universidade de Sao Paulo (USP), SP (Brazil). Dept. de Engenharia Quimica. Lab. de Simulacao e Controle de Processos; Quina, F.H. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica. Centro de Capacitacao e Pesquisa em Meio Ambiente (CEPEMA)

2008-07-01

The objective of the present work is to evaluate the performance of two spectroscopic techniques employed in the crude oil characterization: NIR spectroscopy and fluorescence spectroscopy (Synchronous fluorescence - SF and Time Resolved Fluorescence - TRF) for the development of correlation models between spectral profiles of crude oil samples and both physical properties (viscosity and API density) and physico-chemical properties (SARA analysis: Saturated, Aromatic, Resins and Asphaltenes). The better results for viscosity and density were obtained using NIR whose prediction capacity was good (1.5 cP and 0.5 deg API, respectively). For SARA analysis, fluorescence spectroscopy revealed its potential in the model calibration showing good results (R2 coefficients greater than 0.85). TRF spectroscopy had better performance than SF spectroscopy. (author)

Spectroscopy of selected metal-containing diatomic molecules

Science.gov (United States)

Gordon, Iouli E.

D molecules were studied by means of Fourier transform spectroscopy and laser-induced fluorescence. The extreme complexity of these transitions made rotational assignments of EuH bands impossible. However, the spin-spin interaction constant, lambda, and Fermi contact parameter, bF, in the ground X9Sigma- electronic state were estimated for the 151EuH and 153EuH isotopologues. Electronic spectra of SmH, SmCl, TmH and ErF molecules were recorded for the first time using Fourier transform spectrometer. The poor signal to noise ratio of the observed bands coupled with their complexity prevented a rotational analysis. The electronic states that may be involved in the observed transitions are discussed.

Laser spectroscopy of molecules: State-of-the-art and possible trends

International Nuclear Information System (INIS)

Demtroeder, W.

1990-01-01

A review is given on different techniques in laser spectroscopy of atoms and molecules, which allow high spectral resolution and a very high detection sensitivity of small samples. Analytical applications of the techniques are discussed for basic scientific research, as well as for environmental problems and technical processes. Possible trends of laser spectroscopy, in particular with respect to applications in biology and medicine are shortly outlined. (orig.)

Time-resolved resonance fluorescence spectroscopy for study of chemical reactions in laser-induced plasmas.

Science.gov (United States)

Liu, Lei; Deng, Leimin; Fan, Lisha; Huang, Xi; Lu, Yao; Shen, Xiaokang; Jiang, Lan; Silvain, Jean-François; Lu, Yongfeng

2017-10-30

Identification of chemical intermediates and study of chemical reaction pathways and mechanisms in laser-induced plasmas are important for laser-ablated applications. Laser-induced breakdown spectroscopy (LIBS), as a promising spectroscopic technique, is efficient for elemental analyses but can only provide limited information about chemical products in laser-induced plasmas. In this work, time-resolved resonance fluorescence spectroscopy was studied as a promising tool for the study of chemical reactions in laser-induced plasmas. Resonance fluorescence excitation of diatomic aluminum monoxide (AlO) and triatomic dialuminum monoxide (Al 2 O) was used to identify these chemical intermediates. Time-resolved fluorescence spectra of AlO and Al 2 O were used to observe the temporal evolution in laser-induced Al plasmas and to study their formation in the Al-O 2 chemistry in air.

Very High Spectral Resolution Imaging Spectroscopy: the Fluorescence Explorer (FLEX) Mission

Science.gov (United States)

Moreno, Jose F.; Goulas, Yves; Huth, Andreas; Middleton, Elizabeth; Miglietta, Franco; Mohammed, Gina; Nedbal, Ladislav; Rascher, Uwe; Verhoef, Wouter; Drusch, Matthias

2016-01-01

The Fluorescence Explorer (FLEX) mission has been recently selected as the 8th Earth Explorer by the European Space Agency (ESA). It will be the first mission specifically designed to measure from space vegetation fluorescence emission, by making use of very high spectral resolution imaging spectroscopy techniques. Vegetation fluorescence is the best proxy to actual vegetation photosynthesis which can be measurable from space, allowing an improved quantification of vegetation carbon assimilation and vegetation stress conditions, thus having key relevance for global mapping of ecosystems dynamics and aspects related with agricultural production and food security. The FLEX mission carries the FLORIS spectrometer, with a spectral resolution in the range of 0.3 nm, and is designed to fly in tandem with Copernicus Sentinel-3, in order to provide all the necessary spectral / angular information to disentangle emitted fluorescence from reflected radiance, and to allow proper interpretation of the observed fluorescence spatial and temporal dynamics.

Light emitting diode excitation emission matrix fluorescence spectroscopy.

Science.gov (United States)

Hart, Sean J; JiJi, Renée D

2002-12-01

An excitation emission matrix (EEM) fluorescence instrument has been developed using a linear array of light emitting diodes (LED). The wavelengths covered extend from the upper UV through the visible spectrum: 370-640 nm. Using an LED array to excite fluorescence emission at multiple excitation wavelengths is a low-cost alternative to an expensive high power lamp and imaging spectrograph. The LED-EEM system is a departure from other EEM spectroscopy systems in that LEDs often have broad excitation ranges which may overlap with neighboring channels. The LED array can be considered a hybrid between a spectroscopic and sensor system, as the broad LED excitation range produces a partially selective optical measurement. The instrument has been tested and characterized using fluorescent dyes: limits of detection (LOD) for 9,10-bis(phenylethynyl)-anthracene and rhodamine B were in the mid parts-per-trillion range; detection limits for the other compounds were in the low parts-per-billion range (LED-EEMs were analyzed using parallel factor analysis (PARAFAC), which allowed the mathematical resolution of the individual contributions of the mono- and dianion fluorescein tautomers a priori. Correct identification and quantitation of six fluorescent dyes in two to six component mixtures (concentrations between 12.5 and 500 ppb) has been achieved with root mean squared errors of prediction (RMSEP) of less than 4.0 ppb for all components.

Single Molecule Spectroelectrochemistry of Interfacial Charge Transfer Dynamics In Hybrid Organic Solar Cell

Energy Technology Data Exchange (ETDEWEB)

Pan, Shanlin [Univ. of Alabama, Tuscaloosa, AL (United States)

2014-11-16

Our research under support of this DOE grant is focused on applied and fundamental aspects of model organic solar cell systems. Major accomplishments are: 1) we developed a spectroelectorchemistry technique of single molecule single nanoparticle method to study charge transfer between conjugated polymers and semiconductor at the single molecule level. The fluorescence of individual fluorescent polymers at semiconductor surfaces was shown to exhibit blinking behavior compared to molecules on glass substrates. Single molecule fluorescence excitation anisotropy measurements showed the conformation of the polymer molecules did not differ appreciably between glass and semiconductor substrates. The similarities in molecular conformation suggest that the observed differences in blinking activity are due to charge transfer between fluorescent polymer and semiconductor, which provides additional pathways between states of high and low fluorescence quantum efficiency. Similar spectroelectrochemistry work has been done for small organic dyes for understand their charge transfer dynamics on various substrates and electrochemical environments; 2) We developed a method of transferring semiconductor nanoparticles (NPs) and graphene oxide (GO) nanosheets into organic solvent for a potential electron acceptor in bulk heterojunction organic solar cells which employed polymer semiconductor as the electron donor. Electron transfer from the polymer semiconductor to semiconductor and GO in solutions and thin films was established through fluorescence spectroscopy and electroluminescence measurements. Solar cells containing these materials were constructed and evaluated using transient absorption spectroscopy and dynamic fluorescence techniques to understand the charge carrier generation and recombination events; 3) We invented a spectroelectorchemistry technique using light scattering and electroluminescence for rapid size determination and studying electrochemistry of single NPs in an

Monitoring the diffusion behavior of Na,K-ATPase by fluorescence correlation spectroscopy (FCS) upon fluorescence labelling with eGFP or Dreiklang

Science.gov (United States)

Junghans, Cornelia; Schmitt, Franz-Josef; Vukojević, Vladana; Friedrich, Thomas

2016-02-01

Measurement of lateral mobility of membraneembedded proteins in living cells with high spatial and temporal precision is a challenging task of optofluidics. Biological membranes are complex structures, whose physico-chemical properties depend on the local lipid composition, cholesterol content and the presence of integral or peripheral membrane proteins, which may be involved in supramolecular complexes or are linked to cellular matrix proteins or the cytoskeleton. The high proteinto- lipid ratios in biomembranes indicate that membrane proteins are particularly subject to molecular crowding, making it difficult to follow the track of individual molecules carrying a fluorescence label. Novel switchable fluorescence proteins such as Dreiklang [1], are, in principle, promising tools to study the diffusion behavior of individual molecules in situations of molecular crowding due to excellent spectral control of the ON- and OFF-switching process. In this work, we expressed an integral membrane transport protein, the Na,K-ATPase comprising the human α2-subunit carrying an N-terminal eGFP or Dreiklang tag and human β1-subunit, in HEK293T cells and measured autocorrelation curves by fluorescence correlation spectroscopy (FCS). Furthermore,we measured diffusion times and diffusion constants of eGFP and Dreiklang by FCS, first, in aqueous solution after purification of the proteins upon expression in E. coli, and, second, upon expression as soluble proteins in the cytoplasm of HEK293T cells. Our data show that the diffusion behavior of the purified eGFP and Dreiklang in solution as well as the properties of the proteins expressed in the cytoplasm are very similar. However, the autocorrelation curves of eGFP- and Dreiklanglabeled Na,K-ATPase measured in the plasma membrane exhibit marked differences, with the Dreiklang-labeled construct showing shorter diffusion times. This may be related to an additional, as yet unrecognized quenching process that occurs on the same time

Quantifying the Assembly of Multicomponent Molecular Machines by Single-Molecule Total Internal Reflection Fluorescence Microscopy.

Science.gov (United States)

Boehm, E M; Subramanyam, S; Ghoneim, M; Washington, M Todd; Spies, M

2016-01-01

Large, dynamic macromolecular complexes play essential roles in many cellular processes. Knowing how the components of these complexes associate with one another and undergo structural rearrangements is critical to understanding how they function. Single-molecule total internal reflection fluorescence (TIRF) microscopy is a powerful approach for addressing these fundamental issues. In this article, we first discuss single-molecule TIRF microscopes and strategies to immobilize and fluorescently label macromolecules. We then review the use of single-molecule TIRF microscopy to study the formation of binary macromolecular complexes using one-color imaging and inhibitors. We conclude with a discussion of the use of TIRF microscopy to examine the formation of higher-order (i.e., ternary) complexes using multicolor setups. The focus throughout this article is on experimental design, controls, data acquisition, and data analysis. We hope that single-molecule TIRF microscopy, which has largely been the province of specialists, will soon become as common in the tool box of biophysicists and biochemists as structural approaches have become today. © 2016 Elsevier Inc. All rights reserved.

Feasibility of Raman spectroscopy in vitro after 5-ALA-based fluorescence diagnosis in the bladder

Science.gov (United States)

Grimbergen, M. C. M.; van Swol, C. F. P.; van Moorselaar, R. J. A.; Mahadevan-Jansen, A.,; Stone, N.

2006-02-01

Photodynamic diagnosis (PDD) has become popular in bladder cancer detection. Several studies have however shown an increased false positive biopsies rate under PDD guidance compared to conventional cystoscopy. Raman spectroscopy is an optical technique that utilizes molecular specific, inelastic scattering of light photons to interrogate biological tissues, which can successfully differentiate epithelial neoplasia from normal tissue and inflammations in vitro. This investigation was performed to show the feasibility of NIR Raman spectroscopy in vitro on biopsies obtained under guidance of 5-ALA induced PPIX fluorescence imaging. Raman spectra of a PPIX solution was measured to obtain a characteristic signature for the photosensitzer without contributions from tissue constituents. Biopsies were obtained from patients with known bladder cancer instilled with 50ml, 5mg 5-ALA two hours prior to trans-urethral resection of tumor (TURT). Additional biopsies were obtained at a fluorescent and non-fluorescent area, snap-frozen in liquid nitrogen and stored at -80 °C. Each biopsy was thawed before measurements (10sec integration time) with a confocal Raman system (Renishaw Gloucestershire, UK). The 830 nm excitation (300mW) source is focused on the tissue by a 20X ultra-long-working-distance objective. Differences in fluorescence background between the two groups were removed by means of a special developed fluorescence subtraction algorithm. Raman spectra from ALA biopsies showed different fluorescence background which can be effectively removed by a fluorescence subtraction algorithm. This investigation shows that the interaction of the ALA induced PPIX with Raman spectroscopy in bladder samples. Combination of these techniques in-vivo may lead to a viable method of optical biopsies in bladder cancer detection.

Solvent induced fluorescence enhancement of graphene oxide studied by ultrafast spectroscopy

Science.gov (United States)

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

2018-05-01

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

High resolution x-ray fluorescence spectroscopy - a new technique for site- and spin-selectivity

International Nuclear Information System (INIS)

Wang, Xin

1996-12-01

X-ray spectroscopy has long been used to elucidate electronic and structural information of molecules. One of the weaknesses of x-ray absorption is its sensitivity to all of the atoms of a particular element in a sample. Through out this thesis, a new technique for enhancing the site- and spin-selectivity of the x-ray absorption has been developed. By high resolution fluorescence detection, the chemical sensitivity of K emission spectra can be used to identify oxidation and spin states; it can also be used to facilitate site-selective X-ray Absorption Near Edge Structure (XANES) and site-selective Extended X-ray Absorption Fine Structure (EXAFS). The spin polarization in K fluorescence could be used to generate spin selective XANES or spin-polarized EXAFS, which provides a new measure of the spin density, or the nature of magnetic neighboring atoms. Finally, dramatic line-sharpening effects by the combination of absorption and emission processes allow observation of structure that is normally unobservable. All these unique characters can enormously simplify a complex x-ray spectrum. Applications of this novel technique have generated information from various transition-metal model compounds to metalloproteins. The absorption and emission spectra by high resolution fluorescence detection are interdependent. The ligand field multiplet model has been used for the analysis of Kα and Kβ emission spectra. First demonstration on different chemical states of Fe compounds has shown the applicability of site selectivity and spin polarization. Different interatomic distances of the same element in different chemical forms have been detected using site-selective EXAFS

Exploring the Dynamics of Cell Processes through Simulations of Fluorescence Microscopy Experiments

Science.gov (United States)

Angiolini, Juan; Plachta, Nicolas; Mocskos, Esteban; Levi, Valeria

2015-01-01

Fluorescence correlation spectroscopy (FCS) methods are powerful tools for unveiling the dynamical organization of cells. For simple cases, such as molecules passively moving in a homogeneous media, FCS analysis yields analytical functions that can be fitted to the experimental data to recover the phenomenological rate parameters. Unfortunately, many dynamical processes in cells do not follow these simple models, and in many instances it is not possible to obtain an analytical function through a theoretical analysis of a more complex model. In such cases, experimental analysis can be combined with Monte Carlo simulations to aid in interpretation of the data. In response to this need, we developed a method called FERNET (Fluorescence Emission Recipes and Numerical routines Toolkit) based on Monte Carlo simulations and the MCell-Blender platform, which was designed to treat the reaction-diffusion problem under realistic scenarios. This method enables us to set complex geometries of the simulation space, distribute molecules among different compartments, and define interspecies reactions with selected kinetic constants, diffusion coefficients, and species brightness. We apply this method to simulate single- and multiple-point FCS, photon-counting histogram analysis, raster image correlation spectroscopy, and two-color fluorescence cross-correlation spectroscopy. We believe that this new program could be very useful for predicting and understanding the output of fluorescence microscopy experiments. PMID:26039162

High resolution UV spectroscopy and laser-focused nanofabrication

NARCIS (Netherlands)

Myszkiewicz, G.

2005-01-01

This thesis combines two at first glance different techniques: High Resolution Laser Induced Fluorescence Spectroscopy (LIF) of small aromatic molecules and Laser Focusing of atoms for Nanofabrication. The thesis starts with the introduction to the high resolution LIF technique of small aromatic

Zero-mode waveguide nanophotonic structures for single molecule characterization

Science.gov (United States)

Crouch, Garrison M.; Han, Donghoon; Bohn, Paul W.

2018-05-01

Single-molecule characterization has become a crucial research tool in the chemical and life sciences, but limitations, such as limited concentration range, inability to control molecular distributions in space, and intrinsic phenomena, such as photobleaching, present significant challenges. Recent developments in non-classical optics and nanophotonics offer promising routes to mitigating these restrictions, such that even low affinity (K D ~ mM) biomolecular interactions can be studied. Here we introduce and review specific nanophotonic devices used to support single molecule studies. Optical nanostructures, such as zero-mode waveguides (ZMWs), are usually fabricated in thin gold or aluminum films and serve to confine the observation volume of optical microspectroscopy to attoliter to zeptoliter volumes. These simple nanostructures allow individual molecules to be isolated for optical and electrochemical analysis, even when the molecules of interest are present at high concentration (µM–mM) in bulk solution. Arrays of ZMWs may be combined with optical probes such as single molecule fluorescence, single molecule fluorescence resonance energy transfer, and fluorescence correlation spectroscopy for distributed analysis of large numbers of single-molecule reactions or binding events in parallel. Furthermore, ZMWs may be used as multifunctional devices, for example by combining optical and electrochemical functions in a single discrete architecture to achieve electrochemical ZMWs. In this review, we will describe the optical properties, fabrication, and applications of ZMWs for single-molecule studies, as well as the integration of ZMWs into systems for chemical and biochemical analysis.

Heat shock-induced interactions among nuclear HSFs detected by fluorescence cross-correlation spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Pack, Chan-Gi, E-mail: changipack@amc.seoul.kr [Asan Institute for Life Sciences, University of Ulsan, College of Medicine, Asan Medical Center, Seoul 138-736 (Korea, Republic of); Ahn, Sang-Gun [Dept. of Pathology, College of Dentistry, Chosun University, Seosuk-dong, Dong-gu, Gwangju 501-759 (Korea, Republic of)

2015-07-31

The cellular response to stress is primarily controlled in cells via transcriptional activation by heat shock factor 1 (HSF1). HSF1 is well-known to form homotrimers for activation upon heat shock and subsequently bind to target DNAs, such as heat-shock elements, by forming stress granules. A previous study demonstrated that nuclear HSF1 and HSF2 molecules in live cells interacted with target DNAs on the stress granules. However, the process underlying the binding interactions of HSF family in cells upon heat shock remains unclear. This study demonstrate for the first time that the interaction kinetics among nuclear HSF1, HSF2, and HSF4 upon heat shock can be detected directly in live cells using dual color fluorescence cross-correlation spectroscopy (FCCS). FCCS analyses indicated that the binding between HSFs was dramatically changed by heat shock. Interestingly, the recovery kinetics of interaction between HSF1 molecules after heat shock could be represented by changes in the relative interaction amplitude and mobility. - Highlights: • The binding interactions among nuclear HSFs were successfully detected. • The binding kinetics between HSF1s during recovery was quantified. • HSF2 and HSF4 strongly formed hetero-complex, even before heat shock. • Nuclear HSF2 and HSF4 bound to HSF1 only after heat shock.

Review of X-ray Tomography and X-ray Fluorescence Spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Shear, Trevor A. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

2017-03-16

This literature review will focus on both laboratory and synchrotron based X-ray tomography of materials and highlight the inner workings of these instruments. X-ray fluorescence spectroscopy will also be reviewed and applications of the tandem use of these techniques will be explored. The real world application of these techniques during the internship will also be discussed.

Differential laser-induced perturbation spectroscopy and fluorescence imaging for biological and materials sensing

Science.gov (United States)

Burton, Dallas Jonathan

The field of laser-based diagnostics has been a topic of research in various fields, more specifically for applications in environmental studies, military defense technologies, and medicine, among many others. In this dissertation, a novel laser-based optical diagnostic method, differential laser-induced perturbation spectroscopy (DLIPS), has been implemented in a spectroscopy mode and expanded into an imaging mode in combination with fluorescence techniques. The DLIPS method takes advantage of deep ultraviolet (UV) laser perturbation at sub-ablative energy fluences to photochemically cleave bonds and alter fluorescence signal response before and after perturbation. The resulting difference spectrum or differential image adds more information about the target specimen, and can be used in combination with traditional fluorescence techniques for detection of certain materials, characterization of many materials and biological specimen, and diagnosis of various human skin conditions. The differential aspect allows for mitigation of patient or sample variation, and has the potential to develop into a powerful, noninvasive optical sensing tool. The studies in this dissertation encompass efforts to continue the fundamental research on DLIPS including expansion of the method to an imaging mode. Five primary studies have been carried out and presented. These include the use of DLIPS in a spectroscopy mode for analysis of nitrogen-based explosives on various substrates, classification of Caribbean fruit flies versus Caribbean fruit flies that have been irradiated with gamma rays, and diagnosis of human skin cancer lesions. The nitrogen-based explosives and Caribbean fruit flies have been analyzed with the DLIPS scheme using the imaging modality, providing complementary information to the spectroscopic scheme. In each study, a comparison between absolute fluorescence signals and DLIPS responses showed that DLIPS statistically outperformed traditional fluorescence techniques

Using Quenching to Detect Corrosion on Sculptural Metalwork: A Real-World Application of Fluorescence Spectroscopy

Science.gov (United States)

Hensen, Cory; Clare, Tami Lasseter; Barbera, Jack

2018-01-01

Fluorescence spectroscopy experiments are a frequently taught as part of upper-division teaching laboratories. To expose undergraduate students to an applied fluorescence technique, a corrosion detection method, using quenching, was adapted from authentic research for an instrumental analysis laboratory. In the experiment, students acquire…

Analysis of DNA interactions using single-molecule force spectroscopy.

Science.gov (United States)

Ritzefeld, Markus; Walhorn, Volker; Anselmetti, Dario; Sewald, Norbert

2013-06-01

Protein-DNA interactions are involved in many biochemical pathways and determine the fate of the corresponding cell. Qualitative and quantitative investigations on these recognition and binding processes are of key importance for an improved understanding of biochemical processes and also for systems biology. This review article focusses on atomic force microscopy (AFM)-based single-molecule force spectroscopy and its application to the quantification of forces and binding mechanisms that lead to the formation of protein-DNA complexes. AFM and dynamic force spectroscopy are exciting tools that allow for quantitative analysis of biomolecular interactions. Besides an overview on the method and the most important immobilization approaches, the physical basics of the data evaluation is described. Recent applications of AFM-based force spectroscopy to investigate DNA intercalation, complexes involving DNA aptamers and peptide- and protein-DNA interactions are given.

Fourier transform infrared and fluorescence spectroscopy for analysis of vegetable oils

Directory of Open Access Journals (Sweden)

Nigri S.

2013-09-01

Full Text Available Fourier transform infrared (FTIR and fluorescence spectroscopy, combined with chemometric approaches have been developed to analysis of extra virgin olive oil adulterated with pomace olive oil. The measurements were made on pure vegetable oils: extra virgin oil, pomace olive oil and that adulterated with varying concentration of pomace olive oil. Today, the application of FTIR spectroscopy has increased in food studied, and particularly has become a powerful analytical tool in the study of edible oils and fats. The spectral regions where the variations were observed chosen for developing models and cross validation was used. The synchronous fluorescence spectrometry takes advantage of the hardware capability to vary both the excitation and emission wavelengths during the analysis with constant wavelength difference is maintained between the two. The region between 300 and 400 nm is attributed to the tocopherols and phenols, the derivatives of vitamin E are associated with the region 400–600 nm and the bands in the region of 600–700 nm are attributed to the chlorophyll and peophytin pigments. The results presented in this study suggest that FTIR and fluorescence may be a useful tool for analysis and detecting adulteration of extra virgin olive oil with pomace oil.

Limitations of fluorescence spectroscopy to characterize organic matter in engineered systems

Science.gov (United States)

Korak, J.

2017-12-01

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

Organic molecule fluorescence as an experimental test-bed for quantum jumps in thermodynamics.

Science.gov (United States)

Browne, Cormac; Farrow, Tristan; Dahlsten, Oscar C O; Taylor, Robert A; Vlatko, Vedral

2017-08-01

We demonstrate with an experiment how molecules are a natural test bed for probing fundamental quantum thermodynamics. Single-molecule spectroscopy has undergone transformative change in the past decade with the advent of techniques permitting individual molecules to be distinguished and probed. We demonstrate that the quantum Jarzynski equality for heat is satisfied in this set-up by considering the time-resolved emission spectrum of organic molecules as arising from quantum jumps between states. This relates the heat dissipated into the environment to the free energy difference between the initial and final state. We demonstrate also how utilizing the quantum Jarzynski equality allows for the detection of energy shifts within a molecule, beyond the relative shift.

Probing the local environment of a single OPE3 molecule using inelastic tunneling electron spectroscopy

NARCIS (Netherlands)

Frisenda, R.; Perrin, M.L.; Van der Zant, H.S.J.

2015-01-01

We study single-molecule oligo(phenylene ethynylene)dithiol junctions by means of inelastic electron tunneling spectroscopy (IETS). The molecule is contacted with gold nano-electrodes formed with the mechanically controllable break junction technique. We record the IETS spectrum of the molecule from

Investigation of polyelectrolyte desorption by single molecule force spectroscopy

International Nuclear Information System (INIS)

Friedsam, C; Seitz, M; Gaub, H E

2004-01-01

Single molecule force spectroscopy has evolved into a powerful method for the investigation of intra- and intermolecular interactions at the level of individual molecules. Many examples, including the investigation of the dynamic properties of complex biological systems as well as the properties of covalent bonds or intermolecular transitions within individual polymers, are reported in the literature. The technique has recently been extended to the systematic investigation of desorption processes of individual polyelectrolyte molecules adsorbed on generic surfaces. The stable covalent attachment of polyelectrolyte molecules to the AFM-tip provides the possibility of performing long-term measurements with the same set of molecules and therefore allows the in situ observation of the impact of environmental changes on the adsorption behaviour of individual molecules. Different types of interactions, e.g. electrostatic or hydrophobic interactions, that determine the adsorption process could be identified and characterized. The experiments provided valuable details that help to understand the nature and the properties of non-covalent interactions, which is helpful with regard to biological systems as well as for technical applications. Apart from this, desorption experiments can be utilized to characterize the properties of surfaces or polymer coatings. Therefore they represent a versatile tool that can be further developed in terms of various aspects

Classification of plum spirit drinks by synchronous fluorescence spectroscopy.

Science.gov (United States)

Sádecká, J; Jakubíková, M; Májek, P; Kleinová, A

2016-04-01

Synchronous fluorescence spectroscopy was used in combination with principal component analysis (PCA) and linear discriminant analysis (LDA) for the differentiation of plum spirits according to their geographical origin. A total of 14 Czech, 12 Hungarian and 18 Slovak plum spirit samples were used. The samples were divided in two categories: colorless (22 samples) and colored (22 samples). Synchronous fluorescence spectra (SFS) obtained at a wavelength difference of 60 nm provided the best results. Considering the PCA-LDA applied to the SFS of all samples, Czech, Hungarian and Slovak colorless samples were properly classified in both the calibration and prediction sets. 100% of correct classification was also obtained for Czech and Hungarian colored samples. However, one group of Slovak colored samples was classified as belonging to the Hungarian group in the calibration set. Thus, the total correct classifications obtained were 94% and 100% for the calibration and prediction steps, respectively. The results were compared with those obtained using near-infrared (NIR) spectroscopy. Applying PCA-LDA to NIR spectra (5500-6000 cm(-1)), the total correct classifications were 91% and 92% for the calibration and prediction steps, respectively, which were slightly lower than those obtained using SFS. Copyright © 2015 Elsevier Ltd. All rights reserved.

Dual-wavelength external cavity laser device for fluorescence suppression in Raman spectroscopy

Science.gov (United States)

Zhang, Xuting; Cai, Zhijian; Wu, Jianhong

2017-10-01

Raman spectroscopy has been widely used in the detection of drugs, pesticides, explosives, food additives and environmental pollutants, for its characteristics of fast measurement, easy sample preparation, and molecular structure analyzing capability. However, fluorescence disturbance brings a big trouble to these applications, with strong fluorescence background covering up the weak Raman signals. Recently shifted excitation Raman difference spectroscopy (SERDS) not only can completely remove the fluorescence background, but also can be easily integrated into portable Raman spectrometers. Usually, SERDS uses two lasers with small wavelength gap to excite the sample, then acquires two spectra, and subtracts one to the other to get the difference spectrum, where the fluorescence background will be rejected. So, one key aspects of successfully applying SERDS method is to obtain a dual-wavelength laser source. In this paper, a dual-wavelength laser device design based on the principles of external cavity diode laser (ECDL) is proposed, which is low-cost and compact. In addition, it has good mechanical stability because of no moving parts. These features make it an ideal laser source for SERDS technique. The experiment results showed that the device can emit narrow-spectral-width lasers of two wavelengths, with the gap smaller than 2 nanometers. The laser power corresponding to each wavelength can be up to 100mW.

Study of high density polyethylene under UV irradiation or mechanical stress by fluorescence spectroscopy

International Nuclear Information System (INIS)

Douminge, L.

