Shifts in the fluorescence lifetime of EGFP during bacterial phagocytosis measured by phase-sensitive flow cytometry

NASA Astrophysics Data System (ADS)

Li, Wenyan; Houston, Kevin D.; Houston, Jessica P.

2017-01-01

Phase-sensitive flow cytometry (PSFC) is a technique in which fluorescence excited state decay times are measured as fluorescently labeled cells rapidly transit a finely focused, frequency-modulated laser beam. With PSFC the fluorescence lifetime is taken as a cytometric parameter to differentiate intracellular events that are challenging to distinguish with standard flow cytometry. For example PSFC can report changes in protein conformation, expression, interactions, and movement, as well as differences in intracellular microenvironments. This contribution focuses on the latter case by taking PSFC measurements of macrophage cells when inoculated with enhanced green fluorescent protein (EGFP)-expressing E. coli. During progressive internalization of EGFP-E. coli, fluorescence lifetimes were acquired and compared to control groups. It was hypothesized that fluorescence lifetimes would correlate well with phagocytosis because phagosomes become acidified and the average fluorescence lifetime of EGFP is known to be affected by pH. We confirmed that average EGFP lifetimes consistently decreased (3 to 2 ns) with inoculation time. The broad significance of this work is the demonstration of how high-throughput fluorescence lifetime measurements correlate well to changes that are not easily tracked by intensity-only cytometry, which is affected by heterogeneous protein expression, cell-to-cell differences in phagosome formation, and number of bacterium engulfed.

Nanostructures Derived from Starch and Chitosan for Fluorescence Bio-Imaging

PubMed Central

Zu, Yinxue; Bi, Jingran; Yan, Huiping; Wang, Haitao; Song, Yukun; Zhu, Bei-Wei; Tan, Mingqian

2016-01-01

Fluorescent nanostructures (NSs) derived from polysaccharides have drawn great attention as novel fluorescent probes for potential bio-imaging applications. Herein, we reported a facile alkali-assisted hydrothermal method to fabricate polysaccharide NSs using starch and chitosan as raw materials. Transmission electron microscopy (TEM) demonstrated that the average particle sizes are 14 nm and 75 nm for starch and chitosan NSs, respectively. Fourier transform infrared (FT-IR) spectroscopy analysis showed that there are a large number of hydroxyl or amino groups on the surface of these polysaccharide-based NSs. Strong fluorescence with an excitation-dependent emission behaviour was observed under ultraviolet excitation. Interestingly, the photostability of the NSs was found to be superior to fluorescein and rhodamine B. The quantum yield of starch NSs could reach 11.12% under the excitation of 360 nm. The oxidative metal ions including Cu(II), Hg(II)and Fe(III) exhibited a quench effect on the fluorescence intensity of the prepared NSs. Both of the two kinds of the multicoloured NSs showed a maximum fluorescence intensity at pH 7, while the fluorescence intensity decreased dramatically when they were put in an either acidic or basic environment (at pH 3 or 11). The cytotoxicity study of starch NSs showed that low cell cytotoxicity and 80% viability was found after 24 h incubation, when their concentration was less than 10 mg/mL. The study also showed the possibility of using the multicoloured starch NSs for mouse melanoma cells and guppy fish imaging. PMID:28335258

Calibration of Wide-Field Deconvolution Microscopy for Quantitative Fluorescence Imaging

PubMed Central

Lee, Ji-Sook; Wee, Tse-Luen (Erika); Brown, Claire M.

2014-01-01

Deconvolution enhances contrast in fluorescence microscopy images, especially in low-contrast, high-background wide-field microscope images, improving characterization of features within the sample. Deconvolution can also be combined with other imaging modalities, such as confocal microscopy, and most software programs seek to improve resolution as well as contrast. Quantitative image analyses require instrument calibration and with deconvolution, necessitate that this process itself preserves the relative quantitative relationships between fluorescence intensities. To ensure that the quantitative nature of the data remains unaltered, deconvolution algorithms need to be tested thoroughly. This study investigated whether the deconvolution algorithms in AutoQuant X3 preserve relative quantitative intensity data. InSpeck Green calibration microspheres were prepared for imaging, z-stacks were collected using a wide-field microscope, and the images were deconvolved using the iterative deconvolution algorithms with default settings. Afterwards, the mean intensities and volumes of microspheres in the original and the deconvolved images were measured. Deconvolved data sets showed higher average microsphere intensities and smaller volumes than the original wide-field data sets. In original and deconvolved data sets, intensity means showed linear relationships with the relative microsphere intensities given by the manufacturer. Importantly, upon normalization, the trend lines were found to have similar slopes. In original and deconvolved images, the volumes of the microspheres were quite uniform for all relative microsphere intensities. We were able to show that AutoQuant X3 deconvolution software data are quantitative. In general, the protocol presented can be used to calibrate any fluorescence microscope or image processing and analysis procedure. PMID:24688321

Fluorescence lifetime imaging ophthalmoscopy.

PubMed

Dysli, Chantal; Wolf, Sebastian; Berezin, Mikhail Y; Sauer, Lydia; Hammer, Martin; Zinkernagel, Martin S

2017-09-01

Imaging techniques based on retinal autofluorescence have found broad applications in ophthalmology because they are extremely sensitive and noninvasive. Conventional fundus autofluorescence imaging measures fluorescence intensity of endogenous retinal fluorophores. It mainly derives its signal from lipofuscin at the level of the retinal pigment epithelium. Fundus autofluorescence, however, can not only be characterized by the spatial distribution of the fluorescence intensity or emission spectrum, but also by a characteristic fluorescence lifetime function. The fluorescence lifetime is the average amount of time a fluorophore remains in the excited state following excitation. Fluorescence lifetime imaging ophthalmoscopy (FLIO) is an emerging imaging modality for in vivo measurement of lifetimes of endogenous retinal fluorophores. Recent reports in this field have contributed to our understanding of the pathophysiology of various macular and retinal diseases. Within this review, the basic concept of fluorescence lifetime imaging is provided. It includes technical background information and correlation with in vitro measurements of individual retinal metabolites. In a second part, clinical applications of fluorescence lifetime imaging and fluorescence lifetime features of selected retinal diseases such as Stargardt disease, age-related macular degeneration, choroideremia, central serous chorioretinopathy, macular holes, diabetic retinopathy, and retinal artery occlusion are discussed. Potential areas of use for fluorescence lifetime imaging ophthalmoscopy will be outlined at the end of this review. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Fluorescence correlation spectroscopy: principles and applications.

PubMed

Bacia, Kirsten; Haustein, Elke; Schwille, Petra

2014-07-01

Fluorescence correlation spectroscopy (FCS) is used to study the movements and the interactions of biomolecules at extremely dilute concentrations, yielding results with good spatial and temporal resolutions. Using a number of technical developments, FCS has become a versatile technique that can be used to study a variety of sample types and can be advantageously combined with other methods. Unlike other fluorescence-based techniques, the analysis of FCS data is not based on the average intensity of the fluorescence emission but examines the minute intensity fluctuations caused by spontaneous deviations from the mean at thermal equilibrium. These fluctuations can result from variations in local concentrations owing to molecular mobility or from characteristic intermolecular or intramolecular reactions of fluorescently labeled biomolecules present at low concentrations. Here, we provide a basic introduction to FCS, including its technical development and theoretical basis, experimental setup of an FCS system, adjustment of a setup, data acquisition, and analysis of FCS measurements. Finally, the application of FCS to the study of lipid bilayer membranes and to living cells is discussed. © 2014 Cold Spring Harbor Laboratory Press.

Optical visualisation of thermogenesis in stimulated single-cell brown adipocytes.

PubMed

Kriszt, Rókus; Arai, Satoshi; Itoh, Hideki; Lee, Michelle H; Goralczyk, Anna G; Ang, Xiu Min; Cypess, Aaron M; White, Andrew P; Shamsi, Farnaz; Xue, Ruidan; Lee, Jung Yeol; Lee, Sung-Chan; Hou, Yanyan; Kitaguchi, Tetsuya; Sudhaharan, Thankiah; Ishiwata, Shin'ichi; Lane, E Birgitte; Chang, Young-Tae; Tseng, Yu-Hua; Suzuki, Madoka; Raghunath, Michael

2017-05-03

The identification of brown adipose deposits in adults has led to significant interest in targeting this metabolically active tissue for treatment of obesity and diabetes. Improved methods for the direct measurement of heat production as the signature function of brown adipocytes (BAs), particularly at the single cell level, would be of substantial benefit to these ongoing efforts. Here, we report the first application of a small molecule-type thermosensitive fluorescent dye, ERthermAC, to monitor thermogenesis in BAs derived from murine brown fat precursors and in human brown fat cells differentiated from human neck brown preadipocytes. ERthermAC accumulated in the endoplasmic reticulum of BAs and displayed a marked change in fluorescence intensity in response to adrenergic stimulation of cells, which corresponded to temperature change. ERthermAC fluorescence intensity profiles were congruent with mitochondrial depolarisation events visualised by the JC-1 probe. Moreover, the averaged fluorescence intensity changes across a population of cells correlated well with dynamic changes such as thermal power, oxygen consumption, and extracellular acidification rates. These findings suggest ERthermAC as a promising new tool for studying thermogenic function in brown adipocytes of both murine and human origins.

Correlation of conformational heterogeneity of the tryptophyl side chain and time-resolved fluorescence intensity decay kinetics

NASA Astrophysics Data System (ADS)

Laws, William R.; Ross, J. B. Alexander

1992-04-01

The time-resolved fluorescence properties of a tryptophan residue should be useful for probing protein structure, function, and dynamics. To date, however, the non-single exponential fluorescence intensity decay kinetics for numerous peptides and proteins having a single tryptophan residue have not been adequately explained. Many possibilities have been considered and include: (1) contributions from the 1La and 1Lb states of indole; (2) excited-state hydrogen exchange; and (3) environmental heterogeneity from (chi) 1 and (chi) 2 rotamers. In addition, it has been suggested that generally many factors contribute to the decay and a distribution of probabilities may be more appropriate. Two recent results support multiple species due to conformational heterogeneity as the major contributor to complex kinetics. First, a rotationally constrained tryptophan analogue has fluorescence intensity decay kinetics that can be described by the sum of two exponentials with amplitudes comparable to the relative populations of the two rotational isomers. Second, the multiple exponentials observed for tyrosine-containing model compounds and peptides correlate with the (chi) 1 rotamer populations independently determined by 1H NMR. We now report similar correlations between rotamer populations and fluorescence intensity decay kinetics for a tryptophan analogue of oxytocin. It appears for this compound that either (chi) 2 rotations do not appreciably alter the indole environment, (chi) 2 rotations are rapid enough to average the observed dependence, or only one of two possible (chi) 2 populations is associated with each (chi) 1 rotamer.

Subnanosecond spectrofluorimetry of new indolocarbazole derivatives in solutions and protein complexes and their dipole moments

NASA Astrophysics Data System (ADS)

Nemkovich, N. A.; Kruchenok, Yu. V.; Sobchuk, A. N.; Detert, H.; Wrobel, N.; Chernyavskiĭ, E. A.

2009-08-01

The spectral and temporal characteristics of new 6,12-dimethoxyindolo[3,2- b]carbazole, 5,11-dimethyl-6,12-dimethoxyindolo[3,2- b]carbazole, and 5,11-dihexyl-6,12-di(hexyloxy)indolo[3,2- b]carbazole fluorescence probes in organic solvents and protein complexes are studied. The dipole moments of indolocarbazoles in 1,4-dioxane were measured by electrooptical absorption method. The measured dipole moments have values within the range of (3.1-3.6) × 10-30 C m in the equilibrium ground state and increase to (4.8-5.6) × 10-30 C m after excitation. The excited state lifetime of indolocarbazole derivatives increases with increasing polarity and viscosity of the environment. The binding of indolocarbazoles with trypsinogen and human serum albumin increases the fluorescence intensity, changes the intensity ratio of fluorescence bands, and increases the average excited state lifetime of indolocarbazoles. The analysis of the instantaneous fluorescence spectra and fluorescence decay parameters at different wavelengths revealed the existence of several types of probe binding sites in proteins. It is found that the fluorescence characteristics of indolocarbazole derivatives depend on the conformation rearrangements of trypsinogen due to its thermal denaturation.

Quantitative analysis of comparative genomic hybridization

DOE Office of Scientific and Technical Information (OSTI.GOV)

Manoir, S. du; Bentz, M.; Joos, S.

1995-01-01

Comparative genomic hybridization (CGH) is a new molecular cytogenetic method for the detection of chromosomal imbalances. Following cohybridization of DNA prepared from a sample to be studied and control DNA to normal metaphase spreads, probes are detected via different fluorochromes. The ratio of the test and control fluorescence intensities along a chromosome reflects the relative copy number of segments of a chromosome in the test genome. Quantitative evaluation of CGH experiments is required for the determination of low copy changes, e.g., monosomy or trisomy, and for the definition of the breakpoints involved in unbalanced rearrangements. In this study, a programmore » for quantitation of CGH preparations is presented. This program is based on the extraction of the fluorescence ratio profile along each chromosome, followed by averaging of individual profiles from several metaphase spreads. Objective parameters critical for quantitative evaluations were tested, and the criteria for selection of suitable CGH preparations are described. The granularity of the chromosome painting and the regional inhomogeneity of fluorescence intensities in metaphase spreads proved to be crucial parameters. The coefficient of variation of the ratio value for chromosomes in balanced state (CVBS) provides a general quality criterion for CGH experiments. Different cutoff levels (thresholds) of average fluorescence ratio values were compared for their specificity and sensitivity with regard to the detection of chromosomal imbalances. 27 refs., 15 figs., 1 tab.« less

Micro-Droplet Detection Method for Measuring the Concentration of Alkaline Phosphatase-Labeled Nanoparticles in Fluorescence Microscopy

PubMed Central

Li, Rufeng; Wang, Yibei; Xu, Hong; Fei, Baowei; Qin, Binjie

2017-01-01

This paper developed and evaluated a quantitative image analysis method to measure the concentration of the nanoparticles on which alkaline phosphatase (AP) was immobilized. These AP-labeled nanoparticles are widely used as signal markers for tagging biomolecules at nanometer and sub-nanometer scales. The AP-labeled nanoparticle concentration measurement can then be directly used to quantitatively analyze the biomolecular concentration. Micro-droplets are mono-dispersed micro-reactors that can be used to encapsulate and detect AP-labeled nanoparticles. Micro-droplets include both empty micro-droplets and fluorescent micro-droplets, while fluorescent micro-droplets are generated from the fluorescence reaction between the APs adhering to a single nanoparticle and corresponding fluorogenic substrates within droplets. By detecting micro-droplets and calculating the proportion of fluorescent micro-droplets to the overall micro-droplets, we can calculate the AP-labeled nanoparticle concentration. The proposed micro-droplet detection method includes the following steps: (1) Gaussian filtering to remove the noise of overall fluorescent targets, (2) a contrast-limited, adaptive histogram equalization processing to enhance the contrast of weakly luminescent micro-droplets, (3) an red maximizing inter-class variance thresholding method (OTSU) to segment the enhanced image for getting the binary map of the overall micro-droplets, (4) a circular Hough transform (CHT) method to detect overall micro-droplets and (5) an intensity-mean-based thresholding segmentation method to extract the fluorescent micro-droplets. The experimental results of fluorescent micro-droplet images show that the average accuracy of our micro-droplet detection method is 0.9586; the average true positive rate is 0.9502; and the average false positive rate is 0.0073. The detection method can be successfully applied to measure AP-labeled nanoparticle concentration in fluorescence microscopy. PMID:29160812

Micro-Droplet Detection Method for Measuring the Concentration of Alkaline Phosphatase-Labeled Nanoparticles in Fluorescence Microscopy.

PubMed

Li, Rufeng; Wang, Yibei; Xu, Hong; Fei, Baowei; Qin, Binjie

2017-11-21

This paper developed and evaluated a quantitative image analysis method to measure the concentration of the nanoparticles on which alkaline phosphatase (AP) was immobilized. These AP-labeled nanoparticles are widely used as signal markers for tagging biomolecules at nanometer and sub-nanometer scales. The AP-labeled nanoparticle concentration measurement can then be directly used to quantitatively analyze the biomolecular concentration. Micro-droplets are mono-dispersed micro-reactors that can be used to encapsulate and detect AP-labeled nanoparticles. Micro-droplets include both empty micro-droplets and fluorescent micro-droplets, while fluorescent micro-droplets are generated from the fluorescence reaction between the APs adhering to a single nanoparticle and corresponding fluorogenic substrates within droplets. By detecting micro-droplets and calculating the proportion of fluorescent micro-droplets to the overall micro-droplets, we can calculate the AP-labeled nanoparticle concentration. The proposed micro-droplet detection method includes the following steps: (1) Gaussian filtering to remove the noise of overall fluorescent targets, (2) a contrast-limited, adaptive histogram equalization processing to enhance the contrast of weakly luminescent micro-droplets, (3) an red maximizing inter-class variance thresholding method (OTSU) to segment the enhanced image for getting the binary map of the overall micro-droplets, (4) a circular Hough transform (CHT) method to detect overall micro-droplets and (5) an intensity-mean-based thresholding segmentation method to extract the fluorescent micro-droplets. The experimental results of fluorescent micro-droplet images show that the average accuracy of our micro-droplet detection method is 0.9586; the average true positive rate is 0.9502; and the average false positive rate is 0.0073. The detection method can be successfully applied to measure AP-labeled nanoparticle concentration in fluorescence microscopy.

Time-resolved laser-induced fluorescence spectroscopy as a diagnostic instrument in head and neck carcinoma.

PubMed

Meier, Jeremy D; Xie, Hongtao; Sun, Yang; Sun, Yinghua; Hatami, Nisa; Poirier, Brian; Marcu, Laura; Farwell, D Gregory

2010-06-01

The objectives of this study were to 1) determine differences in lifetime fluorescence between normal and malignant tissue of the upper aerodigestive tract, and 2) evaluate the potential of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) as a diagnostic instrument for head and neck squamous cell carcinoma (HNSCC). Cross-sectional study. University-based medical center. Nine patients with suspected HNSCC were included. In the operating room, a nitrogen pulse laser (337 nm, 700-picosecond pulse width) was used to induce tissue autofluorescence of normal tissue and suspected malignant lesions. Spectral intensities and time-domain measurements were obtained and compared with the histopathology at each site. A total of 53 sites were measured. The fluorescence parameters that provided the most discrimination were determined. Differences in spectral intensities allowed for discrimination between malignant and normal tissue. The spectral intensity of malignant tissue was lower than that of normal tissue, and a shift of peak intensity to a longer wavelength was observed in the normalized spectrum of malignant tissue in the range of 360 to approximately 660 nm. Multiple time-resolved fluorescence parameters provided the best diagnostic discrimination between normal tissue and carcinoma, including average lifetimes (i.e., at 390 nm: 1.7 +/- 0.06 ns [not significant] for normal and 1.3 +/- 0.06 ns for tumor, P = 0.0025) and the second-order Laguerre expansion coefficient (LEC-2) (i.e., at 460 nm: 0.135 +/- 0.001 for normal and 0.155 +/- 0.007 for tumor, P < 0.05). These findings highlight some of the differences in lifetime fluorescence between normal and malignant tissue. TR-LIFS has potential as a noninvasive diagnostic technique for HNSCC. Copyright 2010 American Academy of Otolaryngology-Head and Neck Surgery Foundation. Published by Mosby, Inc. All rights reserved.

Time-resolved laser-induced fluorescence spectroscopy as a diagnostic instrument in head and neck carcinoma

PubMed Central

Meier, Jeremy D.; Xie, Hongtao; Sun, Yang; Sun, Yinghua; Hatami, Nisa; Poirier, Brian; Marcu, Laura; Farwell, D. Gregory

2011-01-01

OBJECTIVE 1) Determine differences in lifetime fluorescence between normal and malignant tissue of the upper aerodigestive tract. 2) Evaluate the potential of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) as a diagnostic instrument for head and neck squamous cell carcinoma (HNSCC). STUDY DESIGN Cross-sectional study. SETTING University-based medical center. SUBJECTS AND METHODS Nine patients with suspected HNSCC were included. In the operating room, a nitrogen pulse laser (337 nm, 700 ps pulse width) was used to induce tissue autofluorescence of normal tissue and suspected malignant lesions. Spectral intensities and time-domain measurements were obtained and compared to the histopathology at each site. A total of 53 sites were measured. The fluorescence parameters that provided the most discrimination were determined. RESULTS Differences in spectral intensities allowed for discrimination between malignant and normal tissue. The spectral intensity of malignant tissue was lower than the normal tissue, and a shift of peak intensity to a longer wavelength was observed in the normalized spectrum of malignant tissue in the range of 360~660 nm. Multiple time-resolved fluorescence parameters provided the best diagnostic discrimination between normal tissue and carcinoma, including average lifetimes (i.e., at 390 nm: 1.7±0.06 ns for normal and 1.3±0.06 ns for tumor, P=0.0025), and the Laguerre coefficients, LEC-2 (i.e., at 460 nm: 0.135±0.001 for normal and 0.155±0.007 for tumor, P<0.05). CONCLUSION These findings highlight some of the differences in lifetime fluorescence between normal and malignant tissue. TR-LIFS has potential as a non-invasive diagnostic technique for HNSCC. PMID:20493355

Fluorescence and fluorescence-lifetime imaging microscopy (FLIM) to characterize yeast strains by autofluorescence

NASA Astrophysics Data System (ADS)

Bhatta, H.; Goldys, E. M.; Ma, J.

2006-02-01

We characterised populations of wild type baking and brewing yeast cells using intrinsic fluorescence and fluorescence lifetime microscopy, in order to obtain quantitative identifiers of different strains. The cell autofluorescence was excited at 405 nm and observed within 440-540 nm range where strong cell to cell variability was observed. The images were analyzed using customised public domain software, which provided information on cell size, intensity and texture-related features. In light of significant diversity of the data, statistical methods were utilized to assess the validity of the proposed quantitative identifiers for strain differentiation. The Kolmogorov-Smirnov test was applied to confirm that empirical distribution functions for size, intensity and entropy for different strains were statistically different. These characteristics were followed with culture age of 24, 48 and 72 h, (the latter corresponding to a stationary growth phase) and size, and to some extent entropy, were found to be independent of age. The fluorescence intensity presented a distinctive evolution with age, different for each of the examined strains. The lifetime analysis revealed a short decay time component of 1.4 ns and a second, longer one with the average value of 3.5 ns and a broad distribution. High variability of lifetime values within cells was observed however a lifetime texture feature in the studied strains was statistically different.

Temperature dependence of metal-enhanced fluorescence of photosystem I from Thermosynechococcus elongatus.

PubMed

Ashraf, Imran; Konrad, Alexander; Lokstein, Heiko; Skandary, Sepideh; Metzger, Michael; Djouda, Joseph M; Maurer, Thomas; Adam, Pierre M; Meixner, Alfred J; Brecht, Marc

2017-03-23

We report the temperature dependence of metal-enhanced fluorescence (MEF) of individual photosystem I (PSI) complexes from Thermosynechococcus elongatus (T. elongatus) coupled to gold nanoparticles (AuNPs). A strong temperature dependence of shape and intensity of the emission spectra is observed when PSI is coupled to AuNPs. For each temperature, the enhancement factor (EF) is calculated by comparing the intensity of individual AuNP-coupled PSI to the mean intensity of 'uncoupled' PSI. At cryogenic temperature (1.6 K) the average EF was 4.3-fold. Upon increasing the temperature to 250 K the EF increases to 84-fold. Single complexes show even higher EFs up to 441.0-fold. At increasing temperatures the different spectral pools of PSI from T. elongatus become distinguishable. These pools are affected differently by the plasmonic interactions and show different enhancements. The remarkable increase of the EFs is explained by a rate model including the temperature dependence of the fluorescence yield of PSI and the spectral overlap between absorption and emission spectra of AuNPs and PSI, respectively.

Application of novel low-intensity nonscanning fluorescence lifetime imaging microscopy for monitoring excited state dynamics in individual chloroplasts and living cells of photosynthetic organisms

NASA Astrophysics Data System (ADS)

Eckert, Hann-Jörg; Petrášek, Zdeněk; Kemnitz, Klaus

2006-10-01

Picosecond fluorescence lifetime imaging microscopy (FLIM) provides a most valuable tool to analyze the primary processes of photosynthesis in individual cells and chloroplasts of living cells. In order to obtain correct lifetimes of the excited states, the peak intensity of the exciting laser pulses as well as the average intensity has to be sufficiently low to avoid distortions of the kinetics by processes such as singlet-singlet annihilation, closing of the reaction centers or photoinhibition. In the present study this requirement is achieved by non-scanning wide-field FLIM based on time- and space-correlated single-photon counting (TSCSPC) using a novel microchannel plate photomultiplier with quadrant anode (QA-MCP) that allows parallel acquisition of time-resolved images under minimally invasive low-excitation conditions. The potential of the wide-field TCSPC method is demonstrated by presenting results obtained from measurements of the fluorescence dynamics in individual chloroplasts of moss leaves and living cells of the chlorophyll d-containing cyanobacterium Acaryochloris marina.

Enhanced Fluorescence Emission of Me-ADOTA+ by Self-Assembled Silver Nanoparticles on a Gold Film

PubMed Central

Sørensen, Thomas J.; Laursen, Bo W.; Luchowski, Rafal; Shtoyko, Tanya; Akopova, Irina; Gryczynski, Zygmunt; Gryczynski, Ignacy

2009-01-01

We report a multi-fold enhancement of the fluorescence of methyl-azadioxatriangulenium chloride (Me-ADOTA•Cl) in PVA deposited on a 50 nm thick gold mirror carrying an evaporation induced self-assembly of colloidal silver nanoparticles (Ag-SACs). The average measured increase in fluorescence emission of about 50-fold is accompanied by hot spots with a local enhancement in brigthness close to 200. The long lifetime of the dye allows for the first direct determination of the correlation between the enhancement of emission intensity and the decrease in fluorescence lifetime. The Ag-SACs surface preparation and observed enhancements are highly reproducible. We believe that these robust plasmonic surfaces will find use in sensing platforms for ultrasensitive detection. PMID:20161182

Two-color fluorescence analysis of individual virions determines the distribution of the copy number of proteins in herpes simplex virus particles.

PubMed

Clarke, Richard W; Monnier, Nilah; Li, Haitao; Zhou, Dejian; Browne, Helena; Klenerman, David

2007-08-15

We present a single virion method to determine absolute distributions of copy number in the protein composition of viruses and apply it to herpes simplex virus type 1. Using two-color coincidence fluorescence spectroscopy, we determine the virion-to-virion variability in copy numbers of fluorescently labeled tegument and envelope proteins relative to a capsid protein by analyzing fluorescence intensity ratios for ensembles of individual dual-labeled virions and fitting the resulting histogram of ratios. Using EYFP-tagged capsid protein VP26 as a reference for fluorescence intensity, we are able to calculate the mean and also, for the first time to our knowledge, the variation in numbers of gD, VP16, and VP22 tegument. The measurement of the number of glycoprotein D molecules was in good agreement with independent measurements of average numbers of these glycoproteins in bulk virus preparations, validating the method. The accuracy, straightforward data processing, and high throughput of this technique make it widely applicable to the analysis of the molecular composition of large complexes in general, and it is particularly suited to providing insights into virus structure, assembly, and infectivity.

Characterization of chromophoric dissolved organic matter (CDOM) in rainwater using fluorescence spectrophotometry.

PubMed

Salve, P R; Lohkare, H; Gobre, T; Bodhe, G; Krupadam, R J; Ramteke, D S; Wate, S R

2012-02-01

The fluorescence excitation-emission matrix of Chromophoric dissolved organic matter (CDOM) samples from rainwater collected at Rameswaram, Tamilnadu, India are analysed. Total five peaks were observed for humic/marine and protein likes substances respectively. The peak A and C intensities varies form 1.98 ± 0.28 and 0.97 ± 0.11 QSU respectively represents humic like substances. The peak B and T intensities varies from 3.94 ± 0.75 and 7.42 ± 1.43 QSU showed association of protein like substances whereas peak M intensities varies from 1.92 ± 0.37 QSU indicates marine contribution. Among the fluorophores, the following sequence were observed as T > B > A > M > C which indicates dominance of Tryptophan like substances in rainwater. The average peak T/C ratios was observed as 7.88 ± 2.2 indicates microbial contamination by Tryptophan-like substances with the high biological activity and low volatility.

Photophysical Behaviors of Single Fluorophores Localized on Zinc Oxide Nanostructures

PubMed Central

Fu, Yi; Zhang, Jian; Lakowicz, Joseph R.

2012-01-01

Single-molecule fluorescence spectroscopy has now been widely used to investigate complex dynamic processes which would normally be obscured in an ensemble-averaged measurement. In this report we studied photophysical behaviors of single fluorophores in proximity to zinc oxide nanostructures by single-molecule fluorescence spectroscopy and time-correlated single-photon counting (TCSPC). Single fluorophores on ZnO surfaces showed enhanced fluorescence brightness to various extents compared with those on glass; the single-molecule time trajectories also illustrated pronounced fluctuations of emission intensities, with time periods distributed from milliseconds to seconds. We attribute fluorescence fluctuations to the interfacial electron transfer (ET) events. The fluorescence fluctuation dynamics were found to be inhomogeneous from molecule to molecule and from time to time, showing significant static and dynamic disorders in the interfacial electron transfer reaction processes. PMID:23109903

Identification of Atherosclerotic Plaques in Carotid Artery by Fluorescence Spectroscopy

NASA Astrophysics Data System (ADS)

Rocha, Rick; Villaverde, Antonio Balbin; Silveira, Landulfo; Costa, Maricília Silva; Alves, Leandro Procópio; Pasqualucci, Carlos Augusto; Brugnera, Aldo

2008-04-01

The aim of this work was to identify the presence of atherosclerotic plaques in carotid artery using the Fluorescence Spectroscopy. The most important pathogeny in the cardiovascular disorders is the atherosclerosis, which may affect even younger individuals. With approximately 1.2 million heart attacks and 750,000 strokes afflicting an aging American population each year, cardiovascular disease remains the number one cause of death. Carotid artery samples were obtained from the Autopsy Service at the University of São Paulo (São Paulo, SP, Brazil) taken from cadavers. After a histopathological analysis the 60 carotid artery samples were divided into two groups: normal (26) and atherosclerotic plaques (34). Samples were irradiated with the wavelength of 488 nm from an Argon laser. A 600 μm core optical fiber, coupled to the Argon laser, was used for excitation of the sample, whereas another 600 optical fiber, coupled to the spectrograph entrance slit, was used for collecting the fluorescence from the sample. Measurements were taken at different points on each sample and then averaged. Fluorescence spectra showed a single broad line centered at 549 nm. The fluorescence intensity for each sample was calculated by subtracting the intensity at the peak (550 nm) and at the bottom (510 nm) and then data were statistically analyzed, looking for differences between both groups of samples. ANOVA statistical test showed a significant difference (p<0,05) between both types of tissues, with regard to the fluorescence peak intensities. Our results indicate that this technique could be used to detect the presence of the atherosclerotic in carotid tissue.

Polar plot representation of time-resolved fluorescence.

PubMed

Eichorst, John Paul; Wen Teng, Kai; Clegg, Robert M

2014-01-01

Measuring changes in a molecule's fluorescence emission is a common technique to study complex biological systems such as cells and tissues. Although the steady-state fluorescence intensity is frequently used, measuring the average amount of time that a molecule spends in the excited state (the fluorescence lifetime) reveals more detailed information about its local environment. The lifetime is measured in the time domain by detecting directly the decay of fluorescence following excitation by short pulse of light. The lifetime can also be measured in the frequency domain by recording the phase and amplitude of oscillation in the emitted fluorescence of the sample in response to repetitively modulated excitation light. In either the time or frequency domain, the analysis of data to extract lifetimes can be computationally intensive. For example, a variety of iterative fitting algorithms already exist to determine lifetimes from samples that contain multiple fluorescing species. However, recently a method of analysis referred to as the polar plot (or phasor plot) is a graphical tool that projects the time-dependent features of the sample's fluorescence in either the time or frequency domain into the Cartesian plane to characterize the sample's lifetime. The coordinate transformations of the polar plot require only the raw data, and hence, there are no uncertainties from extensive corrections or time-consuming fitting in this analysis. In this chapter, the history and mathematical background of the polar plot will be presented along with examples that highlight how it can be used in both cuvette-based and imaging applications.

Laser-induced differential normalized fluorescence method for cancer diagnosis

DOEpatents

Vo-Dinh, Tuan; Panjehpour, Masoud; Overholt, Bergein F.

1996-01-01

An apparatus and method for cancer diagnosis are disclosed. The diagnostic method includes the steps of irradiating a tissue sample with monochromatic excitation light, producing a laser-induced fluorescence spectrum from emission radiation generated by interaction of the excitation light with the tissue sample, and dividing the intensity at each wavelength of the laser-induced fluorescence spectrum by the integrated area under the laser-induced fluorescence spectrum to produce a normalized spectrum. A mathematical difference between the normalized spectrum and an average value of a reference set of normalized spectra which correspond to normal tissues is calculated, which provides for amplifying small changes in weak signals from malignant tissues for improved analysis. The calculated differential normalized spectrum is correlated to a specific condition of a tissue sample.

Laser-induced differential normalized fluorescence method for cancer diagnosis

DOEpatents

Vo-Dinh, T.; Panjehpour, M.; Overholt, B.F.

1996-12-03

An apparatus and method for cancer diagnosis are disclosed. The diagnostic method includes the steps of irradiating a tissue sample with monochromatic excitation light, producing a laser-induced fluorescence spectrum from emission radiation generated by interaction of the excitation light with the tissue sample, and dividing the intensity at each wavelength of the laser-induced fluorescence spectrum by the integrated area under the laser-induced fluorescence spectrum to produce a normalized spectrum. A mathematical difference between the normalized spectrum and an average value of a reference set of normalized spectra which correspond to normal tissues is calculated, which provides for amplifying small changes in weak signals from malignant tissues for improved analysis. The calculated differential normalized spectrum is correlated to a specific condition of a tissue sample. 5 figs.

A Quantitative Microscopy Technique for Determining the Number of Specific Proteins in Cellular Compartments

PubMed Central

Mutch, Sarah A.; Gadd, Jennifer C.; Fujimoto, Bryant S.; Kensel-Hammes, Patricia; Schiro, Perry G.; Bajjalieh, Sandra M.; Chiu, Daniel T.

2013-01-01

This protocol describes a method to determine both the average number and variance of proteins in the few to tens of copies in isolated cellular compartments, such as organelles and protein complexes. Other currently available protein quantification techniques either provide an average number but lack information on the variance or are not suitable for reliably counting proteins present in the few to tens of copies. This protocol entails labeling the cellular compartment with fluorescent primary-secondary antibody complexes, TIRF (total internal reflection fluorescence) microscopy imaging of the cellular compartment, digital image analysis, and deconvolution of the fluorescence intensity data. A minimum of 2.5 days is required to complete the labeling, imaging, and analysis of a set of samples. As an illustrative example, we describe in detail the procedure used to determine the copy number of proteins in synaptic vesicles. The same procedure can be applied to other organelles or signaling complexes. PMID:22094731

Capacitively Coupled Plasma Discharge of Ionic Liquid Solutions to Synthesize Carbon Dots as Fluorescent Sensors.

PubMed

Ke, Ching-Bin; Lu, Te-Ling; Chen, Jian-Lian

2018-05-26

Oxygen and nitrogen capacitively coupled plasma (CCP) was used to irradiate mixtures of aliphatic acids in high boiling point solvents to synthesize fluorescent carbon dots (C-dots). With a high fluorescence intensity, the C-dots obtained from the O₂/CCP radiation of a 1-ethyl-3-methylimidazolium dicyanamide ionic liquid solution of citric acid were characterized with an average diameter of 8.6 nm (σ = 1.1 nm), nitrogen and oxygen bonding functionalities, excitation-independent emissions, and upconversion fluorescence. Through dialysis of the CCP-treated C-dots, two emissive surface states corresponding to their respective functionalities and emissions were identified. The fluorescence spectrum of the CCP-treated C-dots was different from that of the microwave irradiation and possessed higher intensity than that of hydrothermal pyrolysis. By evaluation of the fluorescence quenching effect on flavonoids and metal ions, the CCP-treated C-dots showed a high selectivity for quercetin and sensitivity to Hg 2+ . Based on the Perrin model, a calibration curve ( R ² = 0.9992) was established for quercetin ranging from 2.4 μM to 119 μM with an LOD (limit of detection) = 0.5 μM. The quercetin in the ethanol extract of the sun-dried peel of Citrus reticulata cv. Chachiensis was determined by a standard addition method to be 4.20 ± 0.15 mg/g with a matrix effect of 8.16%.

A three-step reconstruction method for fluorescence molecular tomography based on compressive sensing

NASA Astrophysics Data System (ADS)

Zhu, Yansong; Jha, Abhinav K.; Dreyer, Jakob K.; Le, Hanh N. D.; Kang, Jin U.; Roland, Per E.; Wong, Dean F.; Rahmim, Arman

2017-02-01

Fluorescence molecular tomography (FMT) is a promising tool for real time in vivo quantification of neurotransmission (NT) as we pursue in our BRAIN initiative effort. However, the acquired image data are noisy and the reconstruction problem is ill-posed. Further, while spatial sparsity of the NT effects could be exploited, traditional compressive-sensing methods cannot be directly applied as the system matrix in FMT is highly coherent. To overcome these issues, we propose and assess a three-step reconstruction method. First, truncated singular value decomposition is applied on the data to reduce matrix coherence. The resultant image data are input to a homotopy-based reconstruction strategy that exploits sparsity via l1 regularization. The reconstructed image is then input to a maximum-likelihood expectation maximization (MLEM) algorithm that retains the sparseness of the input estimate and improves upon the quantitation by accurate Poisson noise modeling. The proposed reconstruction method was evaluated in a three-dimensional simulated setup with fluorescent sources in a cuboidal scattering medium with optical properties simulating human brain cortex (reduced scattering coefficient: 9.2 cm-1, absorption coefficient: 0.1 cm-1 and tomographic measurements made using pixelated detectors. In different experiments, fluorescent sources of varying size and intensity were simulated. The proposed reconstruction method provided accurate estimates of the fluorescent source intensity, with a 20% lower root mean square error on average compared to the pure-homotopy method for all considered source intensities and sizes. Further, compared with conventional l2 regularized algorithm, overall, the proposed method reconstructed substantially more accurate fluorescence distribution. The proposed method shows considerable promise and will be tested using more realistic simulations and experimental setups.

A fluorescence-based imaging approach to pharmacokinetic analysis of intracochlear drug delivery.

PubMed

Ayoob, Andrew M; Peppi, Marcello; Tandon, Vishal; Langer, Robert; Borenstein, Jeffrey T

2018-04-05

Advances in microelectromechanical systems (MEMS) technologies are enhancing the development of intracochlear delivery devices for the treatment of hearing loss with emerging pharmacological therapies. Direct intracochlear delivery addresses the limitations of systemic and intratympanic delivery. However, optimization of delivery parameters for these devices requires pharmacokinetic assessment of the spatiotemporal drug distribution inside the cochlea. Robust methods of measuring drug concentration in the perilymph have been developed, but lack spatial resolution along the tonotopic axis or require complex physiological measurements. Here we describe an approach for quantifying distribution of fluorescent drug-surrogate probe along the cochlea's sensory epithelium with high spatial resolution enabled by confocal fluorescence imaging. Fluorescence from FM 1-43 FX, a fixable endocytosis marker, was quantified using confocal fluorescence imaging of whole mount sections of the organ of Corti from cochleae resected and fixed at several time points after intracochlear delivery. Intracochlear delivery of FM 1-43 FX near the base of the cochlea produces a base-apex gradient of fluorescence in the row of inner hair cells after 1 h post-delivery that is consistent with diffusion-limited transport along the scala tympani. By 3 h post-delivery there is approximately an order of magnitude decrease in peak average fluorescence intensity, suggesting FM 1-43 FX clearance from both the perilymph and inner hair cells. The increase in fluorescence intensity at 72 h post-delivery compared to 3 h post-delivery may implicate a potential radial transport pathway into the scala media. Copyright © 2018 Elsevier B.V. All rights reserved.

Automated method for tracing leading and trailing processes of migrating neurons in confocal image sequences

NASA Astrophysics Data System (ADS)

Kerekes, Ryan A.; Gleason, Shaun S.; Trivedi, Niraj; Solecki, David J.

2010-03-01

Segmentation, tracking, and tracing of neurons in video imagery are important steps in many neuronal migration studies and can be inaccurate and time-consuming when performed manually. In this paper, we present an automated method for tracing the leading and trailing processes of migrating neurons in time-lapse image stacks acquired with a confocal fluorescence microscope. In our approach, we first locate and track the soma of the cell of interest by smoothing each frame and tracking the local maxima through the sequence. We then trace the leading process in each frame by starting at the center of the soma and stepping repeatedly in the most likely direction of the leading process. This direction is found at each step by examining second derivatives of fluorescent intensity along curves of constant radius around the current point. Tracing terminates after a fixed number of steps or when fluorescent intensity drops below a fixed threshold. We evolve the resulting trace to form an improved trace that more closely follows the approximate centerline of the leading process. We apply a similar algorithm to the trailing process of the cell by starting the trace in the opposite direction. We demonstrate our algorithm on two time-lapse confocal video sequences of migrating cerebellar granule neurons (CGNs). We show that the automated traces closely approximate ground truth traces to within 1 or 2 pixels on average. Additionally, we compute line intensity profiles of fluorescence along the automated traces and quantitatively demonstrate their similarity to manually generated profiles in terms of fluorescence peak locations.

[Surface Cleaning and Disinfection in the Hospital. Improvement by Objective Monitoring and Intervention].

PubMed

Woltering, R; Hoffmann, G; Isermann, J; Heudorf, U

2016-11-01

Background and Objective: An assessment of cleaning and disinfection in hospitals by the use of objective surveillance and review of mandatory corrective measures was undertaken. Methods: A prospective examination of the cleaning and disinfection of surfaces scheduled for daily cleaning in 5 general care hospitals by use of an ultraviolet fluorescence targeting method (UVM) was performed, followed by structured educational and procedural interventions. The survey was conducted in hospital wards, operating theatres and intensive care units. Cleaning performance was measured by complete removal of UVM. Training courses and reinforced self-monitoring were implemented after the first evaluation. 6 months later, we repeated the assessment for confirmation of success. Results: The average cleaning performance was 34% (31/90) at base-line with significant differences between the 5 hospitals (11-67%). The best results were achieved in intensive care units (61%) and operating theatres (58%), the worst results in hospital wards (22%). The intervention significantly improved cleaning performance up to an average of 69% (65/94; +34.7%; 95% confidence interval (CI): 21.2-48.3; p<0.05), with differences between the hospitals (20-95%). The largest increase was achieved in hospital wards (+45%; CI 29.2-60.8; p<0.05). Improvements in operating theatres (+22.9%; CI 10.9-56.7) and intensive care units (+5.6%; CI 25.8-36.9) were statistically not significant. Conclusions: The monitoring of cleaning and disinfection of surfaces by fluorescence targeting is appropriate for evaluating hygiene regulations. An intervention can lead to a significant improvement of cleaning performance. As part of a strategy to improve infection control in hospitals, fluorescence targeting enables a simple inexpensive and effective surveillance of the cleaning performance and corrective measures. © Georg Thieme Verlag KG Stuttgart · New York.

N III Bowen Lines and Fluorescence Mechanism in the Symbiotic Star AG Peg

NASA Astrophysics Data System (ADS)

Hyung, Siek; Lee, Seong-Jae; Lee, Kang Hwan

2018-03-01

We have investigated the intensities and full width at half maximum (FWHM) of the high dispersion spectroscopic N III emission lines of AG Peg, observed with the Hamilton Echelle Spectrograph (HES) in three different epochs at Mt. Hamilton's Lick Observatory. The earlier theoretical Bowen line study assumed the continuum fluorescence effect, presenting a large discrepancy with the present data. Hence, we analyzed the observed N III lines assuming line fluorescence as the only suitable source: (1) The O III and N III resonance line profiles near λ 374 were decomposed, using the Gaussian function, and the contributions from various O III line components were determined. (2) Based on the theoretical resonant N III intensities, the expected N III Bowen intensities were obtained to fit the observed values. Our study shows that the incoming line photon number ratio must be considered to balance at each N III Bowen line level in the ultraviolet radiation according to the observed lines in the optical zone. We also found that the average FWHM of the N III Bowen lines was about 5 km·s-1 greater than that of the O III Bowen lines, perhaps due to the inherently different kinematic characteristics of their emission zones.

Enhanced emission of nile red fluorescent nanoparticles embedded in hybrid sol-gel glasses.

PubMed

Ferrer, Maria L; del Monte, Francisco

2005-01-13

Highly fluorescent Nile Red (NR) nanoparticles embedded in a hybrid sol-gel glass are reported. The crystallite growth within the confined system created by the porous hybrid matrix results in NR nanoparticles of averaged dimensions below 36 nm. The preparation process allows for the control of both the conformation adopted by single NR molecules prior to aggregation (e.g., near planar) and the configuration of the aggregates (e.g., oblique with phi < 54.7 degrees) prior to their assembly in the supramolecular architecture which ultimately forms the nanoparticles. The full preservation of the fluorescent configuration of the aggregates in the nanoparticles is confirmed through the application of the exciton theory, and it is responsible for the significant increase of the fluorescence emission intensity (e.g., up to 525- and 70-fold as compared to that obtained for single NR molecules embedded in pure and hybrid silica glasses, respectively).

Energy transfer dynamics from individual semiconductor nanoantennae to dye molecules with implication to light-harvesting nanosystems

NASA Astrophysics Data System (ADS)

Shan, Guangcun; Hu, Mingjun; Yan, Ze; Li, Xin; Huang, Wei

2018-03-01

Semiconductor nanocrystals can be used as nanoscale optical antennae to photoexcite individual dye molecules in an ensemble via energy transfer mechanism. The theoretical framework developed by Förster and others describes how electronic excitation migrates in the photosynthetic apparatus of plants, algae, and bacteria from light absorbing pigments to reaction centers where light energy is utilized for the eventual conversion into chemical energy. Herein we investigate the effect of the average donor-acceptor spacing on the time-resolved fluorescence intensity and dynamics of single donor-acceptor pairs with the dye acceptor concentration decreasing by using quantum Monte-Carlo simulation of FRET dynamics. Our results validated that the spatial disorder controlling the microscopic energy transfer rates accounts for the scatter in donor fluorescence lifetimes and intensities, which provides a new design guideline for artificial light-harvesting nanosystems.

Fluorescent carbon nanodots facilely extracted from Coca Cola for temperature sensing

NASA Astrophysics Data System (ADS)

Li, Feiming; Chen, Qiaoling; Cai, Zhixiong; Lin, Fangyuan; Xu, Wei; Wang, Yiru; Chen, Xi

2017-12-01

A novel method for the fabrication of carbon nanodots (CDs) is introduced: extracting CDs from the well-known soft drink Coca Cola via dialysis. The obtained CDs are of good monodispersity with a narrow size distribution (average diameter of 3.0 nm), good biocompatibility, high solubility (about 180 mg ml-1) and stable fluorescence even at a high salt concentration. Furthermore, they are sensitive to the temperature change with a linear relationship between the fluorescence intensity and temperature from 5 °C-95 °C. The CDs have been applied in high stable temperature sensing. This protocol is quite simple, green, cost-effective and technologically simple, which might be used for a range of applications including sensing, catalysts, drug and gene delivery, and so on.

Fluorescent carbon nanodots facilely extracted from Coca Cola for temperature sensing.

PubMed

Li, Feiming; Chen, Qiaoling; Cai, Zhixiong; Lin, Fangyuan; Xu, Wei; Wang, Yiru; Chen, Xi

2017-10-16

A novel method for the fabrication of carbon nanodots (CDs) is introduced: extracting CDs from the well-known soft drink Coca Cola via dialysis. The obtained CDs are of good monodispersity with a narrow size distribution (average diameter of 3.0 nm), good biocompatibility, high solubility (about 180 mg ml -1 ) and stable fluorescence even at a high salt concentration. Furthermore, they are sensitive to the temperature change with a linear relationship between the fluorescence intensity and temperature from 5 °C-95 °C. The CDs have been applied in high stable temperature sensing. This protocol is quite simple, green, cost-effective and technologically simple, which might be used for a range of applications including sensing, catalysts, drug and gene delivery, and so on.

Excited state intramolecular proton transfer reaction of 4'-N,N-diethylamino-3-hydroxyflavone and solvation dynamics in room temperature ionic liquids studied by optical Kerr gate fluorescence measurement.

PubMed

Kimura, Yoshifumi; Fukuda, Masanori; Suda, Kayo; Terazima, Masahide

2010-09-16

Fluorescence dynamics of 4'-N,N-diethylamino-3-hydroxyflavone (DEAHF) and its methoxy derivative (DEAMF) in various room temperature ionic liquids (RTILs) have been studied mainly by an optical Kerr gate method. DEAMF showed a single band fluorescence whose peak shifted with time by the solvation dynamics. The averaged solvation time determined by the fluorescence peak shift was proportional to the viscosity of the solvent except for tetradecyltrihexylphosphonium bis(trifluoromethanesulfonyl)amide. The solvation times were consistent with reported values determined with different probe molecules. DEAHF showed dual fluorescence due to the normal and tautomer forms produced by the excited state intramolecular proton transfer (ESIPT), and the relative intensities were dependent on the time and the solvent cation or anion species. By using the information of the fluorescence spectrum of DEAMF, the fluorescence spectrum of DEAHF at each delay time after the photoexcitation was decomposed into the normal and the tautomer fluorescence components, respectively. The normal component showed a very fast decay simulated by a biexponential function (2-3 and 20-30 ps) with an additional slower decay component. The tautomer component showed a rise with the time constants corresponding to the faster decay of the normal form with an additional instantaneous rise. The faster dynamics of the normal and tautomer population changes were assigned to the ESIPT process, while the slower decay of the fluorescence was attributed to the population decay from the excited state through the radiative and nonradiative processes. The average ESIPT time was much faster than the averaged solvation time of RTILs. Basically, the ESIPT kinetics in RTILs is similar to those in conventional liquid solvents like acetonitrile (Chou et al. J. Phys. Chem. A 2005, 109, 3777). The faster ESIPT is interpreted in terms of the activation barrierless process from the Franck-Condon state before the solvation of the normal state in the electronic excited state. With the advance of the solvation in the excited state, the normal form becomes relatively more stable than the tautomer form, which makes the ESIPT become an activation process.

Dark proteins: effect of inclusion body formation on quantification of protein expression.

PubMed

Iafolla, Marco A J; Mazumder, Mostafizur; Sardana, Vandit; Velauthapillai, Tharsan; Pannu, Karanbir; McMillen, David R

2008-09-01

Plasmid-borne gene expression systems have found wide application in the emerging fields of systems biology and synthetic biology, where plasmids are used to implement simple network architectures, either to test systems biology hypotheses about issues such as gene expression noise or as a means of exerting artificial control over a cell's dynamics. In both these cases, fluorescent proteins are commonly applied as a means of monitoring the expression of genes in the living cell, and efforts have been made to quantify protein expression levels through fluorescence intensity calibration and by monitoring the partitioning of proteins among the two daughter cells after division; such quantification is important in formulating the predictive models desired in systems and synthetic biology research. A potential pitfall of using plasmid-based gene expression systems is that the high protein levels associated with expression from plasmids can lead to the formation of inclusion bodies, insoluble aggregates of misfolded, nonfunctional proteins that will not generate fluorescence output; proteins caught in these inclusion bodies are thus "dark" to fluorescence-based detection methods. If significant numbers of proteins are incorporated into inclusion bodies rather than becoming biologically active, quantitative results obtained by fluorescent measurements will be skewed; we investigate this phenomenon here. We have created two plasmid constructs with differing average copy numbers, both incorporating an unregulated promoter (P(LtetO-1) in the absence of TetR) expressing the GFP derivative enhanced green fluorescent protein (EGFP), and inserted them into Escherichia coli bacterial cells (a common model organism for work on the dynamics of prokaryotic gene expression). We extracted the inclusion bodies, denatured them, and refolded them to render them active, obtaining a measurement of the average number of EGFP per cell locked into these aggregates; at the same time, we used calibrated fluorescent intensity measurements to determine the average number of active EGFP present per cell. Both measurements were carried out as a function of cellular doubling time, over a range of 45-75 min. We found that the ratio of inclusion body EGFP to active EGFP varied strongly as a function of the cellular growth rate, and that the number of "dark" proteins in the aggregates could in fact be substantial, reaching ratios as high as approximately five proteins locked into inclusion bodies for every active protein (at the fastest growth rate), and dropping to ratios well below 1 (for the slowest growth rate). Our results suggest that efforts to compare computational models to protein numbers derived from fluorescence measurements should take inclusion body loss into account, especially when working with rapidly growing cells. 2008 Wiley-Liss, Inc.

Near-Membrane Refractometry Using Supercritical Angle Fluorescence.

PubMed

Brunstein, Maia; Roy, Lopamudra; Oheim, Martin

2017-05-09

Total internal reflection fluorescence (TIRF) microscopy and its variants are key technologies for visualizing the dynamics of single molecules or organelles in live cells. Yet truly quantitative TIRF remains problematic. One unknown hampering the interpretation of evanescent-wave excited fluorescence intensities is the undetermined cell refractive index (RI). Here, we use a combination of TIRF excitation and supercritical angle fluorescence emission detection to directly measure the average RI in the "footprint" region of the cell during image acquisition. Our RI measurement is based on the determination on a back-focal plane image of the critical angle separating evanescent and far-field fluorescence emission components. We validate our method by imaging mouse embryonic fibroblasts and BON cells. By targeting various dyes and fluorescent-protein chimeras to vesicles, the plasma membrane, as well as mitochondria and the endoplasmic reticulum, we demonstrate local RI measurements with subcellular resolution on a standard TIRF microscope, with a removable Bertrand lens as the only modification. Our technique has important applications for imaging axial vesicle dynamics and the mitochondrial energy state or detecting metabolically more active cancer cells. Copyright © 2017. Published by Elsevier Inc.

CdSe/ZnS quantum dots conjugated with a fluorescein derivative: a FRET-based pH sensor for physiological alkaline conditions.

PubMed

Kurabayashi, Tomokazu; Funaki, Nayuta; Fukuda, Takeshi; Akiyama, Shinnosuke; Suzuki, Miho

2014-01-01

Dual pH-dependent fluorescence peaks from a semiconductor quantum dot (QD) and a pH-dependent fluorescent dye can be measured by irradiating with a single wavelength light, and the pH can be estimated from the ratio of the fluorescent intensity of the two peaks. In this work, ratiometric pH sensing was achieved in an aqueous environment by a fluorescent CdSe/ZnS QD appended with a pH-sensitive organic dye, based on fluorescence resonance energy transfer (FRET). By functionalizing the CdSe/ZnS QD with 5-(and 6)-carboxynaphthofluorescein succinimidyl ester as a pH-dependent fluorescent dye, we succeeded in fabricating sensitive nanocomplexes with a linear response to a broad range of physiological pH levels (7.5-9.5) when excited at 450 nm. We found that a purification process is important for increasing the high-fluorescence intensity ratio of a ratiometric fluorescence pH-sensor, and the fluorescence intensity ratio was improved up to 1.0 at pH 8.0 after the purification process to remove unreacted CdSe/ZnS QDs even though the fluorescence of the dye could not be observed without the purification process. The fluorescence intensity ratio corresponds to the fluorescence intensity of the dye, and this fluorescent dye exhibited pH-dependent fluorescence intensity changes. These facts indicate that the fluorescence intensity ratio linearly increased with increasing pH value of the buffer solution containing the QD and the dye. The FRET efficiencies changed from 0.3 (pH 7.5) to 6.2 (pH 9.5).

Measurement of protein-like fluorescence in river and waste water using a handheld spectrophotometer.

PubMed

Baker, Andy; Ward, David; Lieten, Shakti H; Periera, Ryan; Simpson, Ellie C; Slater, Malcolm

2004-07-01

Protein-like fluorescence intensity in rivers increases with increasing anthropogenic DOM inputs from sewerage and farm wastes. Here, a portable luminescence spectrophotometer was used to investigate if this technology could be used to provide both field scientists with a rapid pollution monitoring tool and process control engineers with a portable waste water monitoring device, through the measurement of river and waste water tryptophan-like fluorescence from a range of rivers in NE England and from effluents from within two waste water treatment plants. The portable spectrophotometer determined that waste waters and sewerage effluents had the highest tryptophan-like fluorescence intensity, urban streams had an intermediate tryptophan-like fluorescence intensity, and the upstream river samples of good water quality the lowest tryptophan-like fluorescence intensity. Replicate samples demonstrated that fluorescence intensity is reproducible to +/- 20% for low fluorescence, 'clean' river water samples and +/- 5% for urban water and waste waters. Correlations between fluorescence measured by the portable spectrophotometer with a conventional bench machine were 0.91; (Spearman's rho, n = 143), demonstrating that the portable spectrophotometer does correlate with tryptophan-like fluorescence intensity measured using the bench spectrophotometer.

Absolute intensity measurements of impurity emissions in a shock tunnel and their consequences for laser-induced fluorescence experiments

NASA Technical Reports Server (NTRS)

Palma, P. C.; Houwing, A. F. P.; Sandeman, R. J.

1993-01-01

Absolute intensity measurements of impurity emissions in a shock tunnel nozzle flow are presented. The impurity emission intensities were measured with a photomultiplier and optical multichannel analyzer and calibrated against an intensity standard. The various metallic contaminants were identified and their intensities measured in the spectral regions 290 to 330 nm and 375 to 385 nm. A comparison with calculated fluorescence intensities for predissociated laser-induced fluorescence signals is made. It is found that the emission background is negligible for most fluorescence experiments.

Spectral dependence of fluorescence near plasmon resonant metal nanoparticles

NASA Astrophysics Data System (ADS)

Chen, Yeechi

The optical properties of fluorophores are significantly modified when placed within the near field (0--100 nm) of plasmon resonant metal nanostructures, due to the competition between increased decay rates and "hotspots" of concentrated electric fields. The decay rates and effective electric field intensities are highly dependent on the relative position of dye and metal and the overlap between plasmon resonance and dye absorption and emission. Understanding these dependencies can greatly improve the performance of biosensing and nanophotonic devices. In this dissertation, the fluorescence intensity of organic dyes and CdSe quantum dots near single metal nanoparticles is studied as a function of the local surface plasmon resonance (LSPR) of the nanoparticle. Single metal nanoparticles have narrow, well-defined, intense local surface plasmon resonances that are tunable across the visible spectrum by changes in size and shape. First, we show that organic dyes can be self-assembled on single silver nanoprisms into known configurations by the hybridization of thiolated DNA oligomers. We correlate the fluorescence intensity of the dyes to the LSPR of the individual nanoprism to which they are attached. For each of three different organic dyes, we observe a strong correlation between the fluorescence intensity of the dye and the degree of spectral overlap with the plasmon resonance of the nanoparticle. On average, we observe the brightest fluorescence from dyes attached to metal nanoparticles that have a LSPR scattering peak 40--120 meV higher in energy than the emission peak of the fluorophore. Second, the plasmon-enhanced fluorescence from CdSe/CdS/CdZnS/ZnS core/shell quantum dots is studied near a variety of silver and gold nanoparticles. With single-particle scattering spectroscopy, the localized surface plasmon resonance spectra of single metal nanoparticles is correlated with the photoluminescence excitation (PLE) spectra of the nearby quantum dots. The PLE spectra closely track the scattering spectra of the metal nanoparticles. By taking advantage of the ability to excite quantum dots across a wide range of wavelengths while detecting a single emission wavelength, we measure an excitation enhancement factor for single metal nanoparticles. The data also provide a calculation of a lower-bound of experimentally attainable enhancement factors solely due to increased near-field excitation. This factor was found to range from ˜3 to 10 for Au spheres, Ag cubes and Ag nanoprisms.

Seasonal characterization of CDOM for lakes in semi-arid regions of Northeast China using excitation-emission matrices fluorescence and parallel factor analysis (EEM-PARAFAC)

NASA Astrophysics Data System (ADS)

Zhao, Y.; Song, K.; Wen, Z.; Li, L.; Zang, S.; Shao, T.; Li, S.; Du, J.

2015-04-01

The seasonal characteristics of fluorescence components in CDOM for lakes in the semi-arid region of Northeast China were examined by excitation-emission matrices fluorescence and parallel factor analysis (EEM-PARAFAC). Two humic-like peaks C1 (Ex/Em = 230, 300/425 nm) and C2 (Ex/Em = 255, 350/460 nm) and two protein-like B (Ex/Em = 220, 275/320 nm) and T (Ex/Em = 225, 290/360 nm) peaks were identified using PARAFAC. The average fluorescence intensity of the four components differed with seasonal variation from June and August 2013 to February and April 2014. The total fluorescence intensity significantly varied from 2.54 ± 0.68 nm-1 in June to the mean value 1.93 ± 0.70 nm-1 in August 2013, and then increased to 2.34 ± 0.92 nm-1 in February and reduced to the lowest 1.57 ± 0.55 nm-1 in April 2014. In general, the fluorescence intensity was dominated by peak C1, indicating that most part of CDOM for inland waters being investigated in this study was originated from phytoplankton degradation. The lowest C2 represents only a small portion of CDOM from terrestrial imported organic matter to water bodies through rainwash and soil leaching. The two protein-like intensities (B and T) formed in situ through microbial activity have almost the same intensity. Especially, in August 2013 and February 2014, the two protein-like peaks showed obviously difference from other seasons and the highest C1 (1.02 nm-1) was present in February 2014. Components 1 and 2 exhibited strong linear correlation (R2 = 0.633). There were significantly positive linear relationships between CDOM absorption coefficients a(254) (R2 = 0.72, 0.46, p < 0.01), a(280) (R2 = 0.77, 0.47, p < 0.01), a(350) (R2 = 0.76, 0.78, p < 0.01) and Fmax for two humic-like components (C1 and C2), respectively. A close relationship (R2 = 0.931) was found between salinity and DOC. However, almost no obvious correlation was found between salinity and EEM-PARAFAC extracted components except for C3 (R2 = 0.469). Results from this investigation demonstrate that the EEM-PARAFAC technique can be used to evaluate the seasonal dynamics of CDOM fluorescence components for inland waters in semi-arid regions of Northeast China.

Application of silver nanoparticles in the detection of SYBR Green I by surface enhanced Raman and surface-enhanced fluorescence

NASA Astrophysics Data System (ADS)

Guo, Wei; Wu, Jian; Wang, Chunyan; Zhang, Tian; Chen, Tao

2018-05-01

Silver nanomaterials have remarkable application in biomedical detection due to their unique surface plasmon resonance (SPR) characteristics. It can be used for surface-enhanced Raman scattering (SERS) and surface-enhanced fluorescence (SEF). Current research elaborates a technique for improvement of SYBR Green I detection obtained from surface-enhanced Raman scattering (SERS) and surface-enhanced fluorescence (SEF) by silver nanoparticles with the average size about 70 nm. Primarily, SYBR Green I is an important fluorescent dye used in polymerase chain reaction (PCR). It is found that both Raman and fluorescence can be used for detection of this dye. Furthermore, the enhanced efficiency of the Raman and fluorescence by SERS and SEF is observed in this study, the enhancement factor for Raman signals is 3.2 × 103, and the fluorescence intensity bincreased two times by SEF. The quantitative detection of SYBR Green I by SERS and SEF can be achieved. The present work can be used to improve the detection of SYBR Green I by SERS and SEF. It would also be employed for high-sensitive detection of other materials in the future.

Decay pattern of the Pygmy Dipole Resonance in 130Te

NASA Astrophysics Data System (ADS)

Isaak, J.; Beller, J.; Fiori, E.; Krtička, M.; Löher, B.; Pietralla, N.; Romig, C.; Rusev, G.; Savran, D.; Scheck, M.; Silva, J.; Sonnabend, K.; Tonchev, A.; Tornow, W.; Weller, H.; Zweidinger, M.

2014-03-01

The electric dipole strength distribution in 130Te has been investigated using the method of Nuclear Resonance Fluorescence. The experiments were performed at the Darmstadt High Intensity Photon Setup using bremsstrahlung as photon source and at the High Intensity overrightarrow γ -Ray Source, where quasi-monochromatic and polarized photon beams are provided. Average decay properties of 130Te below the neutron separation energy are determined. Comparing the experimental data to the predictions of the statistical model indicate, that nuclear structure effects play an important role even at sufficiently high excitation energies. Preliminary results will be presented.

Analysis of hydrocarbon-bearing fluid inclusions (HCFI) using time-resolved fluorescence spectroscopy

NASA Astrophysics Data System (ADS)

Przyjalgowski, Milosz A.; Ryder, Alan G.; Feely, Martin; Glynn, Thomas J.

2005-06-01

Hydrocarbon-bearing fluid inclusions (HCFI) are microscopic cavities within rocks that are filled with petroleum oil, the composition of which may not have changed since the trapping event. Thus, the composition of that entrapped oil can provide information about the formation and evolution of the oil reservoir. This type of information is important to the petroleum production and exploration industries. Crude oil fluorescence originates from the presence of cyclic aromatic compounds and the nature of the emission is governed by the chemical composition of the oil. Fluorescence based methods are widely used for analysis of crude oil because they offer robust, non-contact and non-destructive measurement options. The goal of our group is the development of a non-destructive analytical method for HCFI using time-resolved fluorescence methods. In broad terms, crude oil fluorescence behavior is governed by the concentration of quenching species and the distribution of fluorophores. For the intensity averaged fluorescence lifetime, the best correlations have been found between polar or alkane concentrations, but these are not suitable for robust, quantitative analysis. We have recently started to investigate another approach for characterizing oils by looking at Time-resolved Emission Spectra (TRES). TRES are constructed from intensities sampled at discrete times during the fluorescence decay of the sample. In this study, TRES, from a series of 10 crude oils from the Middle East, have been measured at discrete time gates (0.5 ns, 1 ns, 2 ns, 4 ns) over the 450-700 nm wavelength range. The spectral changes in TRES, such as time gate dependent Stokes' shift and spectral broadening, are analyzed in the context of energy transfer rates. In this work, the efficacy of using TRES for fingerprinting individual oils and HCFI is also demonstrated.

Fluorescent optical position sensor

DOEpatents

Weiss, Jonathan D.

2005-11-15

A fluorescent optical position sensor and method of operation. A small excitation source side-pumps a localized region of fluorescence at an unknown position along a fluorescent waveguide. As the fluorescent light travels down the waveguide, the intensity of fluorescent light decreases due to absorption. By measuring with one (or two) photodetectors the attenuated intensity of fluorescent light emitted from one (or both) ends of the waveguide, the position of the excitation source relative to the waveguide can be determined by comparing the measured light intensity to a calibrated response curve or mathematical model. Alternatively, excitation light can be pumped into an end of the waveguide, which generates an exponentially-decaying continuous source of fluorescent light along the length of the waveguide. The position of a photodetector oriented to view the side of the waveguide can be uniquely determined by measuring the intensity of the fluorescent light emitted radially at that location.

Fluorescence Dynamics in the Endoplasmic Reticulum of a Live Cell: Time-Resolved Confocal Microscopy.

PubMed

Ghosh, Shirsendu; Nandi, Somen; Ghosh, Catherine; Bhattacharyya, Kankan

2016-09-19

Fluorescence dynamics in the endoplasmic reticulum (ER) of a live non-cancer lung cell (WI38) and a lung cancer cell (A549) are studied by using time-resolved confocal microscopy. To selectively study the organelle, ER, we have used an ER-Tracker dye. From the emission maximum (λmaxem) of the ER-Tracker dye, polarity (i.e. dielectric constant, ϵ) in the ER region of the cells (≈500 nm in WI38 and ≈510 nm in A549) is estimated to be similar to that of chloroform (λmaxem =506 nm, ϵ≈5). The red shift by 10 nm in λmaxem in the cancer cell (A549) suggests a slightly higher polarity compared to the non-cancer cell (WI38). The fluorescence intensity of the ER-Tracker dye exhibits prolonged intermittent oscillations on a timescale of 2-6 seconds for the cancer cell (A549). For the non-cancer cell (WI38), such fluorescence oscillations are much less prominent. The marked fluorescence intensity oscillations in the cancer cell are attributed to enhanced calcium oscillations. The average solvent relaxation time (<τs >) of the ER region in the lung cancer cell (A549, 250±50 ps) is about four times faster than that in the non-cancer cell (WI38, 1000±50 ps). © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Analysis of root surface properties by fluorescence/Raman intensity ratio.

PubMed

Nakamura, Shino; Ando, Masahiro; Hamaguchi, Hiro-O; Yamamoto, Matsuo

2017-11-01

The aim of this study is to evaluate the existence of residual calculus on root surfaces by determining the fluorescence/Raman intensity ratio. Thirty-two extracted human teeth, partially covered with calculus on the root surface, were evaluated by using a portable Raman spectrophotometer, and a 785-nm, 100-mW laser was applied for fluorescence/Raman excitation. The collected spectra were normalized to the hydroxyapatite Raman band intensity at 960 cm -1 . Raman spectra were recorded from the same point after changing the focal distance of the laser and the target radiating angle. In seven teeth, the condition of calculus, cementum, and dentin were evaluated. In 25 teeth, we determined the fluorescence/Raman intensity ratio following three strokes of debridement. Raman spectra collected from the dentin, cementum, and calculus were different. After normalization, spectra values were constant. The fluorescence/Raman intensity ratio of calculus region showed significant differences compared to the cementum and dentin (p < 0.05). The fluorescence/Raman intensity ratio decreased with calculus debridement. For this analysis, the delta value was defined as the difference between the values before and after three strokes, with the final 2 delta values close to zero, indicating a gradual asymptotic curve and the change in intensity ratio approximating that of individual constants. Fluorescence/Raman intensity ratio was effectively used to cancel the angle- and distance-dependent fluctuations of fluorescence collection efficiency during measurement. Changes in the fluorescence/Raman intensity ratio near zero suggested that cementum or dentin was exposed, and calculus removed.

Fluorescence Behavior and Dural Infiltration of Meningioma Analyzed by 5-Aminolevulinic Acid-Based Fluorescence: Operating Microscope Versus Mini-Spectrometer.

PubMed

Knipps, Johannes; Beseoglu, Kerim; Kamp, Marcel; Fischer, Igor; Felsberg, Joerg; Neumann, Lisa M; Steiger, Hans-Jakob; Cornelius, Jan F

2017-12-01

To compare fluorescence intensity of tumor specimens, as measured by a fluorescence-guided surgery microscope and a spectrometer, to evaluate tumor infiltration of dura mater around meningiomas with help of these 2 different 5-aminolevulinic acid (5-ALA)-based fluorescence tools, and to correlate fluorescence intensity with histopathologic data. In a clinical series, meningiomas were resected by 5-ALA fluorescence-guided surgery. Fluorescence intensity was semiquantitatively rated by the surgeon at predefined points. Biopsies were harvested and fluorescence intensity measured by a spectrometer and histopathologically analyzed. Sampling was realized at the level of the dura in a centrifugal direction. A total of 104 biopsies (n = 13 tumors) were analyzed. Specificity and sensitivity of the microscope were 0.96 and 0.53 and of the spectrometer 0.95 and 0.93, respectively. Fluorescence intensity as measured by the spectrometer was correlated to histologically confirmed tumor burden. In a centrifugal direction, tumor burden and fluorescence intensity continuously decreased (along the dural tail). Below a threshold value of 639 arbitrary units no tumor was histologically detectable. At the level of the dura the spectrometer was highly sensitive for detection of meningioma cells. The surgical microscope showed false negative results and missed residual tumor cells in more than one half of the cases. The complementary use of both fluorescence tools may improve resection quality. Copyright © 2017 Elsevier Inc. All rights reserved.

Differences in the intensity of light-induced fluorescence emitted by resin composites.

PubMed

Kim, Bo-Ra; Kang, Si-Mook; Kim, Gyung-Min; Kim, Baek-Il

2016-03-01

The aims of this study were to compare the intensities of fluorescence emitted by different resin composites as detected using quantitative light-induced fluorescence (QLF) technology, and to compare the fluorescence intensity contrast with the color contrast between a restored composite and the adjacent region of the tooth. Six brands of light-cured resin composites (shade A2) were investigated. The composites were used to prepare composite discs, and fill holes that had been prepared in extracted human teeth. White-light and fluorescence images of all specimens were obtained using a fluorescence camera based on QLF technology (QLF-D) and converted into 8-bit grayscale images. The fluorescence intensity of the discs as well as the fluorescence intensity contrast and the color contrast between the composite restoration and adjacent tooth region were calculated as grayscale levels. The grayscale levels for the composite discs differed significantly with the brand (p<0.001): DenFil (10.84±0.35, mean±SD), Filtek Z350 (58.28±1.37), Premisa (156.94±1.58), Grandio (177.20±0.81), Charisma (207.05±0.77), and Gradia direct posterior (211.52±1.66). The difference in grayscale levels between a resin restoration and the adjacent tooth was significantly greater in fluorescence images for each brand than in white-light images, except for the Filtek Z350 (p<0.05). However, the Filtek Z350 restoration was distinguishable from the adjacent tooth in a fluorescence image. The intensities of fluorescence detected from the resin composites varied. The differences between the composite and adjacent tooth were greater for the fluorescence intensity contrast than for the colors observed in the white-light images. Copyright © 2016 Elsevier B.V. All rights reserved.

Fluorimetric detection of Sn(2+) ion in aqueous medium using Salicylaldehyde based nanoparticles and application to natural samples analysis.

PubMed

Patil, Kishor S; Mahajan, Prasad G; Patil, Shivajirao R

2017-01-05

The fluorescent 2-[(E)-(2-phenylhydrazinylidene)methyl]phenol nanoparticles (PHPNPs) were prepared by a simple reprecipitation method. The prepared PHPNPs examined by Dynamic Light Scattering show narrower particle size distribution having an average particle size of 93.3nm. The Scanning Electron Microphotograph shows distinct spherical shaped morphology of nanoparticles. The blue shift in UV-absorption and fluorescence spectra of PHPNPs with respect to corresponding spectra of PHP in acetone solution indicates H- aggregates and Aggregation Induced Enhanced Emission (AIEE) for nanoparticles. The nanoparticles show selective tendency towards the recognition of Sn(2+) ions by enhancing the fluorescence intensity preference to Cu(2+), Fe(3+), Fe(2+), Ni(2+), NH4(+), Ca(2+), Pb(2+), Hg(2+) and Zn(2+) ions, which actually seem to quench the fluorescence of nanoparticles. The studies on Langmuir adsorption plot, fluorescence lifetime of PHPNPs, DLS-Zeta sizer, UV-visible and fluorescence titration with and without Sn(2+) helped to propose a suitable mechanism of fluorescence enhancement of nanoparticles by Sn(2+) and their binding ability during complexation. The fluorescence enhancement effect of PHPNPs induced by Sn(2+) is further used to develop an analytical method for detection of Sn(2+) from aqueous medium in environmental samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Fluorimetric detection of Sn2 + ion in aqueous medium using Salicylaldehyde based nanoparticles and application to natural samples analysis

NASA Astrophysics Data System (ADS)

Patil, Kishor S.; Mahajan, Prasad G.; Patil, Shivajirao R.

2017-01-01

The fluorescent 2-[(E)-(2-phenylhydrazinylidene)methyl]phenol nanoparticles (PHPNPs) were prepared by a simple reprecipitation method. The prepared PHPNPs examined by Dynamic Light Scattering show narrower particle size distribution having an average particle size of 93.3 nm. The Scanning Electron Microphotograph shows distinct spherical shaped morphology of nanoparticles. The blue shift in UV-absorption and fluorescence spectra of PHPNPs with respect to corresponding spectra of PHP in acetone solution indicates H- aggregates and Aggregation Induced Enhanced Emission (AIEE) for nanoparticles. The nanoparticles show selective tendency towards the recognition of Sn2 + ions by enhancing the fluorescence intensity preference to Cu2 +, Fe3 +, Fe2 +, Ni2 +, NH4+, Ca2 +, Pb2 +, Hg2 + and Zn2 + ions, which actually seem to quench the fluorescence of nanoparticles. The studies on Langmuir adsorption plot, fluorescence lifetime of PHPNPs, DLS-Zeta sizer, UV-visible and fluorescence titration with and without Sn2 + helped to propose a suitable mechanism of fluorescence enhancement of nanoparticles by Sn2 + and their binding ability during complexation. The fluorescence enhancement effect of PHPNPs induced by Sn2 + is further used to develop an analytical method for detection of Sn2 + from aqueous medium in environmental samples.

Resonant fluorescence for multilevel systems in intense nonmonochromatic fields: possibilities for applications in laser medicine

NASA Astrophysics Data System (ADS)

Karagodova, Tamara Y.

1999-03-01

The theory of resonant fluorescence of multilevel system in two monochromatic intense laser fields has been applied for investigating the temporal decay of magnetic sublevels of an atom. As for two-level system the triplet of resonant fluorescence is observed, for real atom being the multilevel system the multiplet of resonant fluorescence can be observed. The excitation spectra, defining the intensities of lines in the multiplet of resonant fluorescence, and shifts of components of spectra are shown. Typical temporal dependence of fluorescence intensity for magnetic sublevels of an atom having different relaxation constants is shown. The computer simulation of resonant fluorescence for simple systems can help to understand the regularities in temporal decay curves of atherosclerotic plaque, malignant tumor compared to normal surrounding tissue.

Theoretical investigation for excitation light and fluorescence signal of fiber optical sensor using tapered fiber tip.

PubMed

Yuan, Yinquan; Ding, Liyun

2011-10-24

For fiber optical sensor made of tapered fiber tip, the effects of the geometrical parameters of tapered tip on two important factors have been investigated. One factor is the intensity of the evanescent wave into fluorescent layer through core-medium interface; the other is the intensity of fluorescence signal transmitted from fluorescent layer to measurement end. A dependence relation of the intensity of fluorescence signal transmitted from fluorescent layer to measurement end upon the geometrical parameters of tapered tip has been obtained. Theoretical results show that the intensity of the evanescent wave into fluorescent layer rises with the decrease of the end diameter of tapered tip, and the increase of the tip length; and the transmitted power of fluorescence signal increases linearly with the increase of the tip length due to the contribution of the side area of tapered tip. © 2011 Optical Society of America

Determination of Residual Stress Distributions in Polycrystalline Alumina using Fluorescence Microscopy

PubMed Central

Michaels, Chris A.; Cook, Robert F.

2016-01-01

Maps of residual stress distributions arising from anisotropic thermal expansion effects in a polycrystalline alumina are generated using fluorescence microscopy. The shifts of both the R1 and R2 ruby fluorescence lines of Cr in alumina are used to create maps with sub-µm resolution of either the local mean and shear stresses or local crystallographic a- and c-stresses in the material, with approximately ± 1 MPa stress resolution. The use of single crystal control materials and explicit correction for temperature and composition effects on line shifts enabled determination of the absolute values and distributions of values of stresses. Temperature correction is shown to be critical in absolute stress determination. Experimental determinations of average stress parameters in the mapped structure are consistent with assumed equilibrium conditions and with integrated large-area measurements. Average crystallographic stresses of order hundreds of MPa are determined with characteristic distribution widths of tens of MPa. The stress distributions reflect contributions from individual clusters of stress in the structure; the cluster size is somewhat larger than the grain size. An example application of the use of stress maps is shown in the calculation of stress-intensity factors for fracture in the residual stress field. PMID:27563163

[Investigation of exciting light and plant leaves age effects on chlorophyll fluorescense of radish plants].

PubMed

Nesterenko, T V; Tikhomirov, A A; Shikhov, V N

2012-01-01

The effect of exciting light intensity and leaves age on characteristics of slow stage of chlorophyll fluorescence induction (CFI) of radish leaves has been investigated. Light dependence of the relationship of maximum fluorescence intensity in the peak P and the stationary fluorescence level (F(P)/F(S)) and also light dependence of temporal characteristics of CFI (T0.5 - half decrease of chlorophyll fluorescence intensity during slow stage of fluorescence induction and tmin - summarized CFI characteristics derived by calculating via integral proportional to variable part of illuminated in the result of chlorophyll fluorescence energy during slow stage of CFI) have been studied. Plants were grown in controlled conditions of light culture at 100 Wt/m2 of photosynthetic active radiation (PAR). It has been shown that variability of the characteristics under study, associated with the effect of leaves age, significantly decreases at exciting light intensity equal to 40 Wt/m2 of PAR and more. The lowest effect of leaves age on the value of fluorescence characteristics for T0.5 and tmin and also for F(P)/F(S) ratio was observed at the intensity of exciting fluorescence light of 60 Wt/m2 of PAR. In the researched range of light intensities the temporal characteristics of T0.5 and tmin for uneven-aged radish leaves appeared to be by an order less responsive to the intensity changes of exciting fluorescence light as compared with F(P)/F(S) ratio.

Light propagation analysis for fluorescence measurements of a molecular probe in the brain

NASA Astrophysics Data System (ADS)

Asai, Kota; Togashi, Takuya; Okada, Eiji

2017-04-01

Light propagation in the slab head model that consists of five types of tissues was calculated to estimate the fluorescent intensity emerged from a molecular probe in the brain by a Monte Carlo simulation. The thickness of the scalp, skull and cerebrospinal fluid layer was varied to analyze the influence of the thickness of the superficial tissues on the fluorescent intensity detected on the scalp surface. The fluorescent intensity is exponentially reduced with increasing the depth of the brain surface. The thickness of the cerebrospinal fluid layer more significantly affects the fluorescent intensity than that of the scalp and skull.

Development of ultrasound-assisted fluorescence imaging of indocyanine green.

PubMed

Morikawa, Hiroyasu; Toyota, Shin; Wada, Kenji; Uchida-Kobayashi, Sawako; Kawada, Norifumi; Horinaka, Hiromichi

2017-01-01

Indocyanine green (ICG) accumulation in hepatocellular carcinoma means tumors can be located by fluorescence. However, because of light scattering, it is difficult to detect ICG fluorescence from outside the body. We propose a new fluorescence imaging method that detects changes in the intensity of ICG fluorescence by ultrasound-induced temperature changes. ICG fluorescence intensity decreases as the temperature rises. Therefore, it should theoretically be possible to detect tissue distribution of ICG using ultrasound to heat tissue, moving the point of ultrasound transmission, and monitoring changes in fluorescence intensity. A new probe was adapted for clinical application. It consisted of excitation light from a laser, fluorescence sensing through a light pipe, and heating by ultrasound. We applied the probe to bovine liver to image the accumulation of ICG. ICG emits fluorescence (820 nm) upon light irradiation (783 nm). With a rise in temperature, the fluorescence intensity of ICG decreased by 0.85 %/°C. The distribution of fluorescent ICG was detected using an ultrasonic warming method in a new integrated probe. Modulating fluorescence by changing the temperature using ultrasound can determine where ICG accumulates at a depth, highlighting its potential as a means to locate hepatocellular carcinoma.

Decreasing photobleaching by silver island films: application to muscle⋆

PubMed Central

Muthu, P.; Gryczynski, I.; Gryczynski, Z.; Talent, J.; Akopova, I.; Jain, K.; Borejdo, J.

2007-01-01

Recently it has become possible to study interactions between proteins at the level of single molecules. This requires collecting data from an extremely small volume, small enough to contain one molecule—typically of the order of attoliters (10−18 L). Collection of data from such a small volume with sufficiently high signal-to-noise ratio requires that the rate of photon detection per molecule be high. This calls for a large illuminating light flux, which in turn leads to rapid photobleaching of the fluorophores that are labeling the proteins. To decrease photobleaching, we measured fluorescence from a sample placed on coverslips coated with silver island films (SIF). SIF reduce photobleaching because they enhance fluorescence brightness and significantly decrease fluorescence lifetime. Increase in the brightness effectively decreases photobleaching because illumination can be attenuated to obtain the same fluorescence intensity. Decrease of lifetime decreases photobleaching because short lifetime minimizes the probability of oxygen attack while the fluorophore is in the excited state. The decrease of photobleaching was demonstrated in skeletal muscle. Myofibrils were labeled lightly with rhodamine–phalloidin, placed on coverslips coated with SIF, illuminated by total internal reflection, and observed through a confocal aperture. We show that SIF causes the intensity of phalloidin fluorescence to increase 4- to 5- fold and its fluorescence lifetime to decrease on average 23-fold. As a consequence, the rate of photobleaching of four or five molecules of actin of a myofibril on Olympus coverslips coated with SIF decreased at least 30-fold in comparison with photobleaching on an uncoated coverslip. Significant decrease of photobleaching makes the measurement of signal from a single cross-bridge of contracting muscle feasible. PMID:17531183

Analysis of rainwater dissolved organic carbon compounds using fluorescence spectrophotometry

NASA Astrophysics Data System (ADS)

Muller, Catherine L.; Baker, Andy; Hutchinson, Robert; Fairchild, Ian J.; Kidd, Chris

Global rainwater dissolved organic carbon (DOC) flux was recently estimated as 430 × 10 12 g C yr -1, yet little is known about the wide range of chemical compounds present, their sources, temporal patterns of variation, and the subsequent impact on climate and the environment. Precipitation events were sampled in Birmingham, UK between April 2005 and May 2007. Rainwater DOC compounds were analysed using fluorescence spectrophotometry. Three fluorophores were identified: HUmic-LIke Substances (HULIS), TYrosine-LIke Substances (TYLIS) and TRYptophan-LIke Substances (TRYLIS). Peak fluorescence intensities and locations for each substance were examined, and their variations with various meteorological parameters were investigated. The mean HULIS fluorescence intensity from all events was 209 a.u. (with sample fluorescence ranging from 37 a.u. to 995 a.u); mean fluorescence intensity was 469 a.u. (214-988 a.u) and 265 a.u. (50-876 a.u.) for TYLIS and TRYLIS, respectively. Results indicate that highest HULIS fluorescence intensities are experienced during convective events and events of continental origin, suggesting terrestrial/anthropogenic sources. Under well-mixed conditions, HULIS fluorescence intensity decreases, whereas during low wind speed, stagnation of the atmosphere results in higher fluorescence intensities, attributed to a build up of localised sources, particularly anthropogenic. TYLIS and TRYLIS did not show any significant trends for the meteorological variables. Fluorescence spectrophotometry is a fast, non-invasive technique which is demonstrated to be a powerful means of fingerprinting rainfall DOC compounds in real time for small sample volumes.

Conditions for NIR fluorescence-guided tumor resectioning in preclinical lung cancer model (Conference Presentation)

NASA Astrophysics Data System (ADS)

Kim, Minji; Quan, Yuhua; Choi, Byeong Hyun; Choi, Yeonho; Kim, Hyun Koo; Kim, Beop-Min

2016-03-01

Pulmonary nodule could be identified by intraoperative fluorescence imaging system from systemic injection of indocyanine green (ICG) which achieves enhanced permeability and retention (EPR) effects. This study was performed to evaluate optimal injection time of ICG for detecting cancer during surgery in rabbit lung cancer model. VX2 carcinoma cell was injected in rabbit lung under fluoroscopic computed tomography-guidance. Solitary lung cancer was confirmed on positron emitting tomography with CT (PET/CT) 2 weeks after inoculation. ICG was administered intravenously and fluorescent intensity of lung tumor was measured using the custom-built intraoperative color and fluorescence merged imaging system (ICFIS) for 15 hours. Solitary lung cancer was resected through thoracoscopic version of ICFIS. ICG was observed in all animals. Because Lung has fast blood pulmonary circulation, Fluorescent signal showed maximum intensity earlier than previous studies in other organs. Fluorescent intensity showed maximum intensity within 6-9 hours in rabbit lung cancer. Overall, Fluorescent intensity decreased with increasing time, however, all tumors were detectable using fluorescent images until 12 hours. In conclusion, while there had been studies in other organs showed that optimal injection time was at least 24 hours before operation, this study showed shorter optimal injection time at lung cancer. Since fluorescent signal showed the maximum intensity within 6-9 hours, cancer resection could be performed during this time. This data informed us that optimal injection time of ICG should be evaluated in each different solid organ tumor for fluorescent image guided surgery.

Zwitterion functionalized gold nanoclusters for multimodal near infrared fluorescence and photoacoustic imaging

NASA Astrophysics Data System (ADS)

Shen, Danjin; Henry, Maxime; Trouillet, Vanessa; Comby-Zerbino, Clothilde; Bertorelle, Franck; Sancey, Lucie; Antoine, Rodolphe; Coll, Jean-Luc; Josserand, Véronique; Le Guével, Xavier

2017-05-01

Gold nanoclusters (Au NCs) are an emerging type of theranostic agents combining therapeutic and imaging features with reduced toxicity. Au NCs stabilized by a zwitterion ligand with a fine control of the metal core size and the ligand coverage were synthesized by wet chemistry. Intense fluorescence signal is reported for the highest ligand coverage, whereas photoacoustic signal is stronger for the largest metal core. The best Au NC candidate with an average molecular weight of 17 kDa could be detected with high sensitivity on a 2D-near-infrared imaging instrument (limit of detection (LOD) = 2.3 μ M ) and by photoacoustic imaging. In vitro and in vivo experiments demonstrate an efficient cell uptake in U87 cell lines, a fast renal clearance (t1 /2 α = 6.5 ± 1.3 min), and a good correlation between near infrared fluorescence and photoacoustic measurements to follow the early uptake of Au NCs in liver.

Molecular dynamics in an optical trap of glutamate receptors labeled with quantum-dots on living neurons

NASA Astrophysics Data System (ADS)

Kishimoto, Tatsunori; Maezawa, Yasuyo; Kudoh, Suguru N.; Taguchi, Takahisa; Hosokawa, Chie

2017-04-01

Molecular dynamics of glutamate receptor, which is major neurotransmitter receptor at excitatory synapse located on neuron, is essential for synaptic plasticity in the complex neuronal networks. Here we studied molecular dynamics in an optical trap of AMPA-type glutamate receptor (AMPAR) labeled with quantum-dot (QD) on living neuronal cells with fluorescence imaging and fluorescence correlation spectroscopy (FCS). When a 1064-nm laser beam for optical trapping was focused on QD-AMPARs located on neuronal cells, the fluorescence intensity of QD-AMPARs gradually increased at the focal spot. Using single-particle tracking of QD-AMPARs on neurons, the average diffusion coefficient decreased in an optical trap. Moreover, the decay time obtained from FCS analysis increased with the laser power and the initial assembling state of AMPARs depended on culturing day, suggesting that the motion of QD-AMPAR was constrained in an optical trap.

Enhanced labeling of microalgae cellular lipids by application of an electric field generated by alternating current.

PubMed

Su, Li-Chien; Hsu, Yi-Hsiang; Wang, Hsiang-Yu

2012-05-01

An alternating current was used to generate an electric field to enhance the fluorescent labeling of microalgae cellular lipids with Nile red and LipidTOX. The decay of the fluorescence intensity of Chlorella vulgaris cells in 0 V/cm was more than 50% after 10 min, and the intensity variation was as high as 7% in 20s. At 2000 V/cm, the decay rate decreased to 1.22% per minute and the intensity fluctuation was less than 1% for LipidTOX-labeled cells. For Spirulina sp. cells at 0 V/cm, the fluorescence intensity increased by 10% after 10 min, whereas at 2000 V/cm, labeling was more rapid and fluorescence intensity doubled. These results show that applying an electric field can improve the quality of fluorescence detection by alleviating decay and fluctuation or by enhancing signal intensity. Copyright © 2012 Elsevier Ltd. All rights reserved.

Sensitive fluorescence detection of mercury(ii) in aqueous solution by the fluorescence quenching effect of MoS2 with DNA functionalized carbon dots.

PubMed

Srinivasan, K; Subramanian, K; Murugan, K; Dinakaran, K

2016-10-24

A rapid and sensitive fluorescent sensor based on the MoS 2 nanosheet/DNA/carbon dot nanoassembly has been developed towards the detection of mercury(ii) present in environmental samples. Bio-carbon dots (CDs) having strong fluorescence maxima at 451 nm were synthesized via one-step treatment with honey under low temperature carbonization. These CDs were nearly spherical with good size distribution and excellent monodispersity, and the average sizes of CD were around 2-4 nm as evidenced from transmission electron microscopy. The conjugation of DNA strands on the surface of the carbon dots provided an efficient fluorescent probe. The fluorescence of the MoS 2 nanosheet/DNA/carbon dot nanoassembly enhanced gradually with the increase in the concentration of Hg 2+ ions and the detection limit was found to be 1.02 nM. Furthermore, the fluorescence intensity was found to be linear with the concentration of Hg 2+ ions in the range from 0 to 10 nM and their respective coefficient of determination was found to be 0.93676 and 0.98178. The present MoS 2 nanosheet/DNA/carbon dot nanoassembly is highly selective toward Hg 2+ ions over a wide range of metal ions tested.

A Semi-Empirical Formula of the Dependence of the Fluorescence Intensity of Naphthalene on Temperature and the Oxygen Concentration

NASA Astrophysics Data System (ADS)

An, B.; Wang, Z.-G.; Yang, L.-C.; Li, X.-P.

2017-09-01

Two-ring aromatics, such as naphthalene, are important fluorescent components of kerosene in the planar laser-induced fluorescent (PLIF) technique. Quantifying measurements of kerosene vapor concentrations by PLIF require a prior knowledge of the fluorescence intensity of naphthalene over a wide temperature and oxygen concentration range. To promote the application of PLIF, a semi-empirical formula based on the collision theory and experimental data at the laser wavelength of 266 nm and a pressure of 0.1 MPa is established to predict the fluorescence intensity of naphthalene at different temperatures and oxygen concentrations. This formula takes vibrational states, temperature, and oxygen quenching into account. Verified by published experimental data, the formula can predict the fluorescence intensity of naphthalene with an error less than 9%.

Chromosome characterization using single fluorescent dye

DOEpatents

Crissman, Harry A.; Hirons, Gregory T.

1995-01-01

Chromosomes are characterized by fluorescent emissions from a single fluorescent dye that is excited over two different wavelengths. A mixture containing chromosomes is stained with a single dye selected from the group consisting of TOTO and YOYO and the stained chromosomes are placed in a flow cytometer. The fluorescent dye is excited sequentially by a first light having a wavelength in the ultraviolet range to excite the TOTO or YOYO to fluoresce at a first intensity and by a second light having a wavelength effective to excite the TOTO or YOYO dye to fluoresce at a second intensity. Specific chromosomes may be identified and sorted by intensity relationships between the first and second fluorescence emissions.

Cell cycle-dependent protein fingerprint from a single cancer cell: image cytometry coupled with single-cell capillary sieving electrophoresis.

PubMed

Hu, Shen; Le, Zhang; Krylov, Sergey; Dovichi, Norman J

2003-07-15

Study of cell cycle-dependent protein expression is important in oncology, stem cell research, and developmental biology. In this paper, we report the first protein fingerprint from a single cell with known phase in the cell cycle. To determine that phase, we treated HT-29 colon cancer cells with Hoescht 33342, a vital nuclear stain. A microscope was used to measure the fluorescence intensity from one treated cell; in this form of image cytometry, the fluorescence intensity is proportional to the cell's DNA content, which varies in a predictable fashion during the cell cycle. To generate the protein fingerprint, the cell was aspirated into the separation capillary and lysed. Proteins were fluorescently labeled with 3-(2-furoylquinoline-2-carboxaldehyde, separated by capillary sieving electrophoresis, and detected by laser-induced fluorescence. This form of electrophoresis is the capillary version of SDS-PAGE. The single-cell electropherogram partially resolved approximately 25 components in a 30-min separation, and the dynamic range of the detector exceeded 5000. There was a large cell-to-cell variation in protein expression, averaging 40% relative standard deviation across the electropherogram. The dominant source of variation was the phase of the cell in the cell cycle; on average, approximately 60% of the cell-to-cell variance in protein expression was associated with the cell cycle. Cells in the G1 and G2/M phases of the cell cycle had 27 and 21% relative standard deviations in protein expression, respectively. Cells in the G2/M phase generated signals that were twice the amplitude of the signals generated by G1 phase cells, as expected for cells that are soon to divide into two daughter cells. When electropherograms were normalized to total protein content, the expression of only one component was dependent on cell cycle at the 99% confidence limit. That protein is tentatively identified as cytokeratin 18 in a companion paper.