2010-05-01

Due to their diversity and their wide range of applications, polymers have emerged in our environment. For technical applications, these materials can be exposed to aggressive environment leading to an alteration of their properties. The effects of this degradation are linked to the concept of life duration, corresponding to the time required for a property to reach a threshold below which the material becomes unusable. Monitoring the ageing of polymer materials constitute a major challenge. Fluorescence spectroscopy is a technique able to provide accurate information concerning this issue. In this study, emphasis was placed on the use of fluorescence spectroscopy to study the phenomena involved in either the UV radiation or mechanical stresses of a polymer. In the case of high density polyethylene, the lack of intrinsic fluorescent signal leads to the use of a dye. This dye gives a fluorescent response depending on its microenvironment. All modifications in the macromolecular chain generate a shift of the fluorescent peak. This work can be dissociated in two major parts, on one hand the influence of UV aging on the fluorescent response and in another hand the influence of mechanical stresses. In the first part, complementary analyses like FTIR or DSC are used to correlate fluorescent results with known photo degradation mechanisms. The results show the great sensibility of the technique to the microstructural rearrangement in the polymer. In the second part, the dependence between the stress and the fluorescence emission gives opportunity to evaluate internal stresses in the material during cyclic solicitations. (author)

Large Molecule Structures by Broadband Fourier Transform Molecular Rotational Spectroscopy

Science.gov (United States)

Evangelisti, Luca; Seifert, Nathan A.; Spada, Lorenzo; Pate, Brooks

2016-06-01

Fourier transform molecular rotational resonance spectroscopy (FT-MRR) using pulsed jet molecular beam sources is a high-resolution spectroscopy technique that can be used for chiral analysis of molecules with multiple chiral centers. The sensitivity of the molecular rotational spectrum pattern to small changes in the three dimensional structure makes it possible to identify diastereomers without prior chemical separation. For larger molecules, there is the additional challenge that different conformations of each diastereomer may be present and these need to be differentiated from the diastereomers in the spectral analysis. Broadband rotational spectra of several larger molecules have been measured using a chirped-pulse FT-MRR spectrometer. Measurements of nootkatone (C15H22O), cedrol (C15H26O), ambroxide (C16H28O) and sclareolide (C16H26O2) are presented. These spectra are measured with high sensitivity (signal-to-noise ratio near 1,000:1) and permit structure determination of the most populated isomers using isotopic analysis of the 13C and 18O isotopologues in natural abundance. The accuracy of quantum chemistry calculations to identify diastereomers and conformers and to predict the dipole moment properties needed for three wave mixing measurements is examined.

Tetrairon(III) Single Molecule Magnet Studied by Scanning Tunneling Microscopy and Spectroscopy

Science.gov (United States)

Oh, Youngtek; Jeong, Hogyun; Lee, Minjun; Kwon, Jeonghoon; Yu, Jaejun; Mamun, Shariful Islam; Gupta, Gajendra; Kim, Jinkwon; Kuk, Young

2011-03-01

Tetrairon(III) single-molecule magnet (SMM) on a clean Au(111) has studied using scanning tunneling microscopy (STM) and spectroscopy (STS) to understand quantum mechanical tunneling of magnetization and hysteresis of pure molecular origin. Before the STM studies, elemental analysis, proton nuclear magnetic resonance (NMR) measurement and Energy Dispersive X- ray Spectroscopy (EDS) were carried out to check the robustness of the sample. The STM image of this molecule shows a hexagonal shape, with a phenyl ring at the center and surrounding six dipivaloylmethane ligands. Two peaks are observed at 0.5 eV, 1.5 eV in the STS results, agreeing well with the first principles calculations. Spin-polarized scanning tunneling microscopy (SPSTM) measurements have been performed with a magnetic tip to get the magnetization image of the SMM. We could observe the antiferromagnetic coupling and a centered- triangular topology with six alkoxo bridges inside the molecule while applying external magnetic fields.

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.

Fluorescence spectroscopy and confocal microscopy of the mycotoxin citrinin in condensed phase and hydrogel films.

Science.gov (United States)

Lauer, Milena H; Gehlen, Marcelo H; de Jesus, Karen; Berlinck, Roberto G S

2014-05-01

The emission spectra, quantum yields and fluorescence lifetimes of citrinin in organic solvents and hydrogel films have been determined. Citrinin shows complex fluorescence decays due to the presence of two tautomers in solution and interconversion from excited-state double proton transfer (ESDPT) process. The fluorescence decay times associated with the two tautomers have values near 1 and 5 ns depending on the medium. In hydrogel films of agarose and alginate, fluorescence imaging showed that citrinin is not homogeneously dispersed and highly emissive micrometer spots may be formed. Fluorescence spectrum and decay analysis are used to recognize the presence of citrinin in hydrogel films using confocal fluorescence microscopy and spectroscopy.

Study of tryptophan assisted synthesis of gold nanoparticles by combining UV-Vis, fluorescence, and SERS spectroscopy

International Nuclear Information System (INIS)

Iosin, Monica; Baldeck, Patrice; Astilean, Simion

2010-01-01

We developed a rapid and non-toxic method for the preparation of colloidal gold nanoparticles (GNPs) by using tryptophan (Trp) as reducing/stabilizing agent. We show that the temperature has a major influence on the kinetics of gold ion reduction and the crystal growth, higher temperatures favoring the synthesis of anisotropic nanoparticles (triangles and hexagons). The as-synthesized nanostructures were characterized by UV-Vis absorption spectroscopy, transmission electron microscopy (TEM), X-ray diffraction (XRD), fluorescence, and surface-enhanced Raman scattering (SERS) spectroscopy. The UV-Vis measurements confirmed that temperature is a critical factor in the synthesis process, having a major effect on the shape of the synthesized GNPs. Moreover, fluorescence spectroscopy was able to monitor the quenching of the Trp fluorescence during the in situ synthesis of GNPs. Using Trp as molecular analyte to evaluate the SERS efficiency of as-prepared GNPs at different temperatures, we demonstrated that the Raman enhancement of the synthesized gold nanoplates is higher than that of the gold spherical nanoparticles.

Automatic classification of fluorescence and optical diffusion spectroscopy data in neuro-oncology

Science.gov (United States)

Savelieva, T. A.; Loshchenov, V. B.; Goryajnov, S. A.; Potapov, A. A.

2018-04-01

The complexity of the biological tissue spectroscopic analysis due to the overlap of biological molecules' absorption spectra, multiple scattering effect, as well as measurement geometry in vivo has caused the relevance of this work. In the neurooncology the problem of tumor boundaries delineation is especially acute and requires the development of new methods of intraoperative diagnosis. Methods of optical spectroscopy allow detecting various diagnostically significant parameters non-invasively. 5-ALA induced protoporphyrin IX is frequently used as fluorescent tumor marker in neurooncology. At the same time analysis of the concentration and the oxygenation level of haemoglobin and significant changes of light scattering in tumor tissues have a high diagnostic value. This paper presents an original method for the simultaneous registration of backward diffuse reflectance and fluorescence spectra, which allows defining all the parameters listed above simultaneously. The clinical studies involving 47 patients with intracranial glial tumors of II-IV Grades were carried out in N.N. Burdenko National Medical Research Center of Neurosurgery. To register the spectral dependences the spectroscopic system LESA- 01-BIOSPEC was used with specially developed w-shaped diagnostic fiber optic probe. The original algorithm of combined spectroscopic signal processing was developed. We have created a software and hardware, which allowed (as compared with the methods currently used in neurosurgical practice) to increase the sensitivity of intraoperative demarcation of intracranial tumors from 78% to 96%, specificity of 60% to 82%. The result of analysis of different techniques of automatic classification shows that in our case the most appropriate is the k Nearest Neighbors algorithm with cubic metrics.

Uptake Of Trivalent Actinides (Cm(III)) And Lanthanides (Eu(III)) By Cement-Type Minerals: A Wet Chemistry And Time-Resolved Laser Fluorescence Spectroscopy (TRLFS) Study

Energy Technology Data Exchange (ETDEWEB)

Tits, J.; Stumpf, T; Wieland, E.; Fanghaenel, T

2003-03-01

The interaction of the two chemical homologues Cm (III) and Eu(III) with calcium silicate hydrates at pH 13.3 has been investigated in batch-type sorption studies using Eu(III), and complemented with time-resolved laser fluorescence spectroscopy using Cm(III). The sorption data for Eu(III) reveal fast sorption kinetics, and a strong uptake by CSH phases, with distribution ratios of 6({+-}3)*105 L kg-1. Three different types of sorbed Cm(III) species have been identified: a non-fluorescing species, which was identified as Cm cluster present either as surface precipitate or as Cm(III) colloid in solution, and two sorbed fluorescing species. The sorbed fluorescing species have characteristic emission spectra (main peak maxima at 618.9 nm and 620.9 nm) and fluorescence emission lifetimes (289 {+-} 11 ms and 1482{+-} 200 ms). From the fluorescence lifetimes, it appears that the two fluorescing Cm(III) species have, respectively, one to two or no water molecules left in their first coordination sphere, suggesting that these species are incorporated into the CSH structure. A structural model for Cm(III) and Eu(III) incorporation into CSH phases is proposed based on the substitution of Ca at two different types of sites in the CSH structure. (author)

Fluorescent Affibody Molecule Administered In Vivo at a Microdose Level Labels EGFR Expressing Glioma Tumor Regions.

Science.gov (United States)

de Souza, Ana Luiza Ribeiro; Marra, Kayla; Gunn, Jason; Samkoe, Kimberley S; Hoopes, P Jack; Feldwisch, Joachim; Paulsen, Keith D; Pogue, Brian W

2017-02-01

Fluorescence guidance in surgical oncology provides the potential to realize enhanced molecular tumor contrast with dedicated targeted tracers, potentially with a microdose injection level. For most glioma tumors, the blood brain barrier is compromised allowing some exogenous drug/molecule delivery and accumulation for imaging. The aberrant overexpression and/or activation of epidermal growth factor receptor (EGFR) is associated with many types of cancers, including glioblastoma, and so the use of a near-infrared (NIR) fluorescent molecule targeted to the EGFR receptor provides the potential for improving tumor contrast during surgery. Fluorescently labeled affibody molecule (ABY-029) has high EGFR affinity and high potential specificity with reasonably fast plasma clearance. In this study, ABY-29 was evaluated in glioma versus normal brain uptake from intravenous injection at a range of doses, down to a microdose injection level. Nude rats were inoculated with the U251 human glioma cell line in the brain. Tumors were allowed to grow for 3-4 weeks. ABY-029 fluorescence ex vivo imaging of brain slices was acquired at different time points (1-48 h) and varying injection doses from 25 to 122 μg/kg (from human protein microdose equivalent to five times microdose levels). The tumor was most clearly visualized at 1-h post-injection with 8- to 16-fold average contrast relative to normal brain. However, the tumor still could be identified after 48 h. In all cases, the ABY-029 fluorescence appeared to localize preferentially in EGFR-positive regions. Increasing the injected dose from a microdose level to five times, a microdose level increased the signal by 10-fold, and the contrast was from 8 to 16, showing that there was value in doses slightly higher than the microdose restriction. Normal tissue uptake was found to be affected by the tumor size, indicating that edema was a likely factor affecting the expected tumor to normal tissue contrast. These results suggest

Two-photon-excited fluorescence spectroscopy of atomic fluorine at 170 nm

Science.gov (United States)

Herring, G. C.; Dyer, Mark J.; Jusinski, Leonard E.; Bischel, William K.

1988-01-01

Two-photon-excited fluorescence spectroscopy of atomic fluorine is reported. A doubled dye laser at 286-nm is Raman shifted in H2 to 170 nm (sixth anti-Stokes order) to excite ground-state 2P(0)J fluorine atoms to the 2D(0)J level. The fluorine atoms are detected by one of two methods: observing the fluorescence decay to the 2PJ level or observing F(+) production through the absorption of an additional photon by the excited atoms. Relative two-photon absorption cross sections to and the radiative lifetimes of the 2D(0)J states are measured.

Strong overtones modes in inelastic electron tunneling spectroscopy with cross-conjugated molecules

DEFF Research Database (Denmark)

Jørgensen, Jacob Lykkebo; Gagliardi, Alessio; Pecchia, Alessandro

2013-01-01

. With this in mind, we investigate a spectroscopic method capable of providing insight into these junctions for cross-conjugated molecules: inelastic electron tunneling spectroscopy (IETS). IETS has the advantage that the molecule interface is probed directly by the tunneling current. Previously, it has been thought...... and leading to suppressed levels of elastic current. In most theoretical studies, only the elastic contributions to the current are taken into account. In this paper, we study the inelastic contributions to the current in cross-conjugated molecules and find that while the inelastic contribution to the current...

[Fluorescent and Raman scattering by molecules embedded in small particles]: Annual report, 1983

International Nuclear Information System (INIS)

Chew, H.; McNulty, P.J.

1983-01-01

An overview is given of the model formulated for fluorescent and Raman scattering by molecules embedded in or in the vicinity of small particles. The model takes into account the size, shape, refractive index, and morphology of the host particles. Analytic and numerical results have been obtained for spherical (one and more layers, including magnetic dipole transitions), cylindrical, and spheroidal particles. Particular attention has been given to the spherical case with fluorescent/Raman scatterers uniformly distributed in the particles radiating both coherently and incoherently. Depolarization effects have been studied with suitable averaging process, and good agreement with experiment has been obtained. Analytic and numerical results have been obtained for the elastic scattering of evanescent waves; these results are useful for the study of fluorescence under excitation by evanescent waves

The development of methods of analysis of documents on the basis of the methods of Raman spectroscopy and fluorescence analysis

Science.gov (United States)

Gorshkova, Kseniia O.; Tumkin, Ilya I.; Kirillova, Elizaveta O.; Panov, Maxim S.; Kochemirovsky, Vladimir A.

2017-05-01

The investigation of natural aging of writing inks printed on paper using Raman spectroscopy was performed. Based on the obtained dependencies of the Raman peak intensities ratios on the exposure time, the dye degradation model was proposed. It was suggested that there are several competing bond breaking and bond forming reactions corresponding to the characteristic vibration frequencies of the dye molecule that simultaneously occur during ink aging process. Also we propose a methodology based on the study of the optical properties of paper, particularly changes in the fluorescence of optical brighteners included in its composition as well as the paper reflectivity using spectrophotometric methods. These results can be implemented to develop the novel and promising method of criminology.

Applications of photon-in, photon-out spectroscopy with third-generation, synchrotron-radiation sources

International Nuclear Information System (INIS)

Lindle, D.W.; Perera, R.C.C.

1991-01-01

This report discusses the following topics: Mother nature's finest test probe; soft x-ray emission spectroscopy with high-brightness synchrotron radiation sources; anisotropy and polarization of x-ray emission from atoms and molecules; valence-hole fluorescence from molecular photoions as a probe of shape-resonance ionization: progress and prospects; structural biophysics on third-generation synchrotron sources; ultra-soft x-ray fluorescence-yield XAFS: an in situ photon-in, photon-out spectroscopy; and x-ray microprobe: an analytical tool for imaging elemental composition and microstructure

Interaction of antihypertensive drug amiloride with metal ions in micellar medium using fluorescence spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Gujar, Varsha; Pundge, Vijaykumar; Ottoor, Divya, E-mail: divya@chem.unipune.ac.in

2015-05-15

Steady state and life time fluorescence spectroscopy have been employed to study the interaction of antihypertensive drug amiloride with biologically important metal ions i.e. Cu{sup 2+}, Fe{sup 2+}, Ni{sup 2+} and Zn{sup 2+} in various micellar media (anionic SDS (sodium dodecyl sulfate), nonionic TX-100 (triton X-100) and cationic CTAB (cetyl trimethyl ammonium bromide)). It was observed that fluorescence properties of drug remain unaltered in the absence of micellar media with increasing concentration of metal ions. However, addition of Cu{sup 2+}, Fe{sup 2+} and Ni{sup 2+} caused fluorescence quenching of amiloride in the presence of anionic micelle, SDS. Binding of drug with metal ions at the charged micellar interface could be the possible reason for this pH-dependent metal-mediated fluorescence quenching. There were no remarkable changes observed due to metal ions addition when drug was present in cationic and nonionic micellar medium. The binding constant and bimolecular quenching constant were evaluated and compared for the drug–metal complexes using Stern–Volmer equation and fluorescence lifetime values. - Highlights: • Interaction of amiloride with biologically important metal ions, Fe{sup 2+}, Cu{sup 2+}, Ni{sup 2+} and Zn{sup 2+}. • Monitoring the interaction in various micelle at different pH by fluorescence spectroscopy. • Micelles acts as receptor, amiloride as transducer and metal ions as analyte in the present system. • Interaction study provides pH dependent quenching and binding mechanism of drug with metal ions.

Interaction of antihypertensive drug amiloride with metal ions in micellar medium using fluorescence spectroscopy

International Nuclear Information System (INIS)

Gujar, Varsha; Pundge, Vijaykumar; Ottoor, Divya

2015-01-01

Steady state and life time fluorescence spectroscopy have been employed to study the interaction of antihypertensive drug amiloride with biologically important metal ions i.e. Cu 2+ , Fe 2+ , Ni 2+ and Zn 2+ in various micellar media (anionic SDS (sodium dodecyl sulfate), nonionic TX-100 (triton X-100) and cationic CTAB (cetyl trimethyl ammonium bromide)). It was observed that fluorescence properties of drug remain unaltered in the absence of micellar media with increasing concentration of metal ions. However, addition of Cu 2+ , Fe 2+ and Ni 2+ caused fluorescence quenching of amiloride in the presence of anionic micelle, SDS. Binding of drug with metal ions at the charged micellar interface could be the possible reason for this pH-dependent metal-mediated fluorescence quenching. There were no remarkable changes observed due to metal ions addition when drug was present in cationic and nonionic micellar medium. The binding constant and bimolecular quenching constant were evaluated and compared for the drug–metal complexes using Stern–Volmer equation and fluorescence lifetime values. - Highlights: • Interaction of amiloride with biologically important metal ions, Fe 2+ , Cu 2+ , Ni 2+ and Zn 2+ . • Monitoring the interaction in various micelle at different pH by fluorescence spectroscopy. • Micelles acts as receptor, amiloride as transducer and metal ions as analyte in the present system. • Interaction study provides pH dependent quenching and binding mechanism of drug with metal ions

Elastin-like Polypeptide Linkers for Single-Molecule Force Spectroscopy.

Science.gov (United States)

Ott, Wolfgang; Jobst, Markus A; Bauer, Magnus S; Durner, Ellis; Milles, Lukas F; Nash, Michael A; Gaub, Hermann E

2017-06-27

Single-molecule force spectroscopy (SMFS) is by now well established as a standard technique in biophysics and mechanobiology. In recent years, the technique has benefitted greatly from new approaches to bioconjugation of proteins to surfaces. Indeed, optimized immobilization strategies for biomolecules and refined purification schemes are being steadily adapted and improved, which in turn has enhanced data quality. In many previously reported SMFS studies, poly(ethylene glycol) (PEG) was used to anchor molecules of interest to surfaces and/or cantilever tips. The limitation, however, is that PEG exhibits a well-known trans-trans-gauche to all-trans transition, which results in marked deviation from standard polymer elasticity models such as the worm-like chain, particularly at elevated forces. As a result, the assignment of unfolding events to protein domains based on their corresponding amino acid chain lengths is significantly obscured. Here, we provide a solution to this problem by implementing unstructured elastin-like polypeptides as linkers to replace PEG. We investigate the suitability of tailored elastin-like polypeptides linkers and perform direct comparisons to PEG, focusing on attributes that are critical for single-molecule force experiments such as linker length, monodispersity, and bioorthogonal conjugation tags. Our results demonstrate that by avoiding the ambiguous elastic response of mixed PEG/peptide systems and instead building the molecular mechanical systems with only a single bond type with uniform elastic properties, we improve data quality and facilitate data analysis and interpretation in force spectroscopy experiments. The use of all-peptide linkers allows alternative approaches for precisely defining elastic properties of proteins linked to surfaces.

Microscopie de fluorescence de protéines autofluorescentes uniques pour la biologie cellulaire

Science.gov (United States)

Cognet, Laurent; Coussen, Françoise; Choquet, Daniel; Lounis, Brahim

In this paper we review the applicability of autofluorescent proteins for single-molecule imaging in biology. The photophysical characteristics of several mutants of the Green Fluorescent Protein (GFP) and those of DsRed are compared and critically discussed for their use in cellular biology. The alternative use of two-photon excitation at the single-molecule level or Fluorescence Correlation Spectroscopy is envisaged for the study of individual autofluorescent proteins. Single-molecule experiments performed in live cells using eGFP and preferably eYFP fusion proteins are reviewed. Finally, the first use at the single-molecule level of citrine, a more photostable variant of the eYFP is reported when fused to a receptor for neurotransmitter in live cells. To cite this article: L. Cognet et al., C. R. Physique 3 (2002) 645-656.

Fluorescence excitation-emission matrix spectroscopy for degradation monitoring of machinery lubricants

Science.gov (United States)

Sosnovski, Oleg; Suresh, Pooja; Dudelzak, Alexander E.; Green, Benjamin

2018-02-01

Lubrication oil is a vital component of heavy rotating machinery defining the machine's health, operational safety and effectiveness. Recently, the focus has been on developing sensors that provide real-time/online monitoring of oil condition/lubricity. Industrial practices and standards for assessing oil condition involve various analytical methods. Most these techniques are unsuitable for online applications. The paper presents the results of studying degradation of antioxidant additives in machinery lubricants using Fluorescence Excitation-Emission Matrix (EEM) Spectroscopy and Machine Learning techniques. EEM Spectroscopy is capable of rapid and even standoff sensing; it is potentially applicable to real-time online monitoring.

Characterization of type I, II, III, IV, and V collagens by time-resolved laser-induced fluorescence spectroscopy

Science.gov (United States)

Marcu, Laura; Cohen, David; Maarek, Jean-Michel I.; Grundfest, Warren S.

2000-04-01

The relative proportions of genetically distinct collagen types in connective tissues vary with tissue type and change during disease progression, development, wound healing, aging. This study aims to 1) characterize the spectro- temporal fluorescence emission of fiber different types of collagen and 2) assess the ability of time-resolved laser- induced fluorescence spectroscopy to distinguish between collagen types. Fluorescence emission of commercially available purified samples was induced with nitrogen laser excitation pulses and detected with a MCP-PMT connected to a digital storage oscilloscope. The recorded time-resolved emission spectra displayed distinct fluorescence emission characteristics for each collagen type. The time domain information complemented the spectral domain intensity data for improved discrimination between different collagen types. Our results reveal that analysis of the fluorescence emission can be used to characterize different species of collagen. Also, the results suggest that time-resolved spectroscopy can be used for monitoring of connective tissue matrix composition changes due to various pathological and non-pathological conditions.

Surface-enhanced resonance Raman scattering spectroscopy of single R6G molecules

Institute of Scientific and Technical Information of China (English)

Zhou Zeng-Hui; Liu Li; Wang Gui-Ying; Xu Zhi-Zhan

2006-01-01

Surface-enhanced resonance Raman scattering (SERRS) of Rhodamine 6G (R6G) adsorbed on colloidal silver clusters has been studied. Based on the great enhancement of the Raman signal and the quench of the fluorescence, the SERRS spectra of R6G were recorded for the samples of dye colloidal solution with different concentrations. Spectral inhomogeneity behaviours from single molecules in the dried sample films were observed with complementary evidences, such as spectral polarization, spectral diffusion, intensity fluctuation of vibrational lines and even "breathing" of the molecules. Sequential spectra observed from a liquid sample with an average of 0.3 dye molecules in the probed volume exhibited the expected Poisson distribution for actually measuring 0, 1 or 2 molecules. Difference between the SERRS spectra of R6G excited by linearly and circularly polarized light were experimentally measured.

Biological Interaction of Molybdenocene Dichloride with Bovine Serum Albumin Using Fluorescence Spectroscopy

Science.gov (United States)

Domínguez, Moralba; Cortes-Figueroa, Jose´ E.; Meléndez, Enrique

2018-01-01

Bioinorganic topics are ubiquitous in the inorganic chemistry curriculum; however, experiments to enhance understanding of related topics are scarce. In this proposed laboratory, upper undergraduate students assess the biological interaction of molybdenocene dichloride (Cp2MoCl2) with bovine serum albumin (BSA) by fluorescence spectroscopy.…

Comment on ’Single Pentacene Molecules Detected by Fluorescence Excitation in a P-Terphenyl Crystal’

Science.gov (United States)

1990-12-10

8217 NO 11 TITLE (include Security Classification) Comment on "Single Pentacene Molecules Detected by Fluorescence Excitation in a p-Terphenyl Crystal" 12...8217 {Continue on reverse it necessary and identify by block numboer) Using h--,Ihly efficient Fluorescence excitation spectroscov of individual pentacene ...molecular impurities in p-terphenvl crystals, we have observed that some pentacene defects exhibit spcntaneous spectral jumps in their resonance frequency at

Sizes of water-soluble luminescent quantum dots measured by fluorescence correlation spectroscopy

International Nuclear Information System (INIS)

Zhang Pudun; Li Liang; Dong Chaoqing; Qian Huifeng; Ren Jicun

2005-01-01

In this paper, fluorescence correlation spectroscopy (FCS) was applied to measure the size of water-soluble quantum dots (QDs). The measurements were performed on a home-built FCS system based on the Stokes-Einstein equation. The obtained results showed that for bare CdTe QDs the sizes from FCS were larger than the ones from transmission electron microscopy (TEM). The brightness of QDs was also evaluated using FCS technique. It was found that the stability of the surface chemistry of QDs would be significantly improved by capping it with hard-core shell. Our data demonstrated that FCS is a simple, fast, and effective method for characterizing the fluorescent quantum dots, and is especially suitable for determining the fluorescent nanoparticles less than 10 nm in water solution

Ethanol- and trifluoroethanol-induced changes in phase states of DPPC membranes. Prodan emission-excitation fluorescence spectroscopy supported by PARAFAC analysis

Science.gov (United States)

Horochowska, Martyna; Cieślik-Boczula, Katarzyna; Rospenk, Maria

2018-03-01

It has been shown that Prodan emission-excitation fluorescence spectroscopy supported by Parallel Factor (PARAFAC) analysis is a fast, simple and sensitive method used in the study of the phase transition from the noninterdigitated gel (Lβ‧) state to the interdigitated gel (LβI) phase, triggered by ethanol and 2,2,2-trifluoroethanol (TFE) molecules in dipalmitoylphosphatidylcholines (DPPC) membranes. The relative contribution of lipid phases with spectral characteristics of each pure phase component has been presented as a function of an increase in alcohol concentration. It has been stated that both alcohol molecules can induce a formation of the LβI phase, but TFE is over six times stronger inducer of the interdigitated phase in DPPC membranes than ethanol molecules. Moreover, in the TFE-mixed DPPC membranes, the transition from the Lβ‧ to LβI phase is accompanied by a formation of the fluid phase, which most probably serves as a boundary phase between the Lβ‧ and LβI regions. Contrary to the three phase-state model of TFE-mixed DPPC membranes, in ethanol-mixed DPPC membranes only the two phase-state model has been detected.

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...

Quantum process tomography by 2D fluorescence spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Pachón, Leonardo A. [Grupo de Física Atómica y Molecular, Instituto de Física, Facultad de Ciencias Exactas y Naturales, Universidad de Antioquia UdeA, Calle 70 No. 52-21, Medellín (Colombia); Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138 (United States); Marcus, Andrew H. [Department of Chemistry and Biochemistry, Oregon Center for Optics, Institute of Molecular Biology, University of Oregon, Eugene, Oregon 97403 (United States); Aspuru-Guzik, Alán [Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138 (United States)

2015-06-07

Reconstruction of the dynamics (quantum process tomography) of the single-exciton manifold in energy transfer systems is proposed here on the basis of two-dimensional fluorescence spectroscopy (2D-FS) with phase-modulation. The quantum-process-tomography protocol introduced here benefits from, e.g., the sensitivity enhancement ascribed to 2D-FS. Although the isotropically averaged spectroscopic signals depend on the quantum yield parameter Γ of the doubly excited-exciton manifold, it is shown that the reconstruction of the dynamics is insensitive to this parameter. Applications to foundational and applied problems, as well as further extensions, are discussed.

Quantum process tomography by 2D fluorescence spectroscopy

International Nuclear Information System (INIS)

Pachón, Leonardo A.; Marcus, Andrew H.; Aspuru-Guzik, Alán

2015-01-01

Reconstruction of the dynamics (quantum process tomography) of the single-exciton manifold in energy transfer systems is proposed here on the basis of two-dimensional fluorescence spectroscopy (2D-FS) with phase-modulation. The quantum-process-tomography protocol introduced here benefits from, e.g., the sensitivity enhancement ascribed to 2D-FS. Although the isotropically averaged spectroscopic signals depend on the quantum yield parameter Γ of the doubly excited-exciton manifold, it is shown that the reconstruction of the dynamics is insensitive to this parameter. Applications to foundational and applied problems, as well as further extensions, are discussed

Compact point-detection fluorescence spectroscopy system for quantifying intrinsic fluorescence redox ratio in brain cancer diagnostics

Science.gov (United States)

Liu, Quan; Grant, Gerald; Li, Jianjun; Zhang, Yan; Hu, Fangyao; Li, Shuqin; Wilson, Christy; Chen, Kui; Bigner, Darell; Vo-Dinh, Tuan

2011-03-01

We report the development of a compact point-detection fluorescence spectroscopy system and two data analysis methods to quantify the intrinsic fluorescence redox ratio and diagnose brain cancer in an orthotopic brain tumor rat model. Our system employs one compact cw diode laser (407 nm) to excite two primary endogenous fluorophores, reduced nicotinamide adenine dinucleotide, and flavin adenine dinucleotide. The spectra were first analyzed using a spectral filtering modulation method developed previously to derive the intrinsic fluorescence redox ratio, which has the advantages of insensitivty to optical coupling and rapid data acquisition and analysis. This method represents a convenient and rapid alternative for achieving intrinsic fluorescence-based redox measurements as compared to those complicated model-based methods. It is worth noting that the method can also extract total hemoglobin concentration at the same time but only if the emission path length of fluorescence light, which depends on the illumination and collection geometry of the optical probe, is long enough so that the effect of absorption on fluorescence intensity due to hemoglobin is significant. Then a multivariate method was used to statistically classify normal tissues and tumors. Although the first method offers quantitative tissue metabolism information, the second method provides high overall classification accuracy. The two methods provide complementary capabilities for understanding cancer development and noninvasively diagnosing brain cancer. The results of our study suggest that this portable system can be potentially used to demarcate the elusive boundary between a brain tumor and the surrounding normal tissue during surgical resection.

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

Minimizing pulling geometry errors in atomic force microscope single molecule force spectroscopy.

Science.gov (United States)

Rivera, Monica; Lee, Whasil; Ke, Changhong; Marszalek, Piotr E; Cole, Daniel G; Clark, Robert L

2008-10-01

In atomic force microscopy-based single molecule force spectroscopy (AFM-SMFS), it is assumed that the pulling angle is negligible and that the force applied to the molecule is equivalent to the force measured by the instrument. Recent studies, however, have indicated that the pulling geometry errors can drastically alter the measured force-extension relationship of molecules. Here we describe a software-based alignment method that repositions the cantilever such that it is located directly above the molecule's substrate attachment site. By aligning the applied force with the measurement axis, the molecule is no longer undergoing combined loading, and the full force can be measured by the cantilever. Simulations and experimental results verify the ability of the alignment program to minimize pulling geometry errors in AFM-SMFS studies.