Development of LEDs-based microplate reader for bioanalytical assay measurements

NASA Astrophysics Data System (ADS)

Alaruri, Sami D.; Katzlinger, Michael; Schinwald, Bernhard; Kronberger, Georg; Atzler, Joseph

2013-10-01

The optical design for an LEDs-based microplate reader that can perform fluorescence intensity (top and bottom), absorbance, luminescence and time-resolved fluorescence measurements is described. The microplate reader is the first microplate reader in the marketplace that incorporates LEDs as excitation light sources. Absorbance measurements over the 0-3.5 optical density range for caffeine solution are presented. Additionally, fluorescence intensity readings collected at 535 and 625 nm from a green and a red RediPlateTM are reported. Furthermore, fluorescence decay lifetime measurements obtained for Eu (europium) and Sm (samarium) standard solutions using 370 nm excitation are presented. The microplate reader detection limits for the fluorescence intensity top, fluorescence intensity bottom, fluorescence polarization and time-resolved fluorescence modes are 1.5 fmol 100 µL-1 fluorescein (384-well plate), 25 fmol 100 µL-1 fluorescein (384-well plate), 5 mP at 10 nM fluorescein (black 384-well plate) and 30 amol 100 µL-1 europium solution (white 384-well plate), respectively.

Fluorescence Stability of Mercaptopropionic Acid Capped Cadmium Telluride Quantum Dots in Various Biochemical Buffers.

PubMed

Borse, Vivek; Kashikar, Adisha; Srivastava, Rohit

2018-04-01

Quantum dots are the semiconductor nanocrystals having unique optical and electronic properties. Quantum dots are category of fluorescent labels utilized for biological tagging, biosensing, bioassays, bioimaging and in vivo imaging as they exhibit very small size, signal brightness, photostability, tuning of light emission range, longer photoluminescence decay time as compared to organic dyes. In this work, we have synthesized and characterized mercaptopropionic acid capped cadmium telluride quantum dots (MPA-CdTe QDs) using hydrothermal method. The study further reports fluorescence intensity stability of quantum dots suspended in different buffers of varying concentration (1-100 mM), stored at various photophysical conditions. Fluorescence intensity values were reduced with increase in buffer concentration. When the samples were stored at room temperature in ambient light condition the quantum dots suspended in different buffers lost the fluorescence intensity after day 15 (except TRIS II). Fluorescence intensity values were found stable for more than 30 days when the samples were stored in dark condition. Samples stored in refrigerator displayed modest fluorescence intensity even after 300 days of storage. Thus, storage of MPA-CdTe QDs in refrigerator may be the suitable choice to maintain its fluorescence stability for longer time for further application.

Enhancement of fluorescence intensity by silicon particles and its size effect.

PubMed

Saitow, Ken-ichi; Suemori, Hidemi; Tamamitsu, Hironori

2014-02-04

Fluorescence-intensity enhancement of dye molecules was investigated using silicon submicron particles as a function of the particle size. Silicon particles with a size of 500 nm gave an enhancement factor up to 180. Measurement of scattering spectra revealed that the localized electric field at the particle enhances the fluorescence intensity.

Synthesis and fluorescence properties of some difluoroboron β-diketonate complexes and composite containing PMMA

NASA Astrophysics Data System (ADS)

Xing, Dongye; Hou, Yanjun; Niu, Haijun

2018-03-01

A series of difluoroboron β-diketonate complexes, containing the indon-β-diketonate ligand carrying methyl or methoxyl substituents was synthesized. The crystal structures of the complexes were confirmed by single crystal X-ray diffraction studies. The fluorescence properties of compounds were studied in solution state, solid state and on PMMA polymer matrix. The photophysical data of compounds 2a-2d exhibited strong fluorescence and photostability under the ultraviolet light (Hg lamp). The complex 2b showed higher fluorescence intensity in solution state as compared to other complexes of the series. The complexes 2c and 2d showed higher fluorescence intensity in the solid state, which are ascribed to the stronger π-π interactions between ligands in the solid state. The introduction of methoxyl or methyl groups on the benzene rings enhanced the absorption intensity, emission intensity, quantum yields and fluorescence lifetimes due to their electron-donating nature. Furthermore, the complex 2b was doped into the PMMA to produce hybrid materials, where the PMMA matrix acted as sensitizer for the central boron ion to enhance the fluorescence emission intensity and quantum yields.

Raman scattering and red fluorescence in the photochemical transformation of dry tryptophan particles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lai, Chih Wei; Schwab, Mark; Hill, Steven C.

Tryptophan is a fluorescent amino acid common in proteins. Its absorption is largest for wavelengths λ ≲ 290 nm and its fluorescence emissions peak around 300–350 nm, depending upon the local environment. Here we report the observation of red fluorescence near 600 nm emerging from 488-nm continuous-wave (CW) laser photoexcitation of dry tryptophan (Trp) particles. With an excitation intensity below 0.5 kW/cm 2, dry Trp particles yield distinctive Raman scattering peaks in the presence of relatively weak and spectrally broad emissions with λ ~500–700 nm, allowing estimation of particle temperature at low excitation intensities. When the photoexcitation intensity is increasedmore » to 1 kW/cm 2 or more for a few minutes, fluorescence intensity dramatically increases by more than two orders of magnitude. The fluorescence continues to increase in intensity and gradually shift to the red when photoexcitation intensity and the duration of exposure are increased. The resulting products absorb at visible wavelengths and generate red fluorescence with λ ~ 650–800 nm with 633-nm CW laser excitation. In conclusion, we attribute the emergence of orange and red fluorescence in the Trp products to a photochemical transformation that is instigated by weak optical transitions to triplet states in Trp with 488-nm excitation and which may be expedited by a photothermal effect.« less

Raman scattering and red fluorescence in the photochemical transformation of dry tryptophan particles

DOE PAGES

Lai, Chih Wei; Schwab, Mark; Hill, Steven C.; ...

2016-05-19

Tryptophan is a fluorescent amino acid common in proteins. Its absorption is largest for wavelengths λ ≲ 290 nm and its fluorescence emissions peak around 300–350 nm, depending upon the local environment. Here we report the observation of red fluorescence near 600 nm emerging from 488-nm continuous-wave (CW) laser photoexcitation of dry tryptophan (Trp) particles. With an excitation intensity below 0.5 kW/cm 2, dry Trp particles yield distinctive Raman scattering peaks in the presence of relatively weak and spectrally broad emissions with λ ~500–700 nm, allowing estimation of particle temperature at low excitation intensities. When the photoexcitation intensity is increasedmore » to 1 kW/cm 2 or more for a few minutes, fluorescence intensity dramatically increases by more than two orders of magnitude. The fluorescence continues to increase in intensity and gradually shift to the red when photoexcitation intensity and the duration of exposure are increased. The resulting products absorb at visible wavelengths and generate red fluorescence with λ ~ 650–800 nm with 633-nm CW laser excitation. In conclusion, we attribute the emergence of orange and red fluorescence in the Trp products to a photochemical transformation that is instigated by weak optical transitions to triplet states in Trp with 488-nm excitation and which may be expedited by a photothermal effect.« less

Raman scattering and red fluorescence in the photochemical transformation of dry tryptophan particles.

PubMed

Lai, Chih Wei; Schwab, Mark; Hill, Steven C; Santarpia, Joshua; Pan, Yong-Le

2016-05-30

Tryptophan is a fluorescent amino acid common in proteins. Its absorption is largest for wavelengths λ ≲ 290 nm and its fluorescence emissions peak around 300-350 nm, depending upon the local environment. Here we report the observation of red fluorescence near 600 nm emerging from 488-nm continuous-wave (CW) laser photoexcitation of dry tryptophan (Trp) particles. With an excitation intensity below 0.5 kW/cm², dry Trp particles yield distinctive Raman scattering peaks in the presence of relatively weak and spectrally broad emissions with λ ∼500-700 nm, allowing estimation of particle temperature at low excitation intensities. When the photoexcitation intensity is increased to 1 kW/cm² or more for a few minutes, fluorescence intensity dramatically increases by more than two orders of magnitude. The fluorescence continues to increase in intensity and gradually shift to the red when photoexcitation intensity and the duration of exposure are increased. The resulting products absorb at visible wavelengths and generate red fluorescence with λ ∼ 650-800 nm with 633-nm CW laser excitation. We attribute the emergence of orange and red fluorescence in the Trp products to a photochemical transformation that is instigated by weak optical transitions to triplet states in Trp with 488-nm excitation and which may be expedited by a photothermal effect.

Impact of laser excitation intensity on deep UV fluorescence detection in microchip electrophoresis.

PubMed

Schulze, Philipp; Ludwig, Martin; Belder, Detlev

2008-12-01

A high intensity 266 nm continuous wave (cw-) laser developed for material processing was utilised as an excitation source for sensitive native fluorescence detection of unlabelled compounds in MCE. This 120 mW laser was attached via an optical fibre into a commercial epifluorescence microscope. With this MCE set-up we evaluated the impact of laser power on the S/N of aromatic compounds as well as of proteins. Compared with a previous work which used a 4 mW pulsed laser for excitation, improved S/N for small aromatics and to a lesser extent for proteins could be attained. The LOD of the system was determined down to 24 ng/mL for serotonin (113 nM), 24 ng/mL for propranolol (81 nM), 80 ng/mL for tryptophan (392 nM) and 80 ng/mL for an aromatic diol (475 nM). Sensitive protein detection was obtained at concentrations of 5 microg/mL for lysocyme, trypsinogen and chymotrypsinogen (340, 208 and 195 nM, respectively). Finally, a comparison of the cw- with a pulsed 266 nm laser, operating at the same average power, showed a higher attainable sensitivity of the cw-laser. This can be attributed to fluorescence saturation and photobleaching effects of the pulsed laser at high pulse energies.

Quantitative monitoring of activity-dependent bulk endocytosis of synaptic vesicle membrane by fluorescent dextran imaging

PubMed Central

Clayton, Emma Louise; Cousin, Michael Alan

2012-01-01

Activity-dependent bulk endocytosis (ADBE) is the dominant synaptic vesicle (SV) retrieval mode in central nerve terminals during periods of intense neuronal activity. Despite this fact there are very few real time assays that report the activity of this critical SV retrieval mode. In this paper we report a simple and quantitative assay of ADBE using uptake of large flourescent dextrans as fluid phase markers. We show that almost all dextran uptake occurs in nerve terminals, using co-localisation with the fluorescent probe FM1-43. We also demonstrate that accumulated dextran cannot be unloaded by neuronal stimulation, indicating its specific loading into bulk endosomes and not SVs. Quantification of dextran uptake was achieved by using thresholding analysis to count the number of loaded nerve terminals, since monitoring the average fluorescence intensity of these nerve terminals did not accurately report the extent of ADBE. Using this analysis we showed that dextran uptake occurs very soon after stimulation and that it does not persist when stimulation terminates. Thus we have devised a simple and quantitative method to monitor ADBE in living neurones, which will be ideal for real time screening of small molecule inhibitors of this key SV retrieval mode. PMID:19766140

Comparative evaluation of performance measures for shading correction in time-lapse fluorescence microscopy.

PubMed

Liu, L; Kan, A; Leckie, C; Hodgkin, P D

2017-04-01

Time-lapse fluorescence microscopy is a valuable technology in cell biology, but it suffers from the inherent problem of intensity inhomogeneity due to uneven illumination or camera nonlinearity, known as shading artefacts. This will lead to inaccurate estimates of single-cell features such as average and total intensity. Numerous shading correction methods have been proposed to remove this effect. In order to compare the performance of different methods, many quantitative performance measures have been developed. However, there is little discussion about which performance measure should be generally applied for evaluation on real data, where the ground truth is absent. In this paper, the state-of-the-art shading correction methods and performance evaluation methods are reviewed. We implement 10 popular shading correction methods on two artificial datasets and four real ones. In order to make an objective comparison between those methods, we employ a number of quantitative performance measures. Extensive validation demonstrates that the coefficient of joint variation (CJV) is the most applicable measure in time-lapse fluorescence images. Based on this measure, we have proposed a novel shading correction method that performs better compared to well-established methods for a range of real data tested. © 2016 The Authors Journal of Microscopy © 2016 Royal Microscopical Society.

A molecular-sized optical logic circuit for digital modulation of a fluorescence signal

NASA Astrophysics Data System (ADS)

Nishimura, Takahiro; Tsuchida, Karin; Ogura, Yusuke; Tanida, Jun

2018-03-01

Fluorescence measurement allows simultaneous detection of multiple molecular species by using spectrally distinct fluorescence probes. However, due to the broad spectra of fluorescence emission, the multiplicity of fluorescence measurement is generally limited. To overcome this limitation, we propose a method to digitally modulate fluorescence output signals with a molecular-sized optical logic circuit by using optical control of fluorescence resonance energy transfer (FRET). The circuit receives a set of optical inputs represented with different light wavelengths, and then it switches high and low fluorescence intensity from a reporting molecule according to the result of the logic operation. By using combinational optical inputs in readout of fluorescence signals, the number of biomolecular species that can be identified is increased. To implement the FRET-based circuits, we designed two types of basic elements, YES and NOT switches. An YES switch produces a high-level output intensity when receiving a designated light wavelength input and a low-level intensity without the light irradiation. A NOT switch operates inversely to the YES switch. In experiments, we investigated the operation of the YES and NOT switches that receive a 532-nm light input and modulate the fluorescence intensity of Alexa Fluor 488. The experimental result demonstrates that the switches can modulate fluorescence signals according to the optical input.

Fluorescent and high intensity discharge lamp use in chambers and greenhouses

NASA Technical Reports Server (NTRS)

Langhans, Robert W.

1994-01-01

Fluorescent and High Intensity Discharge lamps have opened up great opportunities for researchers to study plant growth under controlled environment conditions and for commercial growers to increase plant production during low/light periods. Specific technical qualities of fluorescent and HID lamps have been critically reviewed. I will direct my remarks to fluorescent and high intensity discharge (HID) lamps in growth chambers, growth rooms, and greenhouses. I will discuss the advantages and disadvantages of using each lamp in growth chambers, growth rooms and greenhouses.

Quantifying fluorescence enhancement for slowly diffusing single molecules in plasmonic near fields

NASA Astrophysics Data System (ADS)

Caldarola, Martín; Pradhan, Biswajit; Orrit, Michel

2018-03-01

Gold nanorods are extensively used for single-molecule fluorescence enhancement as they are easy to synthesize, bio-compatible, and provide high light confinement at their nanometer-sized tips. The current way to estimate fluorescence enhancement relies on binned time traces or on fluorescence correlation spectroscopy. We report on novel ways to extract the enhancement factor in a single-molecule enhancement experiment, avoiding the arbitrary selection of one or a few high-intensity burst(s). These new estimates for the enhancement factor make use of the whole distribution of intensity bursts or of the interphoton delay distribution, which avoids the arbitrary binning of the fluorescence intensity time traces. We present experimental results on the bi-dimensional case, experimentally achieved using a lipid bilayer to support the diffusion of fluorophores. We support our findings with histograms of fluorescence bursts and with an analytical derivation of the interphoton delay distribution of (nearly) immobilized emitters from the fluorescence intensity profile.

Recovery of photoinduced reversible dark States utilized for molecular diffusion measurements.

PubMed

Chmyrov, Andriy; Sandén, Tor; Widengren, Jerker

2010-12-15

For a spatially restricted excitation volume, the effective modulation of the excitation in time is influenced by the passage times of the molecules through the excitation volume. By applying an additional time-modulated excitation, the buildup of photoinduced reversible dark states in fluorescent molecules can be made to vary significantly with their passage times through the excitation volume. The variations in the dark state populations are reflected by the time-averaged fluorescence intensity, which thus can be used to characterize the mobilities of the molecules. The concept was experimentally verified by measuring the fluorescence response of freely diffusing cyanine fluorophores (Cy5), undergoing trans-cis isomerization when subject to time-modulated excitation in a focused laser beam. From the fluorescence response, and by applying a simple photodynamic model, the transition times of the Cy5 molecules could be well reproduced when applying different laminar flow speeds through the detection volume. The presented approach puts no constraints on sample concentration, no requirements for high time resolution or sensitivity in the detection, nor requires a high fluorescence brightness of the characterized molecules. This can make the concept useful for a broad range of biomolecular mobility studies.

Excitation Anisotropy in Laser-Induced-Fluorescence Spectroscopy —High-Intensity, Broad-Line Excitation

NASA Astrophysics Data System (ADS)

Hirabayashi, Atsumu; Nambu, Yoshihiro; Fujimoto, Takashi

1986-10-01

The problem of excitation anisotropy in laser-induced-fluorescence spectroscopy (LIFS) was investigated for the intense excitation case under the broad-line condition. The depolarization coefficient for the fluorescence light was derived in the intense-excitation limit (linearly-polarized or unpolarized light excitation) and the results are presented in tables. In the region of intermediate intensity, between the weak and intense-excitation limits, the master equation was solved for a specific example of atomic transitions and its result is compared with experimental results.

Fluorescent fiber diagnostics

DOEpatents

Toeppen, John S.

1994-10-04

A fluorescent fiber (13) having a doped core (16) is pumped (11) by light (18) of a relatively short wavelength to produce fluorescence at a longer wavelength that is detected by detector (24). The level of fluorescence is monitored (26) and evaluated to provide information as to the excitation of the fiber (13) or the environment thereof. In particular, the level of intensity of the detected fluorescence may be used to measure the intensity of a light beam (18) passing axially through an optical fiber system (12) (FIG. 1 ), or the intensity of a light beam (46) passing radially through a fluorescent fiber (13) (FIG. 2 ), or the level of a fluid (32) in a tank (31) (FIG. 3 ), or a scintillation event (37) in a fluorescent fiber (13) pumped to produce amplification of the scintillation event (FIG. 4 ).

Fluorescent fiber diagnostics

DOEpatents

Toeppen, John S.

1994-01-01

A fluorescent fiber (13) having a doped core (16) is pumped (11) by light (18) of a relatively short wavelength to produce fluorescence at a longer wavelength that is detected by detector (24). The level of fluorescence is monitored (26) and evaluated to provide information as to the excitation of the fiber (13) or the environment thereof. In particular, the level of intensity of the detected fluorescence may be used to measure the intensity of a light beam (18) passing axially through an optical fiber system (12) (FIG. 1 ), or the intensity of a light beam (46) passing radially through a fluorescent fiber (13) (FIG. 2 ), or the level of a fluid (32) in a tank (31) (FIG. 3 ), or a scintillation event (37) in a fluorescent fiber (13) pumped to produce amplification of the scintillation event (FIG. 4 ).

Direct spectrometry: a new alternative for measuring the fluorescence of composite resins and dental tissues.

PubMed

da Silva, Tm; de Oliveira, Hpm; Severino, D; Balducci, I; Huhtala, Mfrl; Gonçalves, Sep

2014-01-01

The aim of this study was to evaluate the fluorescence intensity of different composite resins and compare those values with the fluorescence intensity of dental tissues. Different composite resins were used to make 10 discs (2 mm in depth and 4 mm in diameter) of each brand, divided into groups: 1) Z (Filtek Z350, 3M ESPE), 2) ES (Esthet-X, Dentsply), 3) A (Amelogen Plus, Ultradent), 4) DVS (Durafill-VS, Heraeus Kulzer) with 2 mm composite resin for enamel (A2), 5) OES ([Esthet-X] opaque-OA [1 mm] + enamel-A2 [1 mm]); 6) ODVSI ([Charisma-Opal/Durafill-VSI], opaque-OM (1 mm) + translucent [1mm]), and 7) DVSI ([Durafill- VSI] translucent [2 mm]). Dental tissue specimens were obtained from human anterior teeth cut in a mesiodistal direction to obtain enamel, dentin, and enamel/dentin samples (2 mm). The fluorescence intensity of specimens was directly measured using an optic fiber associated with a spectrometer (Ocean Optics USB 4000) and recorded in graphic form (Origin 8.0 program). Data were submitted to statistical analysis using Dunnet, Tukey, and Kruskall-Wallis tests. Light absorption of the composite resins was obtained in a spectral range from 250 to 450 nm, and that of dental tissues was between 250 and 300 nm. All composite resins were excited at 398 nm and exhibited maximum emissions of around 485 nm. Fluorescence intensity values for all of the resins showed statistically significant differences (measured in arbitrary units [AUs]), with the exception of groups Z and DVS. Group DVSI had the highest fluorescence intensity values (13539 AU), followed by ODVS (10440 AU), DVS (10146 AU), ES (3946 AU), OES (3841 AU), A (3540 AU), and Z (1146 AU). The fluorescence intensity values for the composite resins differed statistically from those of dental tissues (E=1380 AU; D=6262 AU; E/D=3251 AU). The opacity interfered with fluorescence intensity, and group Z demonstrated fluorescence intensity values closest to that of tooth enamel. It is concluded that the fluorescence intensity values were significantly different among the composite resins and compared with dental tissues. The direct spectrofluorimetric method represents a tool for evaluating the fluorescence of composite resins.

Azadioxatriangulenium: exploring the effect of a 20 ns fluorescence lifetime in fluorescence anisotropy measurements

NASA Astrophysics Data System (ADS)

Bogh, Sidsel A.; Bora, Ilkay; Rosenberg, Martin; Thyrhaug, Erling; Laursen, Bo W.; Just Sørensen, Thomas

2015-12-01

Azaoxatriangulenium (ADOTA) has been shown to be highly emissive despite a moderate molar absorption coefficient of the primary electronic transition. As a result, the fluorescence lifetime is ~20 ns, longer than all commonly used red fluorescent organic probes. The electronic transitions in ADOTA are highly polarised (r 0  =  0.38), which in combination with the long fluorescence lifetime extents the size-range of biomolecular weights that can be detected in fluorescence polarisation-based experiments. Here, the rotational dynamics of bovine serum albumin (BSA) are monitored with three different ADOTA derivatives, differing only in constitution of the reactive linker. A detailed study of the degree of labelling, the steady-state anisotropy, and the time-resolved anisotropy of the three different ADOTA-BSA conjugates are reported. The fluorescence quantum yields (ϕ fl) of the free dyes in PBS solution are determined to be ~55%, which is reduced to ~20% in the ADOTA-BSA conjugates. Despite the reduction in ϕ fl, a ~20 ns intensity averaged lifetime is maintained, allowing for the rotational dynamics of BSA to be monitored for up to 100 ns. Thus, ADOTA can be used in fluorescence polarisation assays to fill the gap between commonly used organic dyes and the long luminescence lifetime transition metal complexes. This allows for efficient steady-state fluorescence polarisation assays for detecting binding of analytes with molecular weights of up to 100 kDa.

4-Pyridoxic Acid in the Spent Dialysate: Contribution to Fluorescence and Optical Monitoring

PubMed Central

Kalle, Sigrid; Tanner, Risto; Arund, Jürgen; Tomson, Ruth; Luman, Merike; Fridolin, Ivo

2016-01-01

Aim In this work we estimated the contribution of the fluorescence of 4-pyridoxic acid (4-PA) to the total fluorescence of spent dialysate with the aim of evaluating the on-line monitoring of removal of this vitamin B-6 metabolite from the blood of patients with end-stage renal disease (ESRD). Methods Spectrofluorometric analysis of spent dialysate, collected from hemodialysis and hemodiafiltration sessions of 10 patients receiving regularly pyridoxine injections after dialysis treatment, was performed in the range of Ex/Em 220–500 nm. 4-PA in dialysate samples was identified and quantified using HPLC with fluorescent and MS/MS detection. Results Averaged HPLC chromatogram of spent dialysate had many peaks in the wavelength region of Ex320/Em430 nm where 4-PA was the highest peak with contribution of 42.2±17.0% at the beginning and 47.7±18.0% in the end of the dialysis. High correlation (R = 0.88–0.95) between 4-PA concentration and fluorescence intensity of spent dialysate was found in the region of Ex310-330/Em415-500 nm, respectively. Conclusion 4-PA elimination from the blood of ESRD patients can be potentially followed using monitoring of the fluorescence of the spent dialysate during dialysis treatments. PMID:27598005

4-Pyridoxic Acid in the Spent Dialysate: Contribution to Fluorescence and Optical Monitoring.

PubMed

Kalle, Sigrid; Tanner, Risto; Arund, Jürgen; Tomson, Ruth; Luman, Merike; Fridolin, Ivo

2016-01-01

In this work we estimated the contribution of the fluorescence of 4-pyridoxic acid (4-PA) to the total fluorescence of spent dialysate with the aim of evaluating the on-line monitoring of removal of this vitamin B-6 metabolite from the blood of patients with end-stage renal disease (ESRD). Spectrofluorometric analysis of spent dialysate, collected from hemodialysis and hemodiafiltration sessions of 10 patients receiving regularly pyridoxine injections after dialysis treatment, was performed in the range of Ex/Em 220-500 nm. 4-PA in dialysate samples was identified and quantified using HPLC with fluorescent and MS/MS detection. Averaged HPLC chromatogram of spent dialysate had many peaks in the wavelength region of Ex320/Em430 nm where 4-PA was the highest peak with contribution of 42.2±17.0% at the beginning and 47.7±18.0% in the end of the dialysis. High correlation (R = 0.88-0.95) between 4-PA concentration and fluorescence intensity of spent dialysate was found in the region of Ex310-330/Em415-500 nm, respectively. 4-PA elimination from the blood of ESRD patients can be potentially followed using monitoring of the fluorescence of the spent dialysate during dialysis treatments.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lindberg, David J.; Wranne, Moa S.; Gilbert Gatty, Mélina

Thioflavin-T (ThT) is one of the most commonly used dyes for amyloid detection, but the origin of its fluorescence enhancement is not fully understood. Herein we have characterised the ThT fluorescence response upon binding to the Aβ(1-40) and Aβ(1-42) variants of the Alzheimer's-related peptide amyloid-β, in order to explore how the photophysical properties of this dye relates to structural and morphological properties of two amyloid fibril types formed by peptides with a high degree of sequence homology. We show that the steady-state ThT fluorescence is 1.7 times more intense with Aβ(1-40) compared to Aβ(1-42) fibrils in concentration matched samples preparedmore » under quiescent conditions. By measuring the excited state lifetime of bound ThT, we also demonstrate a distinct difference between the two fibril isoforms, with Aβ(1-42) fibrils producing a longer ThT fluorescence lifetime compared to Aβ(1-40). The substantial steady-state intensity difference is therefore not explained by differences in fluorescence quantum yield. Further, we find that the ThT fluorescence intensity, but not the fluorescence lifetime, is dependent on the fibril preparation method (quiescent versus agitated conditions). We therefore propose that the fluorescence lifetime is inherent to each isoform and sensitively reports on fibril microstructure in the protofilament whereas the total fluorescence intensity relates to the amount of exposed β-sheet in the mature Aβ fibrils and hence to differences in their morphology. Our results highlight the complexity of ThT fluorescence, and demonstrate its extended use in amyloid fibril characterisation. - Highlights: • ThT emission is more intense with Aβ(1-40) fibrils than with Aβ(1-42) fibrils. • Aβ(1-42) fibrils induce longer ThT fluorescence lifetimes and higher quantum yield. • ThT emission intensity in Aβ fibril samples reports on fibril morphology. • The ThT fluorescence lifetime is a characteristic feature of each Aβ fibril type.« less

A programmable light engine for quantitative single molecule TIRF and HILO imaging.

PubMed

van 't Hoff, Marcel; de Sars, Vincent; Oheim, Martin

2008-10-27

We report on a simple yet powerful implementation of objective-type total internal reflection fluorescence (TIRF) and highly inclined and laminated optical sheet (HILO, a type of dark-field) illumination. Instead of focusing the illuminating laser beam to a single spot close to the edge of the microscope objective, we are scanning during the acquisition of a fluorescence image the focused spot in a circular orbit, thereby illuminating the sample from various directions. We measure parameters relevant for quantitative image analysis during fluorescence image acquisition by capturing an image of the excitation light distribution in an equivalent objective backfocal plane (BFP). Operating at scan rates above 1 MHz, our programmable light engine allows directional averaging by circular spinning the spot even for sub-millisecond exposure times. We show that restoring the symmetry of TIRF/HILO illumination reduces scattering and produces an evenly lit field-of-view that affords on-line analysis of evanescnt-field excited fluorescence without pre-processing. Utilizing crossed acousto-optical deflectors, our device generates arbitrary intensity profiles in BFP, permitting variable-angle, multi-color illumination, or objective lenses to be rapidly exchanged.

Uptake of Fluorescent Gentamicin by Peripheral Vestibular Cells after Systemic Administration

PubMed Central

Liu, Jianping; Kachelmeier, Allan; Dai, Chunfu; Li, Hongzhe; Steyger, Peter S.

2015-01-01

Objective In addition to cochleotoxicity, systemic aminoglycoside pharmacotherapy causes vestibulotoxicity resulting in imbalance and visual dysfunction. The underlying trafficking routes of systemically-administered aminoglycosides from the vasculature to the vestibular sensory hair cells are largely unknown. We investigated the trafficking of systemically-administered gentamicin into the peripheral vestibular system in C56Bl/6 mice using fluorescence-tagged gentamicin (gentamicin-Texas-Red, GTTR) imaged by scanning laser confocal microscopy to determine the cellular distribution and intensity of GTTR fluorescence in the three semicircular canal cristae, utricular, and saccular maculae at 5 time points over 4 hours. Results Low intensity GTTR fluorescence was detected at 0.5 hours as both discrete puncta and diffuse cytoplasmic fluorescence. The intensity of cytoplasmic fluorescence peaked at 3 hours, while punctate fluorescence was plateaued after 3 hours. At 0.5 and 1 hour, higher levels of diffuse GTTR fluorescence were present in transitional cells compared to hair cells and supporting cells. Sensory hair cells typically exhibited only diffuse cytoplasmic fluorescence at all time-points up to 4 hours in this study. In contrast, non-sensory cells rapidly exhibited both intense fluorescent puncta and weaker, diffuse fluorescence throughout the cytosol. The numbers and size of fluorescent puncta in dark cells and transitional cells increased over time. There is no preferential GTTR uptake by the five peripheral vestibular organs’ sensory cells. Control vestibular tissues exposed to Dulbecco’s phosphate-buffered saline or hydrolyzed Texas Red had negligible fluorescence. Conclusions All peripheral vestibular cells rapidly take up systemically-administered GTTR, reaching peak intensity 3 hours after injection. Sensory hair cells exhibited only diffuse fluorescence, while non-sensory cells displayed both diffuse and punctate fluorescence. Transitional cells may act as a primary pathway for trafficking of systemic GTTR from the vasculature to endolymph prior to entering hair cells. PMID:25793391

Means and method of detection in chemical separation procedures

DOEpatents

Yeung, Edward S.; Koutny, Lance B.; Hogan, Barry L.; Cheung, Chan K.; Ma, Yinfa

1993-03-09

A means and method for indirect detection of constituent components of a mixture separated in a chemical separation process. Fluorescing ions are distributed across the area in which separation of the mixture will occur to provide a generally uniform background fluorescence intensity. For example, the mixture is comprised of one or more charged analytes which displace fluorescing ions where its constituent components separate to. Fluorescing ions of the same charge as the charged analyte components cause a displacement. The displacement results in the location of the separated components having a reduced fluorescence intensity to the remainder of the background. Detection of the lower fluorescence intensity areas can be visually, by photographic means and methods, or by automated laser scanning.

Means and method of detection in chemical separation procedures

DOEpatents

Yeung, E.S.; Koutny, L.B.; Hogan, B.L.; Cheung, C.K.; Yinfa Ma.

1993-03-09

A means and method are described for indirect detection of constituent components of a mixture separated in a chemical separation process. Fluorescing ions are distributed across the area in which separation of the mixture will occur to provide a generally uniform background fluorescence intensity. For example, the mixture is comprised of one or more charged analytes which displace fluorescing ions where its constituent components separate to. Fluorescing ions of the same charge as the charged analyte components cause a displacement. The displacement results in the location of the separated components having a reduced fluorescence intensity to the remainder of the background. Detection of the lower fluorescence intensity areas can be visually, by photographic means and methods, or by automated laser scanning.

The fluorescence properties of aerosol larger than 0.8 μm in an urban and a PBA-dominated location

NASA Astrophysics Data System (ADS)

Gabey, A. M.; Stanley, W. R.; Gallagher, M. W.; Kaye, P. H.

2011-01-01

Dual-wavelength Ultraviolet light-induced fluorescence (UV-LIF) measurements were performed on ambient environmental aerosol in Manchester, UK (urban city centre, winter) and Borneo, Malaysia (remote, tropical), which are taken to represent environments with negligible and significant primary biological aerosol (PBA) influences, respectively. Single-particle fluorescence intensity and optical equivalent diameter were measured with a Wide Issue Bioaerosol Sensor, version 3 (WIBS3) in the diameter range 0.8 μm≤DP≤20 μm for 2-3 weeks and filters were analysed using energy dispersive X-ray (EDX) spectroscopy, which revealed mostly non-PBA dominated particle sizes larger than 1 μm in Manchester. The WIBS3 features three fluorescence channels: Fluorescence excited at 280 nm is recorded at 310-400 nm and 400-600 nm and fluorescence excited at 370 nm is detected at 400-600 nm. In Manchester the primary size mode of fluorescent and non-fluorescent material was at 1.2 μm. In Borneo non-fluorescent material peaked at 1.2 μm and fluorescent at 3-4 μm. The fluorescence intensity at 400-600 nm generally increased with DP at both sites, as did the 310-400 nm intensity in Borneo. In Manchester the 310-400 m fluorescence decreased at DP>4 μm, suggesting this channel offers additional discrimination between fluorescent particle types. Finally, the ratio of fluorescence intensity in two pairs of channels was investigated as a function of particle diameter and this varied significantly between the two environments, demonstrating that the fluorescent aerosol in each can in principle be distinguished using a combination of fluorescence and elastic scattering measurements.

Temperature influence on fluorescence intensity and enzyme activity of the fusion protein of GFP and hyperthermophilic xylanase.

PubMed

Zhang, Chong; Liu, Min-Sheng; Xing, Xin-Hui

2009-09-01

By constructing the expression system for fusion protein of GFPmut1 (a green fluorescent protein mutant) with the hyperthermophilic xylanase obtained from Dictyoglomus thermophilum Rt46B.1, the effects of temperature on the fluorescence of GFP and its relationship with the activities of GFP-fused xylanase have been studied. The fluorescence intensities of both GFP and GFP-xylanase have proved to be thermally sensitive, with the thermal sensitivity of the fluorescence intensity of GFP-xylanase being 15% higher than that of GFP. The lost fluorescence intensity of GFP inactivated at high temperature of below 60 degrees C in either single or fusion form can be completely recovered by treatment at 0 degrees C. By the fluorescence recovery of GFP domain at low temperature, the ratios of fluorescence intensity to xylanase activity (Rgfp/Axyl) at 15 degrees C and 37 degrees C have been compared. Even though the numbers of molecules of GFP and xylanase are equivalent, the Rgfp/Axyl ratio at 15 degrees C is ten times of that at 37 degrees C. This is mainly due to the fact that lower temperature is more conducive to the correct folding of GFP than the hyperthermophilic xylanase during the expression. This study has indicated that the ratio of GFP fluorescence to the thermophilic enzyme activity for the fusion proteins expressed at different temperatures could be helpful in understanding the folding properties of the two fusion partners and in design of the fusion proteins.

Amphiphilic inclusion spaces for various guests and regulation of fluorescence intensity of 1,8-bis(4-aminophenyl)anthracene crystals.

PubMed

Sugino, Misa; Hatanaka, Keisuke; Araki, Yusuke; Hisaki, Ichiro; Miyata, Mikiji; Tohnai, Norimitsu

2014-03-10

A host framework for inclusion of various guest molecules was investigated by preparation of inclusion crystals of 1,8-bis(4-aminophenyl)anthracene (1,8-BAPA) with organic solvents. X-ray crystallographic analysis revealed construction of the same inclusion space incorporating 1,8-BAPA and eight guest molecules including both non-polar (benzene) and polar guests (N,N-dimethylformamide, DMF). Fluorescence efficiencies varied depending on guest molecule polarity; DMF inclusion crystals exhibited the highest fluorescence intensity (ΦF=0.40), four times as high as that of a benzene inclusion crystal (ΦF=0.10). According to systematic investigations of inclusion phenomena, strong host–guest interactions and filling of the inclusion space led to a high fluorescence intensity. Temperature-dependent fluorescence spectral measurements revealed these factors effectively immobilised the host framework. Although hydrogen bonding commonly decreases fluorescence intensity, the present study demonstrated that such strong interactions provide excellent conditions for fluorescence enhancement. Thus, this remarkable behaviour has potential application toward sensing of highly polar molecules, such as biogenic compounds.

Fluorescence intensity positivity classification of Hep-2 cells images using fuzzy logic

NASA Astrophysics Data System (ADS)

Sazali, Dayang Farzana Abang; Janier, Josefina Barnachea; May, Zazilah Bt.

2014-10-01

Indirect Immunofluorescence (IIF) is a good standard used for antinuclear autoantibody (ANA) test using Hep-2 cells to determine specific diseases. Different classifier algorithm methods have been proposed in previous works however, there still no valid set as a standard to classify the fluorescence intensity. This paper presents the use of fuzzy logic to classify the fluorescence intensity and to determine the positivity of the Hep-2 cell serum samples. The fuzzy algorithm involves the image pre-processing by filtering the noises and smoothen the image, converting the red, green and blue (RGB) color space of images to luminosity layer, chromaticity layer "a" and "b" (LAB) color space where the mean value of the lightness and chromaticity layer "a" was extracted and classified by using fuzzy logic algorithm based on the standard score ranges of antinuclear autoantibody (ANA) fluorescence intensity. Using 100 data sets of positive and intermediate fluorescence intensity for testing the performance measurements, the fuzzy logic obtained an accuracy of intermediate and positive class as 85% and 87% respectively.

CdTe quantum dots as a novel biosensor for Serratia marcescens and Lipopolysaccharide.

PubMed

Ebrahim, Sh; Reda, M; Hussien, A; Zayed, D

2015-01-01

The main objective of this work is to synthesize CdTe quantum dots (QDs) conjugated with Concanavalin A (Con A) as a novel biosensor to be selective and specific for the detection of Lipopolysaccharide (LPS). In addition, the conjugated CdTe QDs-Con A was used as fluorescence labels to capture Serratia marcescens bacteria through the recognition between CdTe QDs-Con A and LPS of S. marcescens. The appearance of the lattice plans in the high resolution transmission electron photograph indicated a high crystalline with an average size of 4-5 nm for the CdTe QDs. The results showed that the relative fluorescence intensity of CdTe QDs-Con A decreased linearly with LPS concentration in the range from 10 to 90 fg/mL and with correlation coefficient (R(2)) equal to 0.9713. LPS surrounding the S. marcescens bacteria was bound to the CdTe QDs-Con A and leads to quenching of PL intensity. It was found that a good linear relationship between the relative PL intensity and the logarithmic of cell population of S. marcescens in range from 1×10 to 1×10(6) CFU/mL at pH 7 with R(2) of 0.952 was established. Copyright © 2015 Elsevier B.V. All rights reserved.

Correlation fluorescence method of amine detection

NASA Astrophysics Data System (ADS)

Myslitsky, Valentin F.; Tkachuk, Svetlana S.; Rudeichuk, Volodimir M.; Strinadko, Miroslav T.; Slyotov, Mikhail M.; Strinadko, Marina M.

1997-12-01

The amines fluorescence spectra stimulated by UV laser radiation are investigated in this paper. The fluorescence is stimulated by the coherent laser beam with the wavelength 0.337 micrometers . At the sufficient energy of laser stimulation the narrow peaks of the fluorescence spectra are detected besides the wide maximum. The relationship between the fluorescence intensity and the concentration of amines solutions are investigated. The fluorescence intensity temporal dependence on wavelength 0.363 micrometers of the norepinephrine solution preliminarily radiated by UV laser with wavelength 0.337 micrometers was found. The computer stimulated and experimental investigations of adrenaline and norepinephrine mixtures fluorescence spectra were done. The correlation fluorescent method of amines detection is proposed.

Real-time visualization of oxidative stress in a floating macrophyte Lemna minor L. exposed to cadmium, copper, menadione, and AAPH.

PubMed

Razinger, Jaka; Drinovec, Luka; Zrimec, Alexis

2010-12-01

An ultra-sensitive digital imaging system was employed to visualize oxidative stress in intact L. minor plants exposed to Cd, Cu, menadione, AAPH, and ascorbate in real time. The increase of ROS production was assessed by measuring the rate of fluorescence intensity increases of the test medium supplemented with a fluorescing probe (dichlorofluorescein diacetate). The addition of 100 μM CdCl₂ or 100 μM CuSO₄ to the growth medium resulted in a significant increase of medium fluorescence. Additionally, CuSO₄ caused a significantly higher fluorescence intensity than CdCl₂ did. A strong positive correlation (R² = 0.99) between menadione concentration and fluorescence intensity was observed. The positive correlation between AAPH concentration and fluorescence intensity was not as strong as in the case of menadione (R² = 0.81). Menadione induced a stronger oxidative stress than similar concentration of AAPH. The addition of 100 μM ascorbate to L. minor treated with 50 μM menadione significantly reduced the fluorescence intensity increase. A linear trend of the fluorescence increase was observed in all treatments, indicating that chemical-induced oxidative stress is a gradual process and that the applied concentrations of the chemicals caused a constant increased production of ROS with different intensities, depending on the treatment. This is the combined result of a gradual diminishing of antioxidant reserves and accumulating oxidative damage. The observed rates of ROS production were slower than those in the studies using cell cultures. Copyright © 2009 Wiley Periodicals, Inc.

Evaluating fluorescence spectroscopy as a tool to characterize cyanobacteria intracellular organic matter upon simulated release and oxidation in natural water.

PubMed

Korak, Julie A; Wert, Eric C; Rosario-Ortiz, Fernando L

2015-01-01

Intracellular organic matter (IOM) from cyanobacteria may be released into natural waters following cell death in aquatic ecosystems and during oxidation processes in drinking water treatment plants. Fluorescence spectroscopy was evaluated to identify the presence of IOM from three cyanobacteria species during simulated release into natural water and following oxidation processes (i.e. ozone, free chlorine, chloramine, chlorine dioxide). Peak picking and the fluorescence index (FI) were explored to determine which IOM components (e.g., pigments) provide unique and persistent fluorescence signatures with minimal interferences from the background dissolved organic matter (DOM) found in Colorado River water (CRW). When IOM was added to ultrapure water, the fluorescence signature of the three cyanobacteria species showed similarities to each other. Each IOM exhibited a strong protein-like fluorescence and fluorescence at Ex 370 nm and Em 460 nm (FDOM), where commercial fluorescence sensors monitor. All species also had strong phycobiliprotein fluorescence (i.e. phycocyanin or phycoerythrin) in the higher excitation range (500-650 nm). All three IOM isolates had FI values greater than 2. When IOM was added to CRW, phycobiliprotein fluorescence was quenched through interactions between IOM and CRW-DOM. Mixing IOM and CRW demonstrated that protein-like and FDOM intensity responses were not a simple superposition of the starting material intensities, indicating that interactions between IOM and CRW-DOM fluorescing moieties were important. Fluorescence intensity in all regions decreased with exposure to ozone, free chlorine, and chlorine dioxide, but the FI still indicated compositional differences compared to CRW-DOM. The phycobiliproteins in IOM are not promising as a surrogate for IOM release, because their fluorescence intensity is quenched by interactions with DOM and decreased during oxidation processes. Increases in both FDOM intensity and FI are viable qualitative indicators of IOM release in natural waters and following oxidation and may provide a more robust real-time indication of the presence of IOM than conventional dissolved organic carbon or UV absorbance measurements.

Semi-automated discrimination of retinal pigmented epithelial cells in two-photon fluorescence images of mouse retinas.

PubMed

Alexander, Nathan S; Palczewska, Grazyna; Palczewski, Krzysztof

2015-08-01

Automated image segmentation is a critical step toward achieving a quantitative evaluation of disease states with imaging techniques. Two-photon fluorescence microscopy (TPM) has been employed to visualize the retinal pigmented epithelium (RPE) and provide images indicating the health of the retina. However, segmentation of RPE cells within TPM images is difficult due to small differences in fluorescence intensity between cell borders and cell bodies. Here we present a semi-automated method for segmenting RPE cells that relies upon multiple weak features that differentiate cell borders from the remaining image. These features were scored by a search optimization procedure that built up the cell border in segments around a nucleus of interest. With six images used as a test, our method correctly identified cell borders for 69% of nuclei on average. Performance was strongly dependent upon increasing retinosome content in the RPE. TPM image analysis has the potential of providing improved early quantitative assessments of diseases affecting the RPE.

In-vivo characterization of endogenous porphyrin fluorescence from DMBA-treated Swiss Albino mice skin carcinogenesis for measuring tissue transformation

NASA Astrophysics Data System (ADS)

Ganesan, Singaravelu; Ebenezar, Jeyasingh; Hemamalini, Srinivasan; Aruna, Prakasa R.

2002-05-01

Steady state fluorescence spectroscopic characterization of endogenous porphyrin emission from DMBA treated skin carcinogenesis in Swiss albino mice was carried out. The emission of endogenous porphyrin from normal and abnormal skin tissues was studied both in the presence and absence of exogenous ALA to compare the resultant porphyrin emission characterictics. The mice skin is excited at 405nm and emission spectra are scanned from 430 to 700nm. The average fluorescence emission spectra of mice skin at normal and various tissues transformation conditions were found to be different. Two peaks around 460nm and 636nm were observed and they may be attributed to NADH, Elastin and collagen combination and endogenous porphyrin emission. The intensity at 636nm increases as the stage of the cancer increases. Although exogenous ALA enhances the PPIX level in tumor, the synthesis of PPIX was also found in normal surrounding skin, in fact, with higher concentration than that of tumor tissues.

The fluorescence intensities ratio is not a reliable parameter for evaluation of protein unfolding transitions.

PubMed

Žoldák, Gabriel; Jancura, Daniel; Sedlák, Erik

2017-06-01

Monitoring the fluorescence of proteins, particularly the fluorescence of intrinsic tryptophan residues, is a popular method often used in the analysis of unfolding transitions (induced by temperature, chemical denaturant, and pH) in proteins. The tryptophan fluorescence provides several suitable parameters, such as steady-state fluorescence intensity, apparent quantum yield, mean fluorescence lifetime, position of emission maximum that are often utilized for the observation of the conformational/unfolding transitions of proteins. In addition, the fluorescence intensities ratio at different wavelengths (usually at 330 nm and 350 nm) is becoming an increasingly popular parameter for the evaluation of thermal transitions. We show that, under certain conditions, the use of this parameter for the analysis of unfolding transitions leads to the incorrect determination of thermodynamic parameters characterizing unfolding transitions in proteins (e.g., melting temperature) and, hence, can compromise the hit identification during high-throughput drug screening campaigns. © 2017 The Protein Society.

Fluorescence lifetime imaging of skin cancer

NASA Astrophysics Data System (ADS)

Patalay, Rakesh; Talbot, Clifford; Munro, Ian; Breunig, Hans Georg; König, Karsten; Alexandrov, Yuri; Warren, Sean; Neil, Mark A. A.; French, Paul M. W.; Chu, Anthony; Stamp, Gordon W.; Dunsby, Chris

2011-03-01

Fluorescence intensity imaging and fluorescence lifetime imaging microscopy (FLIM) using two photon microscopy (TPM) have been used to study tissue autofluorescence in ex vivo skin cancer samples. A commercially available system (DermaInspect®) was modified to collect fluorescence intensity and lifetimes in two spectral channels using time correlated single photon counting and depth-resolved steady state measurements of the fluorescence emission spectrum. Uniquely, image segmentation has been used to allow fluorescence lifetimes to be calculated for each cell. An analysis of lifetime values obtained from a range of pigmented and non-pigmented lesions will be presented.

Fluorescence from polystyrene - Photochemical processes in polymeric systems, 7

NASA Technical Reports Server (NTRS)

Gupta, M. C.; Gupta, A.

1983-01-01

Results are presented for measurements of the fluorescence spectra of polystyrene in dilute solution and in pure solid films. It is determined that a major potential source of experimental error is the concurrent photooxidative degradation in air which may obscure fluorescence emission from monomeric sites in solid films at 25 C. The fluorescence spectra of oriented films are evaluated in terms of the monomer to excimer fluorescence intensity ratio and the excimer 'red shift'. The monomer to excimer fluorescence intensity ratio is determined to be significantly higher in fluid solution than in solid film.

Single-molecule spectroscopy reveals photosynthetic LH2 complexes switch between emissive states

PubMed Central

Schlau-Cohen, Gabriela S.; Wang, Quan; Southall, June; Cogdell, Richard J.; Moerner, W. E.

2013-01-01

Photosynthetic organisms flourish under low light intensities by converting photoenergy to chemical energy with near unity quantum efficiency and under high light intensities by safely dissipating excess photoenergy and deleterious photoproducts. The molecular mechanisms balancing these two functions remain incompletely described. One critical barrier to characterizing the mechanisms responsible for these processes is that they occur within proteins whose excited-state properties vary drastically among individual proteins and even within a single protein over time. In ensemble measurements, these excited-state properties appear only as the average value. To overcome this averaging, we investigate the purple bacterial antenna protein light harvesting complex 2 (LH2) from Rhodopseudomonas acidophila at the single-protein level. We use a room-temperature, single-molecule technique, the anti-Brownian electrokinetic trap, to study LH2 in a solution-phase (nonperturbative) environment. By performing simultaneous measurements of fluorescence intensity, lifetime, and spectra of single LH2 complexes, we identify three distinct states and observe transitions occurring among them on a timescale of seconds. Our results reveal that LH2 complexes undergo photoactivated switching to a quenched state, likely by a conformational change, and thermally revert to the ground state. This is a previously unobserved, reversible quenching pathway, and is one mechanism through which photosynthetic organisms can adapt to changes in light intensities. PMID:23776245

Single-molecule spectroscopy reveals photosynthetic LH2 complexes switch between emissive states.

PubMed

Schlau-Cohen, Gabriela S; Wang, Quan; Southall, June; Cogdell, Richard J; Moerner, W E

2013-07-02

Photosynthetic organisms flourish under low light intensities by converting photoenergy to chemical energy with near unity quantum efficiency and under high light intensities by safely dissipating excess photoenergy and deleterious photoproducts. The molecular mechanisms balancing these two functions remain incompletely described. One critical barrier to characterizing the mechanisms responsible for these processes is that they occur within proteins whose excited-state properties vary drastically among individual proteins and even within a single protein over time. In ensemble measurements, these excited-state properties appear only as the average value. To overcome this averaging, we investigate the purple bacterial antenna protein light harvesting complex 2 (LH2) from Rhodopseudomonas acidophila at the single-protein level. We use a room-temperature, single-molecule technique, the anti-Brownian electrokinetic trap, to study LH2 in a solution-phase (nonperturbative) environment. By performing simultaneous measurements of fluorescence intensity, lifetime, and spectra of single LH2 complexes, we identify three distinct states and observe transitions occurring among them on a timescale of seconds. Our results reveal that LH2 complexes undergo photoactivated switching to a quenched state, likely by a conformational change, and thermally revert to the ground state. This is a previously unobserved, reversible quenching pathway, and is one mechanism through which photosynthetic organisms can adapt to changes in light intensities.

Characterization of CDOM of river waters in China using fluorescence excitation-emission matrix and regional integration techniques

NASA Astrophysics Data System (ADS)

Zhao, Ying; Song, Kaishan; Shang, Yingxin; Shao, Tiantian; Wen, Zhidan; Lv, Lili

2017-08-01

The spatial characteristics of fluorescent dissolved organic matter (FDOM) components in river waters in China were first examined by excitation-emission matrix spectra and fluorescence regional integration (FRI) with the data collected during September to November between 2013 and 2015. One tyrosine-like (R1), one tryptophan-like (R2), one fulvic-like (R3), one microbial protein-like (R4), and one humic-like (R5) components have been identified by FRI method. Principal component analysis (PCA) was conducted to assess variations in the five FDOM components (FRί (ί = 1, 2, 3, 4, and 5)) and the humification index for all 194 river water samples. The average fluorescence intensities of the five fluorescent components and the total fluorescence intensities FSUM differed under spatial variation among the seven major river basins (Songhua, Liao, Hai, Yellow and Huai, Yangtze, Pearl, and Inflow Rivers) in China. When all the river water samples were pooled together, the fulvic-like FR3 and the humic-like FR5 showed a strong positive linear relationship (R2 = 0.90, n = 194), indicating that the two allochthonous FDOM components R3 and R5 may originate from similar sources. There is a moderate strong positive correlation between the tryptophan-like FR2 and the microbial protein-like FR4 (R2 = 0.71, n = 194), suggesting that parts of two autochthonous FDOM components R2 and R4 are likely from some common sources. However, the total allochthonous substance FR(3+5) and the total autochthonous substances FR(1+2+4) exhibited a weak correlation (R2 = 0.40, n = 194). Significant positive linear relationships between FR3 (R2 = 0.69, n = 194), FR5 (R2 = 0.79, n = 194), and chromophoric DOM (CDOM) absorption coefficient a(254) were observed, which demonstrated that the CDOM absorption was dominated by the allochthonous FDOM components R3 and R5.

Fluorescence lifetime spectroscopy for guided therapy of brain tumors.

PubMed

Butte, Pramod V; Mamelak, Adam N; Nuno, Miriam; Bannykh, Serguei I; Black, Keith L; Marcu, Laura

2011-01-01

This study evaluates the potential of time-resolved laser induced fluorescence spectroscopy (TR-LIFS) as intra-operative tool for the delineation of brain tumor from normal brain. Forty two patients undergoing glioma (WHO grade I-IV) surgery were enrolled in this study. A TR-LIFS prototype apparatus (gated detection, fast digitizer) was used to induce in-vivo fluorescence using a pulsed N2 laser (337 nm excitation, 0.7 ns pulse width) and to record the time-resolved spectrum (360-550 nm range, 10 nm interval). The sites of TR-LIFS measurement were validated by conventional histopathology (H&E staining). Parameters derived from the TR-LIFS data including intensity values and time-resolved intensity decay features (average fluorescence lifetime and Laguerre coefficients values) were used for tissue characterization and classification. 71 areas of tumor and normal brain were analyzed. Several parameters allowed for the differentiation of distinct tissue types. For example, normal cortex (N=35) and normal white matter (N=12) exhibit a longer-lasting fluorescence emission at 390 nm (τ390=2.12±0.10 ns) when compared with 460 nm (τ460=1.16±0.08 ns). High grade glioma (grades III and IV) samples (N=17) demonstrate emission peaks at 460 nm, with large variation at 390 nm while low grade glioma (I and II) samples (N=7) demonstrated a peak fluorescence emission at 460 nm. A linear discriminant algorithm allowed for the classification of low-grade gliomas with 100% sensitivity and 98% specificity. High-grade glioma demonstrated a high degree of heterogeneity thus reducing the discrimination accuracy of these tumors to 47% sensitivity and 94% specificity. Current findings demonstrate that TR-LIFS holds the potential to diagnose brain tumors intra-operatively and to provide a valuable tool for aiding the neurosurgeon-neuropathologist team in to rapidly distinguish between tumor and normal brain during surgery. Copyright © 2010 Elsevier Inc. All rights reserved.

Actin dynamics at the living cell submembrane imaged by total internal reflection fluorescence photobleaching.

PubMed Central

Sund, S E; Axelrod, D

2000-01-01

Although reversible chemistry is crucial to dynamical processes in living cells, relatively little is known about relevant chemical kinetic rates in vivo. Total internal reflection/fluorescence recovery after photobleaching (TIR/FRAP), an established technique previously demonstrated to measure reversible biomolecular kinetic rates at surfaces in vitro, is extended here to measure reversible biomolecular kinetic rates of actin at the cytofacial (subplasma membrane) surface of living cells. For the first time, spatial imaging (with a charge-coupled device camera) is used in conjunction with TIR/FRAP. TIR/FRAP imaging produces both spatial maps of kinetic parameters (off-rates and mobile fractions) and estimates of kinetic correlation distances, cell-wide kinetic gradients, and dependences of kinetic parameters on initial fluorescence intensity. For microinjected rhodamine actin in living cultured smooth muscle (BC3H1) cells, the unbinding rate at or near the cytofacial surface of the plasma membrane (averaged over the entire cell) is measured at 0.032 +/- 0.007 s(-1). The corresponding rate for actin marked by microinjected rhodamine phalloidin is very similar, 0.033 +/- 0.013 s(-1), suggesting that TIR/FRAP is reporting the dynamics of entire filaments or protofilaments. For submembrane fluorescence-marked actin, the intensity, off-rate, and mobile fraction show a positive correlation over a characteristic distance of 1-3 microm and a negative correlation over larger distances greater than approximately 7-14 microm. Furthermore, the kinetic parameters display a statistically significant cell-wide gradient, with the cell having a "fast" and "slow" end with respect to actin kinetics. PMID:10969025

Segmentation and detection of fluorescent 3D spots.

PubMed

Ram, Sundaresh; Rodríguez, Jeffrey J; Bosco, Giovanni

2012-03-01

The 3D spatial organization of genes and other genetic elements within the nucleus is important for regulating gene expression. Understanding how this spatial organization is established and maintained throughout the life of a cell is key to elucidating the many layers of gene regulation. Quantitative methods for studying nuclear organization will lead to insights into the molecular mechanisms that maintain gene organization as well as serve as diagnostic tools for pathologies caused by loss of nuclear structure. However, biologists currently lack automated and high throughput methods for quantitative and qualitative global analysis of 3D gene organization. In this study, we use confocal microscopy and fluorescence in-situ hybridization (FISH) as a cytogenetic technique to detect and localize the presence of specific DNA sequences in 3D. FISH uses probes that bind to specific targeted locations on the chromosomes, appearing as fluorescent spots in 3D images obtained using fluorescence microscopy. In this article, we propose an automated algorithm for segmentation and detection of 3D FISH spots. The algorithm is divided into two stages: spot segmentation and spot detection. Spot segmentation consists of 3D anisotropic smoothing to reduce the effect of noise, top-hat filtering, and intensity thresholding, followed by 3D region-growing. Spot detection uses a Bayesian classifier with spot features such as volume, average intensity, texture, and contrast to detect and classify the segmented spots as either true or false spots. Quantitative assessment of the proposed algorithm demonstrates improved segmentation and detection accuracy compared to other techniques. Copyright © 2012 International Society for Advancement of Cytometry.

Methods on observation of fluorescence micro-imaging for microalgae

NASA Astrophysics Data System (ADS)

Ou, Lin; Zhuang, Hui-ru; Chen, Rong; Lei, Jin-pin; Liao, Xiao-hua; Lin, Wen-suo

2007-11-01

Objective: Auto-fluorescence micro-imaging of microalgae are observed by using of laser scanning confocal microscopy (LSCM) and fluorescence microscopy, so as to investigate the effect of auto fluorescence alteration on growth of irradiated microalgae irradiated, meanwhile, the method of microalgae cells stained also to be studied. Methods: Platymonas subcordiformis, Phaeodactylum tricormutum and Isochyrsis zhanjiangensis cells are stained with acridine orange, and observed by fluorescence microscopy; the three types microalgae mentioned above are irradiated by Nd:YAP laser with 10w at 1341nm, irradiating time:12s, 30s, 35s and 55s, than to be cultured 6 days, and the auto fluorescence images and fluorescence spectra of algae cells are obtained by LSCM on lambda scan mode, at excitation 488nm (Ar + laser). Results: It is showed that the shapes and the structural features of microalgae cells stained can be seen clearly, and the cytoplasm and nucleus also can be observed. The chloroplasts in cell is bigger on promoting effects, conversely, it is to be mutilated, deformation and shrink. Contrast to the CK, the peak positions of fluorescence of algae cells irradiated is similar to the whole while the peak light intensity alters. On irradiation of promoting dose, however, the auto fluorescence intensity is enhanced more than control. Conclusions: The method of cell stained can be used to observed genetic material in microalgae. There are obvious effects for laser irradiating to chloroplasts in cells, the bigger chloroplasts the greater fluorescence intensity. Physiological incentive effects of microalgae irradiated can be given expression on fluorescence characteristics and fluorescence intensity alteration of cells.

Differences in fluorescence characteristics and bioavailability of water-soluble organic matter (WSOM) in sediments and suspended solids in Lihu Lake, China.

PubMed

Wang, Wenwen; Wang, Shuhang; Jiang, Xia; Zheng, Binghui; Zhao, Li; Zhang, Bo; Chen, Junyi

2018-05-01

The spectral characteristics, spatial distribution, and bioavailability of water-soluble organic matter (WSOM) in suspended solids and surface sediments of Lihu Lake, China, were investigated through excitation-emission matrix spectra and parallel factor analysis. The average content of dissolved organic carbon (DOC) in the sediments reached 643.28 ± 58.34 mg C/kg and that in suspended solids was 714.87 ± 69.24 mg C/kg. The fluorescence intensity of WSOM totaled 90.87 ± 5.65 and 115.42 ± 8.02 RU/g for the sediments and suspended solids, respectively. The DOC and fluorescence intensity of the WSOM showed an increasing trend moving from the west to the east of the lake. The WSOM in sediments and suspended solids contained two humic-like (C1 and C2) and one tryptophan-like (C3) components. These components had different fluorescent peaks and relative proportions. In the sediments, the relative proportions of C1, C2, and C3 were 33.71% ± 0.71, 26.83% ± 0.68, and 39.50% ± 0.71%, respectively. Meanwhile, C1 (35.77 ± 0.84%), C2 (34.07 ± 0.61%), and C3 (30.16 ± 0.75%) had similar relative percentages in suspended solids. The sediments had a lower humification index (3.02 ± 0.08) than the suspended solids (4.04 ± 0.15). Exchangeable nitrogen for the sediments and suspended solids was dominated by exchangeable ammonium nitrogen and soluble organic nitrogen, respectively. WSOM plays an important role in migration and transformation of nitrogen in sediments and suspended solids. The sediment-derived WSOM exhibited higher lability and biological activity than did the suspended solid-derived WSOM. The relative ratio of the intensity of protein-like fluorescent component to that of the humic-like one can be used as a reference index to evaluate the lability and biological activity of WSOM in sediments and suspended solids.

Nonradiative transport of atomic excitation in Na vapor

NASA Astrophysics Data System (ADS)

Zajonc, Arthur G.; Phelps, A. V.

1981-05-01

Measurements are reported which show the effect of nonradiative losses at a gas-window interface on the backscattered fluorescence intensity for Na vapor at frequencies in the vicinity of the resonance lines near 589 nm. The Na 3P12,32 states are excited with a low-intensity single-mode tunable dye laser at high Na densities and the frequency integral of the backscattered fluorescence intensity in the D1 and D2 lines is measured. As the laser is tuned through resonance, the loss of atomic excitation to the window appears as a sharp decrease in the frequency-integrated fluorescence intensity. For example, at 7×1020 atoms m-3 the fluorescence intensity decreases by a factor of 4 in a frequency interval of 4 GHz. Measured absolute fluorescence intensities versus laser frequency are compared with predictions made using the theory of Hummer and Kunasz which includes both radiative and nonradiative transport processes. The agreement between theory and experiment is remarkably good when one considers that the theory contains only one unknown coefficient, i.e., the reflection coefficient for excited atoms at the windows. In our case the excited atoms are assumed to be completely destroyed at the window.

Microfluidic flow cytometer for quantifying photobleaching of fluorescent proteins in cells

PubMed Central

Lubbeck, Jennifer L.; Dean, Kevin M.; Ma, Hairong; Palmer, Amy E.; Jimenez, Ralph

2012-01-01

Traditional flow cytometers are capable of rapid cellular assays on the basis of fluorescence intensity and light scatter. Microfluidic flow cytometers have largely followed the same path of technological development as their traditional counterparts, however the significantly smaller transport distance and resulting lower cell speeds in microchannels provides for the opportunity to detect novel spectroscopic signatures based on multiple, non-temporally-coincident excitation beams. Here, we characterize the design and operation of a cytometer with a 3-beam, probe/bleach/probe geometry, employing HeLa suspension cells expressing fluorescent proteins. The data collection rate exceeds 20 cells/s under a range of beam intensities (5 kW – 179 kW/cm2). The measured percent photobleaching (ratio of fluorescence intensities excited by the first and third beams: Sbeam3/Sbeam1) partially resolves a mixture of four red fluorescent proteins in mixed samples. Photokinetic simulations are presented and demonstrate that the percent photobleaching reflects a combination of the reversible and irreversible photobleaching kinetics. By introducing a photobleaching optical signature, which complements traditional fluorescence intensity-based detection, this method adds another dimension to multi-channel fluorescence cytometry, and provides a means for flow-cytometry-based screening of directed libraries of fluorescent protein photobleaching. PMID:22424298

Development and evaluation of a connective tissue phantom model for subsurface visualization of cancers requiring wide local excision

NASA Astrophysics Data System (ADS)

Samkoe, Kimberley S.; Bates, Brent D.; Tselepidakis, Niki N.; DSouza, Alisha V.; Gunn, Jason R.; Ramkumar, Dipak B.; Paulsen, Keith D.; Pogue, Brian W.; Henderson, Eric R.

2017-12-01

Wide local excision (WLE) of tumors with negative margins remains a challenge because surgeons cannot directly visualize the mass. Fluorescence-guided surgery (FGS) may improve surgical accuracy; however, conventional methods with direct surface tumor visualization are not immediately applicable, and properties of tissues surrounding the cancer must be considered. We developed a phantom model for sarcoma resection with the near-infrared fluorophore IRDye 800CW and used it to iteratively define the properties of connective tissues that typically surround sarcoma tumors. We then tested the ability of a blinded surgeon to resect fluorescent tumor-simulating inclusions with ˜1-cm margins using predetermined target fluorescence intensities and a Solaris open-air fluorescence imaging system. In connective tissue-simulating phantoms, fluorescence intensity decreased with increasing blood concentration and increased with increasing intralipid concentrations. Fluorescent inclusions could be resolved at ≥1-cm depth in all inclusion concentrations and sizes tested. When inclusion depth was held constant, fluorescence intensity decreased with decreasing volume. Using targeted fluorescence intensities, a blinded surgeon was able to successfully excise inclusions with ˜1-cm margins from fat- and muscle-simulating phantoms with inclusion-to-background contrast ratios as low as 2∶1. Indirect, subsurface FGS is a promising tool for surgical resection of cancers requiring WLE.

Synthesis of Water Dispersible Fluorescent Carbon Nanocrystals from Syzygium cumini Fruits for the Detection of Fe3+ Ion in Water and Biological Samples and Imaging of Fusarium avenaceum Cells.

PubMed

Bhamore, Jigna R; Jha, Sanjay; Singhal, Rakesh Kumar; Kailasa, Suresh Kumar

2017-01-01

In this work, water dispersible fluorescent carbon nanocrystals (NCs) were synthesized by a simple, green and low cost hydrothermal method using Syzygium cumini (jamun) as a carbon source at 180 °C for 6 h. The average size of carbon NCs was found to be 2.1 ± 0.5 nm and shown bright blue fluorescence when excited at 365 nm under UV lamp. The carbon NCs were characterized by spectroscopic (UV-visible and fluorescence, Fourier transform infrared and dynamic light scattering) and high resolution transmission electron microscopic techniques. The quantum yield of carbon NCs was found to be ~5.9 % at 438 nm emission wavelength when excited at 360 nm. It was noticed that none of the metal ions quenched the fluorescence intensity of carbon NCs at 438 nm except for Fe 3+ , indicating the formation of Fe 3+ ion-carbon NCs complexes. The linear range was observed in the concentration range of 0.01-100 μM with the corresponding detection limits of 0.001 μM, respectively. Furthermore, the carbon NCs were used as probes for imaging of fungal (Fusarium avenaceum) cells.

Quantitative Analysis of Subcellular Distribution of the SUMO Conjugation System by Confocal Microscopy Imaging.

PubMed

Mas, Abraham; Amenós, Montse; Lois, L Maria

2016-01-01

Different studies point to an enrichment in SUMO conjugation in the cell nucleus, although non-nuclear SUMO targets also exist. In general, the study of subcellular localization of proteins is essential for understanding their function within a cell. Fluorescence microscopy is a powerful tool for studying subcellular protein partitioning in living cells, since fluorescent proteins can be fused to proteins of interest to determine their localization. Subcellular distribution of proteins can be influenced by binding to other biomolecules and by posttranslational modifications. Sometimes these changes affect only a portion of the protein pool or have a partial effect, and a quantitative evaluation of fluorescence images is required to identify protein redistribution among subcellular compartments. In order to obtain accurate data about the relative subcellular distribution of SUMO conjugation machinery members, and to identify the molecular determinants involved in their localization, we have applied quantitative confocal microscopy imaging. In this chapter, we will describe the fluorescent protein fusions used in these experiments, and how to measure, evaluate, and compare average fluorescence intensities in cellular compartments by image-based analysis. We show the distribution of some components of the Arabidopsis SUMOylation machinery in epidermal onion cells and how they change their distribution in the presence of interacting partners or even when its activity is affected.

Efficient synthesis of highly fluorescent nitrogen-doped carbon dots for cell imaging using unripe fruit extract of Prunus mume

NASA Astrophysics Data System (ADS)

Atchudan, Raji; Edison, Thomas Nesakumar Jebakumar Immanuel; Sethuraman, Mathur Gopalakrishnan; Lee, Yong Rok

2016-10-01

Highly fluorescent nitrogen-doped carbon dots (N-CDs) were synthesized using the extract of unripe Prunus mume (P. mume) fruit by a simple one step hydrothermal-carbonization method. The N-CDs were synthesized at different pH ranges, 2.3, 5, 7, and 9. The pH of the P. mume extract was adjusted using an aqueous ammonia solution (25%). The optical properties of N-CDs were examined by UV-vis and fluorescence spectroscopy. The N-CDs synthesized at pH 9 emitted high fluorescence intensity compared to other obtained N-CDs. The N-CDs synthesized at pH 9 was further characterized by high resolution transmission electron microscopy (HR-TEM), X-ray diffraction (XRD), Raman spectroscopy, X-ray photoelectron spectroscopy (XPS), and Fourier transform-infra red (FT-IR) spectroscopy. HR-TEM showed that the average size of the synthesized N-CDs was approximately 9 nm and the interlayer distance was 0.21 nm, which was validated by XRD. The graphitic nature of the synthesized N-CDs were confirmed by Raman spectroscopy. XPS and FT-IR spectroscopy confirmed the doping of the nitrogen moiety over the synthesized CDs. The synthesized nitrogen doped CDs (N-CDs) were low toxicity and were used as a staining probe for fluorescence cell imaging.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chekini, M.; Bierwagen, J.; Cunningham, A.

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

Measurement of Zeta-Potential at Microchannel Wall by a Nanoscale Laser Induced Fluorescence Imaging

NASA Astrophysics Data System (ADS)

Kazoe, Yutaka; Sato, Yohei

A nanoscale laser induced fluorescence imaging was proposed by using fluorescent dye and the evanescent wave with total internal reflection of a laser beam. The present study focused on the two-dimensional measurement of zeta-potential at the microchannel wall, which is an electrostatic potential at the wall surface and a dominant parameter of electroosmotic flow. The evanescent wave, which decays exponentially from the wall, was used as an excitation light of the fluorescent dye. The fluorescent intensity detected by a CCD camera is closely related to the zeta-potential. Two kinds of fluorescent dye solution at different ionic concentrations were injected into a T-shaped microchannel, and formed a mixing flow field in the junction area. The two-dimensional distribution of zeta-potential at the microchannel wall in the pressure-driven flow field was measured. The obtained zeta-potential distribution has a transverse gradient toward the mixing flow field and was changed by the difference in the averaged velocity of pressure-driven flow. To understand the ion motion in the mixing flow field, the three-dimensional flow structure was analyzed by the velocity measurement using micron-resolution particle image velocimetry and the numerical simulation. It is concluded that the two-dimensional distribution of zeta-potential at the microchannel wall was dependent on the ion motion in the flow field, which was governed by the convection and molecular diffusion.

[Fluorescence characterization of dissolved organic matter in the East China Sea after diatom red tide dispersion].

PubMed

Zhuo, Peng-ji; Zhao, Wei-hong

2009-05-01

Fluorescence excitation-emission spectroscopy (EEMS) was employed to analyze the 3-dimensional fluorescence of dissolved organic matter in the East China Sea after diatom red tide dispersion. The relationships between fluorescence peak intensity, and salinity and chlorophyll-a were discussed. The centers of protein-like fluorescence peaks dispersed at Exmax/Exmax = 270-280/290-315 nm (Peak B), 220-230/290-305 nm (Peak D), 230-240/335-350 nm (Peak S) and 280/320 nm (Peak T). Two humic-like peaks appeared at 255-270/435-480 nm (Peak A)and 330-350/420-480 nm (Peak C). High tyrosine-like intensity was observed in diatom red tide dispersion area, and tryptophan-like fluorescence was also found which was lower. High FIB/FIS showed that diatom red tide produced much tyrosine-like matter during dispersion. Peaks S, A and C had positive correlation with one another, and their distributions were similar, which decreased with distance increasing away from the shore. Good negative correlations between peaks S, A and C and salinity suggested that Jiangsu-Zhejiang coastal water was the same source of them. Correlations between fluorescence peak intensity and chlorophyll-a were not remarkable enough to clear the relationship between fluorescence and living algal matter. It was supposed that the living algal matter contributed little to the fluorescence intensity of algal dispersion seawater.

Evaluation of microbial globin promoters for oxygen-limited processes using Escherichia coli.

PubMed

Lara, Alvaro R; Jaén, Karim E; Sigala, Juan-Carlos; Regestein, Lars; Büchs, Jochen

2017-01-01

Oxygen-responsive promoters can be useful for synthetic biology applications, however, information on their characteristics is still limited. Here, we characterized a group of heterologous microaerobic globin promoters in Escherichia coli . Globin promoters from Bacillus subtilis , Campylobacter jejuni , Deinococcus radiodurans , Streptomyces coelicolor , Salmonella typhi and Vitreoscilla stercoraria were used to express the FMN-binding fluorescent protein (FbFP), which is a non-oxygen dependent marker. FbFP fluorescence was monitored online in cultures at maximum oxygen transfer capacities (OTR max ) of 7 and 11 mmol L -1  h -1 . Different FbFP fluorescence intensities were observed and the OTR max affected the induction level and specific fluorescence emission rate (the product of the specific fluorescence intensity multiplied by the specific growth rate) of all promoters. The promoter from S. typhi displayed the highest fluorescence emission yields (the quotient of the fluorescence intensity divided by the scattered light intensity at every time-point) and rate, and together with the promoters from D. radiodurans and S. coelicolor , the highest induction ratios. These results show the potential of diverse heterologous globin promoters for oxygen-limited processes using E. coli .

Apparatus and method for determining the optical power passing through an optical fiber

DOEpatents

Toeppen, John S.

1995-01-01

An apparatus and method for determining the optical power transmitted through an optical fiber. The invention is based on measuring the intensity of the fluorescence produced by a doped segment of an optical fiber. The dopant is selected so that it emits light at a different wavelength than that responsible for producing the fluorescence. The doped segment is of sufficient length and dopant concentration to provide a detectable signal, but short enough to prevent the doped segment from serving as a gain medium, resulting in amplified spontaneous emission and excess fluorescence traveling along the optical fiber. The dopant material is excited by the optical signal carried by the fiber, causing a fluorescence. In the preferred embodiment the intensity of the fluorescence is proportional to the intensity of the propagating light. The signal power is then determined from the intensity of the fluorescence. The intensity of the fluorescent signal is measured by a photodetector placed so as to detect the light emitted through the side of the doped segment. The detector may wrap around the circumference of the fiber, or be placed to one side and used in conjunction with a reflector placed on the opposing side of the fiber. Filters may be used to shield the detector from other light sources and assist with accurately determining the optical power of the signal propagating within the fiber.

Apparatus and method for determining the optical power passing through an optical fiber

DOEpatents

Toeppen, John S.

1995-04-04

An apparatus and method for determining the optical power transmitted through an optical fiber. The invention is based on measuring the intensity of the fluorescence produced by a doped segment of an optical fiber. The dopant is selected so that it emits light at a different wavelength than that responsible for producing the fluorescence. The doped segment is of sufficient length and dopant concentration to provide a detectable signal, but short enough to prevent the doped segment from serving as a gain medium, resulting in amplified spontaneous emission and excess fluorescence traveling along the optical fiber. The dopant material is excited by the optical signal carried by the fiber, causing a fluorescence. In the preferred embodiment the intensity of the fluorescence is proportional to the intensity of the propagating light. The signal power is then determined from the intensity of the fluorescence. The intensity of the fluorescent signal is measured by a photodetector placed so as to detect the light emitted through the side of the doped segment. The detector may wrap around the circumference of the fiber, or be placed to one side and used in conjunction with a reflector placed on the opposing side of the fiber. Filters may be used to shield the detector from other light sources and assist with accurately determining the optical power of the signal propagating within the fiber.

Model-independent analysis of the orientation of fluorescent probes with restricted mobility in muscle fibers.

PubMed Central

Dale, R E; Hopkins, S C; an der Heide, U A; Marszałek, T; Irving, M; Goldman, Y E

1999-01-01

The orientation of proteins in ordered biological samples can be investigated using steady-state polarized fluorescence from probes conjugated to the protein. A general limitation of this approach is that the probes typically exhibit rapid orientational motion ("wobble") with respect to the protein backbone. Here we present a method for characterizing the extent of this wobble and for removing its effects from the available information about the static orientational distribution of the probes. The analysis depends on four assumptions: 1) the probe wobble is fast compared with the nanosecond time scale of its excited-state decay; 2) the orientational distributions of the absorption and emission transition dipole moments are cylindrically symmetrical about a common axis c fixed in the protein; 3) protein motions are negligible during the excited-state decay; 4) the distribution of c is cylindrically symmetrical about the director of the experimental sample. In a muscle fiber, the director is the fiber axis, F. All of the information on the orientational order of the probe that is available from measurements of linearly polarized fluorescence is contained in five independent polarized fluorescence intensities measured with excitation and emission polarizers parallel or perpendicular to F and with the propagation axis of the detected fluorescence parallel or perpendicular to that of the excitation. The analysis then yields the average second-rank and fourth-rank order parameters ( and ) of the angular distribution of c relative to F, and and , the average second-rank order parameters of the angular distribution for wobble of the absorption and emission transition dipole moments relative to c. The method can also be applied to other cylindrically ordered systems such as oriented lipid bilayer membranes and to processes slower than fluorescence that may be observed using longer-lived optically excited states. PMID:10049341

Development of a quantitative diagnostic method of estrogen receptor expression levels by immunohistochemistry using organic fluorescent material-assembled nanoparticles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gonda, Kohsuke, E-mail: gonda@med.tohoku.ac.jp; Miyashita, Minoru; Watanabe, Mika

2012-09-28

Highlights: Black-Right-Pointing-Pointer Organic fluorescent material-assembled nanoparticles for IHC were prepared. Black-Right-Pointing-Pointer New nanoparticle fluorescent intensity was 10.2-fold greater than Qdot655. Black-Right-Pointing-Pointer Nanoparticle staining analyzed a wide range of ER expression levels in tissue. Black-Right-Pointing-Pointer Nanoparticle staining enhanced the quantitative sensitivity for ER diagnosis. -- Abstract: The detection of estrogen receptors (ERs) by immunohistochemistry (IHC) using 3,3 Prime -diaminobenzidine (DAB) is slightly weak as a prognostic marker, but it is essential to the application of endocrine therapy, such as antiestrogen tamoxifen-based therapy. IHC using DAB is a poor quantitative method because horseradish peroxidase (HRP) activity depends on reaction time, temperature andmore » substrate concentration. However, IHC using fluorescent material provides an effective method to quantitatively use IHC because the signal intensity is proportional to the intensity of the photon excitation energy. However, the high level of autofluorescence has impeded the development of quantitative IHC using fluorescence. We developed organic fluorescent material (tetramethylrhodamine)-assembled nanoparticles for IHC. Tissue autofluorescence is comparable to the fluorescence intensity of quantum dots, which are the most representative fluorescent nanoparticles. The fluorescent intensity of our novel nanoparticles was 10.2-fold greater than quantum dots, and they did not bind non-specifically to breast cancer tissues due to the polyethylene glycol chain that coated their surfaces. Therefore, the fluorescent intensity of our nanoparticles significantly exceeded autofluorescence, which produced a significantly higher signal-to-noise ratio on IHC-imaged cancer tissues than previous methods. Moreover, immunostaining data from our nanoparticle fluorescent IHC and IHC with DAB were compared in the same region of adjacent tissues sections to quantitatively examine the two methods. The results demonstrated that our nanoparticle staining analyzed a wide range of ER expression levels with higher accuracy and quantitative sensitivity than DAB staining. This enhancement in the diagnostic accuracy and sensitivity for ERs using our immunostaining method will improve the prediction of responses to therapies that target ERs and progesterone receptors that are induced by a downstream ER signal.« less

Near infrared fluorescent chlorophyll nanoscale liposomes for sentinel lymph node mapping

PubMed Central

Fan, Lina; Wu, Qiang; Chu, Maoquan

2012-01-01

Background Sentinel lymph node (SLN) mapping using in vivo near infrared fluorescence imaging has attracted great attention during the past few years. Here we report on the early use of poorly water-soluble chlorophyll with near infrared fluorescence extracted from the leaf of Chimonanthus salicifolius, for mouse axillary SLN mapping. Methods and results To improve the water solubility and SLN targeting of the chlorophyll, we encapsulated the chlorophyll in nanoscale liposomes. The liposome-coated chlorophyll nanocomposites obtained were spherical in shape and had an average diameter of 21.7 ± 6.0 nm. The nanocomposites dispersed well in water, and in aqueous suspension they exhibited brighter near infrared fluorescence than chlorophyll alone. After incubation of the nanocomposites with normal liver cells (QSG-7701) and macrophage cells (Ana-1) for no more than 48 hours, there was no obvious reduction in cell viability. When the nanocomposites were injected intradermally into the paw of a mouse, the axillary SLN was found to be strongly fluorescent and was easily visualized in real time without a requirement for surgery. The intensity of the near infrared fluorescence emitted by the SLN was obviously brighter than that emitted by the SLN of another mouse that had been intradermally injected with chlorophyll alone. Conclusion Our data show that the liposome-coated chlorophyll nanocomposites could have great potential for clinical SLN mapping due to their lack of toxicity, bright near infrared fluorescence, and small diameter. PMID:22787402

Using water raman intensity to determine the effective excitation and emission path lengths of fluorophotometers for correcting fluorescence inner filter effect

USDA-ARS?s Scientific Manuscript database

Fluorescence and Raman inner filter effects (IFE) cause spectral distortion and nonlinearity between spectral signal intensity with increasing analyte concentration. Convenient and effective correction of fluorescence IFE has been an active research goal for decades. Presented herein is the finding ...

Two-photon sensitized recording materials for multilayer optical disk

NASA Astrophysics Data System (ADS)

Akiba, M.; Goto-Takahashi, E.; Takizawa, H.; Sasaki, T.; Mochizuki, H.; Mikami, T.; Kitahara, T.

2010-06-01

Two types of novel two-photon sensitized recording material writable at 405 nm and 522nm were developed. The fluorescent dye generation type (F-type) material consists of at least two-photon absorption dye (TPAD) and fluorescent dye precursor (FDP), which is non-fluorescent before two-photon recording and fluorescent after two-photon recording due to fluorescent dye generation. The fluorescence quench type (Q-type) material, on the other hand, consists of at least TPAD, fluorescent dye (FD) and fluorescent quencher precursor (QP), which is fluorescent before two-photon recording and the fluorescence intensity is reduced after two-photon recording at the recorded spot due to fluorescent quencher generation. Both types of material showed quadratic dependency of recording light intensity at 522 and 405 nm. A twenty-layer two-photon recording media was fabricated with the Q-type material, and two-photon recording and onephoton fluorescent signal readout was successfully conducted.

Highly sensitive and selective detection of Pb2+ using a turn-on fluorescent aptamer DNA silver nanoclusters sensor.

PubMed

Zhang, Baozhu; Wei, Chunying

2018-05-15

A novel turn-on fluorescent biosensor has been constructed using C-PS2.M-DNA-templated silver nanoclusters (Ag NCs) with an average diameter of about 1 nm. The proposed approach presents a low-toxic, simple, sensitive, and selective detection for Pb 2+ . The fluorescence intensity of C-PS2.M-DNA-Ag NCs enhances significantly in the presence of Pb 2+ , which is attributed to the special interaction between Pb 2+ and its aptamer DNA PS2.M. Pb 2+ induces the aptamer to form G-quadruplex and makes two darkish DNA/Ag NCs located at the 3' and 5' terminus close, resulting in the fluorescence light-up. Moreover, Pb 2+ can be detected as low as 3.0 nM within a good linear range from 5 to 50 nM (R = 0.9862). Furthermore, the application for detection of Pb 2+ in real water samples further demonstrates the reliability of the sensor. Thus, this sensor system shows a potential application for monitoring Pb 2+ in environmental samples. Copyright © 2018 Elsevier B.V. All rights reserved.

Deep brain optical measurements of cell type-specific neural activity in behaving mice.

PubMed

Cui, Guohong; Jun, Sang Beom; Jin, Xin; Luo, Guoxiang; Pham, Michael D; Lovinger, David M; Vogel, Steven S; Costa, Rui M

2014-01-01

Recent advances in genetically encoded fluorescent sensors enable the monitoring of cellular events from genetically defined groups of neurons in vivo. In this protocol, we describe how to use a time-correlated single-photon counting (TCSPC)-based fiber optics system to measure the intensity, emission spectra and lifetime of fluorescent biosensors expressed in deep brain structures in freely moving mice. When combined with Cre-dependent selective expression of genetically encoded Ca(2+) indicators (GECIs), this system can be used to measure the average neural activity from a specific population of cells in mice performing complex behavioral tasks. As an example, we used viral expression of GCaMPs in striatal projection neurons (SPNs) and recorded the fluorescence changes associated with calcium spikes from mice performing a lever-pressing operant task. The whole procedure, consisting of virus injection, behavior training and optical recording, takes 3-4 weeks to complete. With minor adaptations, this protocol can also be applied to recording cellular events from other cell types in deep brain regions, such as dopaminergic neurons in the ventral tegmental area. The simultaneously recorded fluorescence signals and behavior events can be used to explore the relationship between the neural activity of specific brain circuits and behavior.

Lanthanide-functionalized silver nanoparticles for detection of an anthrax biomarker and test paper fabrication

NASA Astrophysics Data System (ADS)

Tan, Hongliang; Li, Qian; Ma, Chanjiao; Song, Yonghai; Xu, Fugang; Chen, Shouhui; Wang, Li

2014-01-01

It is highly desirable to develop a simple and sensitive analytical method for detection of anthrax biomarker (dipicolinic acid, DPA) because of its dangerous nature. In this work, we developed a fluorescent sensor for DPA detection based on terbium ion-functionalized silver nanoparticles with an average size of 6.7 nm (AgNPs-Tb3+). The fluorescent intensity of Tb-DPA complex on the surface of AgNPs was two times higher than that of Tb-DPA complex alone in a solution phase due to the metal-enhanced fluorescence (MEF) effect of AgNPs. The proposed fluorescent sensor exhibits excellent selectivity and high sensitivity for DPA. Importantly, a test paper for DPA detection was fabricated for the first time by the integration of AgNPs-Tb3+ onto the nitrocellulose membrane. Owing to the MEF effect of AgNPs, the lowest detectable concentration of the test paper-integrated AgNPs-Tb3+ for DPA by naked eyes is 10 times lower than that of the test paper-integrated Tb3+ alone. We believe that the presented strategy may open up new avenues to the development of portable and robust-sensing platforms based on functional hybrid materials.

Spectral reconstruction for shifted-excitation Raman difference spectroscopy (SERDS).

PubMed

Guo, Shuxia; Chernavskaia, Olga; Popp, Jürgen; Bocklitz, Thomas

2018-08-15

Fluorescence emission is one of the major obstacles to apply Raman spectroscopy in biological investigations. It is usually several orders more intense than Raman scattering and hampers further analysis. In cases where the fluorescence emission is too intense to be efficiently removed via routine mathematical baseline correction algorithms, an alternative approach is needed. One alternative approach is shifted-excitation Raman difference spectroscopy (SERDS), where two Raman spectra are recorded with two slightly different excitation wavelengths. Ideally, the fluorescence emission at the two excitations does not change while the Raman spectrum shifts according to the excitation wavelength. Hence the fluorescence is removed in the difference of the two recorded Raman spectra. For better interpretability a spectral reconstruction procedure is necessary to recover the fluorescence-free Raman spectrum. This is challenging due to the intensity variations between the two recorded Raman spectra caused by unavoidable experimental changes as well as the presence of noise. Existent approaches suffer from drawbacks like spectral resolution loss, fluorescence residual, and artefacts. In this contribution, we proposed a reconstruction method based on non-negative least squares (NNLS), where the intensity variations between the two measurements are utilized in the reconstruction model. The method achieved fluorescence-free reconstruction on three real-world SERDS datasets without significant information loss. Thereafter, we quantified the performance of the reconstruction based on artificial datasets from four aspects: reconstructed spectral resolution, precision of reconstruction, signal-to-noise-ratio (SNR), and fluorescence residual. The artificial datasets were constructed with varied Raman to fluorescence intensity ratio (RFIR), SNR, full-width at half-maximum (FWHM), excitation wavelength shift, and fluorescence variation between the two spectra. It was demonstrated that the NNLS approach provides a faithful reconstruction without significantly changing the spectral resolution. Meanwhile, the reconstruction is almost robust to fluorescence variations between the two spectra. Last but not the least the SNR was improved after reconstruction for extremely noisy SERDS datasets. Copyright © 2018 Elsevier B.V. All rights reserved.

Characterising DOC, DON and DOP flux in two contrasting lowland UK catchments: impacts of contrasting source character and connectivity on reach scale DOM signature

NASA Astrophysics Data System (ADS)

Yates, C. A.; Johnes, P.; Spencer, R. G.

2012-12-01

Riverine DOM is a significant component of C, N and P transport from source to sea. Research to date has focused on characterising DOC in upland and boreal moorland and forested catchments. Here we present the results of an investigation of DOM character and DOC, DON and DOP flux relative to C, N and P flux in two contrasting lowland UK catchments: the Wylye and Millersford Brook. Both were sampled at daily frequency at 3 sites over a 2 year period, (2010-11 WY, 2011-12 WY) with fluorescence EEMs and UV-Vis determined weekly. The Wylye is a Chalk catchment, underlain by a major aquifer. It has predominantly calcareous brown earth soils, intensive arable agriculture, scattered farms and riparian villages with a major settlement in the lower reaches of the river. There are few natural organic sources in the catchment and flows are baseflow dominated with a BFI of 0.93. DOC (NPOC) concentrations averaged 2.59 mg C/l in water year 2011-12, while Total N concentrations averaged 10.0 mg N/l, with DON averaging 0.9 mg N/l, and Total P concentrations averaged 0.18 mg/l with DOP averaging 0.026 mg/l. Millersford Brook drains peaty soils over glacial sands and clays, with moorland and forestry as the dominant land uses. Flows are dominated by overland and subsurface quick flow through with a BFI of 0.34. There is some low intensity grazing, no fertiliser use and no dwellings in the headwaters of the catchment. As a result nutrient concentrations are lower, but C flux is higher, with mean annual concentrations in WY 2011-12 of 0.93 mg TN/l, 0.051 mg TP/l, 8.83 mg DOC/l, 0.55 mg DON/l and 0.029 mg DOP/l. DOM character in Millersford Brook is comparable for that observed in other catchments with peaty soils and low acid neutralising capacity. The character of DOM varies along the length of the river, with HMW compounds dominating the signal in the headwaters, and LMW fluorescence intensities added to the signal in the lower reaches of the river, where septic tank effluent discharges and livestock wastes from small areas of improved grassland contribute to the DOM flux. The signal for the Wylye differs markedly from that for Millersford Brook. DOM character varies markedly along the length of the river to a greater extent than in Millersford Brook, with fluorescence centres indicating a high proportion of LMW material peaking downstream from septic tank and STW discharges. However, in the summer months lack of dilution of bankside septic tank and STW discharges leads to a high concentration of N-rich LMW DOM in the river, with C:N ratios approaching 1:1. This contrasts with data recorded for Millersford Brook, where DOM flux is dominated by HMW matter with low N content and a much higher N:C ratio. The results from this programme illustrate the variability in the composition of DOM relative to source character, both downstream within catchments, and between catchments of differing character. The specific composition of DOM varies markedly between peaty catchments and lowland intensively farmed catchments in sedimentary environments, and along gradients of nutrient enrichment within each catchment, changing the ecological significance of the DOM flux relative to stoichiometric ratios for differing biotic groups

Ultrasound Induced Fluorescence of Nanoscale Liposome Contrast Agents

PubMed Central

Zhang, Qimei; Morgan, Stephen P.; O’Shea, Paul; Mather, Melissa L.

2016-01-01

A new imaging contrast agent is reported that provides an increased fluorescent signal upon application of ultrasound (US). Liposomes containing lipids labelled with pyrene were optically excited and the excimer fluorescence emission intensity was detected in the absence and presence of an ultrasound field using an acousto-fluorescence setup. The acousto-fluorescence dynamics of liposomes containing lipids with pyrene labelled on the fatty acid tail group (PyPC) and the head group (PyPE) were compared. An increase in excimer emission intensity following exposure to US was observed for both cases studied. The increased intensity and time constants were found to be different for the PyPC and PyPE systems, and dependent on the applied US pressure and exposure time. The greatest change in fluorescence intensity (130%) and smallest rise time constant (0.33 s) are achieved through the use of PyPC labelled liposomes. The mechanism underlying the observed increase of the excimer emission intensity in PyPC labelled liposomes is proposed to arise from the “wagging” of acyl chains which involves fast response and requires lower US pressure. This is accompanied by increased lipid lateral diffusivity at higher ultrasound pressures, a mechanism that is also active in the PyPE labelled liposomes. PMID:27467748

In vivo assessment of wound re-epithelialization by UV fluorescence excitation imaging

NASA Astrophysics Data System (ADS)

Wang, Ying; Ortega-Martinez, Antonio; Padilla-Martinez, Juan Pablo; Williams, Maura; Farinelli, William; Anderson, R. R.; Franco, Walfre

2017-02-01

Background and Objectives: We have previously demonstrated the efficacy of a non-invasive, non-contact, fast and simple but robust fluorescence imaging (u-FEI) method to monitor the healing of skin wounds in vitro. This system can image highly-proliferating cellular processes (295/340 nm excitation/emission wavelengths) to study epithelialization in a cultured wound model. The objective of the current work is to evaluate the suitability of u-FEI for monitoring wound re-epithelialization in vivo. Study Design: Full-thickness wounds were created in the tail of rats and imaged weekly using u-FEI at 295/340nm excitation/emission wavelengths. Histology was used to investigate the correlation between the spatial distribution and intensity of fluorescence and the extent of wound epithelialization. In addition, the expression of the nuclear protein Ki67 was used to confirm the association between the proliferation of keratinocyte cells and the intensity of fluorescence. Results: Keratinocytes forming neo-epidermis exhibited higher fluorescence intensity than the keratinocytes not involved in re-epithelialization. In full-thickness wounds the fluorescence first appeared at the wound edge where keratinocytes initiated the epithelialization process. Fluorescence intensity increased towards the center as the keratinocytes partially covered the wound. As the wound healed, fluorescence decreased at the edges and was present only at the center as the keratinocytes completely covered the wound at day 21. Histology demonstrated that changes in fluorescence intensity from the 295/340nm band corresponded to newly formed epidermis. Conclusions: u-FEI at 295/340nm allows visualization of proliferating keratinocyte cells during re-epithelialization of wounds in vivo, potentially providing a quantitative, objective and simple method for evaluating wound closure in the clinic.