Polysulfone as a scintillation material without doped fluorescent molecules

Energy Technology Data Exchange (ETDEWEB)

Nakamura, Hidehito, E-mail: hidehito@rri.kyoto-u.ac.jp [Kyoto University, 2, Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494 (Japan); National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba 263-8555 (Japan); Kitamura, Hisashi [National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba 263-8555 (Japan); Sato, Nobuhiro; Kanayama, Masaya [Kyoto University, 2, Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494 (Japan); Shirakawa, Yoshiyuki [Kobe University, 1-1, Rokkodai, Nada, Kobe 657-8501 (Japan); Takahashi, Sentaro [Kyoto University, 2, Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494 (Japan)

2015-10-11

Scintillation materials made from un-doped aromatic ring polymers can be potentially used for radiation detection. Here we demonstrate that Polysulfone (PSU) works without doped fluorescent guest molecules, and thus broadens the choices available for radiation detection. The transparent PSU substrate (1.24 g/cm{sup 3}) significantly absorbs short-wavelength light below approximately 350 nm. Visible light absorption colours the substrate slightly yellow, and indigo blue fluorescence is emitted. The fluorescence maximum occurs at the intersection of the 340-nm excitation and 380-nm emission spectra; thus the emission is partially absorbed by the substrate. An effective refractive index of 1.70 is derived based on the wavelength dependence of the refractive indices and the emission spectrum. A peak caused by 976-keV internal-conversion electrons from a {sup 207}Bi radioactive source appears in the light yield distribution. The light yield is equivalent to that of poly (phenyl sulfone), which has a similar structure. Overall, un-doped PSU could be a component substrate in polymer blends and be used as an educational tool in radiation detection. - Highlights: • Polysulfone (PSU) is a scintillation material that does not require doping. • PSU is slightly yellow. • Indigo blue light with 380-nm emission maximum is emitted. • An effective refractive index of 1.70 was derived. • A peak caused by mono-energetic internal-conversion electrons appears in the light yield distribution.

Blood perfusion and pH monitoring in organs by laser-induced fluorescence spectroscopy

Science.gov (United States)

Vari, Sandor G.; Papazoglou, Theodore G.; Pergadia, Vani R.; Stavridi, Marigo; Snyder, Wendy J.; Papaioannou, Thanassis; Duffy, J. T.; Weiss, Andrew B.; Thomas, Reem; Grundfest, Warren S.

1994-01-01

Sensitivity of laser-induced fluorescence spectroscopy (LIFS) in detecting a change in tissue pH, and blood perfusion was determined. Rabbits were anesthetized, paralyzed, and mechanically ventilated. The arterial and venous blood supplies of the kidney were isolated and ligated to alter the perfusion. The femoral artery was cannulated to extract samples for blood gas analysis. A 308-nm XeCl was used as an excitation source. A 600 micrometers core diameter fiber was used for fluorescence acquisition, and the spectra analyzed by an optical multichannel analyzer (EG & G, OMA III). the corresponding intensity ratio R equals INADH / ICOLL was used as an index for respiratory acidosis. Blood perfusion was assessed using the following algorithm: (IELAS minus ICOLL) divided by (INADH minus ICOLL). The intensity ratio linearly decreased with the reduction of blood perfusion. When we totally occluded the artery the ratio decreased tenfold when compared to the ratio of a fully perfused kidney. Results of monitoring blood acidosis by laser-induced fluorescence spectroscopy shows a significant trend between pH and intensity ratio. Since all the slopes were negative, there is an obvious significant correlation between the pH and NADH.COLLAGEN RATIO. Blue-light-induced fluorescence measurements and ratio fluorometry is a sensitive method for monitoring blood perfusion and acidity or alkalinity of an organ.

Single-molecule force-conductance spectroscopy of hydrogen-bonded complexes

Science.gov (United States)

Pirrotta, Alessandro; De Vico, Luca; Solomon, Gemma C.; Franco, Ignacio

2017-03-01

The emerging ability to study physical properties at the single-molecule limit highlights the disparity between what is observable in an ensemble of molecules and the heterogeneous contributions of its constituent parts. A particularly convenient platform for single-molecule studies are molecular junctions where forces and voltages can be applied to individual molecules, giving access to a series of electromechanical observables that can form the basis of highly discriminating multidimensional single-molecule spectroscopies. Here, we computationally examine the ability of force and conductance to inform about molecular recognition events at the single-molecule limit. For this, we consider the force-conductance characteristics of a prototypical class of hydrogen bonded bimolecular complexes sandwiched between gold electrodes. The complexes consist of derivatives of a barbituric acid and a Hamilton receptor that can form up to six simultaneous hydrogen bonds. The simulations combine classical molecular dynamics of the mechanical deformation of the junction with non-equilibrium Green's function computations of the electronic transport. As shown, in these complexes hydrogen bonds mediate transport either by directly participating as a possible transport pathway or by stabilizing molecular conformations with enhanced conductance properties. Further, we observe that force-conductance correlations can be very sensitive to small changes in the chemical structure of the complexes and provide detailed information about the behavior of single molecules that cannot be gleaned from either measurement alone. In fact, there are regions during the elongation that are only mechanically active, others that are only conductance active, and regions where both force and conductance changes as the complex is mechanically manipulated. The implication is that force and conductance provide complementary information about the evolution of molecules in junctions that can be used to

Time-resolved fluorescence spectroscopy

International Nuclear Information System (INIS)

Gustavsson, Thomas; Mialocq, Jean-Claude

2007-01-01

This article addresses the evolution in time of light emitted by a molecular system after a brief photo-excitation. The authors first describe fluorescence from a photo-physical point of view and discuss the characterization of the excited state. Then, they explain some basic notions related to fluorescence characterization (lifetime and decays, quantum efficiency, so on). They present the different experimental methods and techniques currently used to study time-resolved fluorescence. They discuss basic notions of time resolution and spectral reconstruction. They briefly present some conventional methods: intensified Ccd cameras, photo-multipliers and photodiodes associated with a fast oscilloscope, and phase modulation. Other methods and techniques are more precisely presented: time-correlated single photon counting (principle, examples, and fluorescence lifetime imagery), streak camera (principle, examples), and optical methods like the Kerr optical effect (principle and examples) and fluorescence up-conversion (principle and theoretical considerations, examples of application)

Spectroscopic and dynamical studies of highly energized small polyatomic molecules

Energy Technology Data Exchange (ETDEWEB)

Field, R.W.; Silbey, R.J. [Massachusetts Institute of Technology, Cambridge (United States)

1993-12-01

The authors have initiated a program to perform spectroscopic and dynamic studies of small molecules. Large amplitude motions in excited acetylene were discussed along with plans to record the dispersed fluorescence (DF) and the stimulated emission pumping (SEP) spectra. SEP spectra were reported for the formyl radical. A Fourier transform spectrometer was discussed with respect to its ability to probe the structure of radicals. This instrument is capable of performing studies using various techniques such as magnetic rotation spectroscopy and sub-Doppler sideband-OODR Zeman (SOODRZ) spectroscopy.

Nanostructural origin of blue fluorescence in the mineral karpatite.

Science.gov (United States)

Potticary, Jason; Jensen, Torsten T; Hall, Simon R

2017-08-29

The colour of crystals is a function of their atomic structure. In the case of organic crystals, it is the spatial relationships between molecules that determine the colour, so the same molecules in the same arrangement should produce crystals of the same colour, regardless of whether they arise geologically or synthetically. There is a naturally-occurring organic crystal known as karpatite which is prized for its beautiful blue fluorescence under ultra-violet illumination. When grown under laboratory conditions however, the crystals fluoresce with an intense green colour. For 20 years, this difference has been thought to be due to chemical impurities in the laboratory-grown material. Using electron microscopy coupled with fluorescence spectroscopy and X-Ray diffraction, we report here that this disparity is instead due to differences in the structure of the crystals at the nanoscale. The results show that in nature, karpatite has a nanotexture that is not present in the synthetic crystals, which enables different photonic pathways and therefore a blue, rather than green colour whilst undergoing fluorescence.

Precision spectroscopy with ultracold 87Rb2 triplet molecules

International Nuclear Information System (INIS)

Strauss, Christoph

2011-01-01

In this thesis I report precision spectroscopy with ultracold 87 Rb 2 triplet molecules where we use lasers to couple the states in different molecular potentials. We study in detail states of the a 3 sum + u and (1) 3 sum + g potentials. These states are of great importance for transferring weakly bound molecules to the ro-vibrational triplet ground state via states of the excited potential. As most experiments start from molecules in their X 1 sum + g ground state, the triplet states were hard to access via dipole transitions and remained largely unexplored. The measurements presented in this thesis are the first detailed study of diatomic 87 Rb 2 molecules in these states. Our experiments start with an ultracold cloud of 87 Rb atoms. We then load this cloud into an optical lattice where we use a magnetic Feshbach resonance at 1007.4 G to perform a Feshbach association. After we have removed all unbound atoms, we end up with a pure sample of weakly bound Feshbach molecules inside the optical lattice. The optical lattice prevents these molecules from colliding with each other which results in molecular lifetimes on the order of a few hundred milliseconds. In the first set of experiments, we use a laser coupling the Feshbach state to the excited (1) 3 sum + g triplet state to map out its low-lying vibrational (v = 0.. 15), rotational, hyperfine, and Zeeman structure. The experimental results are in good agreement with calculations done by Marius Lysebo and Prof. Leif Veseth. We then map out in detail the vibrational, rotational, hyperfine, and Zeeman structure of the a 3 sum + u triplet ground state using dark state spectroscopy with levels in the (1) 3 sum + g potential as an intermediate state. In this scheme we are able to access molecules in triplet states because our Feshbach state has strong triplet character. Interestingly, it happens that some deeply bound states which belong to the X 1 sum + g potential are close to levels in the a 3 sum + u potential. In

Theoretical Foundation for Electric-Dipole-Allowed Chiral-Specific Fluorescence Optical Rotary Dispersion (F-ORD) from Interfacial Assemblies.

Science.gov (United States)

Deng, Fengyuan; Ulcickas, James R W; Simpson, Garth J

2016-11-03

Fluorescence optical rotary dispersion (F-ORD) is proposed as a novel chiral-specific and interface-specific spectroscopic method. F-ORD measurements of uniaxial assemblies are predicted to be fully electric-dipole-allowed, with corresponding increases in sensitivity to chirality relative to chiral-specific measurements in isotropic assemblies that are commonly interpreted through coupling between electric and magnetic dynamic dipoles. Observations of strong chiral sensitivity in prior single-molecule fluorescence measurements of chiral interfacial molecules are in excellent qualitative agreement with the predictions of the F-ORD mechanism and challenging to otherwise explain. F-ORD may provide methods to suppress background fluorescence in studies of biological interfaces, as the detected signal requires both polar local order and interfacial chirality. In addition, the molecular-level descriptions of the mechanisms underpinning F-ORD may also potentially apply to aid in interpreting chiral-specific Raman and surface-enhanced Raman spectroscopy measurements of uniaxially oriented assemblies, opening up opportunities for chiral-specific and interface-specific vibrational spectroscopy.

Pancreatic tumor detection using hypericin-based fluorescence spectroscopy and cytology

Science.gov (United States)

Lavu, Harish; Geary, Kevin; Fetterman, Harold R.; Saxton, Romaine E.

2005-04-01

Hypericin is a novel, highly fluorescent photosensitizer that exhibits selective tumor cell uptake properties and is particularly resistant to photobleaching. In this study, we have characterized hypericin uptake in human pancreatic tumor cells with relation to incubation time, cell number, and drug concentration. Ex vivo hypericin based fluorescence spectroscopy was performed to detect the presence of MIA PaCa-2 pancreatic tumor cells in the peritoneal cavity of BALB/c nude mice, as well as to quantify gross tumor burden. Hypericin based cytology of peritoneal lavage samples, using both one and two photon laser confocal microscopy, demonstrated more than a two-fold increase in fluorescence emission of pancreatic tumor cells as compared to control samples. In vitro treatment of pancreatic cancer cells with hypericin based photodynamic therapy showed tumor cell cytotoxicity in a drug dose, incident laser power, and time dependent manner. For these experiments, a continuous wavelength solid-state laser source (532 nm) was operated at power levels in the range of 100-400 mW. Potential applications of hypericin in tumor diagnosis, staging, and therapy will be presented.

High-Frequency Electron Paramagnetic Resonance Spectroscopy of Nitroxide-Functionalized Nanodiamonds in Aqueous Solution.

Science.gov (United States)

Akiel, R D; Stepanov, V; Takahashi, S

2017-06-01

Nanodiamond (ND) is an attractive class of nanomaterial for fluorescent labeling, magnetic sensing of biological molecules, and targeted drug delivery. Many of those applications require tethering of target biological molecules on the ND surface. Even though many approaches have been developed to attach macromolecules to the ND surface, it remains challenging to characterize dynamics of tethered molecule. Here, we show high-frequency electron paramagnetic resonance (HF EPR) spectroscopy of nitroxide-functionalized NDs. Nitroxide radical is a commonly used spin label to investigate dynamics of biological molecules. In the investigation, we developed a sample holder to overcome water absorption of HF microwave. Then, we demonstrated HF EPR spectroscopy of nitroxide-functionalized NDs in aqueous solution and showed clear spectral distinction of ND and nitroxide EPR signals. Moreover, through EPR spectral analysis, we investigate dynamics of nitroxide radicals on the ND surface. The demonstration sheds light on the use of HF EPR spectroscopy to investigate biological molecule-functionalized nanoparticles.

Determination of antioxidant content in biodiesel by fluorescence spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Magalhaes, Keurison F.; Caires, Anderson R.L. [Universidade Federal da Grande Dourados, MS (Brazil). Grupo de Optica Aplicada; Oliveira, Samuel L. [Universidade Federal de Mato Grosso do Sul (UFMS), MS (Brazil). Grupo de Optica e Fotonica

2011-07-01

Full text. Biodiesel is an alternative fuel composed by mono-alkyl esters obtained from vegetable oils or animal fats. Due to its chemical structure, biodiesel is highly susceptible to oxidation which leads to formation of insoluble gums and sediments that can block the filter system of fuel injection. Biodiesel made from vegetable oils typically has a small amount of natural antioxidants so that it is necessary to add synthetic antioxidants to enhance its stability and retain their properties for a longer period. The main antioxidants are synthetic phenolic compounds such as butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and tert-butyl hydroquinone (TBHQ) as well as natural antioxidants as tocopherols. The fluorescence spectroscopy has been applied for determination of phenolic compounds in oils. Here, a method based on fluorescence is proposed to quantify the BHA and TBHQ antioxidant concentration in biodiesel produced from sunflower and soybean oils. Soybean and sunflower biodiesel were obtained by transesterification of fatty alcohol in the presence of NaOH as catalyst. The reactions were carried out in the molar ratio of 6:1 methanol/oil. After the production and purification, biodiesel samples were stored. Biodiesel samples with BHA and TBHQ concentrations from 1000 to 8000 ppm (m/m) were pre- pared. These samples were diluted in ethanol (95%) in order to measure the fluorescence spectra. Fluorescence and excitation spectra of the solutions were recorded at room temperature using a spectrofluorimeter. The emission spectra were obtained under excitation at about 310nm and fluorescence in the 320-800nm range was evaluated. Biodiesel samples without BHA and TBHQ showed fluorescence band at about 420nm, which can be attributed to tocopherols inherent to the vegetable oils used in the biodiesel production. The addition of BHA and/or TBHQ is responsible for the appearance of a fluorescence band around 330nm. It was verified that the fluorescence

Photoelectron Imaging Spectroscopy as a Window to Unexpected Molecules

Science.gov (United States)

Blackstone, Christopher C.

2017-06-01

Targeting an anion with the formula CH_{3}O_{3} for exploration with photoelectron imaging spectroscopy, we determine its identity to be dihydroxymethanolate, an anion largely absent in the literature, and the conjugate base of the hypothetical species orthoformic acid. Comparing the observed photoelectron spectrum to CCSD-EOM-IP and CCSD-EOM-SF calculations completed in QChem and Franck-Condon overlap simulations in PESCAL, we are able to determine with confidence the connectivity of the atoms in this molecule.

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

Phenylboronic acid functionalized reduced graphene oxide based fluorescence nano sensor for glucose sensing

Energy Technology Data Exchange (ETDEWEB)

Basiruddin, SK; Swain, Sarat K., E-mail: swainsk2@yahoo.co.in

2016-01-01

Reduced graphene has emerged as promising tools for detection based application of biomolecules as it has high surface area with strong fluorescence quenching property. We have used the concept of fluorescent quenching property of reduced graphene oxide to the fluorescent probes which are close vicinity of its surface. In present work, we have synthesized fluorescent based nano-sensor consist of phenylboronic acid functionalized reduced graphene oxide (rGO–PBA) and di-ol modified fluorescent probe for detection of biologically important glucose molecules. This fluorescent graphene based nano-probe has been characterized by high resolution transmission electron microscope (HRTEM), Atomic force microscope (AFM), UV–visible, Photo-luminescence (PL) and Fourier transformed infrared (FT-IR) spectroscopy. Finally, using this PBA functionalized reduced GO based nano-sensor, we were able to detect glucose molecule in the range of 2 mg/mL to 75 mg/mL in aqueous solution of pH 7.4. - Highlights: • Easy and simple synthesis of PBA functionalized reduced GO based nano probe. • PBA functionalized reduced GO graphene based nano-probes are characterized. • PBA functionalized reduced GO nano probe is used to detect glucose molecules. • It is very cost-effective and enzyme-free detection of glucose in solution.

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

Laser fluorescence spectroscopy of sputtered uranium atoms

International Nuclear Information System (INIS)

Wright, R.B.; Pellin, M.J.; Gruen, D.M.; Young, C.E.

1979-01-01

Laser induced fluorescence (LIF) spectroscopy was used to study the sputtering of 99.8% 238 U metal foil when bombarded by normally incident 500 to 3000 eV Ne + , Ar + , Kr + , and O 2 + . A three-level atom model of the LIF processes is developed to interpret the observed fluorescent emission from the sputtered species. The model shows that close attention must be paid to the conditions under which the experiment is carried out as well as to the details of the collision cascade theory of sputtering. Rigorous analysis shows that when properly applied, LIF can be used to investigate the predictions of sputtering theory as regards energy distributions of sputtered particles and for the determination of sputtering yields. The possibility that thermal emission may occur during sputtering can also be tested using the proposed model. It is shown that the velocity distribution (either the number density or flux density distribution, depending upon the experimental conditions) of the sputtered particles can be determined using the LIF technique and that this information can be used to obtain a description of the basic sputtering mechanisms. These matters are discussed using the U-atom fluorescence measurements as a basis. The relative sputtering yields for various incident ions on uranium were also measured for the first time using the LIF technique. A surprisingly high fraction of the sputtered uranium atoms were found to occupy the low lying metastable energy levels of U(I). The population of the sputtered metastable atoms were found approximately to obey a Boltzman distribution with an effective temperature of 920 +- 100 0 K. 41 references

Revealing time bunching effect in single-molecule enzyme conformational dynamics.

Science.gov (United States)

Lu, H Peter

2011-04-21

In this perspective, we focus our discussion on how the single-molecule spectroscopy and statistical analysis are able to reveal enzyme hidden properties, taking the study of T4 lysozyme as an example. Protein conformational fluctuations and dynamics play a crucial role in biomolecular functions, such as in enzymatic reactions. Single-molecule spectroscopy is a powerful approach to analyze protein conformational dynamics under physiological conditions, providing dynamic perspectives on a molecular-level understanding of protein structure-function mechanisms. Using single-molecule fluorescence spectroscopy, we have probed T4 lysozyme conformational motions under the hydrolysis reaction of a polysaccharide of E. coli B cell walls by monitoring the fluorescence resonant energy transfer (FRET) between a donor-acceptor probe pair tethered to T4 lysozyme domains involving open-close hinge-bending motions. Based on the single-molecule spectroscopic results, molecular dynamics simulation, a random walk model analysis, and a novel 2D statistical correlation analysis, we have revealed a time bunching effect in protein conformational motion dynamics that is critical to enzymatic functions. Bunching effect implies that conformational motion times tend to bunch in a finite and narrow time window. We show that convoluted multiple Poisson rate processes give rise to the bunching effect in the enzymatic reaction dynamics. Evidently, the bunching effect is likely common in protein conformational dynamics involving in conformation-gated protein functions. In this perspective, we will also discuss a new approach of 2D regional correlation analysis capable of analyzing fluctuation dynamics of complex multiple correlated and anti-correlated fluctuations under a non-correlated noise background. Using this new method, we are able to map out any defined segments along the fluctuation trajectories and determine whether they are correlated, anti-correlated, or non-correlated; after which, a

Fluorescence spectroscopy: a tool to characterize humic substances in soil colonized by microorganisms?

Czech Academy of Sciences Publication Activity Database

Řezáčová, Veronika; Gryndler, Milan

2006-01-01

Roč. 51, č. 3 (2006), s. 215-221 ISSN 0015-5632 R&D Projects: GA ČR GA526/03/0188 Institutional research plan: CEZ:AV0Z50200510 Keywords : fluorescence spectroscopy * humic substances * microorganism Subject RIV: EE - Microbiology, Virology Impact factor: 0.963, year: 2006

Multimodal fluorescence imaging spectroscopy

NARCIS (Netherlands)

Stopel, Martijn H W; Blum, Christian; Subramaniam, Vinod; Engelborghs, Yves; Visser, Anthonie J.W.G.

2014-01-01

Multimodal fluorescence imaging is a versatile method that has a wide application range from biological studies to materials science. Typical observables in multimodal fluorescence imaging are intensity, lifetime, excitation, and emission spectra which are recorded at chosen locations at the sample.

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

Quantification of Coffea arabica and Coffea canephora var. robusta concentration in blends by means of synchronous fluorescence and UV-Vis spectroscopies.

Science.gov (United States)

Dankowska, A; Domagała, A; Kowalewski, W

2017-09-01

The potential of fluorescence, UV-Vis spectroscopies as well as the low- and mid-level data fusion of both spectroscopies for the quantification of concentrations of roasted Coffea arabica and Coffea canephora var. robusta in coffee blends was investigated. Principal component analysis was used to reduce data multidimensionality. To calculate the level of undeclared addition, multiple linear regression (PCA-MLR) models were used with lowest root mean square error of calibration (RMSEC) of 3.6% and root mean square error of cross-validation (RMSECV) of 7.9%. LDA analysis was applied to fluorescence intensities and UV spectra of Coffea arabica, canephora samples, and their mixtures in order to examine classification ability. The best performance of PCA-LDA analysis was observed for data fusion of UV and fluorescence intensity measurements at wavelength interval of 60nm. LDA showed that data fusion can achieve over 96% of correct classifications (sensitivity) in the test set and 100% of correct classifications in the training set, with low-level data fusion. The corresponding results for individual spectroscopies ranged from 90% (UV-Vis spectroscopy) to 77% (synchronous fluorescence) in the test set, and from 93% to 97% in the training set. The results demonstrate that fluorescence, UV, and visible spectroscopies complement each other, giving a complementary effect for the quantification of roasted Coffea arabica and Coffea canephora var. robusta concentration in blends. Copyright © 2017 Elsevier B.V. All rights reserved.

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

Science.gov (United States)

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

2017-11-15

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

X-ray absorption spectroscopy of the chiral molecules fenchone, α-pinene, limonene and carvone in the C1s excitation region

Energy Technology Data Exchange (ETDEWEB)

Ozga, Christian, E-mail: ozga@physik.uni-kassel.de [Institute for Physics and CINSaT, University of Kassel, Heinrich-Plett Str. 40, 34132 Kassel (Germany); Jänkälä, Kari [Centre for Molecular Materials Research, University of Oulu, PO Box 3000, 90014 Oulu (Finland); Schmidt, Philipp; Hans, Andreas; Reiß, Philipp; Ehresmann, Arno; Knie, André [Institute for Physics and CINSaT, University of Kassel, Heinrich-Plett Str. 40, 34132 Kassel (Germany)

2016-02-15

Highlights: • Determination of the X-ray absorption spectra for two terpenoids and two terpenes. • Allocation of predominant or even site-selective excitation of stereocenters. • Fragment fluorescence spectra of the prototype molecules are identical. • Presented data can be used for future fluorescence circular dichroism experiments. - Abstract: Relative ionization cross sections and fluorescence intensities as functions of the exciting-photon energy were recorded for the chiral molecules carvone, α-pinene, limonene and fenchone after excitation by monochromatized synchrotron radiation with energies of the exciting-photons between 284 eV and 289 eV. At selected exciting-photon energies dispersed fragment fluorescence spectra in the wavelength range between 365 nm and 505 nm were obtained. Time dependent density functional theory (TD-DFT) computations were performed to analyze the experimentally observed resonance-structures. Comparison of the computed and recorded spectra demonstrates the possibility of a predominant or even specific excitation of one particular stereocenter site in a molecule with more than one stereocenter.

Diode-Laser Induced Fluorescence Spectroscopy of an Optically Thick Plasma in Combination with Laser Absorption Spectroscopy

Directory of Open Access Journals (Sweden)

S. Nomura

2013-01-01

Full Text Available Distortion of laser-induced fluorescence profiles attributable to optical absorption and saturation broadening was corrected in combination with laser absorption spectroscopy in argon plasma flow. At high probe-laser intensity, saturated absorption profiles were measured to correct probe-laser absorption. At low laser intensity, nonsaturated absorption profiles were measured to correct fluorescence reabsorption. Saturation broadening at the measurement point was corrected using a ratio of saturated to non-saturated broadening. Observed LIF broadening and corresponding translational temperature without correction were, respectively, 2.20±0.05 GHz and 2510±100 K and corrected broadening and temperature were, respectively, 1.96±0.07 GHz and 1990±150 K. Although this correction is applicable only at the center of symmetry, the deduced temperature agreed well with that obtained by LAS with Abel inversion.

Monitoring of petroleum hydrocarbon pollution in surface waters by a direct comparison of fluorescence spectroscopy and remote sensing techniques

Energy Technology Data Exchange (ETDEWEB)

De Domenico, L.; Crisafi, E. (Consiglio Nazionale delle Ricerche, Messina (Italy). Thalassografic Inst.); Magazzu, G. (Lecce Univ. (Italy). Dept. of Biology); Puglisi, A. (Mediterranean Oceanological Centre (CEOM), Palermo (Italy)); La Rosa, A. (Air-Survey, Italy s.r.l., Catania (Italy))

1994-10-01

Oil pollution levels were estimated using simultaneous acquisition of data from remote sensing by helicopter and fluorescence spectroscopy on surface samples. Laboratory quantitative analysis of hydrocarbons was used to calibrate remotely sensed data. The data were treated using a computer to generate a colour-coded map not attainable with conventional methods representing seawater pollution. Results were in good agreement and indicated that remotely sensed data together with those achieved by fluorescence spectroscopy are applicable for monitoring hydrocarbon pollution. (author)

Monitoring of petroleum hydrocarbon pollution in surface waters by a direct comparison of fluorescence spectroscopy and remote sensing techniques

International Nuclear Information System (INIS)

De Domenico, L.; Crisafi, E.; La Rosa, A.

1994-01-01

Oil pollution levels were estimated using simultaneous acquisition of data from remote sensing by helicopter and fluorescence spectroscopy on surface samples. Laboratory quantitative analysis of hydrocarbons was used to calibrate remotely sensed data. The data were treated using a computer to generate a colour-coded map not attainable with conventional methods representing seawater pollution. Results were in good agreement and indicated that remotely sensed data together with those achieved by fluorescence spectroscopy are applicable for monitoring hydrocarbon pollution. (author)

Chemometric classification of Chinese lager beers according to manufacturer based on data fusion of fluorescence, UV and visible spectroscopies.

Science.gov (United States)

Tan, Jin; Li, Rong; Jiang, Zi-Tao

2015-10-01

We report an application of data fusion for chemometric classification of 135 canned samples of Chinese lager beers by manufacturer based on the combination of fluorescence, UV and visible spectroscopies. Right-angle synchronous fluorescence spectra (SFS) at three wavelength difference Δλ=30, 60 and 80 nm and visible spectra in the range 380-700 nm of undiluted beers were recorded. UV spectra in the range 240-400 nm of diluted beers were measured. A classification model was built using principal component analysis (PCA) and linear discriminant analysis (LDA). LDA with cross-validation showed that the data fusion could achieve 78.5-86.7% correct classification (sensitivity), while those rates using individual spectroscopies ranged from 42.2% to 70.4%. The results demonstrated that the fluorescence, UV and visible spectroscopies complemented each other, yielding higher synergic effect. Copyright © 2015 Elsevier Ltd. All rights reserved.

Fluorescent atom coincidence spectroscopy of extremely neutron-deficient barium isotopes

International Nuclear Information System (INIS)

Wells, S.A.; Evans, D.E.; Griffith, J.A.R.; Eastham, D.A.; Groves, J.; Smith, J.R.H.; Tolfree, D.W.L.; Warner, D.D.; Billowes, J.; Grant, I.S.; Walker, P.M.

1988-01-01

Fluorescent atom coincidence spectroscopy (FACS) has been used to measure the nuclear mean square radii and moments of the extremely neutron-deficient isotopes 120-124 Ba. At N=65 an abrupt change in nuclear mean square charge radii is observed which can be understood in terms of the occupation of the spin-orbit partner g 7/2 5/2[413] neutron and g 9/2 9/2[404] proton orbitals and the consequent enhancement of the n-p interaction. (orig.)

Assessment of drinking water quality at the tap using fluorescence spectroscopy

OpenAIRE

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

2017-01-01

Treated drinking water may become contaminated while travelling in the distribution system on the way to consumers. Elevated dissolved organic matter (DOM) at the tap relative to the water leaving the treatment plant is a potential indicator of contamination, and can be measured sensitively, inexpensively and potentially on-line via fluorescence and absorbance spectroscopy. Detecting elevated DOM requires potential contamination events to be distinguished from natural fluctuations in the syst...

Single-Molecule Fluorescence Reveals the Oligomerization and Folding Steps Driving the Prion-like Behavior of ASC.