Novel functionalized fluorescent polymeric nanoparticles for immobilization of biomolecules

NASA Astrophysics Data System (ADS)

Jain, Swati; Chattopadhyay, Sruti; Jackeray, Richa; Abid, C. K. V. Zainul; Singh, Harpal

2013-07-01

Novel, size controlled fluorescent polymeric nanoparticles (FPNP) were synthesized having acetoacetoxy functionality on the surface for immobilization of biomolecules which can be utilized as biomarkers and labels in fluoroimmunoassays. Core-shell nanoparticles of poly(styrene, St-methyl methacrylate, MMA-acetoacetoxy ethyl methacrylate, AAEM), stabilized by various concentrations of surfactant, sodium lauryl sulphate (SLS), were obtained by facile miniemulsion co-polymerization encapsulated with pyrene molecules in their hydrophobic core. Analytical, spectroscopic and imaging characterization techniques revealed the formation of stable, monodisperse, spherical nano sized particles exhibiting high luminescence properties. Particles with 1% SLS (S1) showed good dispersion stability and fluorescence intensity and were chosen as ideal candidates for further immobilization studies. Steady state fluorescence studies showed 10 times higher fluorescence intensity of S1 nanoparticles than that of pyrene solution in solvent-toluene at the same concentration. Environmental factors such as pH, ionic strength and time were found to have no effect on fluorescence intensity of FPNPs. Surface β-di-ketone groups were utilized for the covalent immobilization of enzyme conjugated antibodies without any activation or pre-treatment of nanoparticles.Novel, size controlled fluorescent polymeric nanoparticles (FPNP) were synthesized having acetoacetoxy functionality on the surface for immobilization of biomolecules which can be utilized as biomarkers and labels in fluoroimmunoassays. Core-shell nanoparticles of poly(styrene, St-methyl methacrylate, MMA-acetoacetoxy ethyl methacrylate, AAEM), stabilized by various concentrations of surfactant, sodium lauryl sulphate (SLS), were obtained by facile miniemulsion co-polymerization encapsulated with pyrene molecules in their hydrophobic core. Analytical, spectroscopic and imaging characterization techniques revealed the formation of stable, monodisperse, spherical nano sized particles exhibiting high luminescence properties. Particles with 1% SLS (S1) showed good dispersion stability and fluorescence intensity and were chosen as ideal candidates for further immobilization studies. Steady state fluorescence studies showed 10 times higher fluorescence intensity of S1 nanoparticles than that of pyrene solution in solvent-toluene at the same concentration. Environmental factors such as pH, ionic strength and time were found to have no effect on fluorescence intensity of FPNPs. Surface β-di-ketone groups were utilized for the covalent immobilization of enzyme conjugated antibodies without any activation or pre-treatment of nanoparticles. Electronic supplementary information (ESI) available: Resulting ATR-FTIR spectrum and procedure to study fluorescence of nanoparticles, effect of particle size, concentration, pH, ionic strength and time on Fl intensity of FPNP. See DOI: 10.1039/c3nr34100c

Optical Properties of Nano-Spherical Gold Doped Dye Solution Hybrid

NASA Astrophysics Data System (ADS)

Hoa, D. Q.; Lien, N. T. H.; Ha, C. V.; Nhung, T. H.; Long, P.

2011-03-01

Gold nanoparticles with average diameter of 16 nm which are coated with Cetrimonium Bromide (CTAB) by chemical method are dissolved in dye solution at different concentrations. The absorption spectra of the dye mixture appeared almost unchanged at low concentrations of gold nanoparticles (around 1×1014 cm-3) despite its fluorescence intensity increased many-fold. Energy transfer from gold nanoparticles to dye molecules occurs through surface plasmon resonance(SPR). The fluorescence of rhodamine 610 (Rh610) dye molecules co-adsorbed within 16 nm gold nanoparticles assemblies can be useful for enhancing gain in lasing emission. An increase in laser efficiency by a factor of one and half times stronger compared to the single Rh610 dye suggest the potential of using the mixture of rhodamine dye with gold nanoparticles as laser medium in the configuration of quenching distributed feedback dye laser.

Effect of Starvation and the Viable-but-Nonculturable State on Green Fluorescent Protein (GFP) Fluorescence in GFP-Tagged Pseudomonas fluorescens A506

PubMed Central

Lowder, M.; Unge, A.; Maraha, N.; Jansson, J. K.; Swiggett, J.; Oliver, J. D.

2000-01-01

The green fluorescent protein (GFP) gene, gfp, of the jellyfish Aequorea victoria is being used as a reporter system for gene expression and as a marker for tracking prokaryotes and eukaryotes. Cells that have been genetically altered with the gfp gene produce a protein that fluoresces when it is excited by UV light. This unique phenotype allows gfp-tagged cells to be specifically monitored by nondestructive means. In this study we determined whether a gfp-tagged strain of Pseudomonas fluorescens continued to fluoresce under conditions under which the cells were starved, viable but nonculturable (VBNC), or dead. Epifluorescent microscopy, flow cytometry, and spectrofluorometry were used to measure fluorescence intensity in starved, VBNC, and dead or dying cells. Results obtained by using flow cytometry indicated that microcosms containing VBNC cells, which were obtained by incubation under stress conditions (starvation at 37.5°C), fluoresced at an intensity that was at least 80% of the intensity of nonstressed cultures. Similarly, microcosms containing starved cells incubated at 5 and 30°C had fluorescence intensities that were 90 to 110% of the intensity of nonstressed cells. VBNC cells remained fluorescent during the entire 6-month incubation period. In addition, cells starved at 5 or 30°C remained fluorescent for at least 11 months. Treatment of the cells with UV light or incubation at 39 or 50°C resulted in a loss of GFP from the cells. There was a strong correlation between cell death and leakage of GFP from the cells, although the extent of leakage varied depending on the treatment. Most dead cells were not GFP fluorescent, but a small proportion of the dead cells retained some GFP at a lower concentration than the concentration in live cells. Our results suggest that gfp-tagged cells remain fluorescent following starvation and entry into the VBNC state but that fluorescence is lost when the cells die, presumably because membrane integrity is lost. PMID:10919764

Polymerase chain reaction amplification of DNA from aged blood stains: quantitative evaluation of the "suitability for purpose" of four filter papers as archival media.

PubMed

Kline, Margaret C; Duewer, David L; Redman, Janette W; Butler, John M; Boyer, David A

2002-04-15

In collaboration with the Armed Forces Institute of Pathology's Department of Defense DNA Registry, the National Institute of Standards and Technology recently evaluated the performance of a short tandem repeat multiplex with dried whole blood stains on four different commercially available identification card matrixes. DNA from 70 stains that had been stored for 19 months at ambient temperature was extracted or directly amplified and then processed using routine methods. All four storage media provided fully typeable (qualitatively identical) samples. After standardization, the average among-locus fluorescence intensity (electropherographic peak height or area) provided a suitable metric for quantitative analysis of the relative amounts of amplifiable DNA in an archived sample. The amounts of DNA in Chelex extracts from stains on two untreated high-purity cotton linter pulp papers and a paper treated with a DNA-binding coating were essentially identical. Average intensities for the aqueous extracts from a paper treated with a DNA-releasing coating were somewhat lower but also somewhat less variable than for the Chelex extracts. Average intensities of directly amplified punches of the DNA-binding paper were much larger but somewhat more variable than the Chelex extracts. Approximately 25% of the observed variation among the intensity measurements is shared among the four media and thus can be attributed to intrinsic variation in white blood count among the donors. All of the evaluated media adequately "bank" forensically useful DNA in well-dried whole blood stains for at least 19 months at ambient temperature.

Fluorescence multispectral imaging-based diagnostic system for atherosclerosis.

PubMed

Ho, Cassandra Su Lyn; Horiuchi, Toshikatsu; Taniguchi, Hiroaki; Umetsu, Araya; Hagisawa, Kohsuke; Iwaya, Keiichi; Nakai, Kanji; Azmi, Amalina; Zulaziz, Natasha; Azhim, Azran; Shinomiya, Nariyoshi; Morimoto, Yuji

2016-08-20

Composition of atherosclerotic arterial walls is rich in lipids such as cholesterol, unlike normal arterial walls. In this study, we aimed to utilize this difference to diagnose atherosclerosis via multispectral fluorescence imaging, which allows for identification of fluorescence originating from the substance in the arterial wall. The inner surface of extracted arteries (rabbit abdominal aorta, human coronary artery) was illuminated by 405 nm excitation light and multispectral fluorescence images were obtained. Pathological examination of human coronary artery samples were carried out and thickness of arteries were calculated by measuring combined media and intima thickness. The fluorescence spectra in atherosclerotic sites were different from those in normal sites. Multiple regions of interest (ROI) were selected within each sample and a ratio between two fluorescence intensity differences (where each intensity difference is calculated between an identifier wavelength and a base wavelength) from each ROI was determined, allowing for discrimination of atherosclerotic sites. Fluorescence intensity and thickness of artery were found to be significantly correlated. These results indicate that multispectral fluorescence imaging provides qualitative and quantitative evaluations of atherosclerosis and is therefore a viable method of diagnosing the disease.

Method of quantitating dsDNA

DOEpatents

Stark, Peter C.; Kuske, Cheryl R.; Mullen, Kenneth I.

2002-01-01

A method for quantitating dsDNA in an aqueous sample solution containing an unknown amount of dsDNA. A first aqueous test solution containing a known amount of a fluorescent dye-dsDNA complex and at least one fluorescence-attenutating contaminant is prepared. The fluorescence intensity of the test solution is measured. The first test solution is diluted by a known amount to provide a second test solution having a known concentration of dsDNA. The fluorescence intensity of the second test solution is measured. Additional diluted test solutions are similarly prepared until a sufficiently dilute test solution having a known amount of dsDNA is prepared that has a fluorescence intensity that is not attenuated upon further dilution. The value of the maximum absorbance of this solution between 200-900 nanometers (nm), referred to herein as the threshold absorbance, is measured. A sample solution having an unknown amount of dsDNA and an absorbance identical to that of the sufficiently dilute test solution at the same chosen wavelength is prepared. Dye is then added to the sample solution to form the fluorescent dye-dsDNA-complex, after which the fluorescence intensity of the sample solution is measured and the quantity of dsDNA in the sample solution is determined. Once the threshold absorbance of a sample solution obtained from a particular environment has been determined, any similarly prepared sample solution taken from a similar environment and having the same value for the threshold absorbance can be quantified for dsDNA by adding a large excess of dye to the sample solution and measuring its fluorescence intensity.

Optical gating with organic building blocks. A quantitative model for the fluorescence modulation of photochromic perylene bisimide dithienylcyclopentene triads

PubMed Central

Pärs, Martti; Gradmann, Michael; Gräf, Katja; Bauer, Peter; Thelakkat, Mukundan; Köhler, Jürgen

2014-01-01

We investigated the capability of molecular triads, consisting of two strong fluorophores that were covalently linked to a photochromic molecule, for optical gating. Therefore we monitored the fluorescence intensity of the fluorophores as a function of the isomeric state of the photoswitch. From the analysis of our data we develop a kinetic model that allows us to predict quantitatively the degree of the fluorescence modulation as a function of the mutual intensities of the lasers that are used to induce the fluorescence and the switching of the photochromic unit. We find that the achievable contrast for the modulation of the fluorescence depends mainly on the intensity ratio of the two light beams and appears to be very robust against absolute changes of these intensities. The latter result provides valuable information for the development of all-optical circuits which would require to handle different signal strengths for the input and output levels. PMID:24614963

Intensity and pressure dependence of resonance fluorescence of OH induced by a tunable UV laser

NASA Technical Reports Server (NTRS)

Killinger, D. K.; Wang, C. C.; Hanabusa, M.

1976-01-01

The intensity and pressure dependence of the fluorescence spectrum of OH in the presence of N2 and H2O molecules was studied. Saturation of the absorption transition was observed at low pressures, and the corresponding fluorescence signal was found to vary as the square root of the exciting intensity. This observed dependence agreed with the predicted dependence which took into account the presence of laser modes in the spectrum of the exciting radiation. With full laser power incident, a saturation parameter as high as 3 x 10 to the 5th was observed. The fluorescence spectrum was found to peak at 3145 and at 3090 A, with the relative peak intensities dependent upon gas pressures and upon the particular rotational electronic transition used for excitation. It is concluded that vibrational relaxation of the electronically excited OH due to water vapor in the system plays a dominant role in determining the observed fluorescence spectrum.

A fluorescence spectroscopy study of traditional Chinese medicine Angelica

NASA Astrophysics Data System (ADS)

Zhao, Hongyan; Song, Feng; Liu, Shujing; Chen, Guiyang; Wei, Chen; Liu, Yanling; Liu, Jiadong

2013-10-01

By measuring the fluorescence spectra of Chinese medicine (CM) Angelica water solutions with different concentrations from 0.025 to 2.5 mg/mL, results showed that the fluorescence intensity was proportional to the concentration. Through fluorescence spectra of Angelica solution under different pH values, results indicated coumarin compounds were the active ingredients of Angelica. We also observed fluorescence quenching of the Angelica solution in the presence of spherical silver nanoparticles with radius of 12 nm. Keeping a certain value for the volume of the silver nanoparticles, the fluorescence intensity at 402 nm was linearly proportional to the Angelica in the range of 1-3 mg/mL.

Spectral line discriminator for passive detection of fluorescence

NASA Technical Reports Server (NTRS)

Kebabian, Paul L. (Inventor)

1996-01-01

A method and apparatus for detecting fluorescence from sunlit plants is based on spectral line discrimination using the A-band and B-band absorption of atmospheric oxygen. Light from a plant including scattered sunlight and the fluorescence from chlorophyll is passed through a chopper into a cell containing low-pressure, high-purity oxygen. A-band or B-band wavelengths present in the light are absorbed by the oxygen in the cell. When the chopper is closed, the absorbed light is remitted as fluorescence into a detector. The intensity of the fluorescence from the oxygen is proportional to the intensity of fluorescence from the plant.

Aryl-modified graphene quantum dots with enhanced photoluminescence and improved pH tolerance

NASA Astrophysics Data System (ADS)

Luo, Peihui; Ji, Zhe; Li, Chun; Shi, Gaoquan

2013-07-01

Chemical modification is an important technique to modulate the chemical and optical properties of graphene quantum dots (GQDs). In this paper, we report a versatile diazonium chemistry method to graft aryl groups including phenyl, 4-carboxyphenyl, 4-sulfophenyl and 5-sulfonaphthyl to GQDs via Gomberg-Bachmann reaction. The aryl-modified GQDs are nanocrystals with lateral dimensions in the range of 2-4 nm and an average thickness lower than 1 nm. Upon chemical modification with aryl groups, the photoluminescence (PL) bands of GQDs were tuned in the range of 418 and 447 nm, and their fluorescence quantum yields (QYs) were increased for up to about 6 times. Furthermore, the aryl-modified GQDs exhibited stable PL (both intensity and peak position) in a wide pH window of 1-11. The mechanism of improving the PL properties of GQDs by aryl-modification was also discussed.Chemical modification is an important technique to modulate the chemical and optical properties of graphene quantum dots (GQDs). In this paper, we report a versatile diazonium chemistry method to graft aryl groups including phenyl, 4-carboxyphenyl, 4-sulfophenyl and 5-sulfonaphthyl to GQDs via Gomberg-Bachmann reaction. The aryl-modified GQDs are nanocrystals with lateral dimensions in the range of 2-4 nm and an average thickness lower than 1 nm. Upon chemical modification with aryl groups, the photoluminescence (PL) bands of GQDs were tuned in the range of 418 and 447 nm, and their fluorescence quantum yields (QYs) were increased for up to about 6 times. Furthermore, the aryl-modified GQDs exhibited stable PL (both intensity and peak position) in a wide pH window of 1-11. The mechanism of improving the PL properties of GQDs by aryl-modification was also discussed. Electronic supplementary information (ESI) available: Fluorescence quantum yield measurements, estimation of grafting ratio, TEM images, FTIR spectra, PL spectra and zeta potentials. See DOI: 10.1039/c3nr02156d

Development and evaluation of a connective tissue phantom model for subsurface visualization of cancers requiring wide local excision.

PubMed

Samkoe, Kimberley S; Bates, Brent D; Tselepidakis, Niki N; DSouza, Alisha V; Gunn, Jason R; Ramkumar, Dipak B; Paulsen, Keith D; Pogue, Brian W; Henderson, Eric R

2017-12-01

Wide local excision (WLE) of tumors with negative margins remains a challenge because surgeons cannot directly visualize the mass. Fluorescence-guided surgery (FGS) may improve surgical accuracy; however, conventional methods with direct surface tumor visualization are not immediately applicable, and properties of tissues surrounding the cancer must be considered. We developed a phantom model for sarcoma resection with the near-infrared fluorophore IRDye 800CW and used it to iteratively define the properties of connective tissues that typically surround sarcoma tumors. We then tested the ability of a blinded surgeon to resect fluorescent tumor-simulating inclusions with ∼1-cm margins using predetermined target fluorescence intensities and a Solaris open-air fluorescence imaging system. In connective tissue-simulating phantoms, fluorescence intensity decreased with increasing blood concentration and increased with increasing intralipid concentrations. Fluorescent inclusions could be resolved at ≥1-cm depth in all inclusion concentrations and sizes tested. When inclusion depth was held constant, fluorescence intensity decreased with decreasing volume. Using targeted fluorescence intensities, a blinded surgeon was able to successfully excise inclusions with ∼1-cm margins from fat- and muscle-simulating phantoms with inclusion-to-background contrast ratios as low as 2∶1. Indirect, subsurface FGS is a promising tool for surgical resection of cancers requiring WLE. (2017) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).

Increased metabolic activity detected by FLIM in human breast cancer cells with desmoplastic reaction: a pilot study

NASA Astrophysics Data System (ADS)

Natal, Rodrigo de Andrade; Pelegati, Vitor B.; Bondarik, Caroline; Mendonça, Guilherme R.; Derchain, Sophie F.; Lima, Carmen P.; Cesar, Carlos L.; Sarian, Luís. O.; Vassallo, José

2015-07-01

Introduction: In breast cancer (BC), desmoplastic reaction, assembled primarily by fibroblasts, is associated with unfavorable prognosis, but the reason of this fact remains still unclear. In this context, nonlinear optics microscopy, including Fluorescence Lifetime Imaging Microscopy (FLIM), has provided advancement in cellular metabolism research. In this paper, our purpose is to differentiate BC cells metabolism with or without contact to desmoplastic reaction. Formalin fixed, paraffin embedded samples were used at different points of hematoxylin stained sections. Methodology: Sections from 14 patients with invasive ductal breast carcinoma were analyzed with FLIM methodology to NAD(P)H and FAD fluorescence lifetime on a Confocal Upright LSM780 NLO device (Carl Zeiss AG, Germany). Quantification of the fluorescence lifetime and fluorescence intensity was evaluated by SPC Image software (Becker &Hickl) and ImageJ (NIH), respectively. Optical redox ratio was calculated by dividing the FAD fluorescence intensity by NAD(P)H fluorescence intensity. Data value for FLIM measurements and fluorescence intensities were calculated using Wilcoxon test; p< 0.05 was considered significant. Results: BC cells in contact with desmoplastic reaction presented a significantly lower NAD(P)H and FAD fluorescence lifetime. Furthermore, optical redox ratio was also lower in these tumor cells. Conclusion: Our results suggest that contact of BC cells with desmoplastic reaction increase their metabolic activity, which might explain the adverse prognosis of cases associated with higher peritumoral desmoplastic reaction.

Effects of alcohols on fluorescence intensity and color of a discharged-obelin-based biomarker.

PubMed

Alieva, Roza R; Belogurova, Nadezhda V; Petrova, Alena S; Kudryasheva, Nadezhda S

2014-05-01

Photoproteins are responsible for bioluminescence of marine coelenterates; bioluminescent and fluorescent biomarkers based on photoproteins are useful for monitoring of calcium-dependent processes in medical investigations. Here, we present the analysis of intensity and color of light-induced fluorescence of Ca(2+)-discharged photoprotein obelin in the presence of alcohols (ethanol and glycerol). Complex obelin spectra obtained at different concentrations of the alcohols at 350- and 280-nm excitation (corresponding to polypeptide-bound coelenteramide and tryptophan absorption regions) were deconvoluted into Gaussian components; fluorescent intensity and contributions of the components to experimental spectra were analyzed. Five Gaussian components were found in different spectral regions-ultraviolet (tryptophan emission), blue-green (coelenteramide emission), and red (hypothetical indole-coelenteramide exciplex emission). Inhibition coefficients and contributions of the components to experimental fluorescent spectra showed that presence of alcohols increased contributions of ultraviolet, violet, and red components, but decreased contributions of components in the blue-green region. The effects were related to (1) changes of proton transfer efficiency in fluorescent S*1 state of coelenteramide in the obelin active center and (2) formation of indole-coelenteramide exciplex at 280-nm photoexcitation. The data show that variation of fluorescence color and intensity in the presence of alcohols and dependence of emission spectra on excitation wavelength should be considered while applying the discharged obelin as a fluorescence biomarker.

Segmentation-based retrospective shading correction in fluorescence microscopy E. coli images for quantitative analysis

NASA Astrophysics Data System (ADS)

Mai, Fei; Chang, Chunqi; Liu, Wenqing; Xu, Weichao; Hung, Yeung S.

2009-10-01

Due to the inherent imperfections in the imaging process, fluorescence microscopy images often suffer from spurious intensity variations, which is usually referred to as intensity inhomogeneity, intensity non uniformity, shading or bias field. In this paper, a retrospective shading correction method for fluorescence microscopy Escherichia coli (E. Coli) images is proposed based on segmentation result. Segmentation and shading correction are coupled together, so we iteratively correct the shading effects based on segmentation result and refine the segmentation by segmenting the image after shading correction. A fluorescence microscopy E. Coli image can be segmented (based on its intensity value) into two classes: the background and the cells, where the intensity variation within each class is close to zero if there is no shading. Therefore, we make use of this characteristics to correct the shading in each iteration. Shading is mathematically modeled as a multiplicative component and an additive noise component. The additive component is removed by a denoising process, and the multiplicative component is estimated using a fast algorithm to minimize the intra-class intensity variation. We tested our method on synthetic images and real fluorescence E.coli images. It works well not only for visual inspection, but also for numerical evaluation. Our proposed method should be useful for further quantitative analysis especially for protein expression value comparison.

[Acidity and temperature effect on the fluorescence characteristics of hydraulic oils and lubricants].

PubMed

Deng, Hu; Zhou, Xun; Shang, Li-ping; Zhang, Ze-lin; Wang, Shun-li

2014-12-01

By analyzing HyJet V phosphate ester hydraulic oil environmental impacts (oil, etc.) and confounding factors (pH, temperature, etc.), the feasibility was studied for the fluorescence detection of aircraft hydraulic oil leaks. By using the fluorescence spectrophotometer at various acidities and temperatures, the fluorescence properties of HyJet V phosphate ester hydraulic oil, Jet Oil II lubricant and 2197 lubricant were gained. The experimental results are shown as following: The fluorescence peaks of HyJet V phosphate ester hydraulic oil, Jet Oil II lubricant and 2197 lubricant are at 362, 405 and 456 nm, respectively. The impact of temperature on HyJet V phosphate ester hydraulic oil is less effective; Jet Oil II lubricant and 2197 lubricant fluorescence intensity decreases with increasing temperature. When acidity increases, the fluorescence peak of HyJet V phosphate ester hydraulic oil gradient shifts from 370 to 362 nm, and the fluorescence intensity decreases; the fluorescence peak of Jet Oil II lubricant is always 405 nm, while the fluorescence intensity decreases; the fluorescence peak of 2197 lubricant at 456 nm red shifts to 523 nm, and double fluorescence peaks appeare. The results are shown as following: under the influence of the environment and interference factors, the fluorescence characteristics of HyJet V phosphate ester hydraulic oil remain unchanged, and distinguish from Jet Oil II lubricant and 2197 lubricant. Therefore, the experiments indicate that the detection of HyJet V phosphate ester hydraulic oil leak is feasible by using fluorescence method.

Detection of bacterial infection of agave plants by laser-induced fluorescence

NASA Astrophysics Data System (ADS)

Cervantes-Martinez, Jesus; Flores-Hernandez, Ricardo; Rodriguez-Garay, Benjamin; Santacruz-Ruvalcaba, Fernando

2002-05-01

Greenhouse-grown plants of Agave tequilana Weber var. azul were inoculated with Erwinia carotovora, the causal agent of stem soft rot. We investigated the laser-induced fluorescence (LIF) of agave plants to determine whether LIF can be used as a noninvasive sensing tool for pathological studies. The LIF technique was also investigated as a means of detecting the effect of the polyamine biosynthesis inhibitor beta-hydroxyethylhydrazine as a bactericide against the pathogenic bacterium Erwinia carotovora. A He-Ne laser at 632.8 nm was used as the excitation source, and in vivo fluorescence emission spectra were recorded in the 660-790-range. Fluorescence maxima were at 690 and 740 nm. The infected plants that were untreated with the bactericide showed a definite increase in fluorescence intensity at both maxima within the first three days after infection. Beginning on the fifth day, a steady decrease in fluorescence intensity was observed, with a greater effect at 740 than at 690 nm. After 30 days there was no fluorescence. The infected plants that had been treated with the bactericide showed no significant change in fluorescence compared with that of the uninfected plants. The ratio of fluorescence intensities was determined to be F 690 nm/F 740 nm for all treatments. These studies indicate that LIF measurements of agave plants may be used for the early detection of certain types of disease and for determining the effect of a bactericide on bacteria. The results also showed that fluorescence intensity ratios can be used as a reliable indicator of the progress of disease.

Detection of bacterial infection of agave plants by laser-induced fluorescence.

PubMed

Cervantes-Martínez, Jesús; Flores-Hernández, Ricardo; Rodríguez-Garay, Benjamin; Santacruz-Ruvalcaba, Fernando

2002-05-01

Greenhouse-grown plants of Agave tequilana Weber var. azul were inoculated with Erwinia carotovora, the causal agent of stem soft rot. We investigated the laser-induced fluorescence (LIF) of agave plants to determine whether LIF can be used as a noninvasive sensing tool for pathological studies. The LIF technique was also investigated as a means of detecting the effect of the polyamine biosynthesis inhibitor beta-hydroxyethylhydrazine as a bactericide against the pathogenic bacterium Erwinia carotovora. A He-Ne laser at 632.8 nm was used as the excitation source, and in vivo fluorescence emission spectra were recorded in the 660-790-range. Fluorescence maxima were at 690 and 740 nm. The infected plants that were untreated with the bactericide showed a definite increase in fluorescence intensity at both maxima within the first three days after infection. Beginning on the fifth day, a steady decrease in fluorescence intensity was observed, with a greater effect at 740 than at 690 nm. After 30 days there was no fluorescence. The infected plants that had been treated with the bactericide showed no significant change in fluorescence compared with that of the uninfected plants. The ratio of fluorescence intensities was determined to be F 690 nm/F 740 nm for all treatments. These studies indicate that LIF measurements of agave plants may be used for the early detection of certain types of disease and for determining the effect of a bactericide on bacteria. The results also showed that fluorescence intensity ratios can be used as a reliable indicator of the progress of disease.

Fluorescence kinetics of emission from a small finite volume of a biological system

NASA Astrophysics Data System (ADS)

Dagen, A. J.; Alfano, R. R.; Zilinskas, B. A.; Swenberg, C. E.

1985-07-01

The fluorescence decay, apparent quantum yield and transmission from chromophores constrained to a microscopic volume using a single picosecond laser excitation were measured as a function of incident intensity. The β subunit of phycoeryhthrin aggregate isolated from the photosynthetic antenna system of Nostoc sp. was selected since it contains only four chromophores in a volume of less than 5.6×10 4 Å 3. The non-exponential fluorescence decay profiles were intensity independent for the intensity range studied (5 × 10 13 - 2 × 10 15 photon cm -2 per pulse). The apparent decrease in the relative fluorescence quantum yield and increase of the relative transmission with increasing excitation intensity is attributed to the combined effects of ground state depletion and upper excited state absorption. Evidence suggests that exciton annihilation is absent within isolated β subunits.

Intensity calibration of a laser scanning confocal microscope based on concentrated dyes.

PubMed

Model, Michael A; Blank, James L

2006-10-01

To find water-soluble fluorescent dyes with absorption in various regions of the spectrum and investigate their utility as standards for laser scanning confocal microscopy. Several dyes were found to have characteristics required for fluorescence microscopy standards. The intensity of biological fluorescent specimens was measured against the emission of concentrated dyes. Results using different optics and different microscopes were compared. Slides based on concentrated dyes can be prepared in a highly reproducible manner and are stable under laser scanning. Normalized fluorescence of biological specimens remains consistent with different objective lenses and is tolerant to some mismatch in optical filters or imperfect pinhole alignment. Careful choice of scanning parameters is necessary to ensure linearity of intensity measurements. Concentrated dyes provide a robust and inexpensive intensity standard that can be used in basic research or clinical studies.

[Determination of chromphoric dissolved organic matter in water from different sources].

PubMed

Liu, Xian-ping; Li, Lei; Dai, Jin-feng; Wang, Xiao-ru; Lee, Frank S C

2007-10-01

Chromophoric dissolved organic matter (CDOM) represents the fraction of the dissolved organic pool which absorbs light in the visible as well as UV ranges. It could affect the color of the waters. It is necessary to study it during in research on ecosystem, remote sensing of the water color and the cycle of carbon in waters. CDOM can fluoresce when excited, so fluorescence spectrum has been used to study its origin, distribution, and change. In the present article the fluorescence spectrophotometer was used to study the relation between the fluorescence intensity, spectrum area and the concentration of CDOM. When the concentration of CDOM is low (less than 75 mg x L(-1)), there is a better linear relationship (r2 > 0.98) between the fluorescence intensity, the spectrum area and the concentration of CDOM. Meanwhile good linear relations were found between the fluorescence intensity and spectrum area, which showed the same changeable trend of the fluorescence intensity and spectrum area with the concentration change of CDOM. A method was established to quantify the concentration of CDOM in water from different source using the linear relationship between the spectrum area and the concentration. It suits the complicated constituent analysis of CDOM and could really and accurately show the concentration of CDOM in natural water.

Near-infrared (NIR) fluorescence imaging of head and neck squamous cell carcinoma for fluorescence-guided surgery (Conference Presentation)

NASA Astrophysics Data System (ADS)

Moore, Lindsay; Warram, Jason M.; de Boer, Esther; Carroll, William R.; Morlandt, Anthony; Withrow, Kirk P.; Rosenthal, Eben L.

2016-03-01

During fluorescence-guided surgery, a cancer-specific optical probe is injected and visualized using a compatible device intraoperatively to provide visual contrast between diseased and normal tissues to maximize resection of cancer and minimize the resection of precious adjacent normal tissues. Six patients with squamous cell carcinomas of the head and neck region (oral cavity (n=4) or cutaneous (n=2)) were injected with an EGFR-targeting antibody (Cetuximab) conjugated to a near-infrared (NIR) fluorescent dye (IRDye800) 3, 4, or 7 days prior to surgical resection of the cancer. Each patient's tumor was then imaged using a commercially available, open-field NIR fluorescence imaging device each day prior to surgery, intraoperatively, and post-operatively. The mean fluorescence intensity (MFI) of the tumor was calculated for each specimen at each imaging time point. Adjacent normal tissue served as an internal anatomic control for each patient to establish a patient-matched "background" fluorescence. Resected tissues were also imaged using a closed-field NIR imaging device. Tumor to background ratios (TBRs) were calculated for each patient using both devices. Fluorescence histology was correlated with traditional pathology assessment to verify the specificity of antibody-dye conjugate binding. Peak TBRs using the open-field device ranged from 2.2 to 11.3, with an average TBR of 4.9. Peak TBRs were achieved between days 1 and 4. This study demonstrated that a commercially available NIR imaging device suited for intraoperative and clinical use can successfully be used with a fluorescently-labeled dye to delineate between diseased and normal tissue in this single cohort human study, illuminated the potential for its use in fluoresence-guided surgery.

Average M shell fluorescence yields for elements with 70≤Z≤92

NASA Astrophysics Data System (ADS)

Kahoul, A.; Deghfel, B.; Aylikci, V.; Aylikci, N. K.; Nekkab, M.

2015-03-01

The theoretical, experimental and analytical methods for the calculation of average M-shell fluorescence yield (ω¯M ) of different elements are very important because of the large number of their applications in various areas of physical chemistry and medical research. In this paper, the bulk of the average M-shell fluorescence yield measurements reported in the literature, covering the period 1955 to 2005 are interpolated by using an analytical function to deduce the empirical average M-shell fluorescence yield in the atomic range of 70≤Z≤92. The results were compared with the theoretical and fitted values reported by other authors. Reasonable agreement was typically obtained between our result and other works.

Three-dimensional fluorescence analysis of chernozem humic acids and their electrophoretic fractions

NASA Astrophysics Data System (ADS)

Trubetskoi, O. A.; Trubetskaya, O. E.

2017-09-01

Polyacrylamide gel electrophoresis in combination with size-exclusion chromatography (SEC-PAGE) has been used to obtain stable electrophoretic fractions of different molecular size (MS) from chernozem humic acids (HAs). Three-dimensional fluorescence charts of chernozem HAs and their fractions have been obtained for the first time, and all fluorescence excitation-emission maxima have been identified in the excitation wavelength range of 250-500 nm. It has been found that fractionation by the SEC-PAGE method results in a nonuniform distribution of protein- and humin-like fluorescence of the original HA preparation among the electrophoretic fractions. The electrophoretic fractions of the highest and medium MSs have only the main protein-like fluorescence maximum and traces of humin-like fluorescence. In the electrophoretic fraction of the lowest MS, the intensity of protein-like fluorescence is low, but the major part of humin-like fluorescence is localized there. Relationships between the intensity of protein-like fluorescence and the weight distribution of amino acids have been revealed, as well as between the degree of aromaticity and the intensity of humin-like fluorescence in electrophoretic fractions of different MSs. The obtained relationships can be useful in the interpretation of the spatial structural organization and ecological functions of soil HAs.

Optical spectroscopy of the bladder washout fluid to optimize fluorescence cystoscopy with Hexvix®.

PubMed

Martoccia, Carla; Zellweger, Matthieu; Lovisa, Blaise; Jichlinski, Patrice; van den Bergh, Hubert; Wagnières, Georges

2014-09-01

Fluorescence cystoscopy enhances detection of early bladder cancer. Water used to inflate thebladder during the procedure rapidly contains urine, which may contain fluorochromes. This frequently degradesfluorescence images. Samples of bladder washout fluid (BWF) or urine were collected (15 subjects). We studiedtheir fluorescence properties and assessed changes induced by pH (4 to 9) and temperature (15°C to 41°C).A typical fluorescence spectrum of BWF features a main peak (excitation/emission: 320∕420 nm, FWHM =50∕100 nm) and a weaker (5% to 20% of main peak intensity), secondary peak (excitation/emission: 455∕525 nm, FWHM = 80∕50 nm). Interpatient fluctuations of fluorescence intensity are observed. Fluorescence intensity decreases when temperature increases (max 30%) or pH values vary (max 25%). Neither approach is compatible with clinical settings. Fluorescence lifetime measurements suggest that 4-pyridoxic acid/riboflavin is the most likely molecule responsible for urine’s main/secondary fluorescence peak. Our measurements give an insight into the spectroscopy of the detrimental background fluorescence. This should be included in the optical design of fluorescence cystoscopes. We estimate that restricting the excitation range from 370–430 nm to 395–415 nm would reduce the BWF background by a factor 2.

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

NASA Astrophysics Data System (ADS)

Zhong, Xin; Wang, Xinwei; Zhou, Yan

2018-01-01

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

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

PubMed

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

2009-01-01

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

The effects of increasing PGE2 on translocation of labeled albumin into rat brain.

PubMed

Messripour, M; Mesripour, A; Mashayekhie, F J

2015-01-01

Under pathophysiological conditions, infiltration of leukocyte plays a key role in the progression of the neuroinflammatory reaction in the CNS. Prostaglandin E2 (PGE2) is known to accumulate at lesion sites of the post-ischemic brain. Although post-ischemic treatments with cyclooxygenase-2 inhibitors reduce blood-brain barrier (BBB) leukocyte infiltration, the direct effect of PGE2 on BBB has not been fully implemented. Therefore, the direct effect of increasing PGE2 infusion on translocation of labeled albumin into the brain was assessed. Under anesthesia rats were drilled stereo-taxicaly a burr hole in the right forebrain and PGE2 was infused into the forebrain and the hole was occluded. The animals were then injected with fluorescent labeled albumin (FA), via internal right jugular vein and decapitated at different infusion time points. The forebrain was removed and each forebrain hemisphere was homogenized and fluorescence intensities were measured in the supernatant. The fluorescence intensities measured in the right and left forebrain hemispheres of the control group (0.0 μg PGE2) were almost identical. Four hours after infusion of PGE2 at doses higher than 250 μg, fluorescence intensity increased in the right forebrain supernatant, even if it was not statistically significant. The fluorescence intensity was detectable in the brain supernatant 4 h after infusion of PGE2 in doses higher than 250 μg PGE2. The highest fluorescence intensity was 16 h after infusion of 500 μg PGE2, which returned to near control values after 48 h. Increased fluorescence intensity in the brain following PGE2 infusion is concluded to be associated with disruption of the BBB.

Time-resolved and temperature tuneable measurements of fluorescent intensity using a smartphone fluorimeter.

PubMed

Hossain, Md Arafat; Canning, John; Yu, Zhikang; Ast, Sandra; Rutledge, Peter J; Wong, Joseph K-H; Jamalipour, Abbas; Crossley, Maxwell J

2017-05-30

A smartphone fluorimeter capable of time-based fluorescence intensity measurements at various temperatures is reported. Excitation is provided by an integrated UV LED (λ ex = 370 nm) and detection obtained using the in-built CMOS camera. A Peltier is integrated to allow measurements of the intensity over T = 10 to 40 °C. All components are controlled using a smartphone battery powered Arduino microcontroller and a customised Android application that allows sequential fluorescence imaging and quantification every δt = 4 seconds. The temperature dependence of fluorescence intensity for four emitters (rhodamine B, rhodamine 6G, 5,10,15,20-tetraphenylporphyrin and 6-(1,4,8,11-tetraazacyclotetradecane)2-ethyl-naphthalimide) are characterised. The normalised fluorescence intensity over time of the latter chemosensor dye complex in the presence of Zn 2+ is observed to accelerate with an increasing rate constant, k = 1.94 min -1 at T = 15 °C and k = 3.64 min -1 at T = 30 °C, approaching a factor of ∼2 with only a change in temperature of ΔT = 15 °C. Thermally tuning these twist and bend associated rates to optimise sensor approaches and device applications is proposed.

Demonstration of FRET in solutions

NASA Astrophysics Data System (ADS)

Shah, Sunil; Gryczynski, Zygmunt; Chib, Rahul; Fudala, Rafal; Baxi, Aatmun; Borejdo, Julian; Synak, Anna; Gryczynski, Ignacy

2016-03-01

We measured the Förster resonance energy transfer (FRET) from Uranin (U) donor to Rhodamine 101 (R101) acceptor in propylene glycol. Steady-state fluorescence measurements show a significant difference between mixed and unmixed fluorophore solutions. In the solution with mixed fluorophores, fluorescence intensity of the U donor decreases and intensity of R101 fluorescence increases. This is visualized as a color change from green to orange. Fluorescence anisotropy of the mixture solution increases in the donor emission wavelength region and decreases in the acceptor emission wavelengths; which is consistent with FRET occurrence. Time-resolved (lifetime) measurements show a decrease of the U lifetime in the presence of R101 acceptor. In the intensity decay of R101 acceptor appears a negative component indicating excited state process. All these measurements prove the presence of FRET in U/R101 mixture fluorescence.

Listening to membrane potential: photoacoustic voltage-sensitive dye recording.

PubMed

Zhang, Haichong K; Yan, Ping; Kang, Jeeun; Abou, Diane S; Le, Hanh N D; Jha, Abhinav K; Thorek, Daniel L J; Kang, Jin U; Rahmim, Arman; Wong, Dean F; Boctor, Emad M; Loew, Leslie M

2017-04-01

Voltage-sensitive dyes (VSDs) are designed to monitor membrane potential by detecting fluorescence changes in response to neuronal or muscle electrical activity. However, fluorescence imaging is limited by depth of penetration and high scattering losses, which leads to low sensitivity in vivo systems for external detection. By contrast, photoacoustic (PA) imaging, an emerging modality, is capable of deep tissue, noninvasive imaging by combining near-infrared light excitation and ultrasound detection. Here, we show that voltage-dependent quenching of dye fluorescence leads to a reciprocal enhancement of PA intensity. We synthesized a near-infrared photoacoustic VSD (PA-VSD), whose PA intensity change is sensitive to membrane potential. In the polarized state, this cyanine-based probe enhances PA intensity while decreasing fluorescence output in a lipid vesicle membrane model. A theoretical model accounts for how the experimental PA intensity change depends on fluorescence and absorbance properties of the dye. These results not only demonstrate PA voltage sensing but also emphasize the interplay of both fluorescence and absorbance properties in the design of optimized PA probes. Together, our results demonstrate PA sensing as a potential new modality for recording and external imaging of electrophysiological and neurochemical events in the brain.

Listening to membrane potential: photoacoustic voltage-sensitive dye recording

NASA Astrophysics Data System (ADS)

Zhang, Haichong K.; Yan, Ping; Kang, Jeeun; Abou, Diane S.; Le, Hanh N. D.; Jha, Abhinav K.; Thorek, Daniel L. J.; Kang, Jin U.; Rahmim, Arman; Wong, Dean F.; Boctor, Emad M.; Loew, Leslie M.

2017-04-01

Voltage-sensitive dyes (VSDs) are designed to monitor membrane potential by detecting fluorescence changes in response to neuronal or muscle electrical activity. However, fluorescence imaging is limited by depth of penetration and high scattering losses, which leads to low sensitivity in vivo systems for external detection. By contrast, photoacoustic (PA) imaging, an emerging modality, is capable of deep tissue, noninvasive imaging by combining near-infrared light excitation and ultrasound detection. Here, we show that voltage-dependent quenching of dye fluorescence leads to a reciprocal enhancement of PA intensity. We synthesized a near-infrared photoacoustic VSD (PA-VSD), whose PA intensity change is sensitive to membrane potential. In the polarized state, this cyanine-based probe enhances PA intensity while decreasing fluorescence output in a lipid vesicle membrane model. A theoretical model accounts for how the experimental PA intensity change depends on fluorescence and absorbance properties of the dye. These results not only demonstrate PA voltage sensing but also emphasize the interplay of both fluorescence and absorbance properties in the design of optimized PA probes. Together, our results demonstrate PA sensing as a potential new modality for recording and external imaging of electrophysiological and neurochemical events in the brain.

Fluorescence intensity- and lifetime-based glucose sensing using glucose/galactose-binding protein.

PubMed

Pickup, John C; Khan, Faaizah; Zhi, Zheng-Liang; Coulter, Jonathan; Birch, David J S

2013-01-01

We review progress in our laboratories toward developing in vivo glucose sensors for diabetes that are based on fluorescence labeling of glucose/galactose-binding protein. Measurement strategies have included both monitoring glucose-induced changes in fluorescence resonance energy transfer and labeling with the environmentally sensitive fluorophore, badan. Measuring fluorescence lifetime rather than intensity has particular potential advantages for in vivo sensing. A prototype fiber-optic-based glucose sensor using this technology is being tested. © 2013 Diabetes Technology Society.

A fast- and positively photoswitchable fluorescent protein for ultralow-laser-power RESOLFT nanoscopy.

PubMed

Tiwari, Dhermendra K; Arai, Yoshiyuki; Yamanaka, Masahito; Matsuda, Tomoki; Agetsuma, Masakazu; Nakano, Masahiro; Fujita, Katsumasa; Nagai, Takeharu

2015-06-01

Fluorescence nanoscopy has revolutionized our ability to visualize biological structures not resolvable by conventional microscopy. However, photodamage induced by intense light exposure has limited its use in live specimens. Here we describe Kohinoor, a fast-switching, positively photoswitchable fluorescent protein, and show that it has high photostability over many switching repeats. With Kohinoor, we achieved super-resolution imaging of live HeLa cells using biocompatible, ultralow laser intensity (0.004 J/cm(2)) in reversible saturable optical fluorescence transition (RESOLFT) nanoscopy.

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

NASA Astrophysics Data System (ADS)

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 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. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr30578j

Effective transvascular drug delivery to glioma in rats by using a pulsed laser-induced photomechanical wave (Conference Presentation)

NASA Astrophysics Data System (ADS)

Akutsu, Yusuke; Sato, Shunichi; Tomiyama, Arata; Tsunoi, Yasuyuki; Kawauchi, Satoko; Mori, Kentaro; Terakawa, Mitsuhiro

2016-03-01

Glioma is one of the most aggressive cancers, for which efficacy of conventional chemotherapy is often limited due to the blood-tumor barrier (BTB). Thus, the development of a method for enhancing the BTB permeability is strongly desired. In this study, we applied a photomechanical wave (PMW), which was generated by the irradiation of a light-absorbing material with a nanosecond laser pulse, to transiently open the BTB in a rat intracranial glioma model using C6 cells. A tumor was grown in the both hemispheres, and a solution of Evans blue (EB), as a test drug, was injected into the tail vein. Thereafter, we applied a PMW generated at a laser fluence of 0.2 J/cm2 (averaged peak pressure, ~27 MPa), 0.4 J/cm2 (~54 MPa) or 0.6 J/cm2 (~78MPa), to one hemisphere through the cranial window, while the other hemisphere served as a control. Four hours later, the rat was perfused, and we compared intensity distributions of EB fluorescence between the both hemispheres. Intensities of EB fluorescence both in the peritumoral and tumor core regions were increased with increasing the laser fluence, but hemorrhage was observed at the highest fluence. Thus, 0.4 J/cm2 would be optimum for efficient and safe BTB opening. On the basis of fluorescence microscopy with the use of enhanced green fluorescent protein-expressing C6 cells, we confirmed that a drug was delivered into targeted glioma cells in the peritumoral region. These results show the validity of the present transvascular drug delivery method to glioma.

Spatial distribution of fluorescent light emitted from neon and nitrogen excited by low energy electron beams

NASA Astrophysics Data System (ADS)

Morozov, A.; Krücken, R.; Ulrich, A.; Wieser, J.

2006-11-01

Side-view intensity profiles of fluorescent light were measured for neon and nitrogen excited with 12keV electron beams at gas pressures from 250to1400hPa. The intensity profiles were compared with theoretical profiles calculated using the CASINO program which performs Monte Carlo simulations of electron scattering. It was assumed that the spatial distribution of fluorescent intensity is directly proportional to the spatial distribution of energy loss by primary electrons. The comparison shows good correlation of experimental data and the results of numeric simulations.

Effects of hexamethonium, phenothiazines, propranolol and ephedrine on acetylcholinesterase carbamylation by physostigmine, aldicarb and carbaryl: interaction between the active site and the functionally distinct peripheral sites in acetylcholinesterase.

PubMed

Singh, A K; Spassova, D

1998-01-01

Physostigmine, aldicarb and carbaryl were potent inhibitors of acetylcholinesterase (AChE). The physostigmine-inhibited AChE fluoresced at 300 nm excitation and 500 nm emission wavelengths, but the aldicarb and carbaryl inhibited enzyme did not. This suggests that the carbamylated active center is not the fluorescing site in AChE. The fluorescence intensity of physostigmine-inhibited AChE decreased with increasing the substrate (acetylthiocholine) concentration, thus indicating that physostigmine binding to the active site is essential for the development of fluorescence. Thus, the physostigmine-inhibited AChE fluoresces due to the binding of trimethylpyrrolo[2,3-b]indol (TMPI) moiety, formed by the hydrolysis of physostigmine, to a peripheral site in AChE. The fluorescence intensity of the physostigmine-inhibited enzyme decreased when the inhibited-enzyme was dialyzed for either 30 min that poorly reactivated the enzyme or 180 min that fully reactivated the enzyme. This suggests that dialysis dissociates the AChE-TMPI complex much faster than it reactivates the carbamylated AChE. Ephedrine, propranolol and phenothiazines including trifluoparazine (TPZ) caused non-competitive inhibition, while hexamethonium caused an uncompetitive inhibition of AChE activity. TPZ, upon binding with AChE, formed a fluorescent TPZ-enzyme complex. The fluorescence intensity of TPZ-AChE complex was effectively decreased by ephedrine, but not by propranolol or hexamethonium. This indicates that TPZ and ephedrine bind to the same site in AChE which is different from the site/or sites to which propranolol or hexamethonium bind. Hexamethonium protected AChE from inhibition by carbamates and decreased the fluorescence intensity of the physostigmine-inhibited AChE. Phenothiazines and ephedrine did not modulate the enzyme inhibition or the fluorescence intensity of the physostigmine-inhibited AChE. Propranolol and TPZ potentiated the enzyme inhibition and increased the fluorescence intensity in the presence of physostigmine. These compounds, however, did not affect the inhibition of AChE by carbaryl or aldicarb. Ephedrine blocked the effects of TPZ, but did not alter the effects of propranolol on physostigmine-inhibited AChE. AChE, therefore, contains multiple peripheral binding sites which, upon binding to specific ligands, transduce differential signals to the active center.

Fluorescence-based enhanced reality (FLER) for real-time estimation of bowel perfusion in minimally invasive surgery

NASA Astrophysics Data System (ADS)

Diana, Michele

2016-03-01

Pre-anastomotic bowel perfusion is a key factor for a successful healing process. Clinical judgment has limited accuracy to evaluate intestinal microperfusion. Fluorescence videography is a promising tool for image-guided intraoperative assessment of the bowel perfusion at the future anastomotic site in the setting of minimally invasive procedures. The standard configuration for fluorescence videography includes a Near-Infrared endoscope able to detect the signal emitted by a fluorescent dye, more frequently Indocyanine Green (ICG), which is administered by intravenous injection. Fluorescence intensity is proportional to the amount of fluorescent dye diffusing in the tissue and consequently is a surrogate marker of tissue perfusion. However, fluorescence intensity alone remains a subjective approach and an integrated computer-based analysis of the over-time evolution of the fluorescence signal is required to obtain quantitative data. We have developed a solution integrating computer-based analysis for intra-operative evaluation of the optimal resection site, based on the bowel perfusion as determined by the dynamic fluorescence intensity. The software can generate a "virtual perfusion cartography", based on the "fluorescence time-to-peak". The virtual perfusion cartography can be overlapped onto real-time laparoscopic images to obtain the Enhanced Reality effect. We have defined this approach FLuorescence-based Enhanced Reality (FLER). This manuscript describes the stepwise development of the FLER concept.

Temporal Characteristics of Sodium Fluorescein in the Tear Meniscus.

PubMed

Markoulli, Maria; Isa, Nur Amalina M D; Papas, Eric B

2017-02-01

To observe the emission intensity profile of sodium fluorescein in the human tear film as a function of time and concentration. Twenty-two participants with no dry eye signs or symptoms were randomly allocated to receive 1 μL of either a 2 or 10% concentration of fluorescein to one eye. Images of the inferior tear meniscus were captured at regular intervals over 30 minutes and the process repeated for the other eye with the alternate concentration. Fluorescence intensity was quantified on the basis of the grayscale pixel values in the tear meniscus images. The fluorescein-decay profile over time and between concentrations was determined. Peak fluorescence intensity was reached in 3.9 ± 3.0 and 8.7 ± 4.4 minutes after instillation for the 2 and 10% concentrations, respectively. The 10% concentration of fluorescein maintained its peak fluorescence intensity longer than the 2% concentration (about 9 and 2 minutes, respectively). The peak fluorescence intensity was not significantly different between the higher and lower concentrations (44 ± 37 vs. 38 ± 32 units, P = .22). For both concentrations, the observed intensity did not return to baseline levels by the end of the 30-minute observation time. The fluorescence intensity of fluorescein in a clinical setting varies with time such that both the onset and duration of maximum brightness are concentration dependent. At low concentration (2%), maximum brightness occurs almost immediately after instillation and lasts about 2 minutes. With a higher concentration (10%), the effective working window is delayed for about 7 to 8 minutes. Irrespective of initial concentration, observable fluorescence remains in the tear film beyond 30 minutes post-instillation.

Angular shaping of fluorescence from synthetic opal-based photonic crystal.

PubMed

Boiko, Vitalii; Dovbeshko, Galyna; Dolgov, Leonid; Kiisk, Valter; Sildos, Ilmo; Loot, Ardi; Gorelik, Vladimir

2015-01-01

Spectral, angular, and temporal distributions of fluorescence as well as specular reflection were investigated for silica-based artificial opals. Periodic arrangement of nanosized silica globules in the opal causes a specific dip in the defect-related fluorescence spectra and a peak in the reflectance spectrum. The spectral position of the dip coincides with the photonic stop band. The latter is dependent on the size of silica globules and the angle of observation. The spectral shape and intensity of defect-related fluorescence can be controlled by variation of detection angle. Fluorescence intensity increases up to two times at the edges of the spectral dip. Partial photobleaching of fluorescence was observed. Photonic origin of the observed effects is discussed.

Visualization of Porphyrin-Based Photosensitizer Distribution from Fluorescence Images In Vivo Using an Optimized RGB Camera

NASA Astrophysics Data System (ADS)

Liu, L.; Huang, Zh.; Qiu, Zh.; Li, B.

2018-01-01

A handheld RGB camera was developed to monitor the in vivo distribution of porphyrin-based photosensitizer (PS) hematoporphyrin monomethyl ether (HMME) in blood vessels during photodynamic therapy (PDT). The focal length, f-number, International Standardization Organization (ISO) sensitivity, and shutter speed of the camera were optimized for the solution sample with various HMME concentrations. After the parameter optimization, it was found that the red intensity value of the fluorescence image was linearly related to the fluorescence intensity under investigated conditions. The RGB camera was then used to monitor the in vivo distribution of HMME in blood vessels in a skin-fold window chamber model. The red intensity value of the recorded RGB fluorescence image was found to be linearly correlated to HMME concentrations in the range 0-24 μM. Significant differences in the red to green intensity ratios were observed between the blood vessels and the surrounding tissue.

Molecular insight into the inclusion of the dietary plant flavonol fisetin and its chromophore within a chemically modified γ-cyclodextrin: Multi-spectroscopic, molecular docking and solubility studies.

PubMed

Pahari, Biswapathik; Chakraborty, Sandipan; Sengupta, Pradeep K

2018-09-15

We explored the encapsulation of dietary plant flavonols fisetin and its chromophore 3-hydroxyflavone, within 2-hydroxypropyl-γ-cyclodextrin (HPγ-CDx) nano-cavity in aqueous solution using multi-spectroscopic approaches and molecular docking. Upon addition of HPγ-CDx, dramatic changes occur in the intrinsic 'two color' fluorescence behavior of the fluorophores. This is manifested by significant increase in the steady state fluorescence intensities, anisotropies, average fluorescence lifetimes and rotational correlation times. Furthermore, in the CDx environment, intrinsically achiral flavonols exhibit prominent induced circular dichroism bands. These findings indicate that the flavonol molecules spontaneously enter the relatively hydrophobic, chiral environment of the HPγ-CDx nano-cavities. Molecular docking computations corroborate the spectroscopic findings, and predict selectivity in orientation of the encapsulated flavonols. HPγ-CDx inclusion increases the aqueous solubility of individual flavonols ∼100-1000 times. The present study demonstrates that the hydroxypropyl substituent in γ-CDx controls the inclusion mode of the flavonols, leading to their enhanced solubilization and altered spectral signatures. Copyright © 2018 Elsevier Ltd. All rights reserved.

Synthesis and spectroscopic properties of some new difluoroboron bis-β-diketonate derivatives.

PubMed

Pi, Yan; Wang, Dun-Jia; Liu, Hua; Hu, Yan-Jun; Wei, Xian-Hong; Zheng, Jing

2014-10-15

Six new bis-β-diketones (RCOCH2CO-C7H7N-COCH2COR) were synthesized from 3,5-diacetyl-2,6-dimethylpyridine via Claisen condensation with the corresponding esters, and then reacted with boron trifluoride etherate to afford difluoroboron bis-β-diketonate derivatives. Their spectroscopic properties were investigated by UV-vis, FTIR, (1)H NMR and fluorescence spectroscopic techniques. It was found that these boron complexes exhibited violet or blue fluorescence emission at 422-445nm and possessed high extinction coefficients. The results indicate that the extending π-conjugation can increase the fluorescence intensity and quantum yield for these boron complexes. Especially, the compound 2b displayed the stronger fluorescence intensity and the highest fluorescence quantum yield (Φu=0.94) in these boron compounds. However, compounds 2c and 2d had the lower fluorescence intensity and quantum yield as a result of the heavy atom effect of the chlorine atom in the molecules. Copyright © 2014 Elsevier B.V. All rights reserved.

Synthesis and spectroscopic properties of some new difluoroboron bis-β-diketonate derivatives

NASA Astrophysics Data System (ADS)

Pi, Yan; Wang, Dun-Jia; Liu, Hua; Hu, Yan-Jun; Wei, Xian-Hong; Zheng, Jing

2014-10-01

Six new bis-β-diketones (RCOCH2CO-C7H7N-COCH2COR) were synthesized from 3,5-diacetyl-2,6-dimethylpyridine via Claisen condensation with the corresponding esters, and then reacted with boron trifluoride etherate to afford difluoroboron bis-β-diketonate derivatives. Their spectroscopic properties were investigated by UV-vis, FTIR, 1H NMR and fluorescence spectroscopic techniques. It was found that these boron complexes exhibited violet or blue fluorescence emission at 422-445 nm and possessed high extinction coefficients. The results indicate that the extending π-conjugation can increase the fluorescence intensity and quantum yield for these boron complexes. Especially, the compound 2b displayed the stronger fluorescence intensity and the highest fluorescence quantum yield (Φu = 0.94) in these boron compounds. However, compounds 2c and 2d had the lower fluorescence intensity and quantum yield as a result of the heavy atom effect of the chlorine atom in the molecules.

Classification of corn kernels contaminated with aflatoxins using fluorescence and reflectance hyperspectral images analysis

NASA Astrophysics Data System (ADS)

Zhu, Fengle; Yao, Haibo; Hruska, Zuzana; Kincaid, Russell; Brown, Robert; Bhatnagar, Deepak; Cleveland, Thomas

2015-05-01

Aflatoxins are secondary metabolites produced by certain fungal species of the Aspergillus genus. Aflatoxin contamination remains a problem in agricultural products due to its toxic and carcinogenic properties. Conventional chemical methods for aflatoxin detection are time-consuming and destructive. This study employed fluorescence and reflectance visible near-infrared (VNIR) hyperspectral images to classify aflatoxin contaminated corn kernels rapidly and non-destructively. Corn ears were artificially inoculated in the field with toxigenic A. flavus spores at the early dough stage of kernel development. After harvest, a total of 300 kernels were collected from the inoculated ears. Fluorescence hyperspectral imagery with UV excitation and reflectance hyperspectral imagery with halogen illumination were acquired on both endosperm and germ sides of kernels. All kernels were then subjected to chemical analysis individually to determine aflatoxin concentrations. A region of interest (ROI) was created for each kernel to extract averaged spectra. Compared with healthy kernels, fluorescence spectral peaks for contaminated kernels shifted to longer wavelengths with lower intensity, and reflectance values for contaminated kernels were lower with a different spectral shape in 700-800 nm region. Principal component analysis was applied for data compression before classifying kernels into contaminated and healthy based on a 20 ppb threshold utilizing the K-nearest neighbors algorithm. The best overall accuracy achieved was 92.67% for germ side in the fluorescence data analysis. The germ side generally performed better than endosperm side. Fluorescence and reflectance image data achieved similar accuracy.

The exploration of the characteristics of the hyperglycemia serum fluorescence spectrum

NASA Astrophysics Data System (ADS)

Wang, Lexin; Zhao, Zhimin; Chen, Hui; Li, Peng; Xin, Yujun

2008-12-01

Now, spectra technology is widely used in the biomedicine research,so this study investigates variation of the fluorescence spectra in different excitation wavelength, and the spectra of serum with different glucose concentration is tested in the excitation wavelength of 240nm to 280nm. The experimental result shows that the correlation between the serum fluorescence intensity and the excitation light is very close, when the excitation light is in the ultraviolet wave band, the fluorescence of serum is intensive. There is a violent fluorescence emission wavelength, which is 300nm to 410nm, while the excitation wavelength ranges from 220nm to 290nm, and the peaks wavelength are 330nm and 370nm. From 240nm to 280nm, the serum fluorescence intensity increases synchronously with the glucose concentration, and the relationship between the fluorescence peak wavelength and the glucose concentration is almost in line. In this way the blood sugar concentration can be estimated by the fluorescence spectra peak wavelength when the excitation wavelength is from 240nm to 280nm, which is effective. It provides experimental foundation for the wide use of spectra technology in medical diagnose, and the effectiv method to test the blood sugar concentration.

Enhanced fluorescence microscope and its application

NASA Astrophysics Data System (ADS)

Wang, Susheng; Li, Qin; Yu, Xin

1997-12-01

A high gain fluorescence microscope is developed to meet the needs in medical and biological research. By the help of an image intensifier with luminance gain of 4 by 104 the sensitivity of the system can achieve 10-6 1x level and be 104 times higher than ordinary fluorescence microscope. Ultra-weak fluorescence image can be detected by it. The concentration of fluorescent label and emitting light intensity of the system are decreased as much as possible, therefore, the natural environment of the detected call can be kept. The CCD image acquisition set-up controlled by computer obtains the quantitative data of each point according to the gray scale. The relation between luminous intensity and output of CCD is obtained by using a wide range weak photometry. So the system not only shows the image of ultra-weak fluorescence distribution but also gives the intensity of fluorescence of each point. Using this system, we obtained the images of distribution of hypocrellin A (HA) in Hela cell, the images of Hela cell being protected by antioxidant reagent Vit. E, SF and BHT. The images show that the digitized ultra-sensitive fluorescence microscope is a useful tool for medical and biological research.

Dummy molecularly imprinted polymers-capped CdTe quantum dots for the fluorescent sensing of 2,4,6-trinitrotoluene.

PubMed

Xu, Shoufang; Lu, Hongzhi; Li, Jinhua; Song, Xingliang; Wang, Aixiang; Chen, Lingxin; Han, Shaobo

2013-08-28

Molecularly imprinted polymers (MIPs) with trinitrophenol (TNP) as a dummy template molecule capped with CdTe quantum dots (QDs) were prepared using 3-aminopropyltriethoxy silane (APTES) as the functional monomer and tetraethoxysilane (TEOS) as the cross linker through a seed-growth method via a sol-gel process (i.e., DMIP@QDs) for the sensing of 2,4,6-trinitrotoluene (TNT) on the basis of electron-transfer-induced fluorescence quenching. With the presence and increase of TNT in sample solutions, a Meisenheimer complex was formed between TNT and the primary amino groups on the surface of the QDs. The energy of the QDs was transferred to the complex, resulting in the quenching of the QDs and thus decreasing the fluorescence intensity, which allowed the TNT to be sensed optically. DMIP@QDs generated a significantly reduced fluorescent intensity within less than 10 min upon binding TNT. The fluorescence-quenching fractions of the sensor presented a satisfactory linearity with TNT concentrations in the range of 0.8-30 μM, and its limit of detection could reach 0.28 μM. The sensor exhibited distinguished selectivity and a high binding affinity to TNT over its possibly competing molecules of 2,4-dinitrophenol (DNP), 4-nitrophenol (4-NP), phenol, and dinitrotoluene (DNT) because there are more nitro groups in TNT and therefore a stronger electron-withdrawing ability and because it has a high similarity in shape and volume to TNP. The sensor was successfully applied to determine the amount of TNT in soil samples, and the average recoveries of TNT at three spiking levels ranged from 90.3 to 97.8% with relative standard deviations below 5.12%. The results provided an effective way to develop sensors for the rapid recognition and determination of hazardous materials from complex matrices.

Red fluorescent biofilm: the thick, the old, and the cariogenic

PubMed Central

Volgenant, Catherine M.C.; Hoogenkamp, Michel A.; Buijs, Mark J.; Zaura, Egija; ten Cate, Jacob (Bob) M.; van der Veen, Monique H.

2016-01-01

Background Some dental plaque fluoresces red. The factors involved in this fluorescence are yet unknown. Objective The aim of this study was to assess systematically the effect of age, thickness, and cariogenicity on the extent of red fluorescence produced by in vitro microcosm biofilms. Design The effects of biofilm age and thickness on red fluorescence were tested in a constant depth film fermentor (CDFF) by growing biofilms of variable thicknesses that received a constant supply of defined mucin medium (DMM) and eight pulses of sucrose/day. The influence of cariogenicity on red fluorescence was tested by growing biofilm on dentin disks receiving DMM, supplemented with three or eight pulses of sucrose/day. The biofilms were analyzed at different time points after inoculation, up to 24 days. Emission spectra were measured using a fluorescence spectrophotometer (λexc405 nm) and the biofilms were photographed with a fluorescence camera. The composition of the biofilms was assessed using 454-pyrosequecing of the 16S rDNA gene. Results From day 7 onward, the biofilms emitted increasing intensities of red fluorescence as evidenced by the combined red fluorescence peaks. The red fluorescence intensity correlated with biofilm thickness but not in a linear way. Biofilm fluorescence also correlated with the imposed cariogenicity, evidenced by the induced dentin mineral loss. Increasing the biofilm age or increasing the sucrose pulsing frequency led to a shift in the microbial composition. These shifts in composition were accompanied by an increase in red fluorescence. Conclusions The current study shows that a thicker, older, or more cariogenic biofilm results in a higher intensity of red fluorescence. PMID:27060056

A fluorescent molecular rotor probes the kinetic process of degranulation of mast cells.

PubMed

Furuno, T; Isoda, R; Inagaki, K; Iwaki, T; Noji, M; Nakanishi, M

1992-08-01

A confocal fluorescence microscope was used to study the exocytotic secretory processes of mast cells in combination with an fluorescent molecular rotor, 9-(dicyanovinyl)julolidine (DCVJ). DCVJ is known to be an unique fluorescent dye which increases its quantum yield with decreasing intramolecular rotation. Here, DCVJ-loaded peritoneal rat mast cells were stimulated with compound 48/80 and their fluorescence images were compared with fluorescence calcium images of fluo-3-loaded mast cells. Subsequent to transient increases in intracellular free calcium ion concentration, DCVJ fluorescence increased dramatically in the cytoplasm and formed a ring-like structure around the nucleus, suggesting the possibility that the dye bound to the proteins composing the cytoskeletal architecture. Furthermore, the increases of DCVJ fluorescence intensities were mostly blocked in the presence of cytochalasin D (10 microM). However, fluo-3 fluorescence intensities still increased after addition of compound 48/80.

Dextran sulfate sodium-induced acute colitis impairs dermal lymphatic function in mice.

PubMed

Agollah, Germaine D; Wu, Grace; Peng, Ho-Lan; Kwon, Sunkuk

2015-12-07

To investigate whether dermal lymphatic function and architecture are systemically altered in dextran sulfate sodium (DSS)-induced acute colitis. Balb/c mice were administered 4% DSS in lieu of drinking water ad libitum for 7 d and monitored to assess disease activity including body weight, diarrhea severity, and fecal bleeding. Control mice received standard drinking water with no DSS. Changes in mesenteric lymphatics were assessed following oral administration of a fluorescently-labelled fatty acid analogue, while dermal lymphatic function and architecture was longitudinally characterized using dynamic near-infrared fluorescence (NIRF) imaging following intradermal injection of indocyanine green (ICG) at the base of the tail or to the dorsal aspect of the left paw prior to, 4, and 7 d after DSS administration. We also measured dye clearance rate after injection of Alexa680-bovine serum albumin (BSA). NIRF imaging data was analyzed to reveal lymphatic contractile activity after selecting fixed regions of interest (ROIs) of the same size in fluorescent lymphatic vessels on fluorescence images. The averaged fluorescence intensity within the ROI of each fluorescence image was plotted as a function of imaging time and the lymphatic contraction frequency was computed by assessing the number of fluorescent pulses arriving at a ROI. Mice treated with DSS developed acute inflammation with clinical symptoms of loss of body weight, loose feces/watery diarrhea, and fecal blood, all of which were aggravated as disease progressed to 7 d. Histological examination of colons of DSS-treated mice confirmed acute inflammation, characterized by segmental to complete loss of colonic mucosa with an associated chronic inflammatory cell infiltrate that extended into the deeper layers of the wall of the colon, compared to control mice. In situ intravital imaging revealed that mice with acute colitis showed significantly fewer fluorescent mesenteric lymphatic vessels, indicating impaired uptake of a lipid tracer within mesenteric lymphatics. Our in vivo NIRF imaging data demonstrated dilated dermal lymphatic vessels, which were confirmed by immunohistochemical staining of lymphatic vessels, and significantly reduced lymphatic contractile function in the skin of mice with DSS-induced acute colitis. Quantification of the fluorescent intensity remaining in the depot as a function of time showed that there was significantly higher Alexa680-BSA fluorescence in mice with DSS-induced acute colitis compared to pre-treatment with DSS, indicative of impaired lymphatic drainage. The lymphatics are locally and systemically altered in acute colitis, and functional NIRF imaging is useful for noninvasively monitoring systemic lymphatic changes during inflammation.

Average M shell fluorescence yields for elements with 70≤Z≤92

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kahoul, A., E-mail: ka-abdelhalim@yahoo.fr; LPMRN laboratory, Department of Materials Science, Faculty of Sciences and Technology, Mohamed El Bachir El Ibrahimi University, Bordj-Bou-Arreridj 34030; Deghfel, B.

2015-03-30

The theoretical, experimental and analytical methods for the calculation of average M-shell fluorescence yield (ω{sup ¯}{sub M}) of different elements are very important because of the large number of their applications in various areas of physical chemistry and medical research. In this paper, the bulk of the average M-shell fluorescence yield measurements reported in the literature, covering the period 1955 to 2005 are interpolated by using an analytical function to deduce the empirical average M-shell fluorescence yield in the atomic range of 70≤Z≤92. The results were compared with the theoretical and fitted values reported by other authors. Reasonable agreement wasmore » typically obtained between our result and other works.« less

Noninvasive control of the transport function of fluorescent coloured liposomal nanoparticles

NASA Astrophysics Data System (ADS)

Stelmashchuk, O.; Zherebtsov, E.; Zherebtsova, A.; Kuznetsova, E.; Vinokurov, A.; Dunaev, A.; Mamoshin, A.; Snimshchikova, I.; Borsukov, A.; Bykov, A.; Meglinski, I.

2017-06-01

The use of liposomal nanoparticles with an incorporated active substance is an innovative and promising approach to diagnostics and therapy. The application of liposomal nanoparticle-based drugs allows for targeted localized delivery, overcomes the natural barriers within the body effectively, and minimizes possible side effects. Liposomes are able to contain a variety of ingredients with practically no limitations to their chemical composition, chemical properties, or size of constituent molecules. This study evaluated the ability to control the passage of fluorescent dye-filled liposomes through the intestinal mucosal barrier after oral administration. For this purpose, the increase in transcutaneous registered fluorescence from tetrabromofluorescein dye was recorded and analysed. Fluorescence intensity was measured at the proximal end of the tail of an animal model after oral administration of the liposomes. Measurements were taken at the excitation wavelengths of 365 and 450 nm. The fluorescence intensity in the group treated with the fluorescent contrast agent encapsulated in liposomal particles increased 140% of the initial level, but in the group treated with pure contrast agent, the increase in detected fluorescence intensity did not exceed 110%. Mice that received empty liposomes as well as the control group did not demonstrate statistically significant changes in fluorescence intensity. A potential application of our results is an express laser optical method of monitoring the transport of orally administered liposomal particles. The results can be used to help create new optical tools for use in the development of new drugs and in high-throughput screening used during their testing.

Synthesis of novel β-cyclodextrin functionalized S, N codoped carbon dots for selective detection of testosterone.

PubMed

Luo, Mai; Hua, Yifan; Liang, Yiran; Han, Jiajun; Liu, Donghui; Zhao, Wenting; Wang, Peng

2017-12-15

A novel functionalized carbon dot has been synthesized by covalently linking β-cyclodextrin to the surface of N, S codoped carbon dots (β-CD-CDs). The characterization was confirmed by transmission electron microscopy, X-ray photoelectron spectroscopy, infrared spectra, ultraviolet-visible, and fluorescence emission spectra. On the basis of this carbon dot and (ferrocenylmethyl) trimethylammonium iodide (Fc + ), a photo-induced electron transfer (PET) fluorescent probe system was developed to determine the concentration of testosterone in water and identify testosterone in cell by fluorescence imaging as a visible biomarker. Under the optimum condition, the fluorescent intensity of the probe system linearly responded to the concentration of testosterone from 0μM to 280μM and the limit of detection was 0.51μM. This probe system also performed well at determining testosterone in groundwater with average recoveries of testosterone ranging from 96% to 107% at spiking levels of 0.5-100μM, and the relative standard deviation remained below 13%, which provided a reliable, rapid and easy method to determine testosterone in environmental water. Furthermore, the low cytotoxicity, high anti-interference ability, and excellent biocompatibility of β-CD-CDs made this probe system successfully used in cell fluorescence imaging to monitor levels of testosterone in the cytoplasm of cells with a promising application value in medical research. Copyright © 2017 Elsevier B.V. All rights reserved.

Hydrogen-bonding dynamics of free flavins in benzene and FAD in electron-transferring flavoprotein upon excitation.

PubMed

Sato, Kyousuke; Nishina, Yasuzou; Shiga, Kiyoshi; Tanaka, Fumio

2003-01-01

The dynamic natures of two hydrogen-bonding model systems, riboflavin tetrabutylate (RFTB)-trichloroacetic acid (TCA) and RFTB-phenol in benzene, and of electron-transferring flavoprotein (ETF) from pig kidney upon excitation of flavins was investigated by means of steady state and time-resolved fluorescence spectroscopy. In both model systems fluorescence intensities of RFTB decreased as TCA or phenol was added. The spectral characteristics of ETF under steady state excitation were quite similar to those of the RFTB-TCA system, but not to those of the RFTB-phenol system. The observed fluorescence decay curves of ETF fit well with the calculated decay curves with two lifetime components, as in the model systems. Averaged lifetime was 0.9 ns. The time-resolved fluorescence spectrum of ETF shifted toward longer wavelength with time after pulsed excitation, which was also observed in the RFTB-TCA system. In the RFTB-phenol system the emission spectrum did not shift at all with time. These results reveal that the dynamic nature of ETF can be ascribed to aliphatic hydrogen-bonding(s) of the isoalloxazine ring with surrounding amino acid(s). From the fluorescence characteristics of ETF in comparison with the model systems, human ETF and other flavoproteins, it was suggested that ETF from pig kidney does not contain Tyr-16 in the beta subunit, unlike human ETF.

Recording membrane potential changes through photoacoustic voltage sensitive dye

NASA Astrophysics Data System (ADS)

Zhang, Haichong K.; Kang, Jeeun; Yan, Ping; Abou, Diane S.; Le, Hanh N. D.; Thorek, Daniel L. J.; Kang, Jin U.; Gjedde, Albert; Rahmim, Arman; Wong, Dean F.; Loew, Leslie M.; Boctor, Emad M.

2017-03-01

Monitoring of the membrane potential is possible using voltage sensitive dyes (VSD), where fluorescence intensity changes in response to neuronal electrical activity. However, fluorescence imaging is limited by depth of penetration and high scattering losses, which leads to low sensitivity in vivo systems for external detection. In contrast, photoacoustic (PA) imaging, an emerging modality, is capable of deep tissue, noninvasive imaging by combining near infrared light excitation and ultrasound detection. In this work, we develop the theoretical concept whereby the voltage-dependent quenching of dye fluorescence leads to a reciprocal enhancement of PA intensity. Based on this concept, we synthesized a novel near infrared photoacoustic VSD (PA-VSD) whose PA intensity change is sensitive to membrane potential. In the polarized state, this cyanine-based probe enhances PA intensity while decreasing fluorescence output in a lipid vesicle membrane model. With a 3-9 μM VSD concentration, we measured a PA signal increase in the range of 5.3 % to 18.1 %, and observed a corresponding signal reduction in fluorescence emission of 30.0 % to 48.7 %. A theoretical model successfully accounts for how the experimental PA intensity change depends on fluorescence and absorbance properties of the dye. These results not only demonstrate the voltage sensing capability of the dye, but also indicate the necessity of considering both fluorescence and absorbance spectral sensitivities in order to optimize the characteristics of improved photoacoustic probes. Together, our results demonstrate photoacoustic sensing as a potential new modality for sub-second recording and external imaging of electrophysiological and neurochemical events in the brain.

Fluorescent pH sensor based on Ag@SiO2 core-shell nanoparticle.

PubMed

Bai, Zhenhua; Chen, Rui; Si, Peng; Huang, Youju; Sun, Handong; Kim, Dong-Hwan

2013-06-26

We have demonstrated a novel method for the preparation of a fluorescence-based pH sensor by combining the plasmon resonance band of Ag core and pH sensitive dye (HPTS). A thickness-variable silica shell is placed between Ag core and HPTS dye to achieve the maximum fluorescence enhancement. At the shell thickness of 8 nm, the fluorescence intensity increases 4 and 9 times when the sensor is excited at 405 and 455 nm, respectively. At the same time, the fluorescence intensity shows a good sensitivity toward pH value in the range of 5-9, and the ratio of emission intensity at 513 nm excited at 455 nm to that excited at 405 nm versus the pH value in the range of 5-9 is determined. It is believed that the present pH sensor has the potential for determining pH real time in the biological sample.

A novel fluorescent retrograde neural tracer: cholera toxin B conjugated carbon dots

NASA Astrophysics Data System (ADS)

Zhou, Nan; Hao, Zeyu; Zhao, Xiaohuan; Maharjan, Suraj; Zhu, Shoujun; Song, Yubin; Yang, Bai; Lu, Laijin

2015-09-01

The retrograde neuroanatomical tracing method is a key technique to study the complex interconnections of the nervous system. Traditional tracers have several drawbacks, including time-consuming immunohistochemical or immunofluorescent staining procedures, rapid fluorescence quenching and low fluorescence intensity. Carbon dots (CDs) have been widely used as a fluorescent bio-probe due to their ultrasmall size, excellent optical properties, chemical stability, biocompatibility and low toxicity. Herein, we develop a novel fluorescent neural tracer: cholera toxin B-carbon dot conjugates (CTB-CDs). It can be taken up and retrogradely transported by neurons in the peripheral nervous system of rats. Our results show that CTB-CDs possess high photoluminescence intensity, good optical stability, a long shelf-life and non-toxicity. Tracing with CTB-CDs is a direct and more economical way of performing retrograde labelling experiments. Therefore, CTB-CDs are reliable fluorescent retrograde tracers.The retrograde neuroanatomical tracing method is a key technique to study the complex interconnections of the nervous system. Traditional tracers have several drawbacks, including time-consuming immunohistochemical or immunofluorescent staining procedures, rapid fluorescence quenching and low fluorescence intensity. Carbon dots (CDs) have been widely used as a fluorescent bio-probe due to their ultrasmall size, excellent optical properties, chemical stability, biocompatibility and low toxicity. Herein, we develop a novel fluorescent neural tracer: cholera toxin B-carbon dot conjugates (CTB-CDs). It can be taken up and retrogradely transported by neurons in the peripheral nervous system of rats. Our results show that CTB-CDs possess high photoluminescence intensity, good optical stability, a long shelf-life and non-toxicity. Tracing with CTB-CDs is a direct and more economical way of performing retrograde labelling experiments. Therefore, CTB-CDs are reliable fluorescent retrograde tracers. Electronic supplementary information (ESI) available: PL spectra of CTB; absorption spectra of dialysate; fluorescence signal and immunohistochemical staining of CTB-CDs in L4 DRG. See DOI: 10.1039/c5nr04361a

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

NASA Astrophysics Data System (ADS)

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

2018-03-01

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

Excitation anisotropy in laser-induced-fluorescence spectroscopy: Broad-line excitation case

NASA Astrophysics Data System (ADS)

Hirabayashi, A.; Nambu, Y.; Fujimoto, T.

1986-01-01

Treatment of excitation anisotropy for Laser-Induced-Fluorescence Spectroscopy (LIFS) is extended to the intense excitation case. The depolarization coefficient is derived for intense excitation limit (linearly-polarized or unpolarized light excitation), and the result is presented in tables. For the region of intermediate intensity between the weak and intense excitation limits, the master equation is solved for specific example of transitions and its result is compared with experiment.

The fluorescence properties of aerosol larger than 0.8 μm in urban and tropical rainforest locations

NASA Astrophysics Data System (ADS)

Gabey, A. M.; Stanley, W. R.; Gallagher, M. W.; Kaye, P. H.

2011-06-01

UV-LIF measurements were performed on ambient aerosol in Manchester, UK (urban city centre, winter) and Borneo, Malaysia (remote, tropical) using a Wide Issue Bioaerosol Spectrometer, version 3 (WIBS3). These sites are taken to represent environments with minor and significant primary biological aerosol (PBA) influences respectively, and the urban dataset describes the fluorescent background aerosol against which PBA must be identified by researchers using LIF. The ensemble aerosol at both sites was characterised over 2-3 weeks by measuring the fluorescence intensity and optical equivalent diameter (DP) of single particles sized 0.8 ≤ DP ≤ 20 μm. Filter samples were also collected for a subset of the Manchester campaign and analysed using energy dispersive X-Ray (EDX) spectroscopy and environmental scanning electron microscopy (ESEM), which revealed mostly non-PBA at D ≤ 1 μm. The WIBS3 features three fluorescence channels: the emission following a 280 nm excitation is recorded at 310-400 nm (channel F1) and 400-600 nm (F2), and fluorescence excited at 350 nm is detected at 400-600 nm (F3). In Manchester the primary size mode of fluorescent and non-fluorescent material was present at 0.8-1.2 μm, with a secondary fluorescent mode at 2-4 μm. In Borneo non-fluorescent material peaked at 0.8-1.2 μm and fluorescent at 3-4 μm. Agreement between fluorescent number concentrations in each channel differed at the two sites, with F1 and F3 reporting similar concentrations in Borneo but F3 outnumbering F1 by a factor of 2-3 across the size spectrum in Manchester. The fluorescence intensity in each channel generally rose with DP at both sites with the exception of F1 intensity in Manchester, which peaked at DP = 4 μm, causing a divergence between F1 and F3 intensity at larger DP. This divergence and the differing fluorescent particle concentrations demonstrate the additional discrimination provided by the F1 channel in Manchester. The relationships between fluorescence intensities in different pairs of channels were also investigated as a function of DP. Differences between these metrics were apparent at each site and provide some distinction between the two datasets. Finally, particle selection criteria based on the Borneo dataset were applied to identify a median concentration of 10 "Borneo-like" fluorescent particles per litre in Manchester.

Fluorescence dynamics of human epidermis (ex vivo) and skin (in vivo)

NASA Astrophysics Data System (ADS)

Salomatina, Elena V.; Pravdin, Alexander B.

2003-10-01

The temporal behavior of autofluorescence of human skin and epidermis under continuous UV-irradiation has been studied. Fluorescence spectra and kinetic curves of fluorescence intensity have been obtained. The fluorescence intensity recovery after dark period also has been examined. The vitiligo skin and epidermis were used for comparing their spectra with reflectance and fluorescence spectra of healthy skin. The epidermal samples were prepared using surface epidermis stripping technique. It has been concluded that fluorophores being undergone the UVA photobleaching are actually present in epidermal layer, and immediate pigment darkening does contribute, no less than a half of magnitude, to the autofluorescence decrease under continuous UVA irradiation.

Light response characteristics of a morphologically diverse group of southern hemisphere conifers as measured by chlorophyll fluorescence.

PubMed

Brodribb, T; Hill, Robert S

1997-03-01

Unlike northern hemisphere conifer families, the southern family, Podocarpaceae, produces a great variety of foliage forms ranging from functionally broad-, to needle-leaved. The production of broad photosynthetic surfaces in podocarps has been linked qualitatively to low-light-environments, and we undertook to assess the validity of this assumption by measuring the light response of a morphologically diverse group of podocarps. The light response, as apparent photochemical electron transport rate (ETR), was measured by modulated fluorescence in ten species of this family and six associated species (including five Cupressaceae and one functionally needle-leaved angiosperm) all grown under identical glasshouse conditions. In all species, ETR was found to increase as light intensity increased, reaching a peak value (ETR max ) at saturating quantum flux (PPFD sat ), and decreasing thereafter. ETR max ranged from 217 μmol electrons · m -2  · s -1 at a PPFD sat of 1725 μmol photons · m -2  · s -1 in Actinostrobus acuminatus to an ETR of 60 μmol electrons · m -2  · s -1 at a PPFD sat of 745 μmol electrons · m -2  · s -1 in Podocarpus dispermis. Good correlations were observed between ETR max and both PPFD sat and maximum assimilation rate measured by gas-exchange analysis. The effective quantum yield at light saturation remained constant in all species with an average value of 0.278 ± 0.0035 determined for all 16 species. Differences in the shapes of light response curves were related to differences in the response of non-photochemical quenching (q n ), with q n saturating faster in species with low PPFD sat . Amongst the species of Podocarpaceae, the log of average shoot width was well correlated with PPFD sat , wider leaves saturating at lower light intensities. This suggests that broadly flattened shoots in the Podocarpaceae are an adaptation to low light intensity.

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

PubMed

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

2015-09-15

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

Variations of attractors and wavelet spectra of the immunofluorescence distributions for women in the pregnant period

NASA Astrophysics Data System (ADS)

Galich, Nikolay E.

2008-07-01

Communication contains the description of the immunology data treatment. New nonlinear methods of immunofluorescence statistical analysis of peripheral blood neutrophils have been developed. We used technology of respiratory burst reaction of DNA fluorescence in the neutrophils cells nuclei due to oxidative activity. The histograms of photon count statistics the radiant neutrophils populations' in flow cytometry experiments are considered. Distributions of the fluorescence flashes frequency as functions of the fluorescence intensity are analyzed. Statistic peculiarities of histograms set for women in the pregnant period allow dividing all histograms on the three classes. The classification is based on three different types of smoothing and long-range scale averaged immunofluorescence distributions, their bifurcation and wavelet spectra. Heterogeneity peculiarities of long-range scale immunofluorescence distributions and peculiarities of wavelet spectra allow dividing all histograms on three groups. First histograms group belongs to healthy donors. Two other groups belong to donors with autoimmune and inflammatory diseases. Some of the illnesses are not diagnosed by standards biochemical methods. Medical standards and statistical data of the immunofluorescence histograms for identifications of health and illnesses are interconnected. Peculiarities of immunofluorescence for women in pregnant period are classified. Health or illness criteria are connected with statistics features of immunofluorescence histograms. Neutrophils populations' fluorescence presents the sensitive clear indicator of health status.

Energy dispersive X-ray fluorescence determination of cadmium in uranium matrix using Cd Kα line excited by continuum

NASA Astrophysics Data System (ADS)

Dhara, Sangita; Misra, N. L.; Aggarwal, S. K.; Venugopal, V.

2010-06-01

An energy dispersive X-ray fluorescence method for determination of cadmium (Cd) in uranium (U) matrix using continuum source of excitation was developed. Calibration and sample solutions of cadmium, with and without uranium were prepared by mixing different volumes of standard solutions of cadmium and uranyl nitrate, both prepared in suprapure nitric acid. The concentration of Cd in calibration solutions and samples was in the range of 6 to 90 µg/mL whereas the concentration of Cd with respect to U ranged from 90 to 700 µg/g of U. From the calibration solutions and samples containing uranium, the major matrix uranium was selectively extracted using 30% tri-n-butyl phosphate in dodecane. Fixed volumes (1.5 mL) of aqueous phases thus obtained were taken directly in specially designed in-house fabricated leak proof Perspex sample cells for the energy dispersive X-ray fluorescence measurements and calibration plots were made by plotting Cd Kα intensity against respective Cd concentration. For the calibration solutions not having uranium, the energy dispersive X-ray fluorescence spectra were measured without any extraction and Cd calibration plots were made accordingly. The results obtained showed a precision of 2% (1 σ) and the results deviated from the expected values by < 4% on average.

Experimental assessment of fluorescence microscopy signal enhancement by stimulated emission

NASA Astrophysics Data System (ADS)

Dake, Fumihiro; Yazawa, Hiroki

2017-10-01

The quantity of photons generated during fluorescence microscopy is principally determined by the quantum yield of the fluorescence dyes and the optical power of the excitation beam. However, even though low quantum yields can produce poor images, it is challenging to tune this parameter, while increasing the power of the excitation beam often results in photodamage. Here, we propose the use of stimulated emission (SE) as a means of enhancing both the signal intensity and signal-to-noise ratio during confocal fluorescence microscopy. This work experimentally confirmed that both these factors can be enhanced by SE radiation, through generating a greater number of photons than are associated with the standard fluorescence signal. We also propose the concept of stimulated emission enhancing fluorescence (SEEF) microscopy, which employs both the SE and fluorescence signals, and demonstrate that the intensity of an SEEF signal is greater than those of the individual SE and fluorescence signals.

Position-specific incorporation of fluorescent non-natural amino acids into maltose-binding protein for detection of ligand binding by FRET and fluorescence quenching.

PubMed

Iijima, Issei; Hohsaka, Takahiro

2009-04-17

Position-specific incorporation of fluorescent groups is a useful method for analysis of the functions and structures of proteins. We have developed a method for the incorporation of visible-wavelength-fluorescent non-natural amino acids into proteins in a cell-free translation system. Using this technique, we introduced one or two BODIPY-linked amino acids into maltose-binding protein (MBP) to obtain MBP derivatives showing ligand-dependent changes in fluorescence intensity or intensity ratio. BODIPY-FL-aminophenylalanine was incorporated in place of 15 tyrosines, as well as the N-terminal Lys1, and the C-terminal Lys370 of MBP. Fluorescence measurements revealed that MBP containing a BODIPY-FL moiety in place of Tyr210 showed a 13-fold increase in fluorescence upon binding of maltose. Tryptophan-to-phenylalanine substitutions suggest that the increase in fluorescence was the result of a decrease in the quenching of BODIPY-FL by tryptophan located around the binding site. MBP containing a BODIPY-558 moiety also showed a maltose-dependent increase in fluorescence. BODIPY-FL was then additionally incorporated in place of Lys1 of the BODIPY-558-containing MBP as a response to the amber codon. Fluorescence measurements with excitation of BODIPY-FL showed a large change in fluorescence intensity ratio (0.13 to 1.25) upon binding of maltose; this change can be attributed to fluorescence resonance energy transfer (FRET) and maltose-dependent quenching of BODIPY-558. These results demonstrate the usefulness of the position-specific incorporation of fluorescent amino acids in the fluorescence-based detection of protein functions.

Hybrid phosphorescence and fluorescence native spectroscopy for breast cancer detection.

PubMed

Alimova, Alexandra; Katz, A; Sriramoju, Vidyasagar; Budansky, Yuri; Bykov, Alexei A; Zeylikovich, Roman; Alfano, R R

2007-01-01

Fluorescence and phosphorescence measurements are performed on normal and malignant ex vivo human breast tissues using UV LED and xenon lamp excitation. Tryptophan (trp) phosphorescence intensity is higher in both normal glandular and adipose tissue when compared to malignant tissue. An algorithm based on the ratio of trp fluorescence intensity at 345 nm to phosphorescence intensity at 500 nm is successfully used to separate normal from malignant tissue types. Normal specimens consistently exhibited a low I(345)I(500) ratio (<10), while for malignant specimens, the I(345)I(500) ratio is consistently high (>15). The ratio analysis correlates well with histopathology. Intensity ratio maps with a spatial resolution of 0.5 mm are generated in which local regions of malignancy could be identified.

Measurement of intracellular nitric oxide (NO) production in shrimp haemocytes by flow cytometry.

PubMed

Xian, Jian-An; Guo, Hui; Li, Bin; Miao, Yu-Tao; Ye, Jian-Min; Zhang, Sheng-Peng; Pan, Xun-Bin; Ye, Chao-Xia; Wang, An-Li; Hao, Xuan-Ming

2013-12-01

A flow cytometric method to measure the production of intracellular nitric oxide (NO) was adapted for use with shrimp haemocytes. We applied fluorescent probe 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM DA) for NO detection in haemocytes from the tiger shrimp Penaeus monodon, and used flow cytometry to quantify fluorescence intensity in individual haemocyte. The optimized protocol for intracellular NO analysis consists to incubate haemocytes with DAF-FM DA at 10 μM for 60 min to determine the mean fluorescence intensity. Result showed that NO was also produced in the untreated shrimp haemocytes. NO level in granular cells and semigranular cells were much higher than that in hyaline cells. Defined by different characteristic of NO content, three subsets of haemocytes were observed. Zymosan A at dose of 10 or 100 particles per haemocyte triggered higher DAF-FM fluorescence intensity in granular and semigranular cells, than PMA that had no significant impact on all three cell types. These results indicate that granular and semigranular cells are the primary cells for NO generation. Cytochalasin B significantly inhibited the NO level induced by zymosan A. NG-Monomethyl-L-arginine (L-NMMA) and diphenylene iodonium chloride (DPI) significantly suppressed the DAF-FM fluorescence in haemocytes, but apocynin could not modulate it, indicating that the DAF-FM fluorescence was closely related to the activity of NO-synthase pathway. The NO donor sodium nitroprusside (SNP) improved the DAF-FM fluorescence in haemocytes, while the NO scavenger C-PTIO (2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide) significantly decreased the fluorescence, demonstrating that the fluorescence intensity of DAF-FM is mainly dependent on the intracellular NO level.

Fluorescent Approaches to High Throughput Crystallography

NASA Technical Reports Server (NTRS)

Pusey, Marc L.; Forsythe, Elizabeth; Achari, Aniruddha

2006-01-01

We have shown that by covalently modifying a subpopulation, less than or equal to 1%, of a macromolecule with a fluorescent probe, the labeled material will add to a growing crystal as a microheterogeneous growth unit. Labeling procedures can be readily incorporated into the final stages of purification, and the presence of the probe at low concentrations does not affect the X-ray data quality or the crystallization behavior. The presence of the trace fluorescent label gives a number of advantages when used with high throughput crystallizations. The covalently attached probe will concentrate in the crystal relative to the solution, and under fluorescent illumination crystals show up as bright objects against a dark background. Non-protein structures, such as salt crystals, will not incorporate the probe and will not show up under fluorescent illumination. Brightly fluorescent crystals are readily found against less bright precipitated phases, which under white light illumination may obscure the crystals. Automated image analysis to find crystals should be greatly facilitated, without having to first define crystallization drop boundaries as the protein or protein structures is all that shows up. Fluorescence intensity is a faster search parameter, whether visually or by automated methods, than looking for crystalline features. We are now testing the use of high fluorescence intensity regions, in the absence of clear crystalline features or "hits", as a means for determining potential lead conditions. A working hypothesis is that kinetics leading to non-structured phases may overwhelm and trap more slowly formed ordered assemblies, which subsequently show up as regions of brighter fluorescence intensity. Preliminary experiments with test proteins have resulted in the extraction of a number of crystallization conditions from screening outcomes based solely on the presence of bright fluorescent regions. Subsequent experiments will test this approach using a wider range of proteins. The trace fluorescently labeled crystals will also emit with sufficient intensity to aid in the automation of crystal alignment using relatively low cost optics, further increasing throughput at synchrotrons.