Science.gov (United States)

Gambin, Yann; Giles, Nichole; O'Carroll, Ailís; Polinkovsky, Mark; Hunter, Dominic; Sierecki, Emma

2018-02-16

Single-molecule fluorescence has the unique ability to quantify small oligomers and track conformational changes at a single-protein level. Here we tackled one of the most extreme protein behaviors, found recently in an inflammation pathway. Upon danger recognition in the cytosol, NLRP3 recruits its signaling adaptor, ASC. ASC start polymerizing in a prion-like manner and the system goes in "overdrive" by producing a single micron-sized "speck." By precisely controlling protein expression levels in an in vitro translation system, we could trigger the polymerization of ASC and mimic formation of specks in the absence of inflammasome nucleators. We utilized single-molecule spectroscopy to fully characterize prion-like behaviors and self-propagation of ASC fibrils. We next used our controlled system to monitor the conformational changes of ASC upon fibrillation. Indeed, ASC consists of a PYD and CARD domains, separated by a flexible linker. Individually, both domains have been found to form fibrils, but the structure of the polymers formed by the full-length ASC proteins remains elusive. For the first time, using single-molecule Förster resonance energy transfer, we studied the relative positions of the CARD and PYD domains of full-length ASC. An unexpectedly large conformational change occurred upon ASC fibrillation, suggesting that the CARD domain folds back onto the PYD domain. However, contradicting current models, the "prion-like" conformer was not initiated by binding of ASC to the NLRP3 platform. Rather, using a new method, hybrid between Photon Counting Histogram and Number and Brightness analysis, we showed that NLRP3 forms hexamers with self-binding affinities around 300nM. Overall our data suggest a new mechanism, where NLRP3 can initiate ASC polymerization simply by increasing the local concentration of ASC above a supercritical level. Copyright © 2017. Published by Elsevier Ltd.

Fluorescent scattering by molecules embedded in small particles. Progress report, February 1, 1981-January 31, 1982

International Nuclear Information System (INIS)

Chew, H.; McNulty, P.J.

1982-01-01

In earlier work a model of fluorescent and Raman scattering by active molecules represented as classical electric dipoles embedded in small particles was developed. The intensity and angular distribution of the inelastically scattered radiation was shown to depend on the geometric and optical properties of the particle. The model was originally developed for particles having spherical shape and later extended to concentric spheres, cylinders, and prolate spheroids. The active molecules were originally assumed to be isotropically polarizable. The model has been recently extended to certain types of anisotropically polarizable molecules. The model had also been applied to particles having internal structure

Evaluating Activated Carbon Adsorption of Dissolved Organic Matter and Micropollutants Using Fluorescence Spectroscopy.

Science.gov (United States)

Shimabuku, Kyle K; Kennedy, Anthony M; Mulhern, Riley E; Summers, R Scott

2017-03-07

Dissolved organic matter (DOM) negatively impacts granular activated carbon (GAC) adsorption of micropollutants and is a disinfection byproduct precursor. DOM from surface waters, wastewater effluent, and 1 kDa size fractions were adsorbed by GAC and characterized using fluorescence spectroscopy, UV-absorption, and size exclusion chromatography (SEC). Fluorescing DOM was preferentially adsorbed relative to UV-absorbing DOM. Humic-like fluorescence (peaks A and C) was selectively adsorbed relative to polyphenol-like fluorescence (peaks T and B) potentially due to size exclusion effects. In the surface waters and size fractions, peak C was preferentially removed relative to peak A, whereas the reverse was found in wastewater effluent, indicating that humic-like fluorescence is associated with different compounds depending on DOM source. Based on specific UV-absorption (SUVA), aromatic DOM was preferentially adsorbed. The fluorescence index (FI), if interpreted as an indicator of aromaticity, indicated the opposite but exhibited a strong relationship with average molecular weight, suggesting that FI might be a better indicator of DOM size than aromaticity. The influence of DOM intermolecular interactions on adsorption were minimal based on SEC analysis. Fluorescence parameters captured the impact of DOM size on the fouling of 2-methylisoborneol and warfarin adsorption and correlated with direct competition and pore blockage indicators.

Method for rapid multidiameter single-fiber reflectance and fluorescence spectroscopy through a fiber bundle

NARCIS (Netherlands)

Amelink, A.; Hoy, C.L.; Gamm, U.A.; Sterenborg, H.J.C.M.; Robinson, D.J.

2014-01-01

We have recently demonstrated a means for quantifying the absorption and scattering properties of biological tissue through multidiameter single-fiber reflectance (MDSFR) spectroscopy. These measurements can be used to correct single-fiber fluorescence (SFF) spectra for the influence of optical

Recent Progress on Plasmon-Enhanced Fluorescence

Directory of Open Access Journals (Sweden)

Dong Jun

2015-12-01

Full Text Available The optically generated collective electron density waves on metal–dielectric boundaries known as surface plasmons have been of great scientific interest since their discovery. Being electromagnetic waves on gold or silver nanoparticle’s surface, localised surface plasmons (LSP can strongly enhance the electromagnetic field. These strong electromagnetic fields near the metal surfaces have been used in various applications like surface enhanced spectroscopy (SES, plasmonic lithography, plasmonic trapping of particles, and plasmonic catalysis. Resonant coupling of LSPs to fluorophore can strongly enhance the emission intensity, the angular distribution, and the polarisation of the emitted radiation and even the speed of radiative decay, which is so-called plasmon enhanced fluorescence (PEF. As a result, more and more reports on surface-enhanced fluorescence have appeared, such as SPASER-s, plasmon assisted lasing, single molecule fluorescence measurements, surface plasmoncoupled emission (SPCE in biological sensing, optical orbit designs etc. In this review, we focus on recent advanced reports on plasmon-enhanced fluorescence (PEF. First, the mechanism of PEF and early results of enhanced fluorescence observed by metal nanostructure will be introduced. Then, the enhanced substrates, including periodical and nonperiodical nanostructure, will be discussed and the most important factor of the spacer between molecule and surface and wavelength dependence on PEF is demonstrated. Finally, the recent progress of tipenhanced fluorescence and PEF from the rare-earth doped up-conversion (UC and down-conversion (DC nanoparticles (NPs are also commented upon. This review provides an introduction to fundamentals of PEF, illustrates the current progress in the design of metallic nanostructures for efficient fluorescence signal amplification that utilises propagating and localised surface plasmons.

Characterization of nanostructures in the live cell plasma membrane utilizing advanced single molecule fluorescence techniques

International Nuclear Information System (INIS)

Brameshuber, M.

2009-01-01

Unrevealing the detailed structure of the cellular plasma membrane at a nanoscopic length scale is the key for understanding the regulation of various signaling pathways or interaction mechanism. Hypotheses postulate the existence of nanoscopic lipid platforms in the cell membrane which are termed lipid- or membrane rafts. Based on biochemical studies, rafts are believed to play a crucial role in many signaling processes. However, there is currently not much information on their size, shape, stability, surface density, composition and heterogeneity. In this thesis I present an ultra-sensitive fluorescence based method which allows for the first time the direct imaging of single mobile rafts in the live cell plasma membrane. The method senses rafts by their property to assemble a characteristic set of fluorescent marker-proteins or lipids on a time-scale of seconds. A special photobleaching protocol was developed and used to reduce the surface density of labeled mobile rafts down to the level of well-isolated diffraction-limited spots, without altering the single spot brightness. The statistical distribution of probe molecules per raft was determined by single molecule brightness analysis. For demonstration, I used the consensus markers Bodipy-GM1, a fluorescent lipid analogue, and glycosylphosphatidyl-inositol-anchored monomeric GFP. For both markers I found cholesterol-dependent association in the plasma membrane of living CHO and Jurkat T cells in the resting state, indicating the presence of mobile, stable rafts hosting these probes. I further characterized these structures by taking cell-to-cell variations under consideration. By comparing Bodipy-GM1 with mGFP-GPI homo-association upon temperature variation, two different states - a non-equilibrated and an equilibrated state - could be identified. I conclude that rafts are loaded non-randomly; the characteristic load is maintained during its lifetime in the plasma membrane of a non-activated cell. Beside these

Optical spectroscopy of laser-produced plasmas for standoff isotopic analysis

Science.gov (United States)

Harilal, S. S.; Brumfield, B. E.; LaHaye, N. L.; Hartig, K. C.; Phillips, M. C.

2018-06-01

Rapid, in-field, and non-contact isotopic analysis of solid materials is extremely important to a large number of applications, such as nuclear nonproliferation monitoring and forensics, geochemistry, archaeology, and biochemistry. Presently, isotopic measurements for these and many other fields are performed in laboratory settings. Rapid, in-field, and non-contact isotopic analysis of solid material is possible with optical spectroscopy tools when combined with laser ablation. Laser ablation generates a transient vapor of any solid material when a powerful laser interacts with a sample of interest. Analysis of atoms, ions, and molecules in a laser-produced plasma using optical spectroscopy tools can provide isotopic information with the advantages of real-time analysis, standoff capability, and no sample preparation requirement. Both emission and absorption spectroscopy methods can be used for isotopic analysis of solid materials. However, applying optical spectroscopy to the measurement of isotope ratios from solid materials presents numerous challenges. Isotope shifts arise primarily due to variation in nuclear charge distribution caused by different numbers of neutrons, but the small proportional nuclear mass differences between nuclei of various isotopes lead to correspondingly small differences in optical transition wavelengths. Along with this, various line broadening mechanisms in laser-produced plasmas and instrumental broadening generated by the detection system are technical challenges frequently encountered with emission-based optical diagnostics. These challenges can be overcome by measuring the isotope shifts associated with the vibronic emission bands from molecules or by using the techniques of laser-based absorption/fluorescence spectroscopy to marginalize the effect of instrumental broadening. Absorption and fluorescence spectroscopy probe the ground state atoms existing in the plasma when it is cooler, which inherently provides narrower

Highly sensitive immunoassay of protein molecules based on single nanoparticle fluorescence detection in a nanowell

Science.gov (United States)

Han, Jin-Hee; Kim, Hee-Joo; Lakshmana, Sudheendra; Gee, Shirley J.; Hammock, Bruce D.; Kennedy, Ian M.

2011-03-01

A nanoarray based-single molecule detection system was developed for detecting proteins with extremely high sensitivity. The nanoarray was able to effectively trap nanoparticles conjugated with biological sample into nanowells by integrating with an electrophoretic particle entrapment system (EPES). The nanoarray/EPES is superior to other biosensor using immunoassays in terms of saving the amounts of biological solution and enhancing kinetics of antibody binding due to reduced steric hindrance from the neighboring biological molecules. The nanoarray patterned onto a layer of PMMA and LOL on conductive and transparent indium tin oxide (ITO)-glass slide by using e-beam lithography. The suspension of 500 nm-fluorescent (green emission)-carboxylated polystyrene (PS) particles coated with protein-A followed by BDE 47 polyclonal antibody was added to the chip that was connected to the positive voltage. The droplet was covered by another ITO-coated-glass slide and connected to a ground terminal. After trapping the particles into the nanowells, the solution of different concentrations of anti-rabbit- IgG labeled with Alexa 532 was added for an immunoassay. A single molecule detection system could quantify the anti-rabbit IgG down to atto-mole level by counting photons emitted from the fluorescent dye bound to a single nanoparticle in a nanowell.

Interaction of Chelerythrine with Keyhole Limpet Hemocyanin: a Fluorescence Spectroscopy and Molecular Docking Study

Science.gov (United States)

Zhong, M.; Long, R. Q.; Wang, Y. H.; Chen, C. L.

2018-05-01

The quenching mechanism between chelerythrine (CHE) and keyhole limpet hemocyanin (KLH) was investigated using fluorescence spectroscopy and molecular docking. The experiments were conducted at three different temperatures (293, 298, and 303 K). The results revealed that the intrinsic fluorescence of KLH was strongly quenched by CHE through a static quenching mechanism. The thermodynamic parameters (ΔG, ΔH, and ΔS) of the interaction were calculated, indicating that the interaction between CHE and KLH was spontaneous and that van der Waals forces and hydrogen bond formation played major roles in the binding process. The intrinsic fluorescence of the tyrosine and tryptophan residues in KLH was studied by synchronous fluorescence, which suggested that CHE changed the conformation of KLH. Finally, molecular docking was used to obtain detailed information on the binding sites and binding affinities between CHE and KLH.

PyCorrFit-generic data evaluation for fluorescence correlation spectroscopy.

Science.gov (United States)

Müller, Paul; Schwille, Petra; Weidemann, Thomas

2014-09-01

We present a graphical user interface (PyCorrFit) for the fitting of theoretical model functions to experimental data obtained by fluorescence correlation spectroscopy (FCS). The program supports many data file formats and features a set of tools specialized in FCS data evaluation. The Python source code is freely available for download from the PyCorrFit web page at http://pycorrfit.craban.de. We offer binaries for Ubuntu Linux, Mac OS X and Microsoft Windows. © The Author 2014. Published by Oxford University Press.

Measurement of the spectrum of electric-field fluctuations in a plasma by laser-fluorescence spectroscopy

International Nuclear Information System (INIS)

Hildebrandt, J.; Kunze, H.

1980-01-01

Laser-fluorescence spectroscopy has been applied to measure the spectrum of electric wave fields with high temporal resolution in a pulsed hollow-cathode discharge. A low-frequency and a high-frequency component can be identified

Quantitative fluorescence lifetime spectroscopy in turbid media: comparison of theoretical, experimental and computational methods

International Nuclear Information System (INIS)

Vishwanath, Karthik; Mycek, Mary-Ann; Pogue, Brian

2002-01-01

A Monte Carlo model developed to simulate time-resolved fluorescence propagation in a semi-infinite turbid medium was validated against previously reported theoretical and computational results. Model simulations were compared to experimental measurements of fluorescence spectra and lifetimes on tissue-simulating phantoms for single and dual fibre-optic probe geometries. Experiments and simulations using a single probe revealed that scattering-induced artefacts appeared in fluorescence emission spectra, while fluorescence lifetimes were unchanged. Although fluorescence lifetime measurements are generally more robust to scattering artefacts than are measurements of fluorescence spectra, in the dual-probe geometry scattering-induced changes in apparent lifetime were predicted both from diffusion theory and via Monte Carlo simulation, as well as measured experimentally. In all cases, the recovered apparent lifetime increased with increasing scattering and increasing source-detector separation. Diffusion theory consistently underestimated the magnitude of these increases in apparent lifetime (predicting a maximum increase of ∼15%), while Monte Carlo simulations and experiment were closely matched (showing increases as large as 30%). These results indicate that quantitative simulations of time-resolved fluorescence propagation in turbid media will be important for accurate recovery of fluorophore lifetimes in biological spectroscopy and imaging applications. (author)

Application of spectroscopy and super-resolution microscopy: Excited state

Energy Technology Data Exchange (ETDEWEB)

Bhattacharjee, Ujjal [Iowa State Univ., Ames, IA (United States)

2016-02-19

Photophysics of inorganic materials and organic molecules in complex systems have been extensively studied with absorption and emission spectroscopy.1-4 Steady-state and time-resolved fluorescence studies are commonly carried out to characterize excited-state properties of fluorophores. Although steady-state fluorescence measurements are widely used for analytical applications, time-resolved fluorescence measurements provide more detailed information about excited-state properties and the environment in the vicinity of the fluorophore. Many photophysical processes, such as photoinduced electron transfer (PET), rotational reorientation, solvent relaxation, and energy transfer, occur on a nanosecond (10^-9 s) timescale, thus affecting the lifetime of the fluorophores. Moreover, time-resolved microscopy methods, such as lifetimeimaging, combine the benefits of the microscopic measurement and information-rich, timeresolved data. Thus, time-resolved fluorescence spectroscopy combined with microscopy can be used to quantify these processes and to obtain a deeper understanding of the chemical surroundings of the fluorophore in a small area under investigation. This thesis discusses various photophysical and super-resolution microscopic studies of organic and inorganic materials, which have been outlined below.

Monitoring the process of purification of crude glycerol derived from biodiesel production: a method based on fluorescence spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Magalhaes, Keurison F.; Caires, Anderson R.L. [Universidade Federal da Grande Dourados, MS (Brazil). Grupo de Optica Aplicada; Oliveira, Samuel L. [Universidade Federal de Mato Grosso do Sul (UFMS), MS (Brazil). Grupo de Optica e Fotonica

2011-07-01

Full text. The use of biodiesel has increased worldwide. The biodiesel production on an industrial scale has been based on the transesterification of vegetable oils and fats with methanol in the presence of an alkaline catalyst. During the transesterification, one molecule of triglyceride reacts with three molecules of alcohol to produce glycerol and molecules of alkyl esters (biodiesel). As a result, an increase in biodiesel production also enhances the availability of glycerol on the market. However, crude glycerin has about 30% of impurities which are inherent to biodiesel production such as catalyst, alcohol and fatty acids. The present study evaluated the usefulness of the fluorescence spectroscopy as a tool to monitor the glycerol purification process. Glycerol samples were obtained from transesterification of soybean, canola, and sunflower oils in the presence of NaOH. After stirring time, the solutions were let to stand in separating funnels, then two phases were observed: one containing mainly biodiesel and other consisting of glycol. Then, the respective glycerol samples were collected, henceforth called G1. After that, it was added H2SO4 (20%) in the crude glycerol samples to reduce their pH to 4 in order to remove fatty acids. The solutions were stored for 24 hours in separating funnels. The glycerol (heavy phase), hereafter named G2, was then separated and filtered. To remove other impurities from G2 samples by means of ionic exchange columns, the samples were neutralized and diluted using Milli-Q water (G3 samples). Aliquots of 20 mL were then passed through cationic and anionic resins (G4 and G5 samples, respectively). Emission and excitation spectra of the G1-G5 samples as well as of the glycerol PA-ACS (reference) were recorded at room temperature using a spectrofluorimeter. The emission spectra were obtained setting the excitation at 325nm and monitoring the emission in the 330-800nm range. Fluorimetric maps were also achieved by pumping the

Speciation of actinides in aqueous solution by time-resolved laser-induced fluorescence spectroscopy (TRLFS)

International Nuclear Information System (INIS)

Kimura, Takaumi; Kato, Yoshiharu; Meinrath, G.; Yoshida, Zenko; Choppin, G.R.

1995-01-01

Time-resolved laser-induced fluorescence spectroscopy (TRLFS) as a sensitive and selective method has been applied to the speciation of actinides in aqueous solution. Studies on hydrolysis and carbonate complexation of U(VI) and on determination of hydration number of Cm(III) are reported. (author)

Temperature Dependence of Charge Localization in High-Mobility, Solution-Crystallized Small Molecule Semiconductors Studied by Charge Modulation Spectroscopy

DEFF Research Database (Denmark)

Meneau, Aurélie Y. B.; Olivier, Yoann; Backlund, Tomas

2016-01-01

In solution-processable small molecule semiconductors, the extent of charge carrier wavefunction localization induced by dynamic disorder can be probed spectroscopically as a function of temperature using charge modulation spectroscopy (CMS). Here, it is shown based on combined fi eld-effect tran......In solution-processable small molecule semiconductors, the extent of charge carrier wavefunction localization induced by dynamic disorder can be probed spectroscopically as a function of temperature using charge modulation spectroscopy (CMS). Here, it is shown based on combined fi eld......-effect transistor and CMS measurements as a function of temperature that in certain molecular semiconductors, such as solution-processible pentacene, charge carriers become trapped at low temperatures in environments in which the charges become highly localized on individual molecules, while in some other molecules...

Binding of Coumarin 334 with β-Cyclodextrin and with C-Hexylpyrogallol[4]arene: Opposite Fluorescence Behavior

Directory of Open Access Journals (Sweden)

Chandrasekaran Sowrirajan

2013-01-01

Full Text Available We report here the structure of the host-guest complexes of Coumarin 334 (C334 with β-cyclodextrin (β-CD and with C-hexylpyrogallol[4]arene (C-HPA and the effect of acidity on the neutral-cation equilibrium of C334 in water and in the presence of the host molecules. The structures of the host-guest complexes are proposed on the basis of the change of fluorescence on the addition of β-CD or C-HPA to C334 and by 2D ROESY spectroscopy. Opposite fluorescence behaviors, that is, quenching of fluorescence in β-CD and enhancement of fluorescence in C-HPA are observed. Time-resolved fluorescence analysis is done for the complexation, and biexponential decay pattern is observed. The possible strong inclusion complexation with C-HPA is explained. The ground and the excited state pKa values for the protonation equilibrium of C334 in water and the difficulty of protonation in the presence of the host molecules are discussed.

Fluorescence spectroscopy of dental calculus

International Nuclear Information System (INIS)

Bakhmutov, D; Gonchukov, S; Sukhinina, A

2010-01-01

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

Fluorescence spectroscopy of dental calculus

Science.gov (United States)

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

2010-05-01

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

Analysis of photon count data from single-molecule fluorescence experiments

Science.gov (United States)

Burzykowski, T.; Szubiakowski, J.; Rydén, T.

2003-03-01

We consider single-molecule fluorescence experiments with data in the form of counts of photons registered over multiple time-intervals. Based on the observation schemes, linking back to works by Dehmelt [Bull. Am. Phys. Soc. 20 (1975) 60] and Cook and Kimble [Phys. Rev. Lett. 54 (1985) 1023], we propose an analytical approach to the data based on the theory of Markov-modulated Poisson processes (MMPP). In particular, we consider maximum-likelihood estimation. The method is illustrated using a real-life dataset. Additionally, the properties of the proposed method are investigated through simulations and compared to two other approaches developed by Yip et al. [J. Phys. Chem. A 102 (1998) 7564] and Molski [Chem. Phys. Lett. 324 (2000) 301].

Remote imaging laser-induced breakdown spectroscopy and laser-induced fluorescence spectroscopy using nanosecond pulses from a mobile lidar system.

Science.gov (United States)

Grönlund, Rasmus; Lundqvist, Mats; Svanberg, Sune

2006-08-01

A mobile lidar system was used in remote imaging laser-induced breakdown spectroscopy (LIBS) and laser-induced fluorescence (LIF) experiments. Also, computer-controlled remote ablation of a chosen area was demonstrated, relevant to cleaning of cultural heritage items. Nanosecond frequency-tripled Nd:YAG laser pulses at 355 nm were employed in experiments with a stand-off distance of 60 meters using pulse energies of up to 170 mJ. By coaxial transmission and common folding of the transmission and reception optical paths using a large computer-controlled mirror, full elemental imaging capability was achieved on composite targets. Different spectral identification algorithms were compared in producing thematic data based on plasma or fluorescence light.

Application of terahertz spectroscopy for characterization of biologically active organic molecules in natural environment

Science.gov (United States)

Karaliūnas, Mindaugas; Jakštas, Vytautas; Nasser, Kinan E.; Venckevičius, Rimvydas; Urbanowicz, Andrzej; Kašalynas, Irmantas; Valušis, Gintaras

2016-09-01

In this work, a comparative research of biologically active organic molecules in its natural environment using the terahertz (THz) time domain spectroscopy (TDS) and Fourier transform spectroscopy (FTS) systems is carried out. Absorption coefficient and refractive index of Nicotiana tabacum L. leaves containing nicotine, Cannabis sativa L. leaves containing tetrahydrocannabinol, and Humulu lupulus L. leaves containing α-acids, active organic molecules that obtain in natural environment, were measured in broad frequency range from 0.1 to 13 THz at room temperature. In the spectra of absorption coefficient the features were found to be unique for N. tabacum, C. sativa and H. lupulus. Moreover, those features can be exploited for identification of C. sativa sex and N. tabacum origin. The refractive index can be also used to characterize different species.

Determination of Concentration of Living Immobilized Yeast Cells by Fluorescence Spectroscopy

Czech Academy of Sciences Publication Activity Database

Podrazký, Ondřej; Kuncová, Gabriela

2005-01-01

Roč. 107, č. 1 (2005), s. 126-134 ISSN 0925-4005. [European Conference on Optical Chemical Sensors and Biosensors EUROPT(R)ODE /7./. Madrid, 04.04.2004-07.04.2004] R&D Projects: GA ČR GA104/01/0461; GA MŠk(CZ) OC 840.10 Institutional research plan: CEZ:AV0Z40720504 Keywords : immobilization of cells * 2-D fluorescence spectroscopy * sol–gel Subject RIV: CE - Biochemistry Impact factor: 2.646, year: 2005

Spectroscopy of molecules in very high rotational states using an optical centrifuge.

Science.gov (United States)

Yuan, Liwei; Toro, Carlos; Bell, Mack; Mullin, Amy S

2011-01-01

We have developed a high power optical centrifuge for measuring the spectroscopy of molecules in extreme rotational states. The optical centrifuge has a pulse energy that is more than 2 orders of magnitude greater than in earlier instruments. The large pulse energy allows us to drive substantial number densities of molecules to extreme rotational states in order to measure new spectroscopic transitions that are not accessible with traditional methods. Here we demonstrate the use of the optical centrifuge for measuring IR transitions of N2O from states that have been inaccessible until now. In these studies, the optical centrifuge drives N2O molecules into states with J ~ 200 and we use high resolution transient IR probing to measure the appearance of population in states with J = 93-99 that result from collisional cooling of the centrifuged molecules. High resolution Doppler broadened line profile measurements yield information about the rotational and translational energy distributions in the optical centrifuge.

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

Sub-threshold excited Cl Kβ (K-V) x-ray fluorescence from CF3Cl molecule

International Nuclear Information System (INIS)

Perera, R.C.C.; Cowan, P.L.; Lindle, D.W.; LaVilla, R.E.

1987-10-01

With the availability of tunable synchrotron radiation sources, unoccupied molecular orbits (below vacuum level) can be selectively populated producing highly excited neutral molecules. X-ray fluorescence spectra from molecules were obtained with excitation below the ionization threshold and were observed to have significant intensity changes, absolute and relative energy position shifts and line width changes as compared to fluorescence spectra excited above the threshold. As an example, the Cl Kβ (K-V) emission spectra from CF 3 Cl vapor are presented. The energy shifts and intensity changes are explained in terms of perturbation effects due to the presence of an electron in an unoccupied molecular orbital. The narrow line widths obtained in the spectra excited below threshold are explained in terms of the ''effective'' hole production region in a core state limited by the broadening of the unoccupied level. The change in line widths as a function of below-threshold excitation energy is proposed as a novel technique to study the localized properties and reorganization effects of a hole in a core level. 10 refs., 4 figs., 1 tab

Detection of ultra-low oxygen concentration based on the fluorescence blinking dynamics of single molecules

Science.gov (United States)

Wu, Ruixiang; Chen, Ruiyun; Zhou, Haitao; Qin, Yaqiang; Zhang, Guofeng; Qin, Chengbing; Gao, Yan; Gao, Yajun; Xiao, Liantuan; Jia, Suotang

2018-01-01

We present a sensitive method for detection of ultra-low oxygen concentrations based on the fluorescence blinking dynamics of single molecules. The relationship between the oxygen concentration and the fraction of time spent in the off-state, stemming from the population and depopulation of triplet states and radical cationic states, can be fitted with a two-site quenching model in the Stern-Volmer plot. The oxygen sensitivity is up to 43.42 kPa-1 in the oxygen partial pressure region as low as 0.01-0.25 kPa, which is seven times higher than that of the fluorescence intensity indicator. This method avoids the limitation of the sharp and non-ignorable fluctuations that occur during the measurement of fluorescence intensity, providing potential applications in the field of low oxygen-concentration monitoring in life science and industry.

Characterization of the photoreaction between DNA and aminomethyl-trimethylpsoralen using absorption and fluorescence spectroscopy

International Nuclear Information System (INIS)

Johnston, B.H.; Hearst, J.E.

1981-01-01

The use of absorption and fluorescence spectroscopy for following the progress of the photoreaction between DNA and 4'-aminomethyl-4,5',8-trimethylpsoralen (AMT) has been investigated. Absorption at long wavelengths and fluorescence both decline upon intercalation of AMT into the DNA helix. The loss of fluorescence from AMT and the accompanying appearance of monoadduct fluorescence upon irradiation by UV light can be easily followed by using the excitation beam of a spectrofluorometer as the source of irradiation and monitoring the changing emission spectrum. Where cross-link formation is possible, the subsequent decline of monoadduct fluorescence is seen as well. This suggests that the 4',5'-monoadduct is a precursor of cross-links. Both monoaddition and cross-linking are more rapid with poly d(A-T) than with calf thymus DNA or poly d(A.T). Excitation spectra can be helpful in resolving the levels of AMT and 4',5'-monoadduct when both are contributing to the emission spectrum. Some changes are observed in the emission spectrum of AMT-poly d(A.T) monoadducts after prolonged irradiation which indicate further photoreaction. (author)

Accessibility of nucleic acid-complexed biomolecules to hydroxyl radicals correlates with their conformation: a fluorescence polarization spectroscopy study

International Nuclear Information System (INIS)

Makrigiorgos, G.M.; Bump, E.; Huang, C.; Kassis, A.I.; Baranowska-Kortylewicz, J.

1994-01-01

A fluorescence methodology has been developed to examine the relationship between the conformational state of specific biomolecules in simple chromatin models and their accessibility to hydroxyl radicals ( . OH). Polylysine and histone H1 were labelled with SECCA, the succinimidyl ester of coumarin-3-carboxylic acid, which generates the fluorescent derivative 7-OH-SECCA following its interaction with radiation-induced . OH in aqueous solution. The fluorescence induced per unit γ-ray dose reflecting the accessibility of . OH to such SECCA-conjugated biomolecules was recorded. The biomolecules were also labelled with the fluorescent derivative 7-OH-SECCA in trace amounts to study their conformation under identical conditions via fluorescence polarization spectroscopy. (author)

Sample preparation of waste water to determine metallic contaminants by X-ray fluorescence spectroscopy

International Nuclear Information System (INIS)

Gonzalez Olivos, Javier.