The Fluorescent Properties of Dissolved Organic Matter and Assessment of Total Nitrogen in Overlying Water with Different Dissolved Oxygen Conditions.

PubMed

Zhang Hua; Kuan, Wang; Song, Jian; Zhang, Yong; Huang, Ming; Huang, Jian; Zhu, Jing; Huang, Shan; Wang, Meng

2016-03-01

This paper used excitation-emission matrix spectroscopy (EEMs) to probe the fluorescence properties of dissolved organic matter (DOM) in the overlying water with different dissolved oxygen (DO) conditions, investigating the relationship between protein-like fluorescence intensity and total nitrogen concentration. The resulting fluorescence spectra revealed three protein-like components (high-excitation wavelength tyrosine, low-excitation wavelength tyrosine, low-excitation wavelength tryptophan) and two fulvic-like components (ultraviolet fulvic-like components, visible fulvic-like components) in the overlying water. Moreover, the protein-like components were dominant in the overlying water's DOM. The fluorescence intensity of the protein-like components decreased significantly after aeration. Two of the protein-like components--the low-excitation wavelength tyrosine and the low-excitation wavelength tryptophan--were more susceptible to degradation by microorganisms within the degradable organic matter with respect to the high-excitation wavelength tyrosine. In contrast, the ultraviolet and visible fulvic-like fluorescence intensity increased along with increasing DO concentration, indicating that the fulvic-like components were part of the refractory organics. The fluorescence indices of the DOM in the overlying water were between 1.65-1.80, suggesting that the sources of the DOM were related to terrigenous sediments and microbial metabolic processes, with the primary source being the contribution from microbial metabolism. The fluorescence indices increased along with DO growth, which showed that microbial biomass and microbial activity gradually increased with increasing DO while microbial metabolism also improved, which also increased the biogenic components in the overlying water. The fluorescence intensity of the high-excitation wavelength tyrosine peak A showed a good linear relationship with the total nitrogen concentration at higher DO concentrations of 2.5, 3.5, and 5.5 mg x L(-1), with r2 being 0.956, 0.946, and 0.953, respectively. This study demonstrated that excitation-emission matrix spectroscopy can distinguish the transformation characteristics of the DOM and identify the linear relationship between the fluorescence intensity of the high-excitation wavelength tyrosine peak A and total nitrogen concentration, thus providing a quick and effective technique and theoretical support for river water monitoring and water restoration.

IR-stimulated visible fluorescence in pink and brown diamond.

PubMed

Byrne, K S; Chapman, J G; Luiten, A N

2014-03-19

Irradiation of natural pink and brown diamond by middle-ultraviolet light (photon energy ϵ ≥ 4.1 eV ) is seen to induce anomalous fluorescence phenomena at N3 defect centres (structure N3-V). When diamonds primed in this fashion are subsequently exposed to infrared light (even with a delay of many hours), a transient burst of blue N3 fluorescence is observed. The dependence of this IR-triggered fluorescence on pump wavelength and intensity suggest that this fluorescence phenomena is intrinsically related to pink diamond photochromism. An energy transfer process between N3 defects and other defect species can account for both the UV-induced fluorescence intensity changes, and the apparent optical upconversion of IR light. From this standpoint, we consider the implications of this N3 fluorescence behaviour for the current understanding of pink diamond photochromism kinetics.

Fluorescence properties of Schiff base - N,N‧-bis(salicylidene) - 1,2-Phenylenediamine in presence of bile acid host

NASA Astrophysics Data System (ADS)

Roy, Nayan; Paul, Pradip C.; Singh, T. Sanjoy

2015-05-01

Fluorescence properties of Schiff base - N,N‧-bis(salicylidene) - 1,2-phenylenediamine (LH2) is used to study the micelles formed by aggregation of different important bile acids like cholic acid, deoxycholic acid, chenodeoxycholic acid and glycocholic acid by steady state and picosecond time-resolved fluorescence spectroscopy. The fluorescence band intensity was found out to increase with concomitant red shift with gradual addition of different bile acids. Binding constant of the probe with different bile acids as well as critical micelle concentration was obtained from the variation of fluorescence intensity on increasing concentration of bile acids in the medium. The increase in fluorescence quantum yields, fluorescence decay times and substantial decrease in nonradiative decay rate constants in bile acids micellar environment points to the restricted motion of the fluorophore inside the micellar subdomains.

Critical analysis of commonly used fluorescence metrics to characterize dissolved organic matter.

PubMed

Korak, Julie A; Dotson, Aaron D; Summers, R Scott; Rosario-Ortiz, Fernando L

2014-02-01

The use of fluorescence spectroscopy for the analysis and characterization of dissolved organic matter (DOM) has gained widespread interest over the past decade, in part because of its ease of use and ability to provide bulk DOM chemical characteristics. However, the lack of standard approaches for analysis and data evaluation has complicated its use. This study utilized comparative statistics to systematically evaluate commonly used fluorescence metrics for DOM characterization to provide insight into the implications for data analysis and interpretation such as peak picking methods, carbon-normalized metrics and the fluorescence index (FI). The uncertainty associated with peak picking methods was evaluated, including the reporting of peak intensity and peak position. The linear relationship between fluorescence intensity and dissolved organic carbon (DOC) concentration was found to deviate from linearity at environmentally relevant concentrations and simultaneously across all peak regions. Comparative analysis suggests that the loss of linearity is composition specific and likely due to non-ideal intermolecular interactions of the DOM rather than the inner filter effects. For some DOM sources, Peak A deviated from linearity at optical densities a factor of 2 higher than that of Peak C. For carbon-normalized fluorescence intensities, the error associated with DOC measurements significantly decreases the ability to distinguish compositional differences. An in-depth analysis of FI determined that the metric is mostly driven by peak emission wavelength and less by emission spectra slope. This study also demonstrates that fluorescence intensity follows property balance principles, but the fluorescence index does not. Copyright © 2013 Elsevier Ltd. All rights reserved.

Evaluation of natural organic matter changes from Lake Hohloh by three-dimensional excitation-emission matrix fluorescence spectroscopy during TiO(2)/UV process.

PubMed

Valencia, Sergio; Marín, Juan M; Restrepo, Gloria; Frimmel, Fritz H

2014-03-15

This study shows the changes of natural organic matter (NOM) from Lake Hohloh, (Black Forest, Germany) during heterogeneous photocatalysis with TiO2 (TiO2/UV). The effect of pH on the adsorption of NOM onto TiO2 in the dark and TiO2/UV degradation of NOM was followed using three-dimensional excitation-emission matrix (EEM) fluorescence. At pH values between 4 and 9, the NOM was adsorbed onto TiO2 in the dark with a greater decrease in the fluorescence intensity and in the spectral shapes, especially under acidic pH conditions. However, at pH = 10 there was not adsorption on NOM which led to a negligible changes the fluorescence intensity. A significant high linear correlation was observed between the DOC adsorption onto TiO2 and the maximum fluorescence intensity. Additionally, the NOM adsorption onto TiO2 and its TiO2/UV degradation shifted the fluorescence maxima toward shorter wavelengths in the EEM contour plots, with a decrease in aromaticity. These changes were accompanied by a substantial decrease in the organically bound halogens adsorbable on activated carbon (AOXFP) and the trihalomethane formation potential (THMFP). Thus, the decrease in maximum fluorescence intensity can be used as an indicator of AOXFP and TTHMFP removal efficiency. Therefore, fluorescence spectroscopy is a robust analytical technique for evaluate TiO2/UV removal of NOM. Copyright © 2013 Elsevier Ltd. All rights reserved.

Red Fluorescent Line Emission from Hydrogen Molecules in Diffuse Molecular Clouds

NASA Technical Reports Server (NTRS)

Neufeld, David A.; Spaans, Marco

1996-01-01

We have modeled the fluorescent pumping of electronic and vibrational emissions of molecular hydrogen (H2) within diffuse molecular clouds that are illuminated by ultraviolet continuum radiation. Fluorescent line intensities are predicted for transitions at ultraviolet, infrared, and red visible wavelengths as functions of the gas density, the visual extinction through the cloud, and the intensity of the incident UV continuum radiation. The observed intensity in each fluorescent transition is roughly proportional to the integrated rate of H2 photodissociation along the line of sight. Although the most luminous fluorescent emissions detectable from ground-based observatories lie at near-infrared wavelengths, we argue that the lower sky brightness at visible wavelengths makes the red fluorescent transitions a particularly sensitive probe. Fabry-Perot spectrographs of the type that have been designed to observe very faint diffuse Ha emissions are soon expected to yield sensitivities that will be adequate to detect H2 vibrational emissions from molecular clouds that are exposed to ultraviolet radiation no stronger than the mean radiation field within the Galaxy. Observations of red H2 fluorescent emission together with cospatial 21 cm H I observations could serve as a valuable probe of the gas density in diffuse molecular clouds.

AN INVESTIGATION OF THE BENZOIN METHOD FOR THE FLUORIMETRIC DETERMINATION OF BORON

DOE Office of Scientific and Technical Information (OSTI.GOV)

Elliott, G.; Radley, J.A.

1961-01-01

The development of the boron -benzoin fluorescence at microgram concentrations of boron was investigated; a simple, but sensitive, fluorimeter was used. The development and decay of fluorescence intensity with time were observed in various solvents in the presence of different basic compounds. The fluorescence produced when formamide and its N-derivatives are used as the solvent media is stronger than that found when ethanol is used. A glycine buffer solution of pH 12.8 is effective in producing the correct conditions for developing fluorescence with ethanol as solvent, but is not effective in the formamide series of solvents. Isopropylamine and isobutylamine aremore » effective bases in both ethanol and the formamide series. For a series of solvents of a given chemical type, e.g., the formamides, there may be an increase in fluorescence intensity with dielectric constant, although this is not true for the alcohols. Oxygen has a pronounced inhibiting action on the development of fluorescence in ethanol, but has much less effect in formamide. There is a linear relationship between fluorescence intensity and amount of boron present in the range studied (0.05 to 0.5 - g). (auth)« less

Ethidium bromide as a marker of mtDNA replication in living cells

NASA Astrophysics Data System (ADS)

Villa, Anna Maria; Fusi, Paola; Pastori, Valentina; Amicarelli, Giulia; Pozzi, Chiara; Adlerstein, Daniel; Doglia, Silvia Maria

2012-04-01

Mitochondrial DNA (mtDNA) in tumor cells was found to play an important role in maintaining the malignant phenotype. Using laser scanning confocal fluorescence microscopy (LSCFM) in a recent work, we reported a variable fluorescence intensity of ethidium bromide (EB) in mitochondria nucleoids of living carcinoma cells. Since when EB is bound to nucleic acids its fluorescence is intensified; a higher EB fluorescence intensity could reflect a higher DNA accessibility to EB, suggesting a higher mtDNA replication activity. To prove this hypothesis, in the present work we studied, by LSCFM, the EB fluorescence in mitochondria nucleoids of living neuroblastoma cells, a model system in which differentiation affects the level of mtDNA replication. A drastic decrease of fluorescence was observed after differentiation. To correlate EB fluorescence intensity to the mtDNA replication state, we evaluated the mtDNA nascent strands content by ligation-mediated real-time PCR, and we found a halved amount of replicating mtDNA molecules in differentiating cells. A similar result was obtained by BrdU incorporation. These results indicate that the low EB fluorescence of nucleoids in differentiated cells is correlated to a low content of replicating mtDNA, suggesting that EB may be used as a marker of mtDNA replication in living cells.

Assessment of tissue ischemia of nail fold precapillary zones using a fluorescence capillaroscopy

NASA Astrophysics Data System (ADS)

Dremin, Viktor V.; Margaryants, Nikita B.; Volkov, Mikhail V.; Zhukova, Ekaterina V.; Zherebtsov, Evgeny A.; Dunaev, Andrey V.; Rafailov, Edik U.

2017-07-01

An optical instrument for nailfold fluorescence capillaroscopy and image registration has been developed. With this instrument, an effect of increasing fluorescence intensity in the spectral range of NADH fluorescence during ischemia was detected.

Variations in the endogenous fluorescence of rabbit corneas after mechanical property alterations

NASA Astrophysics Data System (ADS)

Ortega-Martinez, Antonio; Touchette, Genna; Zhu, Hong; Kochevar, Irene E.; Franco, Walfre

2017-09-01

Keratoconus is an eye disease in which the cornea progressively deforms due to loss of cornea mechanical rigidity, and thus causes deterioration of visual acuity. Techniques to characterize the mechanical characteristics of the cornea are important to better monitor changes and response to treatments. To investigate the feasibility of using the endogenous fluorescence of cornea for monitoring alterations of its mechanical rigidity, linear tensiometry was used to quantitate stiffness and Young's modulus (YM) after treatments that increase cornea stiffness (collagen photocross-linking) or decrease stiffness (enzymatic digestion). The endogenous ultraviolet fluorescence of cornea was also measured before and after these treatments. The fluorescence excitation/emission spectral ranges were 280 to 430/390 to 520 nm, respectively. A correlation analysis was carried out to identify fluorescence excitation/emission pairs whose intensity changes correlated with the stiffness. A positive correlation was found between variations in fluorescence intensity of the 415-/485-nm excitation/emission pair and YM of photocross-linked corneas. After treatment of corneas with pepsin, the YM decreased as the fluorescence intensity at 290-/390-nm wavelengths decreased. For weakening of corneas with collagenase, only qualitative changes in the fluorescence spectrum were observed. Changes in the concentration of native or newly created fluorescent molecular species contain information that may be directly or indirectly related to the mechanical structure of the cornea.

Conjugated polyelectrolyte based real-time fluorescence assay for phospholipase C.

PubMed

Liu, Yan; Ogawa, Katsu; Schanze, Kirk S

2008-01-01

A fluorescence turnoff assay for phospholipase C (PLC) from Clostridium perfringens is developed based on the reversible interaction between the natural substrate, phosphatidylcholine, and a fluorescent, water-soluble conjugated polyelectrolyte (CPE). The fluorescence intensity of the CPE in water is increased substantially by the addition of the phospholipid due to the formation of a CPE-lipid complex. Incubation of the CPE-lipid complex with the enzyme PLC causes the fluorescence intensity to decrease (turnoff sensor); the response arises due to PLC-catalyzed hydrolysis of the phosphatidylcholine, which effectively disrupts the CPE-lipid complex. The PLC assay operates with phospholipid substrate concentrations in the micromolar range, and the analytical detection limit for PLC is <1 nM. The optimized assay provides a convenient, rapid, and real-time sensor for PLC activity. The real-time fluorescence intensity from the CPE can be converted to substrate concentration by using an ex situ calibration curve, allowing PLC-catalyzed reaction rates and kinetic parameters to be determined. PLC activation by Ca2+ and inhibition by EDTA and fluoride ion are demonstrated using the optimized sensor.

Time-resolved fluorescence and FCS studies of dye-doped DNA

NASA Astrophysics Data System (ADS)

Nicolaou, N.; Marsh, R. J.; Blacker, T.; Armoogum, D. A.; Bain, A. J.

2009-08-01

Fluorescence lifetime, anisotropy and intensity dependent single molecule fluorescence correlation spectroscopy (I-FCS) are used to investigate the mechanism of fluorescence saturation in a free and nucleotide bound fluorophore (NR6104) in an antioxidising ascorbate buffer. Nucleotide attachment does not appreciably affect the fluorescence lifetime of the probe and there is a decrease in the rate of intersystem crossing relative to that of triplet state deactivation. The triplet state fraction is seen to plateau at 72% (G-attached) and 80% (free fluorophore) in agreement with these observations. Measurements of translational diffusion times show no intensity dependence for excitation intensities between 1 and 105kW cm-2 and photobleaching is therefore negligible. The dominant mechanism of fluorescence saturation is thus triplet state formation. I-FCS measurements for Rhodamine 6G in water were compared with those in the ascorbate buffer. In water the triplet fraction was saturated at considerably higher powers (45% at ca. 1.5 × 103kW cm-2) than in the ascorbate buffer (55%ca. 1 1kW cm-2)

Two-dimensional fluorescence correlation spectroscopy: resolution of fluorescence of tryptophan residues in horse heart myoglobin.

PubMed

Nakashima, Kenichi; Yuda, Kazuki; Ozaki, Yukihiro; Noda, Isao

2003-11-01

Generalized two-dimensional (2D) fluorescence correlation spectroscopy has been used to resolve fluorescence of two tryptophan (Trp) residues in horse heart myoglobin. Fluorescence quenching is employed as a perturbation mode for causing intensity changes in the fluorescence (quenching perturbation). Two kinds of quenchers, iodide ion and acrylamide, are used for inducing fluorescence intensity change. This technique works because the Trp residue located at the 7th position (W7) is known to be easily accessible to the quencher, whereas that located at the 14th position (W14) is not. By this technique, the fluorescence spectra of the two Trp residues were clearly resolved. From asynchronous maps, it was also shown that the quenching of W7 fluorescence is brought about prior to the quenching of W14 fluorescence. This result is consistent with the structure of horse heart myoglobin that was proposed earlier. Furthermore, it was elucidated that the present 2D analysis is not interfered with by Raman bands of the solvents, which sometimes brings difficulty into conventional fluorescence analysis.

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

PubMed

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

2018-03-15

In this paper, a novel fluorescent probe, dtpa-bis(cytosine), was designed and synthesized for detecting europium (Eu 3+ ) ion. Upon addition of Eu 3+ ions into the dtpa-bis(cytosine) solution, the fluorescence intensity can strongly be enhanced. Conversely, adding other rare earth metal ions, such as Y 3+ , Ce 3+ , Pr 3+ , Nd 3+ , Sm 3+ , Gd 3+ , Tb 3+ , Dy 3+ , Ho 3+ , Er 3+ , Yb 3+ and Lu 3+ , into dtpa-bis(cytosine) solution, the fluorescence intensity is decreased slightly. Some parameters affecting the fluorescence intensity of dtpa-bis(cytosine) solution in the presence of Eu 3+ ions were investigated, including solution pH value, Eu 3+ ion concentration and interfering substances. The detection mechanism of Eu 3+ ion using dtpa-bis(cytosine) as fluorescent probe was proposed. Under optimum conditions, the fluorescence emission intensities of Eu III -dtpa-bis(cytosine) at 375nm in the concentration range of 0.50×10 -5 mol∙L -1 -5.00×10 -5 mol∙L -1 of Eu 3+ ion display a better linear relationship. The limit of detection (LOD) was determined as 8.65×10 -7 mol∙L -1 and the corresponding correlation coefficient (R 2 ) of the linear equation is 0.9807. It is wished that the proposed method could be applied for sensitively and selectively detecting Eu 3+ ion. Copyright © 2017 Elsevier B.V. All rights reserved.

Chromatic shifts in the fluorescence emitted by murine thymocytes stained with Hoechst 33342.

PubMed

Petersen, Timothy W; Ibrahim, Sherrif F; Diercks, Alan H; van den Engh, Ger

2004-08-01

Many methods in flow cytometry rely on staining DNA with a fluorescent dye to gauge DNA content. From the relative intensity of the fluorescence signature, one can then infer position in cell cycle, amount of DNA (i.e., for sperm selection), or, as in the case of flow karyotyping, to distinguish individual chromosomes. This work examines the staining of murine thymocytes with a common DNA dye, Hoechst 33342, to investigate nonlinearities in the florescence intensity as well as chromatic shifts. Murine thymocytes were stained with Hoechst 33342 and measured in a flow cytometer at two fluorescence emission bands. In other measurements, cells were stained at different dye concentrations, and then centrifuged. The supernatant was then used for a second round of staining to test the amount of dye uptake. Finally, to test for resonant energy transfer, we measured fluorescence anisotropy at two different wavelengths. The fluorescence of cells stained with Hoechst 33342 is a nonlinear process that shows an overall decrease in intensity with increased dye uptake, and spectral shift to the red. Along with the spectral shift of the fluorescence to the longer wavelengths, we document decreases in the fluorescence anisotropy that may indicate resonant energy transfer. At low concentrations, Hoechst 33342 binds to the minor groove of DNA and shows an increase in fluorescence and a blue shift upon binding. At higher concentrations, at which the dye molecules can no longer bind without overlapping, the blue fluorescence decreases and the red fluorescence increases until there is approximately one dye molecule per DNA base pair. The ratio of the blue fluorescence to the red fluorescence is an accurate indicator of the cellular dye concentration.

Chair-side detection of Prevotella Intermedia in mature dental plaque by its fluorescence.

PubMed

Nomura, Yoshiaki; Takeuchi, Hiroaki; Okamoto, Masaaki; Sogabe, Kaoru; Okada, Ayako; Hanada, Nobuhiro

2017-06-01

Prevotella intermedia/nigrescens is one of the well-known pathogens causing periodontal diseases, and the red florescence excited by the visible blue light caused by the protoporphyrin IX in the bacterial cells could be useful for the chair-side detection. The aim of this study was to evaluated levels of periodontal pathogen, especially P. intermedia in clinical samples of red fluorescent dental plaque. Thirty two supra gingival plaque samples from six individuals were measured its fluorescence at 640nm wavelength excited by 409nm. Periodontopathic bacteria were counted by the Invader PLUS PCR assay. Co-relations the fluorescence intensity and bacterial counts were analyzed by Person's correlation coefficient and simple and multiple regression analysis. Positive and negative predictive values of the fluorescence intensities for with or without P. intermedia in supragingival plaque was calculated. When relative fluorescence unit (RFU) were logarithmic transformed, statistically significant linear relations between RFU and bacterial counts were obtained for P. intermedia, Porphyromonas gingivalis and Tannerella forsythia. By the multiple regression analysis, only P. intermedia had statistically significant co-relation with fluorescence intensities. All of the fluorescent dental plaque contained P. intermedia m. In contrast, 28% of non-fluorescent plaques contained P. intermedia. To check the fluorescence dental plaque in the oral cavity could be the simple chair-side screening of the mature dental plaque before examining the periodontal pathogens especially P. intermedia by the PCR method. Copyright © 2017 Elsevier B.V. All rights reserved.

Method and apparatus for evaluating structural weakness in polymer matrix composites

DOEpatents

Wachter, E.A.; Fisher, W.G.

1996-01-09

A method and apparatus for evaluating structural weaknesses in polymer matrix composites is described. An object to be studied is illuminated with laser radiation and fluorescence emanating therefrom is collected and filtered. The fluorescence is then imaged and the image is studied to determine fluorescence intensity over the surface of the object being studied and the wavelength of maximum fluorescent intensity. Such images provide a map of the structural integrity of the part being studied and weaknesses, particularly weaknesses created by exposure of the object to heat, are readily visible in the image. 6 figs.

Method and apparatus for evaluating structural weakness in polymer matrix composites

DOEpatents

Wachter, Eric A.; Fisher, Walter G.

1996-01-01

A method and apparatus for evaluating structural weaknesses in polymer matrix composites is described. An object to be studied is illuminated with laser radiation and fluorescence emanating therefrom is collected and filtered. The fluorescence is then imaged and the image is studied to determine fluorescence intensity over the surface of the object being studied and the wavelength of maximum fluorescent intensity. Such images provide a map of the structural integrity of the part being studied and weaknesses, particularly weaknesses created by exposure of the object to heat, are readily visible in the image.

Does ozone enhance the remineralizing potential of nanohydroxyapatite on artificially demineralized enamel? A laser induced fluorescence study

NASA Astrophysics Data System (ADS)

Srinivasan, Samuelraj; Prabhu, Vijendra; Chandra, Subhash; Koshy, Shalini; Acharya, Shashidhar; Mahato, Krishna K.

2014-02-01

The present era of minimal invasive dentistry emphasizes the early detection and remineralization of initial enamel caries. Ozone has been shown to reverse the initial demineralization before the integrity of the enamel surface is lost. Nano-hydroxyapatite is a proven remineralizing agent for early enamel caries. In the present study, the effect of ozone in enhancing the remineralizing potential of nano-hydroxyapatite on artificially demineralized enamel was investigated using laser induced fluorescence. Thirty five sound human premolars were collected from healthy subjects undergoing orthodontic treatment. Fluorescence was recorded by exciting the mesial surfaces using 325 nm He-Cd laser with 2 mW power. Tooth specimens were subjected to demineralization to create initial enamel caries. Following which the specimens were divided into three groups, i.e ozone (ozonated water for 2 min), without ozone and artificial saliva. Remineralization regimen was followed for 3 weeks. The fluorescence spectra of the specimens were recorded from all the three experimental groups at baseline, after demineralization and remineralization. The average spectrum for each experimental group was used for statistical analysis. Fluorescence intensities of Ozone treated specimens following remineralization were higher than that of artificial saliva, and this difference was found to be statistically significant (P<0.0001). In a nutshell, ozone enhanced the remineralizing potential of nanohydroxyapatite, and laser induced fluorescence was found to be effective in assessing the surface mineral changes in enamel. Ozone can be considered an effective agent in reversing the initial enamel caries there by preventing the tooth from entering into the repetitive restorative cycle.

Fluorescence fingerprinting properties for exploring water-soluble organic compounds in PM2.5 in an industrial city of northwest China

NASA Astrophysics Data System (ADS)

Qin, Juanjuan; Zhang, Leiming; Zhou, Xueming; Duan, Jingchun; Mu, Situ; Xiao, Kang; Hu, Jingnan; Tan, Jihua

2018-07-01

Excitation-emission matrix (EEM) spectra were used to analyze the fluorescence properties of water-soluble organic compounds (WSOC) in PM2.5 during winter and summer seasons in Lanzhou city, northwest China. Protein-like substances, humic-like substances and microbial by-products were found to be the main fluorophoric organic matter. Humification index (HIX), biological index (BIX) and fluorescence index (FI) were 1.2 ± 0.1, 1.4 ± 0.1 and 1.7 ± 0.2 in winter and 2.0 ± 0.3, 1.3 ± 0.1 and 1.2 ± 0.4 in summer, respectively, indicating higher aromaticity of WSOC in summer and terrestrial biogenic sources of WSOC in both seasons. Strong correlations were found between regional average fluorescence intensities (RAFI) at the fluorescence regions of II-V and water-soluble inorganic ions (K+, Cl-, NO3- and SO42-) in winter with the Spearman correlation coefficients ρ being larger than 0.7 and mostly around 0.9, suggesting significant contributions of bioaerosols, coal combustion and vehicle exhaust to PM2.5. Moderate correlations were found between RAFI and K+, Cl-, and NO3- in summer, indicating the more important roles of biogenic and vehicle exhaust than coal combustion sources in summer. High temperature in winter was conducive to and in summer prohibitive to polycondensation of WSOC, indicating the existence of an optimum temperature for such a process to occur. High relative humidity was unfavorable to the formation of fluorescent WSOC.

Real-time monitoring the distribution of antibody-photo-absorber conjugates during photoimmunotherapy in vivo(Conference Presentation)

NASA Astrophysics Data System (ADS)

Tang, Qinggong; Lin, Jonathan; Nagaya, Tadanobu; Liu, Yi; Kobayashi, Hisataka; Chen, Yu

2017-02-01

Photo-immunotherapy (PIT) is an emerging low-side-effect cancer therapy based on monoclonal antibody (mAb) conjugated with a near-infrared (NIR) phthalocyanine dye IRDye700DX (IR700 is not only fluorescent which can be used as an imaging agent, but also phototoxic) that induces rapid cell death after exposure to NIR light. PIT induces highly-selective cancer cell death while leaving most of tumor blood vessels unharmed, leading to an effect termed super-enhanced permeability and retention (SUPR), which significantly improve the effectiveness of anti-cancer drug. Currently, the therapeutic effects of PIT were monitored using IR700 fluorescent signal based on macroscopic fluorescence reflectance imager, which lacks the resolution and depth information to reveal the intra-tumor heterogeneity of mAb-IR700 distribution. We developed a minimally-invasive two-channel fluorescence fiber imaging system by combining the traditional fluorescence imaging microscope with two imaging fiber bundles ( 0.85 mm) to monitor mAb-IR700 distribution and therapeutic effects during PIT at different intra-tumor locations (e.g. tumor periphery vs. tumor rim) in situ and in real time simutaneously, thereby enabling evaluation of the therapeutic effects in vivo and optimization of treatment regimens accordingly. Experiments were carried out on ten mice. The average fluorescence intensity recovery after PIT in tumor rim is 91.50% while 100.63% in tumor periphery. Significantly higher fluorescence redistribution (P=0.0371) in tumor periphery than tumor rim after PIT treatment were observed. In order to verify the results, two-photon microscopy combining with micro-prism was also used to record the mAb-IR700 distribution at different depth locations of the tumor during PIT.

Infection-Mediated Vasoactive Peptides Modulate Cochlear Uptake of Fluorescent Gentamicin

PubMed Central

Koo, Ja-Won; Wang, Qi; Steyger, Peter S.

2011-01-01

Inflammatory mediators released during bacterial infection include vasoactive peptides such as histamine and serotonin, and their serum levels are frequently elevated. These peptides also modulate the vascular permeability of endothelial cells lining the blood-brain and blood-labyrinth barriers (BLB). These peptides may also modulate the permeability of the BLB to ototoxic aminoglycoside antibiotics prescribed to resolve bacterial sepsis. To test this hypothesis, we compared the effect of histamine and serotonin on the cochlear distribution of fluorescently conjugated gentamicin (GTTR) in control animals at 0.5, 1 and 3 h after injection of GTTR. The intensity of GTTR fluorescence was attenuated at 1 h in the histamine group compared to control mice, and more intense 3 h after injection (p < 0.05). In the serotonin group, the intensity of GTTR fluorescence was attenuated at 0.5 and 1 h (p < 0.05) and was increased at 3 h compared to control animals, where GTTR intensities peaked at 1 h and then plateaued or was slightly decreased at 3 h. This biphasic pattern of modulation was statistically significant in the apical turn of the cochlea. No difference in the intensity of GTTR fluorescence was observed in kidney proximal tubules. Systemic increases in serum levels of vasoactive peptides can modulate cochlear uptake of gentamicin, likely via permeability changes in the BLB. Conditions that influence serum levels of vasoactive peptides may potentiate aminoglycoside ototoxicity. PMID:21196726

Fluorescence and chemiluminescence behavior of distyrylbenzene bearing two arms of dipicolylaminomethyl groups: Interactions with zinc ion and ATP

NASA Astrophysics Data System (ADS)

Motoyoshiya, Jiro; Wada, Jun-ya; Itoh, Keiko; Wakabayashi, Kazuaki; Maruyama, Takayuki; Ono, Kazuki; Fukasawa, Kota; Fujimoto, Tetsuya; Akaiwa, Yuji; Nonaka, Eiji

2018-04-01

The absorption and fluorescence spectral study of the distyrylbenzene bearing two arms of the dipicolylaminomethyl groups, the effective ligands for Zn2+, was studied in the presence of Zn2+ and ATP. Upon complexation of the distyrylbenzene with zinc ions in acetonitrile, enhancement of the fluorescence intensity was observed due to inhibition of intramolecular PET (photo-induced electron transfer) quenching, but no effect was found in aqueous media because the equilibrium laid to the free form of the ligands. In contrast, the addition of ATP disodium salt was effective to enhance the fluorescence intensity of the combination of the distyrylbenzne and Zn2+ in aqueous media. This assembly was applied to the peroxyoxalate chemiluminescence system and a significant increase in the intensity was observed, which provides a potential detection for ATP by chemiluminescence.

Fluorescent biosensor for the detection of hyaluronidase: intensity-based ratiometric sensing and fluorescence lifetime-based sensing using a long lifetime azadioxatriangulenium (ADOTA) fluorophore.

PubMed

Chib, Rahul; Mummert, Mark; Bora, Ilkay; Laursen, Bo W; Shah, Sunil; Pendry, Robert; Gryczynski, Ignacy; Borejdo, Julian; Gryczynski, Zygmunt; Fudala, Rafal

2016-05-01

In this report, we have designed a rapid and sensitive, intensity-based ratiometric sensing as well as lifetime-based sensing probe for the detection of hyaluronidase activity. Hyaluronidase expression is known to be upregulated in various pathological conditions. We have developed a fluorescent probe by heavy labeling of hyaluronic acid with a new orange/red-emitting organic azadioxatriangulenium (ADOTA) fluorophore, which exhibits a long fluorescence lifetime (∼20 ns). The ADOTA fluorophore in water has a peak fluorescence lifetime of ∼20 ns and emission spectra centered at 560 nm. The heavily ADOTA-labeled hyaluronic acid (HA-ADOTA) shows a red shift in the peak emission wavelength (605 nm), a weak fluorescence signal, and a shorter fluorescence lifetime (∼4 ns) due to efficient self-quenching and formation of aggregates. In the presence of hyaluronidase, the brightness and fluorescence lifetime of the sample increase with a blue shift in the peak emission to its original wavelength at 560 nm. The ratio of the fluorescence intensity of the HA-ADOTA probe at 560 and 605 nm can be used as the sensing method for the detection of hyaluronidase. The cleavage of the hyaluronic acid macromolecule reduces the energy migration between ADOTA molecules, as well as the degree of self-quenching and aggregation. This probe can be efficiently used for both intensity-based ratiometric sensing as well as fluorescence lifetime-based sensing of hyaluronidase. The proposed method makes it a rapid and sensitive assay, useful for analyzing levels of hyaluronidase in relevant clinical samples like urine or plasma. Graphical Abstract Scheme showing cleavage of HA-ADOTA probe by hyaluronidase and the change in the emission spectrum of HA-ADOTA probe before and after cleavage by hyaluronidase.

Characterization and standardization of tissue-simulating protoporphyrin IX optical phantoms

NASA Astrophysics Data System (ADS)

Marois, Mikael; Bravo, Jaime; Davis, Scott C.; Kanick, Stephen Chad

2016-03-01

Optical devices for measuring protoporphryin IX (PpIX) fluorescence in tissue are routinely validated by measurements in optical phantoms. Yet there exists limited data to form a consensus on the recipe for phantoms that both mimic the optical properties found in tissue and yield a reliable and stable relationship between PpIX concentration and the fluorescence remission intensity. This study characterizes the influence of multiple phantom components on PpIX fluorescence emission intensity, using Intralipid as the scattering source, bovine whole blood as the background absorber, and Tween as a surfactant to prevent PpIX aggregation. Optical measurements showed a linear proportionality (r>0.99) between fluorescence intensity and PpIX concentration (0.1 to 10 μg/mL) over a range of Intralipid (1 to 2%) and whole blood (0.5 to 3%) for phantoms containing low surfactant (≤0.1%), with fluorescence intensities and scattering and absorption properties stable for 5 h after mixing. The role of surfactant in PpIX phantoms was found to be complex, as aggregation was evident in aqueous nonturbid phantoms with no surfactant (0% Tween), and avoided in phantoms containing Intralipid as the scattering source with no additional or low amounts of added surfactant (≤0.1% Tween). Conversely, phantoms containing higher surfactant content (>0.1% Tween) and whole blood showed interactions that distorted the fluorescence emissions.

A long lifetime chemical sensor: study on fluorescence property of fluorescein isothiocyanate and preparation of pH chemical sensor.

PubMed

Ma, Li Ying; Wang, Huai You; Xie, Hui; Xu, Li Xiao

2004-07-01

The fluorescence property of fluorescein isothiocyanate (FITC) in acid-alkaline medium was studied by spectrofluorimetry. The characteristic of FITC response to hydrogen ion has been examined in acid-alkaline solution. A novel pH chemical sensor was prepared based on the relationship between the relative fluorescence intensity of FITC and pH. The measurement of relative fluorescence intensity was carried out at 362 nm with excitation at 250 nm. The excellent linear relationship was obtained between relative fluorescence intensity and pH in the range of pH 1-5. The linear regression equation of the calibration graph is F = 66.871 + 6.605 pH (F is relative fluorescence intensity), with a correlation coefficient of linear regression of 0.9995. Effects of temperature, concentration of FITC on the response to hydrogen ion had been examined. It was important that this chemical sensor was long lifetime, and the property of response to hydrogen ion was stable for at least 70 days. This pH sensor can be used for measuring pH value in water solution. The accuracy is 0.01 pH unit. The results obtained by the pH sensor agreed with those by the pH meter. Obviously, this pH sensor is potential for determining pH change real time in biological system.

A long lifetime chemical sensor: study on fluorescence property of fluorescein isothiocyanate and preparation of pH chemical sensor

NASA Astrophysics Data System (ADS)

Ma, Li Ying; Wang, Huai You; Xie, Hui; Xu, Li Xiao

2004-07-01

The fluorescence property of fluorescein isothiocyanate (FITC) in acid-alkaline medium was studied by spectrofluorimetry. The characteristic of FITC response to hydrogen ion has been examined in acid-alkaline solution. A novel pH chemical sensor was prepared based on the relationship between the relative fluorescence intensity of FITC and pH. The measurement of relative fluorescence intensity was carried out at 362 nm with excitation at 250 nm. The excellent linear relationship was obtained between relative fluorescence intensity and pH in the range of pH 1-5. The linear regression equation of the calibration graph is F=66.871+6.605 pH ( F is relative fluorescence intensity), with a correlation coefficient of linear regression of 0.9995. Effects of temperature, concentration of FITC on the response to hydrogen ion had been examined. It was important that this chemical sensor was long lifetime, and the property of response to hydrogen ion was stable for at least 70 days. This pH sensor can be used for measuring pH value in water solution. The accuracy is 0.01 pH unit. The results obtained by the pH sensor agreed with those by the pH meter. Obviously, this pH sensor is potential for determining pH change real time in biological system.

Sub-annual paleoenvironmental information evaluated from intensity variations of fluorescent annual layers in a stalagmite from Ryuo-do Cave, Nagasaki Prefecture, western Japan

NASA Astrophysics Data System (ADS)

Sasaki, Hana; Onishi, Yuri; Ishihara, Yoshiro; Yoshimura, Kazuhisa

2017-04-01

Stalagmites can provide various types of paleoenvironmental information such as information on vegetation and climate changes. Fluorescent annual layers formed by humic substances (mainly fulvic acids: FA) in these stalagmites can also provide a time proxy, and a time series on precipitation. Fluorescence intensity patterns in these annual layers can be classified into symmetric, gradually increasing and gradually decreasing types. Onishi et al. (EGU2016) demonstrated the existence of these fluorescence intensity patterns in the annual layers, and their stratigraphic changes, by numerical simulations, and suggested that the patterns could provide paleoenvironmental information at a sub-annual resolution. In this study, we carried out an analysis of fluorescence intensity patterns in the annual layers of a stalagmite from Ryuo-do Cave, Nagasaki Prefecture, western Japan, and also simulated the patterns in the stalagmite, to obtain paleoenvironmental information. Fluorescence intensity patterns in the annual layers are strongly affected by annual variations in FA concentration and precipitation rates of calcite. As the result of simulations of fluorescence intensity patterns, cumulative variations and various types of pattern are reproduced. These differences are depending on time lags between the variation of the FA concentration in the drip waters, and that of the growth rate of the stalagmite. Co-precipitation models of FA are divided into the "Hiatus model" in which FA are preferentially preserved in the stalagmite when its growth rate is relatively low, and the "Partition coefficient (PC) model" in which FA concentrations in the stalagmite increase when the calcite precipitation rate is relatively high. However, various fluorescence intensity patterns in the annual layers could be formed under a combination or either of both of the models. Fluorescence intensity patterns in an annual layer in the stalagmite from Ryuo-do Cave, Nagasaki Prefecture, western Japan vary stratigraphically, and multiple types of fluorescence intensity pattern are observed in the stalagmite. When the co-precipitation of FA is governed by the hiatus model, it is suggested that a gradual increase in the annual layers will result from a large accumulation of calcite after the annual peak in the FA concentration, whereas there will be a gradual decrease if the main growth occurs before the annual peak in FA concentration. However, in the case of the PC model, a gradually increasing type of pattern is formed if the main growth occurs before the annual peak in FA concentration, and a gradually decreasing type is formed if the main growth occurs afterwards. If the annual peak of FA concentration occurs several months after high summer, it is suggested that intervals showing a gradually increasing type were formed in winter, and intervals showing a gradually decreasing type were formed in the early summer, in the case of the hiatus model. In the case of PC model, the seasons are reversed. In the climatic environment around the Ryuo-do Cave, the growth rates of stalagmites are affected by cave air circulation in winter and by rainfall (rainy season) in early summer.

Construction of a 'turn-on' fluorescent probe system for His-tagged proteins.

PubMed

Murata, Atsushi; Arai, Satoshi; Yoon, Su-In; Takabayashi, Masao; Ozaki, Miwako; Takeoka, Shinji

2010-12-01

Hexahistidine ((His)(6)) tags are used to purify genetically engineered proteins. Herein, we describe the construction of a 'turn-on' fluorescent probe system that consists of the fluorescence quencher, Dabcyl, conjugated to (His)(6), and fluorescent tetramethylrhodamine conjugated to nitrilotriacetic acid, which, in the presence of Ni(2+), can bind (His)(6). The system is turned off when Dabcyl-(His)(6) is bound to the fluorescent nitrilotriacetic acid derivative. The binding strength of this system was assessed using electrospray ionization mass spectrometry, fluorescence correlation spectroscopy, and fluorescence intensity distribution analysis-polarization. Although there was no significant enhancement in fluorescence after addition of an equimolar amount of ubiquitin, the fluorescence increased from 14% to 40% of its initial intensity when an equimolar amount of (His)(6)-ubiquitin was added. Therefore, this system should be able to specifically recognize (His)(6)-proteins with good resolution and has the additional advantage that a washing step is not required to remove fluorescent probe, that is, not bound to the (His)(6)-protein. Copyright © 2010 Elsevier Ltd. All rights reserved.

Iterative Correction Scheme Based on Discrete Cosine Transform and L1 Regularization for Fluorescence Molecular Tomography With Background Fluorescence.

PubMed

Zhang, Jiulou; Shi, Junwei; Guang, Huizhi; Zuo, Simin; Liu, Fei; Bai, Jing; Luo, Jianwen

2016-06-01

High-intensity background fluorescence is generally encountered in fluorescence molecular tomography (FMT), because of the accumulation of fluorescent probes in nontarget tissues or the existence of autofluorescence in biological tissues. The reconstruction results are affected or even distorted by the background fluorescence, especially when the distribution of fluorescent targets is relatively sparse. The purpose of this paper is to reduce the negative effect of background fluorescence on FMT reconstruction. After each iteration of the Tikhonov regularization algorithm, 3-D discrete cosine transform is adopted to filter the intermediate results. And then, a sparsity constraint step based on L1 regularization is applied to restrain the energy of the objective function. Phantom experiments with different fluorescence intensities of homogeneous and heterogeneous background are carried out to validate the performance of the proposed scheme. The results show that the reconstruction quality can be improved with the proposed iterative correction scheme. The influence of background fluorescence in FMT can be reduced effectively because of the filtering of the intermediate results, the detail preservation, and noise suppression of L1 regularization.

Formation and Fluorimetric Characterization of Micelles in a Micro-flow Through System with Static Micro Mixer

PubMed Central

Schuch, Michael; Gross, G. Alexander; Köhler, J. Michael

2007-01-01

The formation and behaviour of micelles of sodium dodecylsulfate in water by use of a static micro mixer were studied. Trisbipyridylruthenium(II) was applied as indicator dye, 9-methylanthracene was used for fluorescence quenching. All experiments were carried out by a micro fluid arrangement with three syringe pumps, a 2+1 two-step static micro mixer (IPHT Jena) and a on-line micro fluorimetry including a luminescence diode for excitation, a blue glass filter (BG 7, Linos), two edge filters (RG 630, Linos) and a photo counting module (MP 900, Perkin Elmer). It was possible to measure the fluorescence inside the PTFE tube (inner diameter 0.5 mm) directly. A linear dependence of fluorescence intensity from dye concentration was observed in absence of quencher and surfactant as expected. An aggregation number of about 62 was found in the flow rate range between 300 and 800 μL/min. The fluorescence intensity increases slightly, but significant with increasing flow rate, if no quencher is present. In the presence of quencher, the fluorescence intensity decreases with decreasing surfactant concentration and with enhanced flow rate. The strength of the flow rate effect on the fluorescence increases with decreasing surfactant concentration. The size of micelles was determined in micro channels by the micro fluorimetric method in analogy to the conventional system. The micelles extract the quencher from the solution and lower, this way, the quenching effect. The size of micelles was estimated and it could be shown, that the flow rate has only low effect on the aggregation number at the investigated flow rates. The effect of flow rate and surfactant concentration on the fluorescence in the presence of quencher was interpreted as a shift in the micelle concentration due to the shear forces. It is expected, that the fluorescence intensity is lowered, if more quencher molecules are molecular disperse distributed inside the solution. Obviously, the lowered fluorescence intensity at higher flow rates suggests a reduction of the micelle density causing an increase of quencher concentration outside the micelles. PMID:28903241

The Rate Constant for Fluorescence Quenching

ERIC Educational Resources Information Center

Legenza, Michael W.; Marzzacco, Charles J.

1977-01-01

Describes an experiment that utilizes fluorescence intensity measurements from a Spectronic 20 to determine the rate constant for the fluorescence quenching of various aromatic hydrocarbons by carbon tetrachloride in an ethanol solvent. (MLH)

Production and degradation of fluorescent dissolved organic matter in surface waters of the eastern north Atlantic ocean

NASA Astrophysics Data System (ADS)

Lønborg, Christian; Yokokawa, Taichi; Herndl, Gerhard J.; Antón Álvarez-Salgado, Xosé

2015-02-01

The distribution and fate of coloured dissolved organic matter (CDOM) in the epipelagic Eastern North Atlantic was investigated during a cruise in the summer 2009 by combining field observations and culture experiments. Dissolved organic carbon (DOC) and nitrogen (DON), the absorption spectra of CDOM and the fluorescence intensity of proteins (Ex/Em 280/320 nm; F(280/320)) and marine humic-like substances (F(320/410)) were measured in the upper 200 m. DOC and DON showed higher concentrations in the top 20 m than below, and DOC increased southwards, while DON decreased. F(280/320) and F(320/410) showed maxima near the deep chlorophyll maximum (at about 50 m), suggesting that these fluorophores were linked to phytoplankton production and the metabolism of the associated microbial community. The coloured and fluorescent fractions of DOM showed low levels south of the Azores Front, at about 35 °N, likely due to the accumulated photobleaching of the waters transported eastwards by the Azores current into the study area (at 20°W). Twelve culture experiments were also conducted with surface water (5 m) to assess the impact of microbial degradation processes on the bulk, coloured and fluorescent fractions of DOM. After 72 h of incubation in the darkness, 14±9% (average±SD) of the initial DON was consumed at an average rate of 0.24±0.14 μmol l-1 d-1 and the protein-like fluorescence decayed by 29±9% at a net rate of 0.06±0.03 QSU d-1. These rates were significantly lower south of the Azores front, suggesting that DOM in this region was of a more recalcitrant nature. Conversely, the marine humic-like fluorescence increased at a net rate of 0.013±0.003 QSU d-1. The close linear relationship of DON uptake with F(280/320) consumption (R2= 0.91, p <0.0001, n=12) and F(320/410) production (R2= 0.52, p <0.008, n=12) that we found during these incubation experiments suggest that the protein-like fluorescence can be used as a proxy for the dynamics of the labile DON pool and that marine humic-like materials can be produced as a by-product of microbial DOM degradation.