1987-01-01

Trace X-ray fluorescence spectroscopy analysis in liquid samples is preceded by sample preparation, which usually consists in the precipitation of the metallic ions and concentration over a thin cellulose filter. The samples preparation of waste water by this method is not efficient, due to the great amount of organic and insoluble matter that they contain. The purpose of this work was to determine the optimal value of pH in order to adsorbe all the insoluble matter contained in a waste water sample in the activated charcoal, so that the metallic ions could be precipitated and concentrated on a thin filter and determinated by X-ray fluorescence spectroscopy. A survey about the adsorption of some ions in activated charcoal in function of the pH was made for the following: Cr 3+ , Fe 3+ , Ni 2+ , Cu 2+ , Zn 2+ , Se 2+ , Hg 2+ , and Pb 2+ . It was observed that at pH 0, the ions are not adsorbed, but Cu 2+ and Zn 2+ are adsorbed in small amount; at pH 14, the ions are adsorbed, excluding Se, which is not adsorbed at any value of pH. If a waste water sample is treated at pH 0 with activated charcoal to adsorbe the organic and insoluble matter, most of the metallic ions are not adsorbed by the activated charcoal and could be precipitated with APDC (ammonium 1-pirrolidine dithio carbamate salt) and concentrated on a thin filter. The analysis of the metallic ions contained on the filter and those adsorbed in the activated charcoal by X-ray fluorescence spectroscopy, gave the total amount of the ions in the sample. (author)

Synergy Effect of Combining Fluorescence and Mid Infrared Fiber Spectroscopy for Kidney Tumor Diagnostics

Directory of Open Access Journals (Sweden)

Andrey Bogomolov

2017-11-01

Full Text Available Matching pairs of tumor and non-tumor kidney tissue samples of four patients were investigated ex vivo using a combination of two methods, attenuated total reflection mid infrared spectroscopy and fluorescence spectroscopy, through respectively prepared and adjusted fiber probes. In order to increase the data information content, the measurements on tissue samples in both methods were performed in the same 31 preselected positions. Multivariate data analysis revealed a synergic effect of combining the two methods for the diagnostics of kidney tumor compared to individual techniques.

Dynamic fluorescence spectroscopy on single tryptophan mutants of EIImtl in detergent micelles : Effects of substrate binding and phosphorylation on the fluorescence and anisotropy decay

NARCIS (Netherlands)

Swaving Dijkstra, Dolf; Broos, J.; Visser, Antonie J.W.G.; van Hoek, A.; Robillard, George

1997-01-01

The effects of substrate and substrate analogue binding and phosphorylation on the conformational dynamics of the mannitol permease of Escherichia coli were investigated, using time-resolved fluorescence spectroscopy on mutants containing five single tryptophans situated in the membrane-embedded C

Spectroscopy, Manipulation and Trapping of Neutral Atoms, Molecules, and Other Particles Using Optical Nanofibers: A Review

Science.gov (United States)

Morrissey, Michael J.; Deasy, Kieran; Frawley, Mary; Kumar, Ravi; Prel, Eugen; Russell, Laura; Truong, Viet Giang; Chormaic, Síle Nic

2013-01-01

The use of tapered optical fibers, i.e., optical nanofibers, for spectroscopy and the detection of small numbers of particles, such as neutral atoms or molecules, has been gaining interest in recent years. In this review, we briefly introduce the optical nanofiber, its fabrication, and optical mode propagation within. We discuss recent progress on the integration of optical nanofibers into laser-cooled atom and vapor systems, paying particular attention to spectroscopy, cold atom cloud characterization, and optical trapping schemes. Next, a natural extension of this work to molecules is introduced. Finally, we consider several alternatives to optical nanofibers that display some advantages for specific applications. PMID:23945738

Massively Parallel Single-Molecule Manipulation Using Centrifugal Force

Science.gov (United States)

Wong, Wesley; Halvorsen, Ken

2011-03-01

Precise manipulation of single molecules has led to remarkable insights in physics, chemistry, biology, and medicine. However, two issues that have impeded the widespread adoption of these techniques are equipment cost and the laborious nature of making measurements one molecule at a time. To meet these challenges, we have developed an approach that enables massively parallel single- molecule force measurements using centrifugal force. This approach is realized in the centrifuge force microscope, an instrument in which objects in an orbiting sample are subjected to a calibration-free, macroscopically uniform force- field while their micro-to-nanoscopic motions are observed. We demonstrate high- throughput single-molecule force spectroscopy with this technique by performing thousands of rupture experiments in parallel, characterizing force-dependent unbinding kinetics of an antibody-antigen pair in minutes rather than days. Currently, we are taking steps to integrate high-resolution detection, fluorescence, temperature control and a greater dynamic range in force. With significant benefits in efficiency, cost, simplicity, and versatility, single-molecule centrifugation has the potential to expand single-molecule experimentation to a wider range of researchers and experimental systems.

High-Resolution Spectroscopy of Laser Ablation Plumes Using Laser-Induced Fluorescence

Energy Technology Data Exchange (ETDEWEB)

Harilal, Sivanandan S.; LaHaye, Nicole L.; Phillips, Mark C.

2017-02-06

We used a CW laser as a narrow-band (~50kHz) tunable LIF excitation source to probe absorption from selected atomic transitions (Al, U etc. ) in a ns laser ablation plume. A comparison of fluorescence signal with respect to emission spectroscopy show significant increase in the magnitude and persistence from selected Al and U transitions in a LIBS plume. The high spectral resolution provided by the LIF measurement allows peaks to be easily separated even if they overlap in the emission spectra.

Thermally activated delayed fluorescence organic dots for two-photon fluorescence lifetime imaging

Science.gov (United States)

He, Tingchao; Ren, Can; Li, Zhuohua; Xiao, Shuyu; Li, Junzi; Lin, Xiaodong; Ye, Chuanxiang; Zhang, Junmin; Guo, Lihong; Hu, Wenbo; Chen, Rui

2018-05-01

Autofluorescence is a major challenge in complex tissue imaging when molecules present in the biological tissue compete with the fluorophore. This issue may be resolved by designing organic molecules with long fluorescence lifetimes. The present work reports the two-photon absorption (TPA) properties of a thermally activated delayed fluorescence (TADF) molecule with carbazole as the electron donor and dicyanobenzene as the electron acceptor (i.e., 4CzIPN). The results indicate that 4CzIPN exhibits a moderate TPA cross-section (˜9 × 10-50 cm4 s photon-1), high fluorescence quantum yield, and a long fluorescence lifetime (˜1.47 μs). 4CzIPN was compactly encapsulated into an amphiphilic copolymer via nanoprecipitation to achieve water-soluble organic dots. Interestingly, 4CzIPN organic dots have been utilized in applications involving two-photon fluorescence lifetime imaging (FLIM). Our work aptly demonstrates that TADF molecules are promising candidates of nonlinear optical probes for developing next-generation multiphoton FLIM applications.

Serum albumin binding sites properties in donors and in schizophrenia patients: the study of fluorescence decay of the probe K-35 using S-60 synchrotron pulse excitation

Energy Technology Data Exchange (ETDEWEB)

Gryzunov, Y.A. E-mail: grysunov@sci.lebedev.ru; Syrejshchikova, T.I.; Komarova, M.N.; Misionzhnik, E.Yu.; Uzbekov, M.G.; Molodetskich, A.V.; Dobretsov, G.E.; Yakimenko, M.N

2000-06-21

The properties of serum albumin obtained from donors and from paranoid schizophrenia patients were studied with the fluorescent probe K-35 (N-carboxyphenylimide of dimethylaminonaphthalic acid) and time-resolved fluorescence spectroscopy on the SR beam station of the S-60 synchrotron of the Lebedev Physical Institute. The mean fluorescence quantum yield of K-35 in patients serum was decreased significantly by 25-60% comparing with donors. The analysis of pre-exponential factors of fluorescence decay using 'amplitude standard' method has shown that in patient sera the fraction of K-35 molecules bound with albumin and inaccessible to fluorescence quenchers ('bright' K-35 molecules with {tau}{sub 1}=8.0{+-}0.4 ns) is 1.2-3 times less than in the donor sera. The fraction of K-35 molecules with partly quenched fluorescence ({tau}{sub 2}=1.44{+-}0.22 ns) was significantly increased in schizophrenia patients. The results obtained suggest that the properties of binding region in serum albumin molecules of acute paranoid schizophrenia patients change significantly.

Serum albumin binding sites properties in donors and in schizophrenia patients: the study of fluorescence decay of the probe K-35 using S-60 synchrotron pulse excitation

International Nuclear Information System (INIS)

Gryzunov, Y.A.; Syrejshchikova, T.I.; Komarova, M.N.; Misionzhnik, E.Yu.; Uzbekov, M.G.; Molodetskich, A.V.; Dobretsov, G.E.; Yakimenko, M.N.

2000-01-01

The properties of serum albumin obtained from donors and from paranoid schizophrenia patients were studied with the fluorescent probe K-35 (N-carboxyphenylimide of dimethylaminonaphthalic acid) and time-resolved fluorescence spectroscopy on the SR beam station of the S-60 synchrotron of the Lebedev Physical Institute. The mean fluorescence quantum yield of K-35 in patients serum was decreased significantly by 25-60% comparing with donors. The analysis of pre-exponential factors of fluorescence decay using 'amplitude standard' method has shown that in patient sera the fraction of K-35 molecules bound with albumin and inaccessible to fluorescence quenchers ('bright' K-35 molecules with τ 1 =8.0±0.4 ns) is 1.2-3 times less than in the donor sera. The fraction of K-35 molecules with partly quenched fluorescence (τ 2 =1.44±0.22 ns) was significantly increased in schizophrenia patients. The results obtained suggest that the properties of binding region in serum albumin molecules of acute paranoid schizophrenia patients change significantly

Serum albumin binding sites properties in donors and in schizophrenia patients: the study of fluorescence decay of the probe K-35 using S-60 synchrotron pulse excitation

Science.gov (United States)

Gryzunov, Yu. A.; Syrejshchikova, T. I.; Komarova, M. N.; Misionzhnik, E. Yu; Uzbekov, M. G.; Molodetskich, A. V.; Dobretsov, G. E.; Yakimenko, M. N.

2000-06-01

The properties of serum albumin obtained from donors and from paranoid schizophrenia patients were studied with the fluorescent probe K-35 (N-carboxyphenylimide of dimethylaminonaphthalic acid) and time-resolved fluorescence spectroscopy on the SR beam station of the S-60 synchrotron of the Lebedev Physical Institute. The mean fluorescence quantum yield of K-35 in patients serum was decreased significantly by 25-60% comparing with donors. The analysis of pre-exponential factors of fluorescence decay using "amplitude standard" method has shown that in patient sera the fraction of K-35 molecules bound with albumin and inaccessible to fluorescence quenchers ("bright" K-35 molecules with τ1=8.0±0.4 ns) is 1.2-3 times less than in the donor sera. The fraction of K-35 molecules with partly quenched fluorescence ( τ2=1.44±0.22 ns) was significantly increased in schizophrenia patients. The results obtained suggest that the properties of binding region in serum albumin molecules of acute paranoid schizophrenia patients change significantly.

Using a Spectrofluorometer for Resonance Raman Spectra of Organic Molecules

Directory of Open Access Journals (Sweden)

Vadivel Masilamani

2017-01-01

Full Text Available Scattering (Rayleigh and Raman and fluorescence are two common light signals that frequently occur together, confusing the researchers and graduate students experimenting in molecular spectroscopy laboratories. This report is a brief study presenting a clear discrimination between the two signals mentioned, employing a common spectrofluorometer such as the PerkinElmer LS 55. Even better, the resonance Raman signal of a molecule (e.g., acetone can be obtained elegantly using the same instrument.

Interaction of VUV-photons with molecules. Spectroscopy and dynamics of molecular superexcited states

International Nuclear Information System (INIS)

Hatano, Y.

2002-01-01

Complete text of publication follows. A survey is given of recent progress in experimental studies of the interaction of VUV-photons with molecules, i.e., those of photoabsorption, photoionization, and photodissociation of molecules in the excitation photon energy range of 10-50 eV, with a particular emphasis placed on current understanding of the spectroscopy and dynamics of formed molecular superexcited states. These studies are of great importance in understanding the interaction of ionizing radiation with matter. Molecules studied are ranged from simple diatomic and triatomic molecules to polyatomic molecules such as hydrocarbons. Most of the observed molecular superexcited states are assigned to high Rydber states which are vibrationally, doubly, or inner-core excited and converge to each of ion states. Non-Rydberg superexcited states are also observed. Dissociation into neutral fragments in comparison with ionization is of unexpectedly great importance in the observed decay of each of these state-assigned superexcited molecules. Dissociation dynamics as well as its products of superexcited states are remarkably different from those of lower excited states below about ionization thresholds. Some remarks are also presented of molecules in the condensed phase

The role of ABC proteins Aus1p and Pdr11p in the uptake of external sterols in yeast: dehydroergosterol fluorescence study

DEFF Research Database (Denmark)

Kohut, Peter; Wüstner, Daniel; Hronska, L

2011-01-01

of sterol molecules into plasma membrane is not spontaneous but requires assistance of two ABC (ATP-binding cassette) pumps--Aus1p or Pdr11p. DHE taken up by uptake-competent hem1ΔAUS1PDR11 cells could be directly visualized by UV-sensitive wide field fluorescence microscopy. HPLC analysis of sterols......Uptake of external sterols in the yeast Saccharomyces cerevisiae is a multistep process limited to anaerobiosis or heme deficiency. It includes crossing the cell wall, insertion of sterol molecules into plasma membrane and their internalization and integration into intracellular membranes. We...... applied the fluorescent ergosterol analog dehydroergosterol (DHE) to monitor the initial steps of sterol uptake by three independent approaches: fluorescence spectroscopy, fluorescence microscopy and sterol quantification by HPLC. Using specific fluorescence characteristics of DHE we showed that the entry...

Reverse engineering of an affinity-switchable molecular interaction characterized by atomic force microscopy single-molecule force spectroscopy.

Science.gov (United States)

Anselmetti, Dario; Bartels, Frank Wilco; Becker, Anke; Decker, Björn; Eckel, Rainer; McIntosh, Matthew; Mattay, Jochen; Plattner, Patrik; Ros, Robert; Schäfer, Christian; Sewald, Norbert

2008-02-19

Tunable and switchable interaction between molecules is a key for regulation and control of cellular processes. The translation of the underlying physicochemical principles to synthetic and switchable functional entities and molecules that can mimic the corresponding molecular functions is called reverse molecular engineering. We quantitatively investigated autoinducer-regulated DNA-protein interaction in bacterial gene regulation processes with single atomic force microscopy (AFM) molecule force spectroscopy in vitro, and developed an artificial bistable molecular host-guest system that can be controlled and regulated by external signals (UV light exposure and thermal energy). The intermolecular binding functionality (affinity) and its reproducible and reversible switching has been proven by AFM force spectroscopy at the single-molecule level. This affinity-tunable optomechanical switch will allow novel applications with respect to molecular manipulation, nanoscale rewritable molecular memories, and/or artificial ion channels, which will serve for the controlled transport and release of ions and neutral compounds in the future.

Combination of Small Molecule Microarray and Confocal Microscopy Techniques for Live Cell Staining Fluorescent Dye Discovery

Directory of Open Access Journals (Sweden)

Attila Bokros

2013-08-01

Full Text Available Discovering new fluorochromes is significantly advanced by high-throughput screening (HTS methods. In the present study a combination of small molecule microarray (SMM prescreening and confocal laser scanning microscopy (CLSM was developed in order to discover novel cell staining fluorescent dyes. Compounds with high native fluorescence were selected from a 14,585-member library and further tested on living cells under the microscope. Eleven compartment-specific, cell-permeable (or plasma membrane-targeted fluorochromes were identified. Their cytotoxicity was tested and found that between 1–10 micromolar range, they were non-toxic even during long-term incubations.

Investigation into interaction of CO/sub 2/ molecules with zeolites by infrared spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Ignat' eva, L A; Levshin, L V; Chukin, G D; Efimenko, L V; Kozlova, T I [Moskovskij Gosudarstvennyj Univ. (USSR). Kafedra Optiki

1975-07-01

Interaction of CO/sub 2/ molecules with zeolites, particularly with SrNaJ was studied by infrared-spectroscopy. To obtain infrared-spectra the zeolites were pressed into tablets and were calcinated at 500 deg. In the spectra the bands of chemisorbed CO/sub 2/ absorption were found in the range 1300 - 1600 cm/sup -1/. The CO/sub 2/ molecule was found to be strongly deformed due to chemisorption. In terms of electronic structure of the zeolite crystalline skeleton several types of CO/sub 2/ molecules interaction with different active zeolites were found. The position of the high-frequency band of CO/sub 2/ absorption in zeolites spectra was found to be a linear function of electrostatic field of the cations.

X-ray fluorescence/Auger-electron coincidence spectroscopy of vacancy cascades in atomic argon

International Nuclear Information System (INIS)

Arp, U.

1996-01-01

Argon L 2.3 -M 2.3 M 2.3 Auger-electron spectra were measured in coincidence with Kα fluorescent x-rays in studies of Ar K-shell vacancy decays at several photon energies above the K-threshold and on the 1s-4p resonance in atomic argon. The complex spectra recorded by conventional electron spectroscopy are greatly simplified when recorded in coincidence with fluorescent x-rays, allowing a more detailed analysis of the vacancy cascade process. The resulting coincidence spectra are compared with Hartree-Fock calculations which include shake-up transitions in the resonant case. Small energy shifts of the coincidence electron spectra are attributed to post-collision interaction with 1s photoelectrons

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.

Analysis of fluorescence quenching of pyronin B and pyronin Y by molecular oxygen in aqueous solution

Energy Technology Data Exchange (ETDEWEB)

Celebi, Neslihan [Faculty of Arts and Sciences, Department of Chemistry, Atatuerk University, 25240 Erzurum (Turkey); Arik, Mustafa [Faculty of Arts and Sciences, Department of Chemistry, Atatuerk University, 25240 Erzurum (Turkey); Onganer, Yavuz [Faculty of Arts and Sciences, Department of Chemistry, Atatuerk University, 25240 Erzurum (Turkey)]. E-mail: yonganer@atauni.edu.tr

2007-09-15

The fluorescence quenching of pyronin B and pyronin Y molecules by molecular oxygen in aqueous solution was studied by using steady-state and time-resolved fluorescence and UV-Vis absorption spectroscopy techniques. In order to understand the quenching mechanism, fluorescence decays, absorption and fluorescence spectra of the probes were recorded as a function of the oxygen concentration and temperature. The quenching was found to be appreciable and shows positive deviation in the Stern-Volmer representation obtained from the fluorescence intensity ratio. Fluorescence quenching constants (k {sub q}) were calculated from the {tau} {sub o}/{tau} vs. [Q] plots having linear correlation and compared with calculated diffusion-controlled rate constants (k {sub diff}) values. Experimental results were in good agreement with the simultaneous dynamic and static quenching model.

Analysis of fluorescence quenching of pyronin B and pyronin Y by molecular oxygen in aqueous solution

International Nuclear Information System (INIS)

Celebi, Neslihan; Arik, Mustafa; Onganer, Yavuz

2007-01-01

The fluorescence quenching of pyronin B and pyronin Y molecules by molecular oxygen in aqueous solution was studied by using steady-state and time-resolved fluorescence and UV-Vis absorption spectroscopy techniques. In order to understand the quenching mechanism, fluorescence decays, absorption and fluorescence spectra of the probes were recorded as a function of the oxygen concentration and temperature. The quenching was found to be appreciable and shows positive deviation in the Stern-Volmer representation obtained from the fluorescence intensity ratio. Fluorescence quenching constants (k q ) were calculated from the τ o /τ vs. [Q] plots having linear correlation and compared with calculated diffusion-controlled rate constants (k diff ) values. Experimental results were in good agreement with the simultaneous dynamic and static quenching model

Single molecule force spectroscopy at high data acquisition: A Bayesian nonparametric analysis

Science.gov (United States)

Sgouralis, Ioannis; Whitmore, Miles; Lapidus, Lisa; Comstock, Matthew J.; Pressé, Steve

2018-03-01

Bayesian nonparametrics (BNPs) are poised to have a deep impact in the analysis of single molecule data as they provide posterior probabilities over entire models consistent with the supplied data, not just model parameters of one preferred model. Thus they provide an elegant and rigorous solution to the difficult problem encountered when selecting an appropriate candidate model. Nevertheless, BNPs' flexibility to learn models and their associated parameters from experimental data is a double-edged sword. Most importantly, BNPs are prone to increasing the complexity of the estimated models due to artifactual features present in time traces. Thus, because of experimental challenges unique to single molecule methods, naive application of available BNP tools is not possible. Here we consider traces with time correlations and, as a specific example, we deal with force spectroscopy traces collected at high acquisition rates. While high acquisition rates are required in order to capture dwells in short-lived molecular states, in this setup, a slow response of the optical trap instrumentation (i.e., trapped beads, ambient fluid, and tethering handles) distorts the molecular signals introducing time correlations into the data that may be misinterpreted as true states by naive BNPs. Our adaptation of BNP tools explicitly takes into consideration these response dynamics, in addition to drift and noise, and makes unsupervised time series analysis of correlated single molecule force spectroscopy measurements possible, even at acquisition rates similar to or below the trap's response times.

Surface Passivation for Single-molecule Protein Studies

Science.gov (United States)

Chandradoss, Stanley D.; Haagsma, Anna C.; Lee, Young Kwang; Hwang, Jae-Ho; Nam, Jwa-Min; Joo, Chirlmin

2014-01-01

Single-molecule fluorescence spectroscopy has proven to be instrumental in understanding a wide range of biological phenomena at the nanoscale. Important examples of what this technique can yield to biological sciences are the mechanistic insights on protein-protein and protein-nucleic acid interactions. When interactions of proteins are probed at the single-molecule level, the proteins or their substrates are often immobilized on a glass surface, which allows for a long-term observation. This immobilization scheme may introduce unwanted surface artifacts. Therefore, it is essential to passivate the glass surface to make it inert. Surface coating using polyethylene glycol (PEG) stands out for its high performance in preventing proteins from non-specifically interacting with a glass surface. However, the polymer coating procedure is difficult, due to the complication arising from a series of surface treatments and the stringent requirement that a surface needs to be free of any fluorescent molecules at the end of the procedure. Here, we provide a robust protocol with step-by-step instructions. It covers surface cleaning including piranha etching, surface functionalization with amine groups, and finally PEG coating. To obtain a high density of a PEG layer, we introduce a new strategy of treating the surface with PEG molecules over two rounds, which remarkably improves the quality of passivation. We provide representative results as well as practical advice for each critical step so that anyone can achieve the high quality surface passivation. PMID:24797261

Assessment of the Inhibitory Effect of Rifampicin on Amyloid Formation of Hen Egg White Lysozyme: Thioflavin T Fluorescence Assay versus FTIR Difference Spectroscopy

Directory of Open Access Journals (Sweden)

Gang Ma

2014-01-01

Full Text Available The inhibitory effect of rifampicin on the amyloid formation of hen egg white lysozyme was assessed with both Thioflavin T (ThT fluorescence assay and Fourier transform infrared (FTIR difference spectroscopy. We reveal that ThT fluorescence assay gives a false positive result due to rifampicin interference, while FTIR difference spectroscopy provides a reliable assessment. With FTIR, we show that rifampicin only has marginally inhibitory effect. We then propose that FTIR difference spectroscopy can potentially be a convenient method for inhibitor screening in amyloid study.

Conformation and dynamics of nucleotides in bulges and symmetric internal loops in duplex DNA studied by EPR and fluorescence spectroscopies

International Nuclear Information System (INIS)

Cekan, Pavol; Sigurdsson, Snorri Th.

2012-01-01

Highlights: ► Bulges and loops were studied by both EPR and fluorescence spectroscopies using the probe Ç/Ç f . ► One-base bulge was in a temperature-dependent equilibrium between looped-out and stacked states. ► Bases in two- and three-base bulges were stacked at all temperatures, resulting in DNA bending. ► Bases were stacked in symmetrical two- to five-base internal loops, according to EPR data. ► Unexpectedly high fluorescence for the smaller loops indicated local structural perturbations. -- Abstract: The dynamics and conformation of base bulges and internal loops in duplex DNA were studied using the bifunctional spectroscopic probe Ç, which becomes fluorescent (Ç f ) upon reduction of the nitroxide functional group, along with EPR and fluorescence spectroscopies. A one-base bulge was in a conformational equilibrium between looped-out and stacked states, the former favored at higher temperature and the latter at lower temperature. Stacking of bulge bases was favored in two- and three-base bulges, independent of temperature, resulting in DNA bending as evidenced by increased fluorescence of Ç f . EPR spectra of Ç-labeled three-, four- and five-base symmetrical interior DNA bulges at 20 °C showed low mobility, indicating that the spin-label was stacked within the loop. The spin-label mobility at 37 °C increased as the loops became larger. A considerable variation in fluorescence between different loops was observed, as well as a temperature-dependence within constructs. Fluorescence unexpectedly increased as the size of the loop decreased at 2 °C. Fluorescence of the smallest loops, where a single T·T mismatch was located between the stem region and the probe, was even larger than for the single strand, indicating a considerable local structural deformation of these loops from regular B-DNA. These results show the value of combining EPR and fluorescence spectroscopy to study non-helical regions of nucleic acids.

Macromolecule biosynthesis assay and fluorescence spectroscopy methods to explore antimicrobial peptide mode(s) of action

DEFF Research Database (Denmark)

Jana, Bimal; Baker, Kristin Renee; Guardabassi, Luca

2017-01-01

the biosynthesis rate of macromolecules (e.g., DNA, RNA, protein, and cell wall) and the cytoplasmic membrane proton motive force (PMF) energy can help to unravel the diverse modes of action of AMPs. Here, we present an overview of macromolecule biosynthesis rate measurement and fluorescence spectroscopy methods...

Spatially resolved x-ray fluorescence spectroscopy of beryllium capsule implosions at the NIF

Science.gov (United States)

MacDonald, M. J.; Bishel, D. T.; Saunders, A. M.; Scott, H. A.; Kyrala, G.; Kline, J.; MacLaren, S.; Thorn, D. B.; Yi, S. A.; Zylstra, A. B.; Falcone, R. W.; Doeppner, T.

2017-10-01

Beryllium ablators used in indirectly driven inertial confinement fusion implosions are doped with copper to prevent preheat of the cryogenic hydrogen fuel. Here, we present analysis of spatially resolved copper K- α fluorescence spectra from the beryllium ablator layer. It has been shown that K- α fluorescence spectroscopy can be used to measure plasma conditions of partially ionized dopants in high energy density systems. In these experiments, K-shell vacancies in the copper dopant are created by the hotspot emission at stagnation, resulting in K-shell fluorescence at bang time. Spatially resolved copper K- α emission spectra are compared to atomic kinetics and radiation code simulations to infer density and temperature profiles. This work was supported by the US DOE under Grant No. DE-NA0001859, under the auspices of the US DOE by Lawrence Livermore National Laboratory under Contract No. DE-AC52-07NA27344, and by Los Alamos National Laboratory under contract DE-AC52-06NA52396.

Ultraviolet-Visible and Fluorescence Spectroscopy Techniques Are Important Diagnostic Tools during the Progression of Atherosclerosis: Diet Zinc Supplementation Retarded or Delayed Atherosclerosis

Science.gov (United States)

Abdelhalim, Mohamed Anwar K.; Moussa, Sherif A. Abdelmottaleb; AL-Mohy, Yanallah Hussain

2013-01-01

Background. In this study, we examined whether UV-visible and fluorescence spectroscopy techniques detect the progression of atherosclerosis in serum of rabbits fed on high-cholesterol diet (HCD) and HCD supplemented with zinc (HCD + Zn) compared with the control. Methods. The control rabbits group was fed on 100 g/day of normal diet. The HCD group was fed on Purina Certified Rabbit Chow supplemented with 1.0% cholesterol plus 1.0% olive oil (100 g/day) for the same period. The HCD + Zn group was fed on normal Purina Certified Rabbit Chow plus 1.0% cholesterol and 1.0% olive oil supplemented with 470 ppm Zn for the same feeding period. UV-visible and fluorescence spectroscopy and biochemistry in Rabbit's blood serum and blood hematology were measured in Rabbit's blood. Results. We found that the fluorescent peak of HCD shifted toward UV-visible wavelength compared with the control using fluorescent excitation of serum at 192 nm. In addition, they showed that supplementation of zinc (350 ppm) restored the fluorescent peak closely to the control. By using UV-visible spectroscopy approach, we found that the peak absorbance of HCD (about 280 nm) was higher than that of control and that zinc supplementation seemed to decrease the absorbance. Conclusions. This study demonstrates that ultraviolet-visible and fluorescence spectroscopy techniques can be applied as noninvasive techniques on a sample blood serum for diagnosing or detecting the progression of atherosclerosis. The Zn supplementation to rabbits fed on HCD delays or retards the progression of atherosclerosis. Inducing anemia in rabbits fed on HCD delays the progression of atherosclerosis. PMID:24350281

Characterization of dissolved organic matter in fogwater by excitation-emission matrix fluorescence spectroscopy

Science.gov (United States)

Birdwell, J.E.; Valsaraj, K.T.

2010-01-01

Dissolved organic matter (DOM) present in fogwater samples collected in southeastern Louisiana and central-eastern China has been characterized using excitation-emission matrix fluorescence spectroscopy. The goal of the study was to illustrate the utility of fluorescence for obtaining information on the large fraction of organic carbon in fogwaters (typically >40% by weight) that defies characterization in terms of specific chemical compounds without the difficulty inherent in obtaining sufficient fogwater volume to isolate DOM for assessment using other spectroscopic and chemical analyses. Based on the findings of previous studies using other characterization methods, it was anticipated that the unidentified organic carbon fraction would have characteristic peaks associated with humic substances and fluorescent amino acids. Both humic- and protein-like fluorophores were observed in the fogwater spectra and fluorescence-derived indices for the fogwater had similar values to those of soil and sediment porewater. Greater biological character was observed in samples with higher organic carbon concentrations. Fogwaters are shown to contain a mixture of terrestrially- and microbially-derived fluorescent organic material, which is expected to be derived from an array of different sources, such as suspended soil and dust particles, biogenic emissions and organic substances generated by atmospheric processes. The fluorescence results indicate that much of the unidentified organic carbon present in fogwater can be represented by humic-like and biologically-derived substances similar to those present in other aquatic systems, though it should be noted that fluorescent signatures representative of DOM produced by atmospheric processing of organic aerosols may be contributing to or masked by humic-like fluorophores. ?? 2010.

Femtosecond laser irradiation of the fluorescent molecules-loaded poly(lactic-co-glycolic acid)

Science.gov (United States)

Umemoto, Taiga; Shibata, Akimichi; Terakawa, Mitsuhiro

2017-09-01

Molecular release from scaffolds is desired for tailoring cell-compatible tissue engineering. Several methods have been proposed to control molecular release, such as annealing, plasma treatment, and laser processing. In this study, we describe the alteration of Rhodamine B (RhB)-loaded poly(lactic-co-glycolic acid) (PLGA) after femtosecond laser irradiation, which was evaluated on the basis of the water absorption and mass remaining. Fluorescence measurement of released RhB molecules revealed the acceleration of the molecular release upon 400-nm laser irradiation, whereas 800-nm laser irradiation did not induce a comparable degree of change compared with non-irradiated samples. The result of the water absorption measurement indicates that the large amount of water absorption of 400-nm laser-irradiated PLGA sample may accelerate the diffusion of the loaded molecules through absorbing water, which resulted in the faster molecular release.

Other origins for the fluorescence modulation of single dye molecules in open-circuit and short-circuit devices.

Science.gov (United States)

Teguh, Jefri S; Kurniawan, Michael; Wu, Xiangyang; Sum, Tze Chien; Yeow, Edwin K L

2013-01-07

Fluorescence intensity modulation of single Atto647N dye molecules in a short-circuit device and a defective device, caused by damaging an open-circuit device, is due to a variation in the excitation light focus as a result of the formation of an alternating electric current.

Detection of ethanol in alcoholic beverages or vapor phase using fluorescent molecules embedded in a nanofibrous polymer.

Science.gov (United States)

Akamatsu, Masaaki; Mori, Taizo; Okamoto, Ken; Komatsu, Hirokazu; Kumagai, Ken; Shiratori, Seimei; Yamamura, Masaki; Nabeshima, Tatsuya; Sakai, Hideki; Abe, Masahiko; Hill, Jonathan P; Ariga, Katsuhiko

2015-03-25

An alcohol sensor was developed using the solid-state fluorescence emission of terphenyl-ol (TPhOH) derivatives. Admixtures of TPhOH and sodium carbonate exhibited bright sky-blue fluorescence in the solid state upon addition of small quantities of ethanol. A series of terphenol derivatives was synthesized, and the effects of solvent polarities and the structures of these π-conjugated systems on their fluorescence were systematically investigated by using fluorescence spectroscopy. In particular, π-extended TPhOHs and TPhOHs containing electron-withdrawing groups exhibited significant solvatochromism, and fluorescence colors varied from blue to red. Detection of ethanol contents in alcohol beverages (detection limit ∼ 5 v/v %) was demonstrated using different TPhOHs revealing the effect of molecular structure on sensing properties. Ethanol contents in alcoholic beverages could be estimated from the intensity of the fluorescence elicited from the TPhOHs. Moreover, when terphenol and Na2CO3 were combined with a water-absorbent polymer, ethanol could be detected at lower concentrations. Detection of ethanol vapor (8 v/v % in air) was also accomplished using a nanofibrous polymer scaffold as the immobilized sensing film.

Determination of the botanical origin of honey by front-face synchronous fluorescence spectroscopy.

Science.gov (United States)

Lenhardt, Lea; Zeković, Ivana; Dramićanin, Tatjana; Dramićanin, Miroslav D; Bro, Rasmus

2014-01-01

Front-face synchronous fluorescence spectroscopy combined with chemometrics is used to classify honey samples according to their botanical origin. Synchronous fluorescence spectra of three monofloral (linden, sunflower, and acacia), polyfloral (meadow mix), and fake (fake acacia and linden) honey types (109 samples) were collected in an excitation range of 240-500 nm for synchronous wavelength intervals of 30-300 nm. Chemometric analysis of the gathered data included principal component analysis and partial least squares discriminant analysis. Mean cross-validated classification errors of 0.2 and 4.8% were found for a model that accounts only for monofloral samples and for a model that includes both the monofloral and polyfloral groups, respectively. The results demonstrate that single synchronous fluorescence spectra of different honeys differ significantly because of their distinct physical and chemical characteristics and provide sufficient data for the clear differentiation among honey groups. The spectra of fake honey samples showed pronounced differences from those of genuine honey, and these samples are easily recognized on the basis of their synchronous fluorescence spectra. The study demonstrated that this method is a valuable and promising technique for honey authentication.

Putting prions into focus: application of single molecule detection to the diagnosis of prion diseases.

Science.gov (United States)

Giese, A; Bieschke, J; Eigen, M; Kretzschmar, H A

2000-01-01

Prion diseases are characterized by the cerebral deposition of an aggregated pathological isoform of the prion protein (PrP(Sc)) which constitutes the principal component of the transmissible agent termed prion. In order to develop a highly sensitive method for the detection of PrP(Sc) aggregates in biological samples such as cerebrospinal fluid (CSF), we used a method based on Fluorescence Correlation Spectroscopy (FCS), a technique which allows detection of single fluorescently labeled molecules in solution. Within the FCS setup, fluorescent photons emitted by molecules passing an open volume element defined by the beam of an excitation laser focussed into a diffraction-limited spot are imaged confocally onto a single photon counting detector. Aggregates of PrP(Sc) could be labeled by co-aggregation of probe molecules such as monomeric recombinant PrP or PrP-specific antibodies tagged with a fluorescent dye. In addition to slow diffusion, labeled aggregates are characterized by high fluorescence intensity, which allows detection and quantification by analysis of fluorescence intensity distribution. To improve detection of rare target particles, the accessible volume element was increased by scanning for intensely fluorescent targets (SIFT). To further improve sensitivity and specificity, two different probes were used simultaneously in a two-color setup. In a diagnostic model system of CSF spiked with purified prion rods, dual-color SIFT was more sensitive than Western blot analysis. In addition, a PrP(Sc)-specific signal was also detected in a number of CSF samples derived from CJD patients but not in controls.

Analysis of the substrate recognition state of TDP-43 to single-stranded DNA using fluorescence correlation spectroscopy

Directory of Open Access Journals (Sweden)

Akira Kitamura

2018-07-01

Full Text Available Normal function and abnormal aggregation of transactivation response (TAR DNA/RNA-binding protein 43 kDa (TDP-43 are directly associated with the lethal genetic diseases: cystic fibrosis, amyotrophic lateral sclerosis (ALS, and frontotemporal lobar degeneration (FTLD. The binding of TDP-43 to single-stranded DNA (ssDNA or RNA is involved in transcriptional repression, regulation of RNA splicing, and RNA stabilization. Equilibrium dissociation constants (Kd of TDP-43 and ssDNA or RNA have been determined using various methods; however, methods that can measure Kd with high sensitivity in a short time using a small amount of TDP-43 in solution would be advantageous. Here, in order to determine the Kd of TDP-43 and fluorescence-labeled ssDNA as well as the binding stoichiometry, we use fluorescence correlation spectroscopy (FCS, which detects the slowed diffusion of molecular interactions in solution with single-molecule sensitivity, in addition to electrophoretic mobility shift assay (EMSA. Using tandem affinity chromatography of TDP-43 dually tagged with glutathione-S-transferase and poly-histidine tags, highly purified protein was obtained. FCS successfully detected specific interaction between purified TDP-43 and TG ssDNA repeats, with a Kd in the nanomolar range. The Kd of the TDP-43 mutant was not different from the wild type, although mutant oligomers, which did not bind ssDNA, were observed. Analysis of the fluorescence brightness per dimerized TDP-43/ssDNA complex was used to evaluate their binding stoichiometry. The results suggest that an assay combining FCS and EMSA can precisely analyze ssDNA recognition mechanisms, and that FCS may be applied for the rapid and quantitative determination of the interaction strength between TDP-43 and ssDNA or RNA. These methods will aid in the elucidation of the substrate recognition mechanism of ALS- and FTLD-associated variants of TDP-43.

A versatile optical microscope for time-dependent single-molecule and single-particle spectroscopy

Science.gov (United States)

Li, Hao; Yang, Haw

2018-03-01

This work reports the design and implementation of a multi-function optical microscope for time-dependent spectroscopy on single molecules and single nanoparticles. It integrates the now-routine single-object measurements into one standalone platform so that no reconfiguration is needed when switching between different types of sample or spectroscopy modes. The illumination modes include evanescent field through total internal reflection, dark-field illumination, and epi-excitation onto a diffraction-limited spot suitable for confocal detection. The detection modes include spectrally resolved line imaging, wide-field imaging with dual-color capability, and two-color single-element photon-counting detection. The switch between different spectroscopy and data acquisition modes is fully automated and executed through computer programming. The capability of this microscope is demonstrated through selected proof-of-principle experiments.

A versatile optical microscope for time-dependent single-molecule and single-particle spectroscopy.

Science.gov (United States)

Li, Hao; Yang, Haw

2018-03-28

This work reports the design and implementation of a multi-function optical microscope for time-dependent spectroscopy on single molecules and single nanoparticles. It integrates the now-routine single-object measurements into one standalone platform so that no reconfiguration is needed when switching between different types of sample or spectroscopy modes. The illumination modes include evanescent field through total internal reflection, dark-field illumination, and epi-excitation onto a diffraction-limited spot suitable for confocal detection. The detection modes include spectrally resolved line imaging, wide-field imaging with dual-color capability, and two-color single-element photon-counting detection. The switch between different spectroscopy and data acquisition modes is fully automated and executed through computer programming. The capability of this microscope is demonstrated through selected proof-of-principle experiments.

Resonantly enhanced production of excited fragments of gaseous molecules following core-level excitation

International Nuclear Information System (INIS)

Chen, J.M.; Lu, K.T.; Lee, J.M.; Ho, S.C.; Chang, H.W.; Lee, Y.Y.

2005-01-01

State-selective dissociation dynamics for the excited fragments of gaseous Si(CH 3 ) 2 Cl 2 following Cl 2p and Si 2p core-level excitations have been investigated by resonant photoemission spectroscopy and dispersed UV/optical fluorescence spectroscopy. The main features in the gaseous Si(CH 3 ) 2 Cl 2 fluorescence spectrum are identified as the emission from excited Si*, Si + *, CH* and H*. The core-to-Rydberg excitations at both Si 2p and Cl 2p edges lead to a noteworthy production of not only the excited atomic fragments, neutral and ionic (Si*, Si + *) but also the excited diatomic fragments (CH*). In particular, the excited neutral atomic fragments Si* are significantly reinforced. The experimental results provide deeper insight into the state-selective dissociation dynamics for the excited fragments of molecules via core-level excitation

Core excitation and de-excitation spectroscopies of free atoms and molecules

International Nuclear Information System (INIS)

Ueda, Kiyoshi

2006-01-01

This article provides a review of the current status of core excitation and de-excitation spectroscopy studies of free atoms molecules using a high-resolution soft X-ray monochromator and a high-resolution electron energy analyzer, installed in the soft X-ray photochemistry beam line at SPring-8. Experimental results are discussed for 1s excitation of Ne, O 1s excitation of CO and H 2 O, and F 1s excitation of CF 4 . (author)

Infrared Multiphoton Dissociation Spectroscopy with Free-Electron Lasers: On the Road from Small Molecules to Biomolecules.

Science.gov (United States)

Jašíková, Lucie; Roithová, Jana

2018-03-07

Infrared multiphoton dissociation (IRMPD) spectroscopy is commonly used to determine the structure of isolated, mass-selected ions in the gas phase. This method has been widely used since it became available at free-electron laser (FEL) user facilities. Thus, in this Minireview, we examine the use of IRMPD/FEL spectroscopy for investigating ions derived from small molecules, metal complexes, organometallic compounds and biorelevant ions. Furthermore, we outline new applications of IRMPD spectroscopy to study biomolecules. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Single molecule tracking fluorescence microscopy in mitochondria reveals highly dynamic but confined movement of Tom40

Science.gov (United States)

Kuzmenko, Anton; Tankov, Stoyan; English, Brian P.; Tarassov, Ivan; Tenson, Tanel; Kamenski, Piotr; Elf, Johan; Hauryliuk, Vasili

2011-12-01

Tom40 is an integral protein of the mitochondrial outer membrane, which as the central component of the Translocase of the Outer Membrane (TOM) complex forms a channel for protein import. We characterize the diffusion properties of individual Tom40 molecules fused to the photoconvertable fluorescent protein Dendra2 with millisecond temporal resolution. By imaging individual Tom40 molecules in intact isolated yeast mitochondria using photoactivated localization microscopy with sub-diffraction limited spatial precision, we demonstrate that Tom40 movement in the outer mitochondrial membrane is highly dynamic but confined in nature, suggesting anchoring of the TOM complex as a whole.

Statistical Analysis of Bending Rigidity Coefficient Determined Using Fluorescence-Based Flicker-Noise Spectroscopy.

Science.gov (United States)

Doskocz, Joanna; Drabik, Dominik; Chodaczek, Grzegorz; Przybyło, Magdalena; Langner, Marek

2018-06-01

Bending rigidity coefficient describes propensity of a lipid bilayer to deform. In order to measure the parameter experimentally using flickering noise spectroscopy, the microscopic imaging is required, which necessitates the application of giant unilamellar vesicles (GUV) lipid bilayer model. The major difficulty associated with the application of the model is the statistical character of GUV population with respect to their size and the homogeneity of lipid bilayer composition, if a mixture of lipids is used. In the paper, the bending rigidity coefficient was measured using the fluorescence-enhanced flicker-noise spectroscopy. In the paper, the bending rigidity coefficient was determined for large populations of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3-phosphocholine vesicles. The quantity of obtained experimental data allows to perform statistical analysis aiming at the identification of the distribution, which is the most appropriate for the calculation of the value of the membrane bending rigidity coefficient. It has been demonstrated that the bending rigidity coefficient is characterized by an asymmetrical distribution, which is well approximated with the gamma distribution. Since there are no biophysical reasons for that we propose to use the difference between normal and gamma fits as a measure of the homogeneity of vesicle population. In addition, the effect of a fluorescent label and types of instrumental setups on determined values has been tested. Obtained results show that the value of the bending rigidity coefficient does not depend on the type of a fluorescent label nor on the type of microscope used.

Probing the local environment of a single OPE3 molecule using inelastic tunneling electron spectroscopy.

Science.gov (United States)

Frisenda, Riccardo; Perrin, Mickael L; van der Zant, Herre S J

2015-01-01

We study single-molecule oligo(phenylene ethynylene)dithiol junctions by means of inelastic electron tunneling spectroscopy (IETS). The molecule is contacted with gold nano-electrodes formed with the mechanically controllable break junction technique. We record the IETS spectrum of the molecule from direct current measurements, both as a function of time and electrode separation. We find that for fixed electrode separation the molecule switches between various configurations, which are characterized by different IETS spectra. Similar variations in the IETS signal are observed during atomic rearrangements upon stretching of the molecular junction. Using quantum chemistry calculations, we identity some of the vibrational modes which constitute a chemical fingerprint of the molecule. In addition, changes can be attributed to rearrangements of the local molecular environment, in particular at the molecule-electrode interface. This study shows the importance of taking into account the interaction with the electrodes when describing inelastic contributions to transport through single-molecule junctions.

Simultaneous tracking of drug molecules and carriers using aptamer-functionalized fluorescent superstable gold nanorod-carbon nanocapsules during thermo-chemotherapy

Science.gov (United States)

Wang, Xue-Wei; Gao, Wei; Fan, Huanhuan; Ding, Ding; Lai, Xiao-Fang; Zou, Yu-Xiu; Chen, Long; Chen, Zhuo; Tan, Weihong

2016-04-01

Controlling and monitoring the drug delivery process is critical to its intended therapeutic function. Many nanocarrier systems for drug delivery have been successfully developed. However, biocompatibility, stability, and simultaneously tracing drugs and nanocarriers present significant limitations. Herein, we have fabricated a multifunctional nanocomposite by coating the gold nanorod (AuNR) with a biocompatible, superstable and fluorescent carbon layer, obtaining the AuNR@carbon core-shell nanocapsule. In this system, the carbon shell, originally obtained in aqueous glucose solutions and, therefore, biocompatible in physiological environments, could be simply loaded with cell-specific aptamers and therapeutic molecules through π-π interactions, a useful tool for cancer-targeted cellular imaging and therapy. Moreover, such a stable and intrinsic fluorescence effect of the AuNR@carbon enabled simultaneous tracking of released therapeutic molecules and nanocarriers under thermo-chemotherapy. The AuNR@carbons had high surface areas and stable shells, as well as unique optical and photothermal properties, making them promising nanostructures for biomedical applications.Controlling and monitoring the drug delivery process is critical to its intended therapeutic function. Many nanocarrier systems for drug delivery have been successfully developed. However, biocompatibility, stability, and simultaneously tracing drugs and nanocarriers present significant limitations. Herein, we have fabricated a multifunctional nanocomposite by coating the gold nanorod (AuNR) with a biocompatible, superstable and fluorescent carbon layer, obtaining the AuNR@carbon core-shell nanocapsule. In this system, the carbon shell, originally obtained in aqueous glucose solutions and, therefore, biocompatible in physiological environments, could be simply loaded with cell-specific aptamers and therapeutic molecules through π-π interactions, a useful tool for cancer-targeted cellular imaging and

Photoelectron-Auger electron coincidence spectroscopy of free molecules: New experiments

International Nuclear Information System (INIS)

Ulrich, Volker; Barth, Silko; Lischke, Toralf; Joshi, Sanjeev; Arion, Tiberiu; Mucke, Melanie; Foerstel, Marko; Bradshaw, Alex M.; Hergenhahn, Uwe

2011-01-01

Photoelectron-Auger electron coincidence spectroscopy probes the dicationic states produced by Auger decay following the photoionization of core or inner valence levels in atoms, molecules or clusters. Moreover, the technique provides valuable insight into the dynamics of core hole decay. This paper serves the dual purpose of demonstrating the additional information obtained by this technique compared to Auger spectroscopy alone as well as of describing the new IPP/FHI apparatus at the BESSY II synchrotron radiation source. The distinguishing feature of the latter is the capability to record both the photoelectron and Auger electron with good energy and angle resolution, for which purpose a large hemispherical electrostatic analyser is combined with several linear time-of-flight spectrometers. New results are reported for the K-shell photoionization of oxygen (O 2 ) and the subsequent KVV Auger decay. Calculations in the literature for non-coincident O 2 Auger spectra are found to be in moderately good agreement with the new data.

Decay time shortening of fluorescence from donor-acceptor pair proteins using ultrafast time-resolved fluorescence resonance energy transfer spectroscopy

International Nuclear Information System (INIS)

Baba, Motoyoshi; Suzuki, Masayuki; Ganeev, Rashid A.; Kuroda, Hiroto; Ozaki, Tsuneyuki; Hamakubo, Takao; Masuda, Kazuyuki; Hayashi, Masahiro; Sakihama, Toshiko; Kodama, Tatsuhiko; Kozasa, Tohru

2007-01-01

We improved an ultrafast time-resolved fluorescence resonance energy transfer (FRET) spectroscopy system and measured directly the decrease in the fluorescence decay time of the FRET signal, without any entanglement of components in the picosecond time scale from the donor-acceptor protein pairs (such as cameleon protein for calcium ion indicator, and ligand-activated GRIN-Go proteins pair). The drastic decrease in lifetime of the donor protein fluorescence under the FRET condition (e.g. a 47.8% decrease for a GRIN-Go protein pair) proves the deformation dynamics between donor and acceptor fluorescent proteins in an activated state of a mixed donor-acceptor protein pair. This study is the first clear evidence of physical contact of the GRIN-Go proteins pair using time-resolved FRET system. G protein-coupled receptors (GPCRs) are the most important protein family for the recognition of many chemical substances at the cell surface. They are the targets of many drugs. Simultaneously, we were able to observe the time-resolved spectra of luminous proteins at the initial stage under the FRET condition, within 10 ns from excitation. This new FRET system allows us to trace the dynamics of the interaction between proteins at the ligand-induced activated state, molecular structure change and combination or dissociation. It will be a key technology for the development of protein chip technology

Design of ortho-Substituted Donor-Acceptor Molecules as Highly Efficient Green Thermally Activated Delayed Fluorescent Emitters

Science.gov (United States)

Cha, Jae-Ryung; Gong, Myoung-Seon; Lee, Tak Jae; Ha, Tae Hoon; Lee, Chil Won

2018-04-01

The ortho-substituted donor-acceptor molecules 2-(4,6-diphenyl-1, 3, 5-triazin-2-yl)- N,Ndiphenylaniline (DPA- o-Trz) and 2-(4,6-diphenyl-1, 3, 5-triazine-2-yl)- N,N-di- p-tolylaniline (MPA- o-Trz) were designed, synthesized, and found to exhibit green fluorescence characteristics. Notably, the singlet-triplet energy gap was less than 0.1 eV, indicating that reverse intersystem crossing gave rise to thermally activated delayed fluorescence (TADF). The organic light-emitting device performance of MPA- o-Trz showed a high external quantum efficiency of 16.3% and good color stability from 0.1 cd/m2 to 5000 cd/m2.

A novel approach for the detection of early gastric cancer: fluorescence spectroscopy of gastric juice.

Science.gov (United States)

Deng, Kai; Zhou, Li Ya; Lin, San Ren; Li, Yuan; Chen, Mo; Geng, Qiu Ming; Li, Yu Wen

2013-06-01

This study aimed to investigate the efficacy of fluorescence spectroscopy of gastric juice for early gastric cancer (EGC) screening. Gastric juice was collected from 101 participants who underwent endoscopy in the Outpatient Endoscopy Center of Peking University Third Hospital. The participants were divided into three groups: the normal mucosa or chronic non-atrophic gastritis (NM-CNAG) group (n = 35), advanced gastric cancer (AGC) group (n = 33) and EGC group (n = 33). Fluorescence spectroscopic analysis was performed in all the gastric juice samples and the maximum fluorescence intensity of the first peak (P1 FI) was measured. The mean fluorescence intensity of P1 FI of gastric juice in AGC (92.1 ± 10.7) and EGC (90.8 ± 12.0) groups was significantly higher than that in the NM-CNAG group (55.7 ± 7.5) (AGC vs NM-CNAG, P = 0.006 and EGC vs NM-CNAG, P = 0.015, respectively). The areas under the receiver operating characteristic curves for the detection of AGC and EGC were 0.681 (95% confidence interval [CI] 0.553-0.810, P = 0.010) and 0.655 (95% CI 0.522-0.787, P = 0.028). With the P1 FI of ≥47.7, the sensitivity, specificity and accuracy for detecting EGC were 69.7%, 57.1% and 63.2%, respectively. The enhancement of P1 FI of gastric juice occurs at the early stage of gastric cancer. Fluorescence spectroscopy of gastric juice may be used as a novel screening tool for the early detection of gastric cancer. © 2013 The Authors. Journal of Digestive Diseases © 2013 Wiley Publishing Asia Pty Ltd and Chinese Medical Association Shanghai Branch, Chinese Society of Gastroenterology, Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine.

Fluorescence excitation studies of molecular photoionization in external electric fields

International Nuclear Information System (INIS)

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

1985-01-01

Using molecular nitrogen as an example, we show that fluorescence excitation spectroscopy can be used to measure partial photoionization cross sections of free molecules in external electric fields. The production of the N 2 + (B 2 Σ/sub u/ + ) state was studied and the threshold for this process was found to shift linearly with the square root of the applied field. This behavior is compared with the hydrogenic case and with previously studied systems

Fluorescence spectroscopy as a tool for determination of organic matter removal efficiency at water treatment works

Directory of Open Access Journals (Sweden)

M. Z. Bieroza

2010-04-01

Full Text Available Organic matter (OM in drinking water treatment is a common impediment responsible for increased coagulant and disinfectant dosages, formation of carcinogenic disinfection-by products, and microbial re-growth in distribution system. The inherent heterogeneity of OM implies the utilization of advanced analytical techniques for its characterization and assessment of removal efficiency. Here, the application of simple fluorescence excitation-emission technique to OM characterization in drinking water treatment is presented. The fluorescence data of raw and clarified water was obtained from 16 drinking water treatment works. The reduction in fulvic-like fluorescence was found to significantly correlate with OM removal measured with total organic carbon (TOC. Fluorescence properties, fulvic- and tryptophan-like regions, were found to discriminate OM fractions of different removal efficiencies. The results obtained in the study show that fluorescence spectroscopy provides a rapid and accurate characterization and quantification of OM fractions and indication of their treatability in conventional water treatment.

Atoms, molecules and optical physics 2. Molecules and photons - Spectroscopy and collisions

Energy Technology Data Exchange (ETDEWEB)

Hertel, Ingolf V.; Schulz, Claus-Peter [Max-Born-Institut fuer Nichtlineare Optik und Kurzzeitspektroskopie im Forschungsverbund Berlin e.V. (Germany)

2015-09-01

This is the second volume of textbooks on atomic, molecular and optical physics, aiming at a comprehensive presentation of this highly productive branch of modern physics as an indispensable basis for many areas in physics and chemistry as well as in state of the art bio- and material-sciences. It primarily addresses advanced students (including PhD students), but in a number of selected subject areas the reader is lead up to the frontiers of present research. Thus even the active scientist is addressed. This volume 2 introduces lasers and quantum optics, while the main focus is on the structure of molecules and their spectroscopy, as well as on collision physics as the continuum counterpart to bound molecular states. The emphasis is always on the experiment and its interpretation, while the necessary theory is introduced from this perspective in a compact and occasionally somewhat heuristic manner, easy to follow even for beginners.

Dye lasers in atomic spectroscopy

International Nuclear Information System (INIS)

Lange, W.; Luther, J.; Steudel, A.

1974-01-01

The properties of dye lasers which are relevant to atomic spectroscopy are discussed. Several experiments made possible by tunable dye lasers are discussed. Applications of high spectral density dye lasers are covered in areas such as absorption spectroscopy, fluorescence spectroscopy, photoionization and photodetachment, and two- and multi-photon processes. Applications which take advantage of the narrow bandwidth of tunable dye lasers are discussed, including saturation spectroscopy, fluorescence line narrowing, classic absorption and fluorescence spectroscopy, nonoptical detection of optical resonances, heterodyne spectroscopy, and nonlinear coherent resonant phenomena. (26 figures, 180 references) (U.S.)

Production and spectroscopy of ultracold YbRb* molecules

International Nuclear Information System (INIS)

Nemitz, Nils

2008-11-01

This thesis describes the formation of electronically excited but translationally cold molecules formed from rubidium atoms and two isotopes of ytterbium ( 176 Yb and 174 Yb) by means of photoassociation. The experiments were performed in a combined MOT with 10 9 rubidium atoms and 2.10 6 ytterbium atoms at temperatures of less than 1 mK. Photoassociation lines were found by trap loss spectroscopy throughout a wavelength range of 2 nm near the 795 nm D1 transition in rubidium. The majority of lines belong to two vibrational series in the excited YbRb * molecule, converging on a system of a ground state ytterbium atom and an excited rubidium atom. The strong variation of line strength between different vibrational lines is explained through the Franck-Condon principle. An improved version of the Leroy-Bernstein equation was used to extract the leading dispersion coefficient of the potential from the vibrational progression. Most of the observed lines show a resolved rotational structure as expected from a basic quantum mechanical model. The series terminates with the third or forth rotational component due to the ground state centrifugal barrier.The measured rotational constants agree very well with calculations based on the C 6 coefficient. The discovery of a splitting of the rotational components into subcomponents indicates an uncommon angular momentum coupling described by Hund's case. Variations in the depth of the subcomponents indicates a similar splitting in the ground state, with the energies of the substates based on the alignment of the rubidium atom's magnetic dipole moment relative to the angular momentum carried by an approaching ytterbium atom. This creates an additional ground state barrier, partially suppressing some of the subcomponents. Using a rate equation model developed for this purpose, a maximum formation rate of 2.5.10 6 molecules per second was calculated over the volume of the entire trap. The work presented here is an important step on

A statistical strategy to assess cleaning level of surfaces using fluorescence spectroscopy and Wilks’ ratio

DEFF Research Database (Denmark)

Stoica, Iuliana-Madalina; Babamoradi, Hamid; van den Berg, Frans

2017-01-01

•A statistical strategy combining fluorescence spectroscopy, multivariate analysis and Wilks’ ratio is proposed.•The method was tested both off-line and on-line having riboflavin as a (controlled) contaminant.•Wilks’ ratio signals unusual recordings based on shifts in variance and covariance...... structure described in in-control data....

Bilayer Localization of Membrane-Active Peptides Studied in Biomimetic Vesicles by Visible and Fluorescence Spectroscopies

Czech Academy of Sciences Publication Activity Database

Sheynis, T.; Sýkora, Jan; Benda, Aleš; Kolusheva, S.; Hof, Martin; Jelinek, R.

2003-01-01

Roč. 270, č. 22 (2003), s. 4478-4487 ISSN 0014-2956 R&D Projects: GA MŠk LN00A032 Institutional research plan: CEZ:AV0Z4040901 Keywords : solvent relaxation * fluorescence correlation spectroscopy * lipid bilayers Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.001, year: 2003

Photon-HDF5: An Open File Format for Timestamp-Based Single-Molecule Fluorescence Experiments

OpenAIRE

Ingargiola, Antonino; Laurence, Ted; Boutelle, Robert; Weiss, Shimon; Michalet, Xavier

2016-01-01

We introduce Photon-HDF5, an open and efficient file format to simplify exchange and long-term accessibility of data from single-molecule fluorescence experiments based on photon-counting detectors such as single-photon avalanche diode, photomultiplier tube, or arrays of such detectors. The format is based on HDF5, a widely used platform- and language-independent hierarchical file format for which user-friendly viewers are available. Photon-HDF5 can store raw photon data (timestamp, channel n...

In vivo study of the human skin by the method of laser-induced fluorescence spectroscopy

International Nuclear Information System (INIS)

Borisova, E.; Avramov, L.

2000-01-01

The goals of this study are to perform a preliminary evaluation of the diagnostic potential of noninvasive laser-induced auto-fluorescence spectroscopy (LIAFS) for human skin and optimize of detection and diagnosis of hollow organs and skin. In recent years, there has been growing interest in the use of laser-induced fluorescence to discriminate disease from normal surrounding tissue. The most fluorescence studies have used exogenous fluorophores of this discrimination. The laser-induced auto-fluorescence which is used for diagnosis of tissues in the human body avoids administration of any drugs. In this study a technique for optical biopsy of in vivo human skin is presented. The auto-fluorescence characterization of tissue relies on different spectral properties of tissues. It was demonstrated a differentiation between normal skin and skin with vitiligo. Two main endogenous fluorophores in the human skin account for most of the cellular auto-fluorescence for excitation wavelength 337 nm reduced from of nicotinamide adenine dinucleotide and collagen. The auto-fluorescence spectrum of human skin depend on main internal absorbers which are blood and melanin. In this study was described the effect caused by blood and melanin content on the shape of the auto-fluorescence spectrum of human skin. Human skin fluorescence spectrum might provide dermatologists with important information and such investigations are successfully used now in skin disease diagnostics, in investigation of the environmental factor impact or for evaluation of treatment efficiency. (authors)

Effect of tissue scaffold topography on protein structure monitored by fluorescence spectroscopy.

Science.gov (United States)

Portugal, Carla A M; Truckenmüller, Roman; Stamatialis, Dimitrios; Crespo, João G

2014-11-10

The impact of surface topography on the structure of proteins upon adhesion was assessed through non-invasive fluorescence monitoring. This study aimed at obtaining a better understanding about the role of protein structural status on cell-scaffold interactions. The changes induced upon adsorption of two model proteins with different geometries, trypsin (globular conformation) and fibrinogen (rod-shaped conformation) on poly-l-lactic acid (PLLA) scaffolds with different surface topographies, flat, fibrous and surfaces with aligned nanogrooves, were assessed by fluorescence spectroscopy monitoring, using tryptophan as structural probe. Hence, the maximum emission blue shift and the increase of fluorescence anisotropy observed after adsorption of globular and rod-like shaped proteins on surfaces with parallel nanogrooves were ascribed to more intense protein-surface interactions. Furthermore, the decrease of fluorescence anisotropy observed upon adsorption of proteins to scaffolds with fibrous morphology was more significant for rod-shaped proteins. This effect was associated to the ability of these proteins to adjust to curved surfaces. The additional unfolding of proteins induced upon adsorption on scaffolds with a fibrous morphology may be the reason for better cell attachment there, promoting an easier access of cell receptors to initially hidden protein regions (e.g. RGDS sequence), which are known to have a determinant role in cell attaching processes. Copyright © 2014 Elsevier B.V. All rights reserved.

DETECTION OF MERCURIC BROMIDE IN A GAS PHASE FLOW CELL BY LASER PHOTOFRAGMENT FLUORESCENCE SPECTROSCOPY. (R825380)

Science.gov (United States)

Photofragment fluorescence (PFF) spectroscopy offers real-time monitoring capability with high-analytical sensitivity and selectivity for volatile mercury compounds found in process gas streams, such as incinerator stacks. In this work, low concentrations (6 ppb to...

Fluorescence spectroscopy for assessment of liver transplantation grafts concerning graft viability and patient survival

Science.gov (United States)

Vollet Filho, José D.; da Silveira, Marina R.; Castro-e-Silva, Orlando; Bagnato, Vanderlei S.; Kurachi, Cristina

2015-06-01

Evaluating transplantation grafts at harvest is essential for its success. Laser-induced fluorescence spectroscopy (LIFS) can help monitoring changes in metabolic/structural conditions of tissue during transplantation. The aim of the present study is to correlate LIFSobtained spectra of human hepatic grafts during liver transplantation with post-operative patients' mortality rate and biochemical parameters, establishing a method to exclude nonviable grafts before implantation. Orthotopic liver transplantation, piggyback technique was performed in 15 patients. LIFS was performed under 408nm excitation. Collection was performed immediately after opening donor's abdominal cavity, after cold perfusion, end of back-table period, and 5 min and 1 h after warm perfusion at recipient. Fluorescence information was compared to lactate, creatinine, bilirubin and INR levels and to survival status. LIFS was sensitive to liver changes during transplantation stages. Study-in-progress; initial results indicate correlation between fluorescence and life/death status of patients.

Study of the heavy impurity influx into the plasma using laser fluorescence spectroscopy in the TO-2 tokamak with toroidal divertor

International Nuclear Information System (INIS)

Vukolov, K.Yu.; Shvindt, N.N.

1992-01-01

Measurement cycle for determination of iron atom absolute concentrations was carried out in divertor and diaphragm modes of laser fluorescence spectroscopy. The conclusion is made on effective wall shielding by divertor layer as compared to material diaphragm. The basic result of the work consists in creating and testing on the tokamak TO-2 of multichannel diagnostic complex for remote measurement of atom (ion) absolute concentrations of metallic impurities in the near-wall plasma with high spatial and time resolution through laser fluorescence spectroscopy method intended for studies at the Tokamak-15 facility

Fluorescent labeling of NASBA amplified tmRNA molecules for microarray applications

Directory of Open Access Journals (Sweden)

Kaplinski Lauris

2009-05-01

Full Text Available Abstract Background Here we present a novel promising microbial diagnostic method that combines the sensitivity of Nucleic Acid Sequence Based Amplification (NASBA with the high information content of microarray technology for the detection of bacterial tmRNA molecules. The NASBA protocol was modified to include aminoallyl-UTP (aaUTP molecules that were incorporated into nascent RNA during the NASBA reaction. Post-amplification labeling with fluorescent dye was carried out subsequently and tmRNA hybridization signal intensities were measured using microarray technology. Significant optimization of the labeled NASBA protocol was required to maintain the required sensitivity of the reactions. Results Two different aaUTP salts were evaluated and optimum final concentrations were identified for both. The final 2 mM concentration of aaUTP Li-salt in NASBA reaction resulted in highest microarray signals overall, being twice as high as the strongest signals with 1 mM aaUTP Na-salt. Conclusion We have successfully demonstrated efficient combination of NASBA amplification technology with microarray based hybridization detection. The method is applicative for many different areas of microbial diagnostics including environmental monitoring, bio threat detection, industrial process monitoring and clinical microbiology.

Precision spectroscopy with ultracold {sup 87}Rb{sub 2} triplet molecules

Energy Technology Data Exchange (ETDEWEB)

Strauss, Christoph

2011-10-19

In this thesis I report precision spectroscopy with ultracold {sup 87}Rb{sub 2} triplet molecules where we use lasers to couple the states in different molecular potentials. We study in detail states of the a {sup 3} sum {sup +}{sub u} and (1) {sup 3} sum {sup +}{sub g} potentials. These states are of great importance for transferring weakly bound molecules to the ro-vibrational triplet ground state via states of the excited potential. As most experiments start from molecules in their X {sup 1} sum {sup +}{sub g} ground state, the triplet states were hard to access via dipole transitions and remained largely unexplored. The measurements presented in this thesis are the first detailed study of diatomic {sup 87}Rb{sub 2} molecules in these states. Our experiments start with an ultracold cloud of {sup 87}Rb atoms. We then load this cloud into an optical lattice where we use a magnetic Feshbach resonance at 1007.4 G to perform a Feshbach association. After we have removed all unbound atoms, we end up with a pure sample of weakly bound Feshbach molecules inside the optical lattice. The optical lattice prevents these molecules from colliding with each other which results in molecular lifetimes on the order of a few hundred milliseconds. In the first set of experiments, we use a laser coupling the Feshbach state to the excited (1) {sup 3} sum {sup +}{sub g} triplet state to map out its low-lying vibrational (v = 0.. 15), rotational, hyperfine, and Zeeman structure. The experimental results are in good agreement with calculations done by Marius Lysebo and Prof. Leif Veseth. We then map out in detail the vibrational, rotational, hyperfine, and Zeeman structure of the a {sup 3} sum {sup +}{sub u} triplet ground state using dark state spectroscopy with levels in the (1) {sup 3} sum {sup +}{sub g} potential as an intermediate state. In this scheme we are able to access molecules in triplet states because our Feshbach state has strong triplet character. Interestingly, it

Precision spectroscopy with ultracold {sup 87}Rb{sub 2} triplet molecules

Energy Technology Data Exchange (ETDEWEB)

Strauss, Christoph

2011-10-19

In this thesis I report precision spectroscopy with ultracold {sup 87}Rb{sub 2} triplet molecules where we use lasers to couple the states in different molecular potentials. We study in detail states of the a {sup 3} sum {sup +}{sub u} and (1) {sup 3} sum {sup +}{sub g} potentials. These states are of great importance for transferring weakly bound molecules to the ro-vibrational triplet ground state via states of the excited potential. As most experiments start from molecules in their X {sup 1} sum {sup +}{sub g} ground state, the triplet states were hard to access via dipole transitions and remained largely unexplored. The measurements presented in this thesis are the first detailed study of diatomic {sup 87}Rb{sub 2} molecules in these states. Our experiments start with an ultracold cloud of {sup 87}Rb atoms. We then load this cloud into an optical lattice where we use a magnetic Feshbach resonance at 1007.4 G to perform a Feshbach association. After we have removed all unbound atoms, we end up with a pure sample of weakly bound Feshbach molecules inside the optical lattice. The optical lattice prevents these molecules from colliding with each other which results in molecular lifetimes on the order of a few hundred milliseconds. In the first set of experiments, we use a laser coupling the Feshbach state to the excited (1) {sup 3} sum {sup +}{sub g} triplet state to map out its low-lying vibrational (v = 0.. 15), rotational, hyperfine, and Zeeman structure. The experimental results are in good agreement with calculations done by Marius Lysebo and Prof. Leif Veseth. We then map out in detail the vibrational, rotational, hyperfine, and Zeeman structure of the a {sup 3} sum {sup +}{sub u} triplet ground state using dark state spectroscopy with levels in the (1) {sup 3} sum {sup +}{sub g} potential as an intermediate state. In this scheme we are able to access molecules in triplet states because our Feshbach state has strong triplet character. Interestingly, it

Assessment of post-implantation integration of engineered tissues using fluorescence lifetime spectroscopy

Science.gov (United States)

Elahi, Sakib F.; Lee, Seung Y.; Lloyd, William R.; Chen, Leng-Chun; Kuo, Shiuhyang; Zhou, Ying; Kim, Hyungjin M.; Kennedy, Robert; Marcelo, Cynthia; Feinberg, Stephen E.; Mycek, Mary-Ann

2018-02-01

Clinical translation of engineered tissue constructs requires noninvasive methods to assess construct health and viability after implantation in patients. However, current practices to monitor post-implantation construct integration are either qualitative (visual assessment) or destructive (tissue histology). As label-free fluorescence lifetime sensing can noninvasively characterize pre-implantation construct viability, we employed a handheld fluorescence lifetime spectroscopy probe to quantitatively and noninvasively assess tissue constructs that were implanted in a murine model. We designed the system to be suitable for intravital measurements: portability, localization with precise maneuverability, and rapid data acquisition. Our model tissue constructs were manufactured from primary human cells to simulate patient variability and were stressed to create a range of health states. Secreted amounts of three cytokines that relate to cellular viability were measured in vitro to assess pre-implantation construct health. In vivo optical sensing assessed tissue integration of constructs at one-week and three-weeks post-implantation. At one-week post-implantation, optical parameters correlated with in vitro pre-implantation secretion levels of all three cytokines (p clinical optical diagnostic tools based on label-free fluorescence lifetime sensing of endogenous tissue fluorophores could noninvasively monitor post-implantation integration of engineered tissues.

Uranium concentrate analysis by X-ray fluorescence spectroscopy

International Nuclear Information System (INIS)

Diaz-Guerra, J.P.; Bayon, A.; Roca, R.

1978-01-01

The determination of As, Ca, Fe, Mo, P, S, Si. Th, V and U in uranium concentrates by X-ray fluorescence spectroscopy has been studied. As and U are determined in nitric solutions and for the rest of elements analysis is performed by a bead fusion technique using Li 2 B 4 O 7 and Li 2 CO 3 as fluxes. Although the uranium matrix minimizes the absorption and enhancement effects, because of the content variations of this element it is advisable to operate at a constant level of U 3 O 8 . Despite the high matrix absorption and the large dilution of the samples, sensitivity and speed are found to be satisfactory as the result of the use of a high sensitivity automatic spectrometer. The spectral interferences of Mo on S and P, and of Pb on As have been particularly considered. (author) [es

Single-Molecule Spectroscopy

Indian Academy of Sciences (India)

IAS Admin

overall absorption spectrum of a molecule is a superposition of many such sharp lines .... dilute solution of the enzyme and the substrate over few drops of silicone oil placed ..... Near-field Scanning Optical Microscopy (NSOM): Development.

Spectroscopy

DEFF Research Database (Denmark)

Berg, Rolf W.

This introductory booklet covers the basics of molecular spectroscopy, infrared and Raman methods, instrumental considerations, symmetry analysis of molecules, group theory and selection rules, as well as assignments of fundamental vibrational modes in molecules.......This introductory booklet covers the basics of molecular spectroscopy, infrared and Raman methods, instrumental considerations, symmetry analysis of molecules, group theory and selection rules, as well as assignments of fundamental vibrational modes in molecules....

A method for measurements of neutral fragments kinetic energies released to a specific dissociation threshold: optical translational spectroscopy

International Nuclear Information System (INIS)

Roney, A.; Frigon, C.; Larzilliere, M.

1999-01-01

The optical translational spectroscopy technique, based on the principles of fast ion beam laser spectroscopy (FIBLAS) and translational spectroscopy, allows the kinetic energies study of neutral fragments released through free dissociation of a neutral molecule. This method presents interesting features such as near-threshold energy measurements and selection of a specific dissociation limit. The fragments resulting from free dissociation (not induced) of neutral molecules, produced by charge exchange processes with a fast ion beam, are probed by laser radiation. Monitoring of the laser-induced fluorescence allows high-resolution spectra due to the kinematic compression of the velocity spread. Measurements of kinetic energies released to the second limit of dissociation H(1s) + H(2l) of H 2 are put forth and compared with those obtained by means of off-axis translational spectroscopy

Multimodal Raman-fluorescence spectroscopy of formalin fixed samples is able to discriminate brain tumors from dysplastic tissue

Science.gov (United States)

Anand, Suresh; Cicchi, Riccardo; Giordano, Flavio; Buccoliero, Anna Maria; Pavone, Francesco Saverio

2014-05-01

In the recent years, there has been a considerable surge in the application of spectroscopy for disease diagnosis. Raman and fluorescence spectra provide characteristic spectral profile related to biochemical and morphological changes when tissues progress from normal state towards malignancy. Spectroscopic techniques offer the advantage of being minimally invasive compared to traditional histopathology, real time and quantitative. In biomedical optical diagnostics, freshly excised specimens are preferred for making ex-vivo spectroscopic measurements. With regard to fresh tissues, if the lab is located far away from the clinic it could pose a problem as spectral measurements have to be performed immediately after dissection. Tissue samples are usually placed in a fixative agent such as 4% formaldehyde to preserve the samples before processing them for routine histopathological studies. Fixation prevents the tissues from decomposition by arresting autolysis. In the present study, we intend to investigate the possibility of using formalin fixed samples for discrimination of brain tumours from dysplastic tissue using Raman and fluorescence spectroscopy. Formalin fixed samples were washed with phosphate buffered saline for about 5 minutes in order to remove the effects of formalin during spectroscopic measurements. In case of fluorescence spectroscopy, changes in spectral profile have been observed in the region between 550-670 nm between dysplastic and tumor samples. For Raman measurements, we found significant differences in the spectral profiles between dysplasia and tumor. In conclusion, formalin fixed samples can be potentially used for the spectroscopic discrimination of tumor against dysplastic tissue in brain samples.

Single-Photon Source for Quantum Information Based on Single Dye Molecule Fluorescence in Liquid Crystal Host

International Nuclear Information System (INIS)

Lukishova, S.G.; Knox, R.P.; Freivald, P.; McNamara, A.; Boyd, R.W.; Stroud, Jr. C.R.; Schmid, A.W.; Marshall, K.L.

2006-01-01

This paper describes a new application for liquid crystals: quantum information technology. A deterministically polarized single-photon source that efficiently produces photons exhibiting antibunching is a pivotal hardware element in absolutely secure quantum communication. Planar-aligned nematic liquid crystal hosts deterministically align the single dye molecules which produce deterministically polarized single (antibunched) photons. In addition, 1-D photonic bandgap cholesteric liquid crystals will increase single-photon source efficiency. The experiments and challenges in the observation of deterministically polarized fluorescence from single dye molecules in planar-aligned glassy nematic-liquid-crystal oligomer as well as photon antibunching in glassy cholesteric oligomer are described for the first time

A small azide-modified thiazole-based reporter molecule for fluorescence and mass spectrometric detection

Directory of Open Access Journals (Sweden)

Stefanie Wolfram

2014-10-01

Full Text Available Molecular probes are widely used tools in chemical biology that allow tracing of bioactive metabolites and selective labeling of proteins and other biomacromolecules. A common structural motif for such probes consists of a reporter that can be attached by copper(I-catalyzed 1,2,3-triazole formation between terminal alkynes and azides to a reactive headgroup. Here we introduce the synthesis and application of the new thiazole-based, azide-tagged reporter 4-(3-azidopropoxy-5-(4-bromophenyl-2-(pyridin-2-ylthiazole for fluorescence, UV and mass spectrometry (MS detection. This small fluorescent reporter bears a bromine functionalization facilitating the automated data mining of electrospray ionization MS runs by monitoring for its characteristic isotope signature. We demonstrate the universal utility of the reporter for the detection of an alkyne-modified small molecule by LC–MS and for the visualization of a model protein by in-gel fluorescence. The novel probe advantageously compares with commercially available azide-modified fluorophores and a brominated one. The ease of synthesis, small size, stability, and the universal detection possibilities make it an ideal reporter for activity-based protein profiling and functional metabolic profiling.

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.

Ultraviolet-visible and fluorescence spectroscopy can be used as a diagnostic tool for gamma irradiation detection in vivo.

Science.gov (United States)

K-Abdelhalim, Mohamed Anwar; Moussa, Sherif A-Abdelmottaleb

2016-09-01

The spectroscopic properties can indicate important features about the nature and severity of the disease. However, no earlier studies have been used the spectroscopic properties as a diagnostic tool for radiation detection. This study was aimed to use ultraviolet-visible and fluorescence spectroscopy as a diagnostic tool for gamma irradiation detection in rats in vivo. Adult male rats were exposed to 25, 50, 75 and 100 Gray as single dose, using Cobalt-60 (Co-60) source with a dose rate of 0.883 centi Gray/sec (cGy/s). Ultraviolet and fluorescence spectroscopy of rat's blood serum were measured. After gamma irradiation of rats in vivo, the blood serum absorbance peaks for 25, 50, 75 and 100 Gray (Gy) decreased and shifted towards the ultra violet wavelength. A maximal change in fluorescence intensity of blood serum at 350 nm was obtained when exciting light at 194 nm after irradiation. The fluorescence intensity also decreased with the dose. The highest radiation gamma dose might be accompanied with the highest oxidative stress. This study suggests that at the above mentioned gamma radiation doses, the blood is highly fragmented; with low aggregation at 25 Gy and with high aggregation at 50-100 Gy.

A Linear Ion Trap with an Expanded Inscribed Diameter to Improve Optical Access for Fluorescence Spectroscopy

Science.gov (United States)

Rajagopal, Vaishnavi; Stokes, Chris; Ferzoco, Alessandra

2018-02-01

We report a custom-geometry linear ion trap designed for fluorescence spectroscopy of gas-phase ions at ambient to cryogenic temperatures. Laser-induced fluorescence from trapped ions is collected from between the trapping rods, orthogonal to the excitation laser that runs along the axis of the linear ion trap. To increase optical access to the ion cloud, the diameter of the round trapping rods is 80% of the inscribed diameter, rather than the roughly 110% used to approximate purely quadrupolar electric fields. To encompass as much of the ion cloud as possible, the first collection optic has a 25.4 mm diameter and a numerical aperture of 0.6. The choice of geometry and collection optics yields 107 detected photons/s from trapped rhodamine 6G ions. The trap is coupled to a closed-cycle helium refrigerator, which in combination with two 50 Ohm heaters enables temperature control to below 25 K on the rod electrodes. The purpose of the instrument is to broaden the applicability of fluorescence spectroscopy of gas-phase ions to cases where photon emission is a minority relaxation pathway. Such studies are important to understand how the microenvironment of a chromophore influences excited state charge transfer processes.

Optical spectroscopy of laser-produced plasmas for standoff isotopic analysis

Energy Technology Data Exchange (ETDEWEB)

Harilal, Sivanandan S.; Brumfield, Brian E.; LaHaye, Nicole L.; Hartig, Kyle C.; Phillips, Mark C.

2018-04-20

This review article covers the present status of isotope detection through emission, absorption, and fluorescence spectroscopy of atoms and molecules in a laser-produced plasma formed from a solid sample. A description of the physics behind isotope shifts in atoms and molecules is presented, followed by the physics behind solid sampling of laser ablation plumes, optical methods for isotope measurements, the suitable physical conditions of laser-produced plasma plumes for isotopic analysis, and the current status. Finally, concluding remarks will be made on the existing gaps between previous works in the literature and suggestions for future work.

Modulation of a fluorescence switch based on photochromic spirooxazine in composite organic nanoparticles

International Nuclear Information System (INIS)

Sheng Xiaohai; Peng Aidong; Fu Hongbing; Liu Yuanyuan; Zhao Yongsheng; Ma Ying; Yao Jiannian

2007-01-01

We describe a versatile and convenient approach to achieve fluorescence modulation by the preparation of composite nanoparticles (CNPs), based on photochromic 5-methoxy-1,3,3-trimethyl-9'-hydroxyspiroindolinenaphthoxazine (SO), fluorescent 4-(dicyanomethylene)-2-methyl-6-(p-dimethyl-aminostyryl)-4H-pyran (DCM), and emissive-assistant 1,3-bis(pyrene) propane (BPP) molecules, employing doping techniques. The mechanism of the fluorescence switch is the intermolecular energy transfer as supported by both steady-state and time-resolved spectroscopy results. The addition of BPP not only enhances the contrast of the fluorescence signal between the 'ON' and 'OFF' state, but also provides a convenient way to tune the excitation wavelength for reading the fluorescence. High-contrast ON/OFF (20:1) fluorescence switching is successfully implemented in the CNPs and also in a more practical PVA film loaded with the CNPs. This system may represent an alternative to the covalent system in potentially rewritable high-density optical data or image storage utilizing luminescence intensity readout schemes

Solution conformation of 2-aminopurine dinucleotide determined by ultraviolet two-dimensional fluorescence spectroscopy

International Nuclear Information System (INIS)

Widom, Julia R; Marcus, Andrew H; Johnson, Neil P; Von Hippel, Peter H

2013-01-01

We have observed the conformation-dependent electronic coupling between the monomeric subunits of a dinucleotide of 2-aminopurine (2-AP), a fluorescent analogue of the nucleic acid base adenine. This was accomplished by extending two-dimensional fluorescence spectroscopy (2D FS)—a fluorescence-detected variation of 2D electronic spectroscopy—to excite molecular transitions in the ultraviolet (UV) regime. A collinear sequence of four ultrafast laser pulses centered at 323 nm was used to resonantly excite the coupled transitions of 2-AP dinucleotide. The phases of the optical pulses were continuously swept at kilohertz frequencies, and the ensuing nonlinear fluorescence was phase-synchronously detected at 370 nm. Upon optimization of a point–dipole coupling model to our data, we found that in aqueous buffer the 2-AP dinucleotide adopts an average conformation in which the purine bases are non-helically stacked (center-to-center distance R 12 = 3.5 ± 0.5 Å , twist angle θ 12 = 5° ± 5° ), which differs from the conformation of such adjacent bases in duplex DNA. These experiments establish UV–2D FS as a method for examining the local conformations of an adjacent pair of fluorescent nucleotides substituted into specific DNA or RNA constructs, which will serve as a powerful probe to interpret, in structural terms, biologically significant local conformational changes within the nucleic acid framework of protein–nucleic acid complexes. (paper)

Detection of Counterfeit Tequila by Fluorescence Spectroscopy

Directory of Open Access Journals (Sweden)

José Manuel de la Rosa Vázquez

2015-01-01

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

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.

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.

Fluorescence lifetime correlation spectroscopy combined with lifetime tuning: New perspectives in supported phospholipid bilayer research

Czech Academy of Sciences Publication Activity Database

Benda, Aleš; Fagulová, Veronika; Deyneka, Alexander; Enderlain, J.; Hof, Martin

2006-01-01

Roč. 22, č. 23 (2006), s. 9580-9585 ISSN 0743-7463 R&D Projects: GA ČR GA203/05/2308; GA MŠk LC06063 Institutional research plan: CEZ:AV0Z40400503; CEZ:AV0Z10100522 Keywords : spectroscopy * fluorescence * FLCS Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.902, year: 2006

Charge Transfer Effect on Raman and Surface Enhanced Raman Spectroscopy of Furfural Molecules.

Science.gov (United States)

Wan, Fu; Shi, Haiyang; Chen, Weigen; Gu, Zhaoliang; Du, Lingling; Wang, Pinyi; Wang, Jianxin; Huang, Yingzhou

2017-08-02

The detection of furfural in transformer oil through surface enhanced Raman spectroscopy (SERS) is one of the most promising online monitoring techniques in the process of transformer aging. In this work, the Raman of individual furfural molecules and SERS of furfural-M x (M = Ag, Au, Cu) complexes are investigated through density functional theory (DFT). In the Raman spectrum of individual furfural molecules, the vibration mode of each Raman peak is figured out, and the deviation from experimental data is analyzed by surface charge distribution. In the SERS of furfural-M x complexes, the influence of atom number and species on SERS chemical enhancement factors (EFs) are studied, and are further analyzed by charge transfer effect. Our studies strengthen the understanding of charge transfer effect in the SERS of furfural molecules, which is important in the online monitoring of the transformer aging process through SERS.

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

International Nuclear Information System (INIS)

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

1994-01-01

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

Photo-induced processes in collagen-hypericin system revealed by fluorescence spectroscopy and multiphoton microscopy

OpenAIRE

Hovhannisyan, V.; Guo, H. W.; Hovhannisyan, A.; Ghukasyan, V.; Buryakina, T.; Chen, Y. F.; Dong, C. Y.

2014-01-01

Collagen is the main structural protein and the key determinant of mechanical and functional properties of tissues and organs. Proper balance between synthesis and degradation of collagen molecules is critical for maintaining normal physiological functions. In addition, collagen influences tumor development and drug delivery, which makes it a potential cancer therapy target. Using second harmonic generation, two-photon excited fluorescence microscopy, and spectrofluorimetry, we show that the ...

Integrated Transmission Electron and Single‐Molecule Fluorescence Microscopy Correlates Reactivity with Ultrastructure in a Single Catalyst Particle

OpenAIRE

Hendriks, Frank C.; Mohammadian, Sajjad; Ristanović, Zoran; Kalirai, Sam; Meirer, Florian; Vogt, Eelco T. C.; Bruijnincx, Pieter C. A.; Gerritsen, Hans C.; Weckhuysen, Bert M.

2017-01-01

Abstract Establishing structure–activity relationships in complex, hierarchically structured nanomaterials, such as fluid catalytic cracking (FCC) catalysts, requires characterization with complementary, correlated analysis techniques. An integrated setup has been developed to perform transmission electron microscopy (TEM) and single‐molecule fluorescence (SMF) microscopy on such nanostructured samples. Correlated structure–reactivity information was obtained for 100 nm thin, microtomed secti...

Communication: atomic force detection of single-molecule nonlinear optical vibrational spectroscopy.

Science.gov (United States)

Saurabh, Prasoon; Mukamel, Shaul

2014-04-28

Atomic Force Microscopy (AFM) allows for a highly sensitive detection of spectroscopic signals. This has been first demonstrated for NMR of a single molecule and recently extended to stimulated Raman in the optical regime. We theoretically investigate the use of optical forces to detect time and frequency domain nonlinear optical signals. We show that, with proper phase matching, the AFM-detected signals closely resemble coherent heterodyne-detected signals. Applications are made to AFM-detected and heterodyne-detected vibrational resonances in Coherent Anti-Stokes Raman Spectroscopy (χ((3))) and sum or difference frequency generation (χ((2))).

Study of adsorption states for lubricant molecule using hard X-ray photoemission spectroscopy

International Nuclear Information System (INIS)

Ikenaga, E.; Kobata, M.; Kim, J.J.; Wakabayashi, A.; Nishino, Y.; Tamasaku, K.; Sakane, Y.; Ishikawa, T.; Komiya, S.; Kobayashi, K.

2007-01-01

The adsorption states for lubricant molecules have been investigated using hard X-ray (hν = 7.95 keV) photoemission spectroscopy (HX-PES). This method has the advantage for the organic molecules to be able to measure damage few. Being aware of the fact that P atoms exist only in cyclotriphosphazene base, we measured the take-off angle dependence of the P1s spectra. Each spectrum consists from two peaks, that is, substrate NiP peak and cyclotriphosphazene P peak. The cyclotriphosphazene P peak rapidly disappears with increasing take-off angle. We have also measured C1s spectra. Combining these experimental results, we have found that the adsorption state of cyclotriphosphazene end group is undergoing

High-resolution flurescence spectroscopy in immunoanalysis

Energy Technology Data Exchange (ETDEWEB)

Grubor, Nenad M. [Iowa State Univ., Ames, IA (United States)

2005-01-01

The work presented in this dissertation combines highly sensitive and selective fluorescence line-narrowing spectroscopy (FLNS) detection with various modes of immunoanalytical techniques. It has been shown that FLNS is capable of directly probing molecules immunocomplexed with antibodies, eliminating analytical ambiguities that may arise from interferences that accompany traditional immunochemical techniques. Moreover, the utilization of highly cross-reactive antibodies for highly specific analyte determination has been demonstrated. Finally, they demonstrate the first example of the spectral resolution of diastereomeric analytes based on their interaction with a cross-reactive antibody.

In situ detection of atomic and molecular iodine using Resonance and Off-Resonance Fluorescence by Lamp Excitation: ROFLEX

Directory of Open Access Journals (Sweden)

J. C. Gómez Martín

2011-01-01

Full Text Available We demonstrate a new instrument for in situ detection of atmospheric iodine atoms and molecules based on atomic and molecular resonance and off-resonance ultraviolet fluorescence excited by lamp emission. The instrument combines the robustness, light weight, low power consumption and efficient excitation of radio-frequency discharge light sources with the high sensitivity of the photon counting technique. Calibration of I₂ fluorescence is achieved via quantitative detection of the molecule by Incoherent Broad Band Cavity-enhanced Absorption Spectroscopy. Atomic iodine fluorescence signal is calibrated by controlled broad band photolysis of known I₂ concentrations in the visible spectral range at atmospheric pressure. The instrument has been optimised in laboratory experiments to reach detection limits of 1.2 pptv for I atoms and 13 pptv for I₂, for S/N = 1 and 10 min of integration time. The ROFLEX system has been deployed in a field campaign in northern Spain, representing the first concurrent observation of ambient mixing ratios of iodine atoms and molecules in the 1–350 pptv range.

Elastic Properties of Nucleic Acids by Single-Molecule Force Spectroscopy.

Science.gov (United States)

Camunas-Soler, Joan; Ribezzi-Crivellari, Marco; Ritort, Felix

2016-07-05

We review the current knowledge on the use of single-molecule force spectroscopy techniques to extrapolate the elastic properties of nucleic acids. We emphasize the lesser-known elastic properties of single-stranded DNA. We discuss the importance of accurately determining the elastic response in pulling experiments, and we review the simplest models used to rationalize the experimental data as well as the experimental approaches used to pull single-stranded DNA. Applications used to investigate DNA conformational transitions and secondary structure formation are also highlighted. Finally, we provide an overview of the effects of salt and temperature and briefly discuss the effects of contour length and sequence dependence.

Evaluation of portable Raman spectroscopy and handheld X-ray fluorescence analysis (hXRF) for the direct analysis of glyptics

Science.gov (United States)

Lauwers, D.; Candeias, A.; Coccato, A.; Mirao, J.; Moens, L.; Vandenabeele, P.

2016-03-01

In archaeometry, the advantages of a combined use of Raman spectroscopy and X-ray fluorescence spectroscopy are extensively discussed for applications such as the analysis of paintings, manuscripts, pottery, etc. Here, we demonstrate for the first time the advantage of using both techniques for analysing glyptics. These engraved gemstones or glass materials were originally used as stamps, to identify the owner, for instance on letters, but also on wine vessels. For this research, a set of 64 glyptics (42 Roman glass specimens and 22 modern ones), belonging to the collection of the museum 'Quinta das Cruzes' in Funchal (Madeira, Portugal), was analysed with portable Raman spectroscopy and handheld X-ray fluorescence (hXRF). These techniques were also used to confirm the gemological identification of these precious objects and can give extra information about the glass composition. Raman spectroscopy identifies the molecular composition as well as on the crystalline phases present. On the other hand, hXRF results show that the antique Roman glass samples are characterised with low Pb and Sn levels and that the modern specimens can be discriminated in two groups: lead-based and non-lead-based ones.

Artificial neural networks for processing fluorescence spectroscopy data in skin cancer diagnostics

International Nuclear Information System (INIS)

Lenhardt, L; Zeković, I; Dramićanin, T; Dramićanin, M D

2013-01-01

Over the years various optical spectroscopic techniques have been widely used as diagnostic tools in the discrimination of many types of malignant diseases. Recently, synchronous fluorescent spectroscopy (SFS) coupled with chemometrics has been applied in cancer diagnostics. The SFS method involves simultaneous scanning of both emission and excitation wavelengths while keeping the interval of wavelengths (constant-wavelength mode) or frequencies (constant-energy mode) between them constant. This method is fast, relatively inexpensive, sensitive and non-invasive. Total synchronous fluorescence spectra of normal skin, nevus and melanoma samples were used as input for training of artificial neural networks. Two different types of artificial neural networks were trained, the self-organizing map and the feed-forward neural network. Histopathology results of investigated skin samples were used as the gold standard for network output. Based on the obtained classification success rate of neural networks, we concluded that both networks provided high sensitivity with classification errors between 2 and 4%. (paper)

A dual-stimuli-responsive fluorescent switch ultrathin film

Science.gov (United States)

Li, Zhixiong; Liang, Ruizheng; Liu, Wendi; Yan, Dongpeng; Wei, Min

2015-10-01

Stimuli-responsive fluorescent switches have shown broad applications in optical devices, biological materials and intelligent responses. Herein, we describe the design and fabrication of a dual-stimuli-responsive fluorescent switch ultrathin film (UTF) via a three-step layer-by-layer (LBL) technique: (i) encapsulation of spiropyran (SP) within an amphiphilic block copolymer (PTBEM) to give the (SP@PTBEM) micelle; (ii) the mixture of riboflavin (Rf) and poly(styrene 4-sulfonate) (PSS) to enhance the adhesion ability of small molecules; (iii) assembly of negatively charged SP@PTBEM and Rf-PSS with cationic layered double hydroxide (LDH) nanoplatelets to obtain the (Rf-PSS/LDH/SP@PTBEM)n UTFs (n: bilayer number). The assembly process of the UTFs and their luminescence properties, as monitored by fluorescence spectroscopy and scanning electron microscopy (SEM), present a uniform and ordered layered structure with stepwise growth. The resulting Rf-PSS/LDH/SP@PTBEM UTF serves as a three-state switchable multicolor (green, yellow, and red) luminescent system based on stimulation from UV/Vis light and pH, with an acceptable reversibility. Therefore, this work provides a facile way to fabricate stimuli-responsive solid-state film switches with tunable-color luminescence, which have potential applications in the areas of displays, sensors, and rewritable optical memory and fluorescent logic devices.Stimuli-responsive fluorescent switches have shown broad applications in optical devices, biological materials and intelligent responses. Herein, we describe the design and fabrication of a dual-stimuli-responsive fluorescent switch ultrathin film (UTF) via a three-step layer-by-layer (LBL) technique: (i) encapsulation of spiropyran (SP) within an amphiphilic block copolymer (PTBEM) to give the (SP@PTBEM) micelle; (ii) the mixture of riboflavin (Rf) and poly(styrene 4-sulfonate) (PSS) to enhance the adhesion ability of small molecules; (iii) assembly of negatively charged SP

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

NARCIS (Netherlands)

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

2011-01-01

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

Surface characterization of selected polymer thin films by total-reflection x-ray fluorescence spectroscopy and x-ray reflectivity

International Nuclear Information System (INIS)

Innis, Vallerie Ann A.

2006-01-01

Development of available x-ray characterizations tools for grazing incidence techniques was done to be able to probe nano-size thin films. Alignment of a Philips x-ray powder diffractometer was improved to let it perform as an x-ray reflectometer. X-ray reflectometry was coupled with total-reflection x-ray fluorescence spectroscopy. Evaluation of the performance of this grazing incidence techniques was done by preparing polymer thin films of carboxymethylcellulose, carrageenan and polyvinylpyrrolidone (PVP). The thickness of the films were varied by varying the process parameters such as concentration, spin speed and spin time. Angle-dispersive total-reflection x-ray fluorescence spectroscopy profiles of three films showed film formation only in carrageenan and PVP. For both carrageenan and PVP, an increase in concentration yielded a corresponding increase in intensity of the fluorescent or scattered peaks. XRR profiles of carrageenan thin films yielded a mean value for the critical angle close to quartz substrate. Thickness measurements of the prepared carrageenan thin films showed that concentration was the main determinant for final film thickness over the other process parameters. Sulfur fluorescent intensity derived from the TXRF measurement showed a linear relationship with the measured thickness by XRR. For PVP, measured critical angle is lower than quartz. Poor adhesion of the polymer onto the substrate yielded a limited number of thickness measurements made from the XRR profiles. (Author)

Energy storage and redistribution in molecules

International Nuclear Information System (INIS)

Hinze, J.

1983-01-01

This book presents information on the following topics: chemistry and spectroscopy of molecules at high levels of excitation; energy and phase randomization in large molecules as probed by laser spectroscopy; intramolecular processes in isolated polyatomic molecules; pulse-probe measurements in low-temperature, low-pressure SF 6 ; the photodissociation dynamics of H 2 S and CF 3 NO; photofragment spectroscopy of the NO 2 dissociation; preparation, laser spectroscopy and predissociation of alkali dimers in supersonic nozzle beams; excited states of small molecules - collisional quenching and photodissociation; quantum-state-resolved scattering of lithium hydride; and molecular negative ions

Spectroscopic characterization of Venus at the single molecule level.

Science.gov (United States)

David, Charlotte C; Dedecker, Peter; De Cremer, Gert; Verstraeten, Natalie; Kint, Cyrielle; Michiels, Jan; Hofkens, Johan

2012-02-01

Venus is a recently developed, fast maturating, yellow fluorescent protein that has been used as a probe for in vivo applications. In the present work the photophysical characteristics of Venus were analyzed spectroscopically at the bulk and single molecule level. Through time-resolved single molecule measurements we found that single molecules of Venus display pronounced fluctuations in fluorescence emission, with clear fluorescence on- and off-times. These fluorescence intermittencies were found to occupy a broad range of time scales, ranging from milliseconds to several seconds. Such long off-times can complicate the analysis of single molecule counting experiments or single-molecule FRET experiments. This journal is © The Royal Society of Chemistry and Owner Societies 2012

Integrated Transmission Electron and Single-Molecule Fluorescence Microscopy Correlates Reactivity with Ultrastructure in a Single Catalyst Particle

OpenAIRE

Hendriks, Frank C.; Mohammadian, Sajjad; Ristanovic, Zoran; Kalirai, Samanbir; Meirer, Florian; Vogt, Eelco T. C.; Bruijnincx, Pieter C. A.; Gerritsen, Hans; Weckhuysen, Bert M.

2018-01-01

Establishing structure–activity relationships in complex, hierarchically structured nanomaterials, such as fluid catalytic cracking (FCC) catalysts, requires characterization with complementary, correlated analysis techniques. An integrated setup has been developed to perform transmission electron microscopy (TEM) and single-molecule fluorescence (SMF) microscopy on such nanostructured samples. Correlated structure–reactivity information was obtained for 100 nm thin, microtomed sections of a ...

On the performance of bioanalytical fluorescence correlation spectroscopy measurements in a multiparameter photon-counting microscope

Energy Technology Data Exchange (ETDEWEB)

Mazouchi, Amir; Liu Baoxu; Bahram, Abdullah [Department of Physics, Institute for Optical Sciences, University of Toronto, Toronto (Canada); Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Rd. N., Mississauga, ON, L5L 1C6 (Canada); Gradinaru, Claudiu C., E-mail: claudiu.gradinaru@utoronto.ca [Department of Physics, Institute for Optical Sciences, University of Toronto, Toronto (Canada); Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Rd. N., Mississauga, ON, L5L 1C6 (Canada)

2011-02-28

Fluorescence correlation spectroscopy (FCS) data acquisition and analysis routines were developed and implemented in a home-built, multiparameter photon-counting microscope. Laser excitation conditions were investigated for two representative fluorescent probes, Rhodamine110 and enhanced green fluorescent protein (EGFP). Reliable local concentrations and diffusion constants were obtained by fitting measured FCS curves, provided that the excitation intensity did not exceed 20% of the saturation level for each fluorophore. Accurate results were obtained from FCS measurements for sample concentrations varying from pM to {mu}M range, as well as for conditions of high background signals. These experimental constraints were found to be determined by characteristics of the detection system and by the saturation behavior of the fluorescent probes. These factors actually limit the average number of photons that can be collected from a single fluorophore passing through the detection volume. The versatility of our setup and the data analysis capabilities were tested by measuring the mobility of EGFP in the nucleus of Drosophila cells under conditions of high concentration and molecular crowding. As a bioanalytical application, we studied by FCS the binding affinity of a novel peptide-based drug to the cancer-regulating STAT3 protein and corroborated the results with fluorescence polarization analysis derived from the same photon data.

UV laser-induced fluorescence spectroscopy and laser Doppler flowmetry in the diagnostics of alopecia

Science.gov (United States)

Skomorokha, Diana P.; Pigoreva, Yulia N.; Salmin, Vladimir V.

2016-04-01

Development of optical biopsy methods has a great interest for medical diagnostics. In clinical and experimental studies it is very important to analyze blood circulation quickly and accurately, thereby laser Doppler flowmetry (LDF) is widely used. UV laser-induced fluorescence spectroscopy (UV LIFS) is express highly sensitive and widely-spread method with no destructive impact, high excitation selectivity and the possibility to use in highly scattering media. The goal of this work was to assess a correlation of UV laser-induced fluorescence spectroscopy and laser Doppler flowmetry parameters, and a possibility to identify or to differentiate various types of pathological changes in tissues according to their autofluorescence spectra. Three groups of patients with diffuse (symptomatic) alopecia, androgenic alopecia, and focal alopecia have been tested. Each groups consisted of not less than 20 persons. The measurements have been done in the parietal and occipital regions of the sculls. We used the original automated spectrofluorimeter to record autofluorescence spectra, and standard laser Doppler flowmeter BLF-21 (Transonic Systems, Inc., USA) to analyze the basal levels of blood circulation. Our results show that UV LIFS accurately distinguishes the zones with different types of alopecia. We found high correlation of the basal levels of blood circulation and the integrated intensity of autofluorescence in the affected tissue.

X-ray Fluorescence Spectroscopy of Pre-Federal American Currency

Science.gov (United States)

Raddell, Mark; Manukyan, Khachatur; Aprahamian, Ani; Wiescher, Michael; Jordan, Louis

2017-09-01

X-ray Fluorescence Spectroscopy (XRF) was used to study 17th and 18th century Mexican, Potosí, and Massachusetts silver colonial coins from the University of Notre Dame's Rare Books and Special Collections. Using different configurations and devices, we have learned more about the limitations and optimizations of the method. We have developed a moveable stand that may be used for XRF mapping of coin surfaces. We created standard silver alloy materials for quantification of the elemental composition of the coins. Inductively coupled plasma (ICP) spectroscopy was applied to determine the precise composition of the standards for accurate and non-destructive analyses of the colonial coins. XRF measurements were performed using two different XRF spectrometers, in both air and vacuum conditions, as well as an x-ray beam tube of varying diameters from 2 mm, 1 mm, and 0.03 mm. We quantified both the major elements and the bulk and surface impurities for 90 coins. We are using PCA to look at possible correlations between compositions of coinage from different geographical regions. Preliminary data analyses suggest that Massachusetts coins were minted using silver from Latin American sources. These results are of great interest to historians in tracing the origins of the currency. This work was made possible by the Notre Dame College of Science Summer Undergraduate Research Fellowships (COS-SURF).

Europium Uptake and Partitioning in Oat (Avena sativa) Roots as studied By Laser-Induced Fluorescence Spectroscopy and Confocal Microscopy Profiling Technique

International Nuclear Information System (INIS)

Fellows, Robert J.; Wang, Zheming; Ainsworth, Calvin C.

2003-01-01

The uptake of Eu3+ by elongating oat plant roots was studied by fluorescence spectroscopy, fluorescence lifetime measurement, as well as laser excitation time-resolved confocal fluorescence profiling technique. The results of this work indicated that the initial uptake of Eu(III) by oat root was most evident within the apical meristem of the root just proximal to the root cap. Distribution of assimilated Eu(III) within the roots differentiation and elongation zone was non-uniform. Higher concentrations were observed within the vascular cylinder, specifically in the phloem and developing xylem parenchyma. Elevated levels of the metal were also observed in the root hairs of the mature root. The concentration of assimilated Eu3+ dropped sharply from the apical meristem to the differentiation and elongation zone and then gradually decreased as the distance from the root cap increased. Fluorescence spectroscopic characteristics of the assimilated Eu3+ suggested that the Eu3+ exists a s inner-sphere mononuclear complexes inside the root. This work has also demonstrated the effectiveness of a time-resolved Eu3+ fluorescence spectroscopy and confocal fluorescence profiling techniques for the in vivo, real-time study of metal[Eu3+] accumulation by a functioning intact plant root. This approach can prove valuable for basic and applied studies in plant nutrition and environmental uptake of actinide radionuclides

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

A method for the measurement of in line pistachio aflatoxin concentration based on the laser induced fluorescence spectroscopy

International Nuclear Information System (INIS)

Paghaleh, Soodeh Jamali; Askari, Hassan Ranjbar; Marashi, Seyed Mohammad Bagher; Rahimi, Mojtaba; Bahrampour, Ali Reza

2015-01-01

Contamination of pistachio nuts with aflatoxin is one of the most significant issues related to pistachio health and expert. A fast pistachio aflatoxin concentration measurement method based on the laser induced fluorescence spectroscopy (LIFS) is proposed. The proposed method from theoretical and experimental points of view is analyzed. In our experiments XeCl Excimer laser is employed as an Ultra Violet (UV) source (λ=308 nm) and a UV–visible (UV–vis) spectrometer is used for fluorescent emission detection. Our setup is employed to measure the concentration of different type of Aflatoxins in pistachio nuts. Measurements results obtained by the LIFS method are compared with those are measured by the standard HPLC method. Aflatoxins concentrations are in good agreement with those are obtained by the HPLC method. The proposed laser induced fluorescence spectroscopy can be used as an in line aflatoxins concentrations measurement instrument for industrial applications. - Highlights: • XeCl Excimer laser is employed as an UV source for measurement of AFs in pistachio nuts. • Results are compared with those are measured by the standard HPLC method. • LIFS is an online AFs concentration measurement method for industrial applications

Analysis of functional organic molecules at noble metal surfaces by means of vibrational spectroscopies

Energy Technology Data Exchange (ETDEWEB)

Leyssner, Felix

2011-10-24

The goal of this work is to optimize the efficiency of photoinduced molecular switching processes on surfaces via controlled variations of the adsorption and electronic properties of the switch. We investigated the influence of external stimuli, i.e. photons and thermal activation, on surface bound molecular switches undergoing trans/cis-isomerizations and ring-opening/closing-reactions, respectively. High resolution electron energy loss spectroscopy (HREELS) and sum-frequency generation (SFG) spectroscopy have been used as the main tools to investigate the adsorption behavior and the molecular switching properties. Two basic concepts of coupling the molecular switch to the surface have been studied: (i) physisorbed or weakly chemisorbed systems deposited on noble metal surfaces under UHV conditions and (ii) molecular switches bound covalently via anchor groups. In the HREELS study following concept (i), we investigated the adsorption geometry and isomerization behavior of various molecular switches on metal substrates which are able to undergo a photoinduced trans/cis-isomerization in solution. We investigated three isoelectronic molecules on Au where we systematically changed the photochemically active group from the diazo-group in an azobenzene-derivative (on Cu(111)) to the imine-group, and the vinylene-group, respectively. Finding the photoisomerization quenched for all systems we observed considerable differences in their thermal isomerization behavior. Comparable we find the photoinduced ring-opening/closing-reaction of spiropyran quenched on Au(111) but a thermally induced ring-opening reaction resulting in the open form being strongly stabilized by the metal. SFG spectroscopy is employed to investigate the reversible, photoinduced trans/cis-isomerization of an azobenzene-functionalized self-assembled monolayer (SAM) on gold using a tripodal linker system. In consequence of the decoupling provided by the tripodal linker, the switching behavior of the

Tip-Enhanced Nano-Spectroscopy, Imaging, and Control: From Single Molecules to van der Waals Materials

Science.gov (United States)

Park, Kyoung-Duck

Photon-induced phenomena in molecules and other materials play a significant role in device applications as well as understanding their physical properties. While a range of device applications using organic and inorganic molecules and soft and hard materials have led striking developments in modern technologies, using bulk systems has reached the limit in their functions, performance, and regarding application range. Recently, low-dimensional systems have emerged as appealing resources for the advanced technologies based on their significantly improved functions and properties. Hence, understanding light-matter interactions at their natural length scale is of fundamental significance, in addition to the next generation device applications. This thesis demonstrates a range of new functions and behaviors of low-dimensional materials revealed and controlled by the advanced tip-enhanced near-field spectroscopy and imaging techniques exceeding the current instrumental limits. To understand the behaviors of zero-dimensional (0D) molecular systems in interacting environments, we explore new regimes in tip-enhanced Raman spectroscopy (TERS) and scanning near-field optical microscopy (SNOM), revealing the fundamental nature of single-molecule dynamics and nanoscale spatial heterogeneity of biomolecules on the cell membranes. To gain insight into intramolecular properties and dynamic processes of single molecules, we use TERS at cryogenic temperatures. From temperature-dependent line narrowing and splitting, we investigate and quantify ultrafast vibrational dephasing, intramolecular coupling, and conformational heterogeneity. Through correlation analysis of fluctuations of individual modes, we observe rotational motion and spectral fluctuations of single-molecule. We extend single-molecule spectroscopy study into in situ nano-biomolecular imaging of cancer cells by developing in-liquid SNOM. We use a new mechanical resonance control, achieving a high-Q force sensing of the

Optimization of fluorescent proteins

NARCIS (Netherlands)

Bindels, D.S.; Goedhart, J.; Hink, M.A.; van Weeren, L.; Joosen, L.; Gadella (jr.), T.W.J.; Engelborghs, Y.; Visser, A.J.W.G.

2014-01-01

Nowadays, fluorescent protein (FP) variants have been engineered to fluoresce in all different colors; to display photoswitchable, or photochromic, behavior; or to show yet other beneficial properties that enable or enhance a still growing set of new fluorescence spectroscopy and microcopy

The Single-Molecule Centroid Localization Algorithm Improves the Accuracy of Fluorescence Binding Assays.

Science.gov (United States)

Hua, Boyang; Wang, Yanbo; Park, Seongjin; Han, Kyu Young; Singh, Digvijay; Kim, Jin H; Cheng, Wei; Ha, Taekjip

2018-03-13

Here, we demonstrate that the use of the single-molecule centroid localization algorithm can improve the accuracy of fluorescence binding assays. Two major artifacts in this type of assay, i.e., nonspecific binding events and optically overlapping receptors, can be detected and corrected during analysis. The effectiveness of our method was confirmed by measuring two weak biomolecular interactions, the interaction between the B1 domain of streptococcal protein G and immunoglobulin G and the interaction between double-stranded DNA and the Cas9-RNA complex with limited sequence matches. This analysis routine requires little modification to common experimental protocols, making it readily applicable to existing data and future experiments.

Reaction mechanism studies of unsaturated molecules using photofragment translational spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Longfellow, C.A. [Univ. of California, Berkeley, CA (United States). Dept. of Chemistry]|[Lawrence Berkeley National Lab., CA (United States). Chemical Sciences Div.

1996-05-01

A number of molecules have been studied using the technique of photofragment translational spectroscopy. In Chapter One a brief introduction to the experimental technique is given. In Chapter Two the infrared multiphoton dissociation (IRMPD) of acetic acid is discussed. Carbon dioxide and methane were observed for the first time as products from dissociation under collisionless conditions. Chapter Three relates an IRMPD experiment of hexafluoropropene. The predominant channel produces CFCF{sub 3} or C{sub 2}F{sub 4} and CF{sub 2}, with the heavier species undergoing further dissociation to two CF{sub 2} fragments. In Chapter Four the ultraviolet (UV) dissociation of hexafluoropropene is investigated. Chapter Five explores the IRMPD of octafluoro-1-butene and octafluoro-2-butene.

Combining total internal reflection sum frequency spectroscopy spectral imaging and confocal fluorescence microscopy.

Science.gov (United States)

Allgeyer, Edward S; Sterling, Sarah M; Gunewardene, Mudalige S; Hess, Samuel T; Neivandt, David J; Mason, Michael D

2015-01-27

Understanding surface and interfacial lateral organization in material and biological systems is critical in nearly every field of science. The continued development of tools and techniques viable for elucidation of interfacial and surface information is therefore necessary to address new questions and further current investigations. Sum frequency spectroscopy (SFS) is a label-free, nonlinear optical technique with inherent surface specificity that can yield critical organizational information on interfacial species. Unfortunately, SFS provides no spatial information on a surface; small scale heterogeneities that may exist are averaged over the large areas typically probed. Over the past decade, this has begun to be addressed with the advent of SFS microscopy. Here we detail the construction and function of a total internal reflection (TIR) SFS spectral and confocal fluorescence imaging microscope directly amenable to surface investigations. This instrument combines, for the first time, sample scanning TIR-SFS imaging with confocal fluorescence microscopy.

Transition probability of the 5971-A line in neutral uranium from collision-induced fluorescence spectroscopy

International Nuclear Information System (INIS)

Gagne, J.M.; Mongeau, B.; Demers, Y.; Pianarosa, P.

1981-01-01

From collision-induced fluorescence spectroscopy measurements, we have determined the transition probability Aof the 5971-A transition in neutral uranium. Our value, A 5971 = (5.9 +- 1.8) x 10 5 sec -1 , is, within experimental error, in good agreement with the previous determination of Corliss, A 5971 = (7.3 +- 3.0) x 10 5 sec -1 [J. Res. Nat. Bur. Stand. Sect. A 80,1 (1976)

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

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'.

Study of the interaction of Tb (III) with dextran through fluorescence spectroscopy and optical rotatory dispersion

International Nuclear Information System (INIS)

Vasconcelos, Sandra S.; Rodrigues, J.F.

1984-01-01

A study of the interaction of Tb(III) with dextran in aqueous solution was perform using fluorescence spectroscopy and optical rotatory dispersion. The results indicate the formation of a complex with the displacent of water from the cation coordinated sphere by hydroxyl groups at the second and third carbon atoms of the monomer unit. (Author) [pt

Electronic structure of molecules of substituted benzenes by x-ray spectroscopy. I. Nitrobenzene

International Nuclear Information System (INIS)

Yumatov, V.D.; Murakhtanov, V.V.; Salakhutdinov, N.F.; Okotrub, A.V.; Mazalov, L.N.; Logunova, L.G.; Koptyug, V.A.; Furin, G.G.

1988-01-01

The electronic structure of the nitrobenzene molecule has been studied by x-ray spectroscopy with the aid of quantum-chemical calculations. The structure of the molecular orbitals of nitrobenzene has been compared with the structure of benzene and nitrogen dioxide. It has been shown in the framework of a fragment-by-fragment analysis that the interaction of the highest occupied π orbitals of the benzene ring and the nitro group is weak

Ninth international conference on hole burning, single molecule and related spectroscopies: science and applications (HBSM 2006)

International Nuclear Information System (INIS)

2006-01-01

This conference was organized around 9 sessions: -) single molecule, -) quantum optics, -) hole-burning materials and mechanisms, -) single nano-particle spectroscopy, -) dephasing and spectral diffusion, -) microwave photonics, -) biological systems, -) rare earth doped materials, -) novel laser sources. This document gathers only the slides of the presentations

Ninth international conference on hole burning, single molecule and related spectroscopies: science and applications (HBSM 2006)

Energy Technology Data Exchange (ETDEWEB)

NONE

2006-07-01

This conference was organized around 9 sessions: -) single molecule, -) quantum optics, -) hole-burning materials and mechanisms, -) single nano-particle spectroscopy, -) dephasing and spectral diffusion, -) microwave photonics, -) biological systems, -) rare earth doped materials, -) novel laser sources. This document gathers only the slides of the presentations.

Fluorescence quenching and photocatalytic degradation of textile dyeing waste water by silver nanoparticles

Science.gov (United States)

Kavitha, S. R.; Umadevi, M.; Janani, S. R.; Balakrishnan, T.; Ramanibai, R.

2014-06-01

Silver nanoparticles (Ag NPs) of different sizes have been prepared by chemical reduction method and characterized using UV-vis spectroscopy and transmission electron microscopy (HRTEM). Fluorescence spectral analysis showed that the quenching of fluorescence of textile dyeing waste water (TDW) has been found to decrease with decrease in the size of the Ag NPs. Experimental results show that the silver nanoparticles can quench the fluorescence emission of adsorbed TDW effectively. The fluorescence interaction between Ag NPs (acceptor) and TDW (donor) confirms the Förster Resonance Energy Transfer (FRET) mechanism. Long range dipole-dipole interaction between the excited donor and ground state acceptor molecules is the dominant mechanism responsible for the energy transfer. Furthermore, photocatalytic degradation of TDW was measured spectrophotometrically by using silver as nanocatalyst under UV light illumination. The kinetic study revealed that synthesized Ag NPs was found to be effective in degrading TDW.

Investigating temperature effects on extra virgin olive oil using fluorescence spectroscopy

Science.gov (United States)

Saleem, M.; Ahmad, Naveed; Ali, H.; Bilal, M.; Khan, Saranjam; Ullah, Rahat; Ahmed, M.; Mahmood, S.

2017-12-01

The potential of fluorescence spectroscopy has been utilized to study the heating effects on extra virgin olive oil (EVOO). Through a series of experiments, a temperature range from 140 °C  -  150 °C has been found where cooking with EVOO is possible without destroying its natural ingredients. Fluorescence emission spectra from all heated and non-heated EVOO samples were recorded using an excitation source at 350 nm, where emission bands in non-heated EVOO at 380, 440, 455, and 525 nm are labelled for vitamin E and a band at 673 nm is assigned for chlorophyll a. The emission band at 525 nm is also responsible for beta carotenoids (vitamin A). As a result of heating, prominent intensity variations have been observed in all spectral bands, but it is particularly affected at 525 nm, indicating the deterioration of vitamin E and beta carotenoids. However, if the temperature of oil can be maintained in the above defined range, then frying food with EVOO is possible by preserving its natural ingredients. The spectral variations resulting from the heating effects have been further highlighted by using principal component analysis for classification purposes.

Fluorescence spectral properties of outer antenna LHC II

CERN Document Server

He Jun Fang; Zhang, Shu; He Fang Tao; Ren Zhao You; Li Liang Bi; Kuang Ting Yun

2002-01-01

Outer antenna LHC II acts to absorb and transfer energy for photosynthesis. The authors studied the fluorescence properties of LHC II of spinach with scanning imaging fluorescence spectroscopy. After it had been excited by 514.5 nm laser, the integral fluorescence spectrum of LHC II was detected. It was shown that energy transfer existed between carotenoid and chlorophyll. Seven bands of LHC II fluorescence emission were resolved by Gauss combination, viz. 656.7, 664.6, 671.5, 677.2, 683.5, 689.6, 695.3 nm, and the percentages of them were 3.0%, 13.1%,13.3%, 21.1%, 13.2%, 33.3%, 3.0% respectively. The emission of 658.7 nm was attributed to chlorophyll b, the other emission bands were produced by chlorophyll a molecules with the maximum absorption 662, 670/671, 676, 680 nm and over 690 nm. The band 656.7 nm, whose percentage was 3.0%, shows that the most energy was absorbed by chlorophyll a. The percentage of band 689.6 nm was the most, which was possibly correlated with one type of self protective mechanism o...