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

PubMed

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

2015-01-01

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

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

PubMed Central

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

2015-01-01

We introduce here MATtrack, an open source MATLAB-based computational platform developed to process multi-Tiff files produced by a photo-conversion time lapse protocol for live cell fluorescent microscopy. MATtrack automatically performs a series of steps required for image processing, including extraction and import of numerical values from Multi-Tiff files, red/green image classification using gating parameters, noise filtering, background extraction, contrast stretching and temporal smoothing. MATtrack also integrates a series of algorithms for quantitative image analysis enabling the construction of mean and standard deviation images, clustering and classification of subcellular regions and injection point approximation. In addition, MATtrack features a simple user interface, which enables monitoring of Fluorescent Signal Intensity in multiple Regions of Interest, over time. The latter encapsulates a region growing method to automatically delineate the contours of Regions of Interest selected by the user, and performs background and regional Average Fluorescence Tracking, and automatic plotting. Finally, MATtrack computes convenient visualization and exploration tools including a migration map, which provides an overview of the protein intracellular trajectories and accumulation areas. In conclusion, MATtrack is an open source MATLAB-based software package tailored to facilitate the analysis and visualization of large data files derived from real-time live cell fluorescent microscopy using photoconvertible proteins. It is flexible, user friendly, compatible with Windows, Mac, and Linux, and a wide range of data acquisition software. MATtrack is freely available for download at eleceng.dit.ie/courtney/MATtrack.zip. PMID:26485569

Effect of surfactant and budesonide on the pulmonary distribution of fluorescent dye in mice.

PubMed

Huang, Liang-Ti; Yeh, Tsu-Fu; Kuo, Yu-Lin; Chen, Pin-Chuan; Chen, Chung-Ming

2015-02-01

Surfactant is a useful vehicle for the intratracheal delivery of medicine to the distal lung. The aim of this study was to analyze the effect of intratracheal surfactant and budesonide instillation on the pulmonary distribution of fluorescent dye in mice. Male athymic nude mice were assigned randomly as controls, fluorescent dye, fluorescent dye + surfactant (50 mg/kg), fluorescent dye + budesonide (0.25 mg/kg), and fluorescent dye + surfactant + budesonide groups. A total volume of 60 μL fluorescent solutions was intratracheally injected and followed by 60 μL of air. We photographed and measured fluorescence in the lungs, from the back, 15 minutes after intratracheal administration using an IVIS Xenogen imaging instrument. The fluorescent dye (1,1'-dioctadecyltetramethyl indotricarbocyanine iodide) was most strongly detected near the trachea and weakly detected in the lungs in mice administered with fluorescent solutions. Almost no fluorescence was seen in the lung region of control mice. Intratracheal administration of surfactant or budesonide increased fluorescent intensity compared with control mice. Combined administration of surfactant and budesonide further increased fluorescent intensity compared with mice given surfactant or budesonide alone. Surfactant and budesonide enhance the pulmonary distribution of fluorescent dye in mice. Copyright © 2014. Published by Elsevier B.V.

Fluorescence Imaging Reveals Surface Contamination

NASA Technical Reports Server (NTRS)

Schirato, Richard; Polichar, Raulf

1992-01-01

In technique to detect surface contamination, object inspected illuminated by ultraviolet light to make contaminants fluoresce; low-light-level video camera views fluorescence. Image-processing techniques quantify distribution of contaminants. If fluorescence of material expected to contaminate surface is not intense, tagged with low concentration of dye.

Determination of ethambutol by a sensitive fluorescent probe

NASA Astrophysics Data System (ADS)

Wu, Wen-Ying; Yang, Ji-Yuan; Du, Li-Ming; Wu, Hao; Li, Chang-Feng

2011-08-01

The competitive reaction between ethambutol and two fluorescent probes (i.e., berberine and palmatine) for occupancy of the cucurbit[7]uril (CB[7]) cavity was studied by spectrofluorometry. The CB[7] reacts with these probes to form stable complexes, and the fluorescence intensity of the complexes is greatly enhanced. In addition, the excitation and emission wavelengths of their complexes moved to wavelengths of 343 nm and 495 nm, respectively. However, the addition of ethambutol dramatically quenches the fluorescence intensity of the two complexes. Accordingly, a couple of new fluorescence quenching methods for the determination of ethambutol were established. The methods can be applied for quantifying ethambutol. A linear relationship between the fluorescence quenching values (Δ F) and ethambutol concentration exists in the range of 5.0-1000.0 ng mL -1, with a correlation coefficient ( r) of 0.9997. The detection limit is 1.7 ng mL -1. The fluorescent probe of berberine has higher sensitivity than palmatine. This paper also discusses the mechanism of fluorescence indicator probes.

Quantifying exciton hopping in disordered media with quenching sites: Application to arrays of quantum dots

NASA Astrophysics Data System (ADS)

Miyazaki, Jun

2013-10-01

We present an analytical method for quantifying exciton hopping in an energetically disordered system with quenching sites. The method is subsequently used to provide a quantitative understanding of exciton hopping in a quantum dot (QD) array. Several statistical quantities that characterize the dynamics (survival probability, average number of distinct sites visited, average hopping distance, and average hopping rate in the initial stage) are obtained experimentally by measuring time-resolved fluorescence intensities at various temperatures. The time evolution of these quantities suggests in a quantitative way that at low temperature an exciton tends to be trapped at a local low-energy site, while at room temperature, exciton hopping occurs repeatedly, leading to a large hopping distance. This method will serve to facilitate highly efficient optoelectronic devices using QDs such as photovoltaic cells and light-emitting diodes, since exciton hopping is considered to strongly influence their operational parameters. The presence of a dark QD (quenching site) that exhibits fast decay is also quantified.

P63 EXPRESSION LEVELS IN SIDE POPULATION AND LOW LIGHT SCATTERING OCULAR SURFACE EPITHELIAL CELLS

PubMed Central

Epstein, Seth P; Wolosin, J. Mario; Asbell, Penny A

2005-01-01

Purpose Because stem cells exhibit high self-renewal capacity, slow cycling, and high proliferative potential, and one of many markers postulated for epithelial stem cells, p63, is challenged by widespread expression within stem cell–free regions, we examined p63 expression in these stem cell–associated cohorts compared with their controls. Methods Rabbit limbocorneal cryosections, cytospun cell-sorted (by fluorescence-activated cell sorter) side population (SP) and low side scatter (LSSC) cells, and limbal epithelial cells over feeders were stained for p63 by indirect immunofluorescence. Clones were fixed and stained daily for 7 days. Image analysis measured p63 intensity, plotting it against colony size. Results All basal limbal cells were positive for p63, yet only 5% to 7% expressed high p63 intensities, 40% intermediate, and the majority low. Side population cells were less than 1% of total cells. The average intensity of SP staining was three times that of controls. Subpopulations displaying stemlike features exhibited highest p63 expression. Replication rates of isolated cells differed. Day 5 colonies contained 256 (16 hours/cycle) to two (96 hours/cycle) cells. Whereas all cells were positive for p63, intensity in slow-cycling cells was three to four times that in rapidly proliferating congeners. Increased cell doublings did not decrease fluorescence. Conclusions Results suggest that p63 concentration is maximal in stem cells and decreases with differentiation. High p63 levels seem to correlate with cells of the SP and LSSC phenotypes, indicating high cell stemness. With identification of stem cells, further studies can elucidate their use in supporting ocular surface health. PMID:17057802

MEH-PPV film thickness influenced fluorescent quenching of tip-coated plastic optical fiber sensors

NASA Astrophysics Data System (ADS)

Yusufu, A. M.; Noor, A. S. M.; Tamchek, N.; Abidin, Z. Z.

2017-12-01

The performance of plastic optical fiber sensors in detecting nitro aromatic explosives 1,4-dinitrobenzene (DNB) have been investigated by fluorescence spectroscopy and analyzed by using fluorescence quenching technique. The plastic optical fiber utilized is 90 degrees cut tip and dip-coated with conjugated polymer MEH-PPV poly[2-methoxy-5-(2-ethylhexyloxy)-1,4-phenylenevinylene] thin films for detection conjugants. The thicknesses of the MEH-PPV coating were varied to improvise the sensitivity whilst slowly reducing the fluorescence intensity. It was shown that fluorescence intensity from thinner film decreased by (82% in 40 s) in the presence of DNB signifying an improvement of 28% reduction with time 13 s less than that of the thicker film.

Photosensitizer-induced fluorescence of the rat adrenal gland and rat pheochromocytoma cells (PC 12) by meso-tetra(hydroxyphenyl)chlorin (mTHPC)

NASA Astrophysics Data System (ADS)

Colombo-Benkmann, Mario; Muhm, Markus; Gahlen, Johannes; Heym, Christine; Senninger, Norbert

1997-12-01

Rat adrenal glands exhibit an intense mTHPC-induced fluorescence. The objective of our study was the identification of adrenal cells exhibiting mTHPC-induced fluorescence under normal conditions and under stimulation of adrenal proliferation by reserpine. Furthermore mTHPC-uptake of rat pheochromocytoma (PC 12) cells was investigated. Four male Wistar rats received 0.5 mg mTHPC/kg iv 48 hours before perfusion. Furthermore four rats received reserpine (2 mg/kg im od), bromo-deoxy-uridine (BrdU; 50 mg/kg ip od) each for one week and mTHPC (0.5 mg/kg) 48 hours before perfusion. BrdU was detected immunohistochemically. PC 12-cells were incubated with 0.5 mg mTHPC/l culture medium for 24 or 48 hours. Cells and tissues were examined by fluorescence microscopy. The adrenal cortex exhibited an intense mTHPC-induced fluorescence. The adrenal medulla fluoresced faintly. Reserpine increased fluorescence of intramedullary cells, not coinciding with adrenal proliferation. Cortical fluorescence remained unchanged. PC 12-cells lying singly or in small groups and differentiating cells showed a more intense mTHPC- induced fluorescence than confluent cells. Differences of cortical and medullary uptake of mTHPC are independent of proliferation and may be explained by lipophilia of mTHPC, since adrenocytes have an uptake mechanism for cholesterol. The difference of mTHPC-uptake between PC 12-cells and chromaffin cells implicate the possibility of photodynamic applications for medullary neoplasia.

Detection of airway ischaemic damage after lung transplantation by using autofluorescence imaging bronchoscopy.

PubMed

Iga, Norichika; Oto, Takahiro; Okada, Masanori; Harada, Masaaki; Nishikawa, Hitoshi; Miyoshi, Kentaroh; Otani, Shinji; Sugimoto, Seiichiro; Yamane, Masaomi; Toyooka, Shinichi; Miyoshi, Shinichiro

2014-03-01

Airway complications related to ischaemia are a major cause of morbidity after lung transplantation. Early detection of airway ischaemia and optimal management of the anastomotic site could reduce the risk of airway complications. Autofluorescence imaging (AFI) bronchoscopy has been increasingly recognized as an effective technique for detecting abnormal mucosal thickening. The aim of this study was to investigate whether AFI bronchoscopy can facilitate the detection of airway ischaemic damage in lung transplant patients. Twenty Landrace pigs were used to create a tracheal autotransplantation model. A four-ring length of trachea was excised and implanted orthotopically. The tracheal autograft was observed on postoperative days 0, 2, 4 and 7 with AFI bronchoscopy. The extent and origin of graft autofluorescence were examined using histology and measured according to fluorescence intensity. The lesions on the tracheal autografts appeared as bright green fluorescence on AFI bronchoscopy. On confocal fluorescence microscopy, high-intensity green fluorescence was observed in the elastin fibre layer of the submucosa. The fluorescence intensity of elastin was significantly higher in the graft showing fluorescence than the graft that did not show fluorescence and that at the control site. Bright green fluorescence was seen in an elastin fibre layer in the submucosa, which was likely a result of epithelial sloughing. There is a close relationship between the bright green fluorescence pattern observed using AFI bronchoscopy and airway ischaemic damage. We conclude that AFI bronchoscopy may detect airway ischaemic damage after lung transplantation.

Effect of solvents on the fluorescence spectra of bacterial luciferase

NASA Astrophysics Data System (ADS)

Sukovataya, Irina E.; Tyulkova, Natalya A.; Kaykova, Elisaveta V.

2006-08-01

Bacteria luciferases catalyze the oxidation reaction of the long-chain aliphatic aldehyde and reduced flavinmononucleotide involving molecular oxygen to a respective fatty acid emitting light quanta in the visible spectrum. Fluorescence emission of luciferases from Photobacterium leiognathi dissolved in organic solvent-water mixtures was investigated. Methanol, acetone, dimethyl sulfoxide and formamide were used as organic solvents. As the methanol and acetone concentration is increased the emission maximum peak is decrease. In contrast, with dimethyl sulfoxide and formamide addition induced a increasing of the emission maximum intensity. The values of wavelength maximum (λ max) at the addition of this solvent can shows the spectra shifted to the red by about 12 nm. These increasing in the fluorescence intensity and in the λ max may be due to luciferase denaturation, resulting from the more intensive contact of chromospheres of luciferase with the solvent. At all used concentrations of methanol, acetone and formamide the shape of the fluorescence spectra was not changed. These studies demonstrate that the luciferase tryptophan fluorescence is sensitive to changes of physical-chemical property of enzyme environment. A comparison of activation/inactivation and fluorescence spectra of luciferase in methanol or acetone solutions shows that the extent of inactivation is larger than the extent of fluorescence changes at the same methanol or acetone concentration.

Light-induced flickering of DsRed provides evidence for distinct and interconvertible fluorescent states.

PubMed Central

Malvezzi-Campeggi, F; Jahnz, M; Heinze, K G; Dittrich, P; Schwille, P

2001-01-01

Green fluorescent protein (GFP) from jellyfish Aequorea victoria, the powerful genetically encoded tag presently available in a variety of mutants featuring blue to yellow emission, has found a red-emitting counterpart. The recently cloned red fluorescent protein DsRed, isolated from Discosoma corals (), with its emission maximum at 583 nm, appears to be the long awaited tool for multi-color applications in fluorescence-based biological research. Studying the emission dynamics of DsRed by fluorescence correlation spectroscopy (FCS), it can be verified that this protein exhibits strong light-dependent flickering similar to what is observed in several yellow-shifted mutants of GFP. FCS data recorded at different intensities and excitation wavelengths suggest that DsRed appears under equilibrated conditions in at minimum three interconvertible states, apparently fluorescent with different excitation and emission properties. Light absorption induces transitions and/or cycling between these states on time scales of several tens to several hundreds of microseconds, dependent on excitation intensity. With increasing intensity, the emission maximum of the static fluorescence continuously shifts to the red, implying that at least one state emitting at longer wavelength is preferably populated at higher light levels. In close resemblance to GFP, this light-induced dynamic behavior implies that the chromophore is subject to conformational rearrangements upon population of the excited state. PMID:11509387

Monitoring the Behavior of Emerging Contaminants in Wastewater-Impacted Rivers Based on the Use of Fluorescence Excitation Emission Matrixes (EEM).

PubMed

Sgroi, Massimiliano; Roccaro, Paolo; Korshin, Gregory V; Vagliasindi, Federico G A

2017-04-18

This study investigated the applicability of fluorescence indexes based on the interpretation of excitation emission matrices (EEMs) by PARAFAC analysis and by selecting fluorescence intensities at a priori defined excitation/emission pairs as surrogates for monitoring the behavior of emerging organic compounds (EOCs) in two catchment basins impacted by wastewater discharges. Relevant EOC and EEM data were obtained for a 90 km stretch of the Simeto River, the main river in Sicily, and the smaller San Leonardo River, which was investigated for a 17 km stretch. The use of fluorescence indexes developed by these two different approaches resulted in similar observations. Changes of the fluorescence indexes that correspond to a group of humic-like fluorescing species were determined to be highly correlated with the concentrations of recalcitrant contaminants such as sucralose, sulfamethoxazole and carbamazepine, which are typical wastewater markers in river water. Changes of the fluorescence indexes related to tyrosine-like substances were well correlated with the concentrations of ibuprofen and caffeine, anthropogenic indicators of untreated wastewater discharges. Chemical oxygen demand and dissolved organic carbon concentrations were correlated with humic-like fluorescence indexes. The observed correlations were site-specific and characterized by different regression parameters for every collection event. Caffeine and carbamazepine showed correlations with florescence indexes in the San Leonardo River and in the alluvial plain stretch of the Simeto River, whereas sucralose, sulfamethoxazole and ibuprofen have always been well correlated in all the investigated river stretches. However, when data of different collection events from river stretches where correlations were observed were combined, good linear correlations were obtained for data sets generated via the normalization of the measured concentrations by the average value for the corresponding collection event. These results show that fluorescence based indexes can be used to monitor the behavior of some trace organic contaminants in wastewater impacted rivers and to track wastewater discharges in streams and rivers.

A Flow-Cytometric Gram-Staining Technique for Milk-Associated Bacteria

PubMed Central

Holm, Claus; Jespersen, Lene

2003-01-01

A Gram-staining technique combining staining with two fluorescent stains, Oregon Green-conjugated wheat germ agglutinin (WGA) and hexidium iodide (HI) followed by flow-cytometric detection is described. WGA stains gram-positive bacteria while HI binds to the DNA of all bacteria after permeabilization by EDTA and incubation at 50°C for 15 min. For WGA to bind to gram-positive bacteria, a 3 M potassium chloride solution was found to give the highest fluorescence intensity. A total of 12 strains representing some of the predominant bacterial species in bulk tank milk and mixtures of these were stained and analyzed by flow cytometry. Overall, the staining method showed a clear differentiation between gram-positive and gram-negative bacterial populations. For stationary-stage cultures of seven gram-positive bacteria and five gram-negative bacteria, an average of 99% of the cells were correctly interpreted. The method was only slightly influenced by the growth phase of the bacteria or conditions such as freezing at −18°C for 24 h. For any of these conditions, an average of at least 95% of the cells were correctly interpreted. When stationary-stage cultures were stored at 5°C for 14 days, an average of 86% of the cells were correctly interpreted. The Gram-staining technique was applied to the flow cytometry analysis of bulk tank milk inoculated with Staphylococcus aureus and Escherichia coli. These results demonstrate that the technique is suitable for analyzing milk samples without precultivation. PMID:12732558

A flow-cytometric gram-staining technique for milk-associated bacteria.

PubMed

Holm, Claus; Jespersen, Lene

2003-05-01

A Gram-staining technique combining staining with two fluorescent stains, Oregon Green-conjugated wheat germ agglutinin (WGA) and hexidium iodide (HI) followed by flow-cytometric detection is described. WGA stains gram-positive bacteria while HI binds to the DNA of all bacteria after permeabilization by EDTA and incubation at 50 degrees C for 15 min. For WGA to bind to gram-positive bacteria, a 3 M potassium chloride solution was found to give the highest fluorescence intensity. A total of 12 strains representing some of the predominant bacterial species in bulk tank milk and mixtures of these were stained and analyzed by flow cytometry. Overall, the staining method showed a clear differentiation between gram-positive and gram-negative bacterial populations. For stationary-stage cultures of seven gram-positive bacteria and five gram-negative bacteria, an average of 99% of the cells were correctly interpreted. The method was only slightly influenced by the growth phase of the bacteria or conditions such as freezing at -18 degrees C for 24 h. For any of these conditions, an average of at least 95% of the cells were correctly interpreted. When stationary-stage cultures were stored at 5 degrees C for 14 days, an average of 86% of the cells were correctly interpreted. The Gram-staining technique was applied to the flow cytometry analysis of bulk tank milk inoculated with Staphylococcus aureus and Escherichia coli. These results demonstrate that the technique is suitable for analyzing milk samples without precultivation.

Thermal maturity of Tasmanites microfossils from confocal laser scanning fluorescence microscopy

USGS Publications Warehouse

Hackley, Paul C.; Kus, Jolanta

2015-01-01

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

Interpretation of the fluorescence signatures from vegetation

NASA Astrophysics Data System (ADS)

Buschmann, C.

Vegetation emits fluorescence as part of the energy taken up by absorption %of solar radiation from UV to the visible. This fluorescence consists of light with low intensity (only few percents of the reflected light) emitted from the leaves. The fluorescence emission of a green leaf is characterized by four bands with maxima in the blue (440 nm), green (520 nm), red (690 nm) and far red (740 nm) spectral region. The intensity of fluorescence in the maxima of the emission spectrum varies depending on the following six basic parameters which must be taken into account for the interpretation of fluorescence signatures from vegetation: (a) content of the fluorophores (ferulic acid, chlorophyll a), (b) temperature of the leaf, (c) penetration of excitation light into the leaf, (d) emission of fluorescence from the leaf (re-absorption inside the leaf tissue), (e) photosynthetic activity of the leaf, (f) non-radiative decay (heat production) parallel to the fluorescence The ratios between the intensities of the maxima (F440/F690, F440/F520, F690/F740) are used as characteristic fluorescence parameter. The wide range of changes of these ratios caused by differences in the leaf tissue (aerial interspaces, variegated/homogeneous green leaves), various types of stress (UV, photoinhibition, sun exposure, heat, water deficiency, N-deficiency) and chemicals (inhibitors, fertilizers) can be explained by changes of the six basic parameters. It will be shown that the interpretation of the fluorescence signatures, in most cases, must be based on a complex consideration of more than one of the basic parameters.

Filter Enhances Fluorescent-Penetrant-Inspecting Borescope

NASA Technical Reports Server (NTRS)

Molina, Orlando G.

1990-01-01

Slip-on eyepiece for commercial ultraviolet-light borescope reduces both amount of short-wave ultraviolet light that reaches viewer's eye and apparent intensity of unwanted reflections of white light from surfaces undergoing inspection. Fits on stock eyepiece of borescope, which illuminates surface inspected with intense ultraviolet light. Surface, which is treated with fluorescent dye, emits bright-green visible light wherever dye penetrates - in cracks and voids. Eyepiece contains deep-yellow Wratten 15 (G) filter, which attenuates unwanted light strongly but passes yellow-green fluorescence so defects seen clearly.

Blinking fluorescence of single donor-acceptor pairs: important role of "dark'' states in resonance energy transfer via singlet levels.

PubMed

Osad'ko, I S; Shchukina, A L

2012-06-01

The influence of triplet levels on Förster resonance energy transfer via singlet levels in donor-acceptor (D-A) pairs is studied. Four types of D-A pair are considered: (i) two-level donor and two-level acceptor, (ii) three-level donor and two-level acceptor, (iii) two-level donor and three-level acceptor, and (iv) three-level donor and three-level acceptor. If singlet-triplet transitions in a three-level acceptor molecule are ineffective, the energy transfer efficiency E=I_{A}/(I_{A}+I_{D}), where I_{D} and I_{A} are the average intensities of donor and acceptor fluorescence, can be described by the simple theoretical equation E(F)=FT_{D}/(1+FT_{D}). Here F is the rate of energy transfer, and T_{D} is the donor fluorescence lifetime. In accordance with the last equation, 100% of the donor electronic energy can be transferred to an acceptor molecule at FT_{D}≫1. However, if singlet-triplet transitions in a three-level acceptor molecule are effective, the energy transfer efficiency is described by another theoretical equation, E(F)=F[over ¯](F)T_{D}/[1+F[over ¯](F)T_{D}]. Here F[over ¯](F) is a function of F depending on singlet-triplet transitions in both donor and acceptor molecules. Expressions for the functions F[over ¯](F) are derived. In this case the energy transfer efficiency will be far from 100% even at FT_{D}≫1. The character of the intensity fluctuations of donor and acceptor fluorescence indicates which of the two equations for E(F) should be used to find the value of the rate F. Therefore, random time instants of photon emission in both donor and acceptor fluorescence are calculated by the Monte Carlo method for all four types of D-A pair. Theoretical expressions for start-stop correlators (waiting time distributions) in donor and acceptor fluorescence are derived. The probabilities w_{N}^{D}(t) and w_{N}^{A}(t) of finding N photons of donor and acceptor fluorescence in the time interval t are calculated for various values of the energy transfer rate F and for all four types of D-A pair. Comparison of the calculated D and A fluorescence trajectories with those measured by Weiss and co-workers proves the important role of triplet levels in energy transfer via singlet levels.

[Laser Induced Fluorescence Spectroscopic Analysis of Aromatics from One Ring to Four Rings].

PubMed

Zhang, Peng; Liu, Hai-feng; Yue, Zong-yu; Chen, Bei-ling; Yao, Ming-fa

2015-06-01

In order to distinguish small aromatics preferably, a Nd : YAG Laser was used to supply an excitation laser, which was adjusted to 0.085 J x cm(-2) at 266 nm. Benzene, toluene, naphthalene, phenanthrene, anthracene, pyrene and chrysene were used as the representative of different rings aromatics. The fluorescence emission spectra were researched for each aromatic hydrocarbon and mixtures by Laser induced fluorescence (LIF). Results showed that the rings number determined the fluorescence emission spectra, and the structure with same rings number did not affect the emission fluorescence spectrum ranges. This was due to the fact that the absorption efficiency difference at 266 nm resulted in that the fluorescence intensities of each aromatic hydrocarbon with same rings number were different and the fluorescence intensities difference were more apparently with aromatic ring number increasing. When the absorption efficiency was similar at 266 nm and the concentrations of each aromatic hydrocarbon were same, the fluorescence intensities were increased with aromatic ring number increasing. With aromatic ring number increasing, the fluorescence spectrum and emission peak wavelength were all red-shifted from ultraviolet to visible and the fluorescence spectrum range was also wider as the absorption efficiency was similar. The fluorescence emission spectra from one to four rings could be discriminated in the following wavelengths, 275 to 320 nm, 320 to 375 nm, 375 to 425 nm, 425 to 556 nm, respectively. It can be used for distinguish the type of the polycyclic aromatic hydrocarbons (PAHs) as it exists in single type. As PAHs are usually exist in a variety of different rings number at the same time, the results for each aromatic hydrocarbon may not apply to the aromatic hydrocarbon mixtures. For the aromatic hydrocarbon mixtures, results showed that the one- or two-ring PAHs in mixtures could not be detected by fluorescence as three- or four-ring PAHs existed in mixture. This was caused by radiation energy transfer mechanism, in which the ultraviolet light was lost in mixtures but the fluorescence intensities were increased with the one- or two-ring PAHs adding. When the mixture only contained three- and four-ring PAHs, the fluorescence emission spectrum showed the both characteristics of three- and four-ring PAHs fluorescence. When three- and four-ring PAHs existed in mixtures at the same time, the fluorescence emission spectra were related to each concentration, so the rings number could be discriminated to a certain extent.

Fluorescence detection system for microfluidic droplets

NASA Astrophysics Data System (ADS)

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

2018-05-01

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

Fluorescence from a single Symbiodinium cell

NASA Astrophysics Data System (ADS)

Guzman, Christine; Han, Xue; Shoguchi, Eiichi; Chormaic, Síle Nic

2018-07-01

The partnership between coral and its algal symbionts, Symbiodinium, is crucial to the global environment. Yet, the regulatory process within the photosynthetic machinery of Symbiodinium is still not clearly understood. Here, we studied the influence of light stress from focussed red and blue lasers on single Symbiodinium cells. Fluorescence signals were measured to show cell response. Increasing the incident laser power or the exposure time resulted in an increase followed by a decline in fluorescence intensity. The trend of fluorescence intensity changes was associated with mechanisms of light use efficiency, non-photochemical quenching, photoinhibition, and repair of the cell. Our study provides new approaches to studying the photobiology and physiology of Symbiodinium cells.

Neoplasm diagnostics based on fluorescence of polymethine dyes

NASA Astrophysics Data System (ADS)

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

2002-05-01

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

A novel small molecule mediate 18F-FDG excited fluorescence molecular imaging

NASA Astrophysics Data System (ADS)

Zhang, Zeyu; Guo, Hongbo; Hu, Zhenhua; Tian, Jie

2018-02-01

Fluorescence molecular imaging (FMI) has been widely used in many medical fields with small molecule indocyanine green (ICG). However, low signal-background ratio and limited specificity to tumor remain big challenges for FMI. In this study, a novel excitation strategy is proposed on the basis of clinical approved ICG and 18F-FDG. A series of in vitro experiments are designed to reveal the mechanism and results show obvious decreasing of ICG fluorescence intensity with the increasing distance to excitation source. Meanwhile, the ICG fluorescence intensity is proportional to the activity of radiopharmaceutical. Results from different respects illustrate the promising of this proposed excitation strategy.

Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes

NASA Astrophysics Data System (ADS)

Chow, Yu Ting; Chen, Shuxun; Wang, Ran; Liu, Chichi; Kong, Chi-Wing; Li, Ronald A.; Cheng, Shuk Han; Sun, Dong

2016-04-01

Cell transfection is a technique wherein foreign genetic molecules are delivered into cells. To elucidate distinct responses during cell genetic modification, methods to achieve transfection at the single-cell level are of great value. Herein, we developed an automated micropipette-based quantitative microinjection technology that can deliver precise amounts of materials into cells. The developed microinjection system achieved precise single-cell microinjection by pre-patterning cells in an array and controlling the amount of substance delivered based on injection pressure and time. The precision of the proposed injection technique was examined by comparing the fluorescence intensities of fluorescent dye droplets with a standard concentration and water droplets with a known injection amount of the dye in oil. Injection of synthetic modified mRNA (modRNA) encoding green fluorescence proteins or a cocktail of plasmids encoding green and red fluorescence proteins into human foreskin fibroblast cells demonstrated that the resulting green fluorescence intensity or green/red fluorescence intensity ratio were well correlated with the amount of genetic material injected into the cells. Single-cell transfection via the developed microinjection technique will be of particular use in cases where cell transfection is challenging and genetically modified of selected cells are desired.

Plasmonic hybrid nanostructure with controlled interaction strength

NASA Astrophysics Data System (ADS)

Grzelak, Justyna K.; Krajnik, Bartosz; Thoreson, Mark D.; Nyga, Piotr; Shalaev, Vladimir M.; Mackowski, Sebastian

2014-03-01

In this report we discuss the influence of plasmon excitations in a silver island film on the fluorescence of photosynthetic complex, peridinin-chlorophyll-protein (PCP). Control of the separation between these two components is obtained by fabricating a wedge layer of silica across the substrate, with a thickness from 0 to 46 nm. Continuous variation of the silica thickness allows for gradual change of interaction strength between plasmon excitations in the metallic film and the excited states of pigments comprising photosynthetic complexes. While the largest separation between the silver film and photosynthetic complexes results in fluorescence featuring a mono-exponential decay and relatively narrow distribution of intensities, the PCP complexes placed on thinner silica spacers show biexponential fluorescence decay and significantly broader distribution of total fluorescence intensities. This broad distribution is a signature of stronger sensitivity of fluorescence enhancement upon actual parameters of a hybrid nanostructure. By gradual change of the silica spacer thickness we are able to reproduce classical distance dependence of fluorescence intensity in plasmonic hybrid nanostructures on ensemble level. Experiments carried out for different excitation wavelengths indicate that the interaction is stronger for excitations resonant with plasmon absorption in the metallic layer.

Nanostructured fluorescent particles for glucose sensing

NASA Astrophysics Data System (ADS)

Grant, Patrick S.; Fang, Ming; Lvov, Yuri; McShane, Michael J.

2002-05-01

Self-assembled thin films containing embedded enzymes and fluorescent indicators are being developed for use as highly specific glucose biosensors. The sensors are fabricated using electrostatic Layer-by-Layer (LBL) adsorption to create oxygen-sensitive (Ruthenium-based) layers, the fluorescent intensity of which responds to changes in local oxygen levels. Oxygen is consumed locally by the reaction between glucose oxidase (GOx) molecules and glucose. Latex particles serve as the templates for our sensors and fabrication is carried out through the alternate adsorption of multiple levels of {GOx/polycation} and {Ruthenium-polycation/polyanion} bilayers. Additional fluorescence layers as well as fluorescent latex are being considered as internal intensity references to allow ratiometric monitoring. Films adsorbed to the nanoparticle templates are being studied to understand the fundamental chemical and optical properties, including enzymatic activity, spectral shape and emission intensity. Enzymatic activity is retained and stability is improved after adsorption, and increased surface area afforded by the particles allows use of increased numbers of molecules. Fluorescence is also maintained, though blue shifts are observed in emission spectra, and indicator activity remains. In vitro characterization studies demonstrate the feasibility of the particles as glucose biosensors, and future work will aim to optimize the response for neural monitoring.

Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes.

PubMed

Chow, Yu Ting; Chen, Shuxun; Wang, Ran; Liu, Chichi; Kong, Chi-Wing; Li, Ronald A; Cheng, Shuk Han; Sun, Dong

2016-04-12

Cell transfection is a technique wherein foreign genetic molecules are delivered into cells. To elucidate distinct responses during cell genetic modification, methods to achieve transfection at the single-cell level are of great value. Herein, we developed an automated micropipette-based quantitative microinjection technology that can deliver precise amounts of materials into cells. The developed microinjection system achieved precise single-cell microinjection by pre-patterning cells in an array and controlling the amount of substance delivered based on injection pressure and time. The precision of the proposed injection technique was examined by comparing the fluorescence intensities of fluorescent dye droplets with a standard concentration and water droplets with a known injection amount of the dye in oil. Injection of synthetic modified mRNA (modRNA) encoding green fluorescence proteins or a cocktail of plasmids encoding green and red fluorescence proteins into human foreskin fibroblast cells demonstrated that the resulting green fluorescence intensity or green/red fluorescence intensity ratio were well correlated with the amount of genetic material injected into the cells. Single-cell transfection via the developed microinjection technique will be of particular use in cases where cell transfection is challenging and genetically modified of selected cells are desired.

The Effect of Temperature on Photoluminescence Enhancement of Quantum Dots in Brain Slices.

PubMed

Zhao, Fei; Kim, Jongsung

2017-04-01

In this paper, we investigated the effect of temperature on photoluminescence of quantum dots immobilized on the surface of an optical fiber in a rat brain slice. The optical fiber was silanized with 3-aminopropyl trimethoxysilane (APTMS), following which quantum dots with carboxyl functional group were immobilized on the optical fiber via amide bond formation. The effect of temperature on the fluorescence intensity of the quantum dots in rat brain slices was studied. This report shows that the fluorescence intensity of quantum dots increases with the increase of temperature of the brain slice. The fluorescence enhancement phenomenon appears to take place via electron transfer related to pH increase. With the gradual increase of temperature, the fluorescence intensity of quantum dots in solution decreased, while that in the brain slice increased. This enhanced thermal performance of QDs in brain slice makes suggestion for the study of QDs-based brain temperature sensors.

Accurate thermometry based on the red and green fluorescence intensity ratio in NaYF₄: Yb, Er nanocrystals for bioapplication.

PubMed

Liu, Lixin; Qin, Feng; Lv, Tianquan; Zhang, Zhiguo; Cao, Wenwu

2016-10-15

A biological temperature measurement method based on the fluorescence intensity ratio (FIR) was developed to reduce uncertainty. The upconversion luminescence of NaYF₄:Yb, Er nanocrystals was studied as a function of temperature around the physiologically relevant range of 300-330 K. We found that the green-green FIR Fe and red-green FIR (I₆₆₀/I₅₄₀) varied linearly as temperature increased. The thermometric uncertainties using the two FIRs were discussed and were determined to be almost constant at 0.6 and 0.09 K for green-green and red-green, respectively. The lower thermometric uncertainty comes from the intense signal-to-noise ratio of the measured FIRs owing to their comparable fluorescence intensities.

Changes in River Organic Matter Through Time.

NASA Astrophysics Data System (ADS)

Hudson, N.; Baker, A.; Ward, D.

2006-12-01

Samples of river water from central England were collected during the summer base-flow period. They were analysed for BOD and filtered at 1.2μm and 0.1μm increments to obtain i) the colloidal and dissolved, and ii) dissolved filter sterilized fractions. Each filtered fraction was plated up for microbiological cell counts and the agar plates and water samples were stored under a range of environmental conditions (4° C dark, 11° C light/ dark, 11° C dark, and 20° C dark) for 26 days. Absorbance, fluorescence, pH, conductivity and total organic carbon (TOC) were measured and colony forming units (CFU) counted on days 1, 2, 3, 4, 5, 12, 19 and 26. The fluorescence intensity was recorded for 5 commonly studied regions: protein like fluorescence, indicative of microbial activity, represented by the fluorescent amino acids tyrosine and tryptophan (which has two clear fluorescence regions) and humic and fulvic acids derived from the break down of terrestrial and aquatic plant material. Humic and fulvic-like fluorescence increased in all samples under all storage conditions suggesting that peaks A and C probably include a microbial element, either a product of the living community or as dead cell material in all fraction sizes including <0.1μm. Tryptophan and tyrosine-like fluorescence intensities demonstrated less clear trends which may be reflective of the intrinsic variation in natural samples. Tryptophan-like fluorescence generally decreased or showed minimal change, except in samples exposed to light in which an increase was observed in line with algal growth. A decrease in intensity may relate to the use of the tryptophan-like material as a microbial substrate. The increase in tryptophan-like fluorescence intensity suggests that this fluorescent material is being produced, either by algae, or bacterial activity associated with algal growth. It may also occur as a result of changing water chemistry causing a change in molecular conformation, and resulting fluorescence, as an increase in pH was also observed in these samples. This work illustrates the dynamic character of river organic matter within a timescale and under conditions that are representative of the natural system.

Relationship between the Fluorescence Lifetime of Chlorophyll 'a' and Primary Productivity within the Mississippi River Plume and Adjacent Shelf Region

NASA Technical Reports Server (NTRS)

Hall, Callie; Miller, Richard L.; Fernandez, Salvador M.; McKee, Brent A.

2000-01-01

In situ measurements of chlorophyll fluorescence intensity have been widely used to estimate phytoplankton biomass. However, because the fluorescence quantum yield of chlorophyll a in vivo can be highly variable, measurements of chlorophyll fluorescence intensity cannot be directly correlated with phytoplankton biomass and do not provide information on the physiological state of the phytoplankton under study. Conversely, lifetime-based measurements of chlorophyll fluorescence provide a framework in which photosynthetic rates of phytoplankton can be analyzed according to phytoplankton physiology. Along with the measurement of primary production and ambient nutrient concentrations within the Mississippi River plume in the northern Gulf of Mexico, phytoplankton fluorescence lifetimes were measured using a Fluorescence Lifetime Phytoplankton Analyzer (developed under a NASA Small Business Innovative Research contract to Ciencia, Inc.). Variability of fluorescence lifetimes within the plume can be used as a background from which to interpret variations in the maximum quantum yield of photochemistry. The extent to which nutrient and effluent loading in this dynamic coastal area affect the photosynthetic performance of phytoplankton will be presented as a function of phytoplankton fluorescence lifetimes.

Simple and effective label-free capillary electrophoretic analysis of sugars by complexation using quinoline boronic acids.

PubMed

Kubo, Takuya; Kanemori, Koichi; Kusumoto, Risa; Kawai, Takayuki; Sueyoshi, Kenji; Naito, Toyohiro; Otsuka, Koji

2015-01-01

An effective separation and detection procedure for sugars by capillary electrophoresis (CE) using a complexation between quinolineboronic acid (QBA) and multiple hydroxyl structure of sugar alcohol is reported. We investigated the variation of fluorescence spectra of a variety of QBAs with sorbitol at a wide range of pH conditions and then found that 5-isoQBA strongly enhanced the fluorescence intensity by the complexation at basic pH conditions. The other sugar alcohols having multiple hydroxyls also revealed the enhancement of the fluorescence intensity with 5-isoQBA, whereas the alternation of the intensity was not found in the sugars such as glucose. After optimization of the 5-isoQBA concentration and pH of the buffered solution in CE analysis, 6 sugar alcohols were successfully separated in the order based on the formation constants with 5-isoQBA, which were calculated from the variation of the fluorescence intensity with each sugar alcohol and 5-isoQBA. Furthermore, the limits of detection for sorbitol and xylitol by the CE method were estimated at 15 and 27 μM, respectively.

Medium effects on fluorescence of ciprofloxacin hydrochloride

NASA Astrophysics Data System (ADS)

Yang, Rui; Fu, Yan; Li, Long-Di; Liu, Jia-Ming

2003-10-01

The medium (pH, organic solvents, cyclodextrin (CD) or surfactants) effects on the fluorescence of ciprofloxacin hydrochloride (CPFX·HCl) were studied in detail. It is found that the three acid constants of ciprofloxacin (CPFX) are near to each other. Therefore the relation curve between pH and fluorescence intensity has no strident change and keeps relative stable in the pH range of 2-7. When pH was in the range of 5.5-6.0, the fluorescence intensity of CPFX reached the max. The kind and amount of organic solvent added to the luminescent system have various effects. Ethanol quenched fluorescence and the fluorescence excitation wavelength is red shift at first and then blue shift. Acetone has complicated effects on the fluorescence properties of CPFX·HCl solution. The experiment result shows that acetone is really a quencher when its volume content in the system is from 0 to 20%, but when its content is 90%, the signal intensity is unexpectedly one and a half times as much as that of no acetone. This means that there is a strong interaction between the acetone and CPFX; CPFX·H + could be included into the γ-CD but the capping effect is not notable. The effect of cationic surfactant cetyltrimethylammonium bromide and non-ionic surfactant TX-100 and TX-80 on CPFX fluorescence was unimpressive, but the anionic surfactant's effect is aberrant. The fluorescence intensity of CPFX·HCl solution experiences three stages of increasing, decreasing and increasing in turn, as sodium dodecyl sulfate is adding gradually. But for sodium lauryl sulfonate, there are only two stages of decreasing and increasing with the concentration increasing. It is problematic to illustrate clearly the effect mechanism of acetone and anionic surfactant at present. Undoubtedly, the experimental results in this paper should be useful in practice works and the research is worth studying still further.

Fluorescent Applications to Crystallization

NASA Technical Reports Server (NTRS)

Pusey, Marc L.; Forsythe, Elizabeth; Achari, Aniruddha

2006-01-01

By covalently modifying a subpopulation, less than or equal to 1%, of a macromolecule with a fluorescent probe, the labeled material will add to a growing crystal as a microheterogeneous growth unit. Labeling procedures can be readily incorporated into the final stages of purification, and tests with model proteins have shown that labeling u to 5 percent of the protein molecules does not affect the X-ray data quality obtained . The presence of the trace fluorescent label gives a number of advantages. Since the label is covalently attached to the protein molecules, it "tracks" the protein s response to the crystallization conditions. The covalently attached probe will concentrate in the crystal relative to the solution, and under fluorescent illumination crystals show up as bright objects against a darker background. Non-protein structures, such as salt crystals, do not show up under fluorescent illumination. Crystals have the highest protein concentration and are readily observed against less bright precipitated phases, which under white light illumination may obscure the crystals. Automated image analysis to find crystals should be greatly facilitated, without having to first define crystallization drop boundaries as the protein or protein structures is all that shows up. Fluorescence intensity is a faster search parameter, whether visually or by automated methods, than looking for crystalline features. Preliminary tests, using model proteins, indicates that we can use high fluorescence intensity regions, in the absence of clear crystalline features or "hits", as a means for determining potential lead conditions. A working hypothesis is that more rapid amorphous precipitation kinetics may overwhelm and trap more slowly formed ordered assemblies, which subsequently show up as regions of brighter fluorescence intensity. Experiments are now being carried out to test this approach using a wider range, of proteins. The trace fluorescently labeled crystals will also emit with sufficient intensity to aid in the automation of crystal alignment using relatively low cost optics, further increasing throughput at synchrotrons.

Nanowire-Intensified Metal-Enhanced Fluorescence in Hybrid Polymer-Plasmonic Electrospun Filaments.

PubMed

Camposeo, Andrea; Jurga, Radoslaw; Moffa, Maria; Portone, Alberto; Cardarelli, Francesco; Della Sala, Fabio; Ciracì, Cristian; Pisignano, Dario

2018-05-01

Hybrid polymer-plasmonic nanostructures might combine high enhancement of localized fields from metal nanoparticles with light confinement and long-range transport in subwavelength dielectric structures. Here, the complex behavior of fluorophores coupling to Au nanoparticles within polymer nanowires, which features localized metal-enhanced fluorescence (MEF) with unique characteristics compared to conventional structures, is reported. The intensification effect when the particle is placed in the organic filaments is remarkably higher with respect to thin films of comparable thickness, thus highlighting a specific, nanowire-related enhancement of MEF effects. A dependence on the confinement volume in the dielectric nanowire is also indicated, with MEF significantly increasing upon reduction of the wire diameter. These findings are rationalized by finite element simulations, predicting a position-dependent enhancement of the quantum yield of fluorophores embedded in the fibers. Calculation of the ensemble-averaged fluorescence enhancement unveils the possibility of strongly enhancing the overall emission intensity for structures with size twice the diameter of the embedded metal particles. These new, hybrid fluorescent systems with localized enhanced emission, and the general nanowire-enhanced MEF effects associated to them, are highly relevant for developing nanoscale light-emitting devices with high efficiency and intercoupled through nanofiber networks, highly sensitive optical sensors, and novel laser architectures. © 2018 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA Weinheim.

Delay, change and bifurcation of the immunofluorescence distribution attractors in health statuses diagnostics and in medical treatment

NASA Astrophysics Data System (ADS)

Galich, Nikolay E.; Filatov, Michael V.

2008-07-01

Communication contains the description of the immunology experiments and the experimental data treatment. New nonlinear methods of immunofluorescence statistical analysis of peripheral blood neutrophils have been developed. We used technology of respiratory burst reaction of DNA fluorescence in the neutrophils cells nuclei due to oxidative activity. The histograms of photon count statistics the radiant neutrophils populations' in flow cytometry experiments are considered. Distributions of the fluorescence flashes frequency as functions of the fluorescence intensity are analyzed. Statistic peculiarities of histograms set for healthy and unhealthy donors allow dividing all histograms on the three classes. The classification is based on three different types of smoothing and long-range scale averaged immunofluorescence distributions and their bifurcation. Heterogeneity peculiarities of long-range scale immunofluorescence distributions allow dividing all histograms on three groups. First histograms group belongs to healthy donors. Two other groups belong to donors with autoimmune and inflammatory diseases. Some of the illnesses are not diagnosed by standards biochemical methods. Medical standards and statistical data of the immunofluorescence histograms for identifications of health and illnesses are interconnected. Possibilities and alterations of immunofluorescence statistics in registration, diagnostics and monitoring of different diseases in various medical treatments have been demonstrated. Health or illness criteria are connected with statistics features of immunofluorescence histograms. Neutrophils populations' fluorescence presents the sensitive clear indicator of health status.

Speciation and Sources of Brown Carbon in Precipitation at Seoul, Korea: Insights from Excitation-Emission Matrix Spectroscopy and Carbon Isotopic Analysis.

PubMed

Yan, Ge; Kim, Guebuem

2017-10-17

Brown carbon (BrC) plays a significant role in the Earth's radiative balance, yet its sources and chemical composition remain poorly understood. In this work, we investigated BrC in the atmospheric environment of Seoul by characterizing dissolved organic matter in precipitation using excitation-emission matrix (EEM) fluorescence spectroscopy coupled with parallel factor analysis (PARAFAC). The two independent fluorescent components identified by PARAFAC were attributed to humic-like substance (HULIS) and biologically derived material based on their significant correlations with measured HULIS isolated using solid-phase extraction and total hydrolyzable tyrosine. The year-long observation shows that HULIS contributes to 66 ± 13% of total fluorescence intensity of our samples on average. By using dual carbon ( 13 C and 14 C) isotopic analysis conducted on isolated HULIS, the HULIS fraction of BrC was found to be primarily derived from biomass burning and emission of terrestrial biogenic gases and particles (>70%), with minor contributions from fossil-fuel combustion. The knowledge derived from this study could contribute to the establishment of a characterizing system of BrC components identified by EEM spectroscopy. Our work demonstrates that, EEM fluorescence spectroscopy is a powerful tool in BrC study, on the basis of its chromophore resolving power, allowing investigation into individual components of BrC by other organic matter characterization techniques.

Synthesis and spectral properties of polymethine-cyanine dye-nitroxide radical hybrid compounds for use as fluorescence probes to monitor reducing species and radicals

NASA Astrophysics Data System (ADS)

Sato, Shingo; Tsunoda, Minoru; Suzuki, Minoru; Kutsuna, Masahiro; Takido-uchi, Kiyomi; Shindo, Mitsuru; Mizuguchi, Hitoshi; Obara, Heitaro; Ohya, Hiroaki

2009-01-01

Various hybrid compounds comprised of two types of nitroxide radicals and either a pentamethine (Cy5) or trimethine cyanine (Cy3) were synthesized. The nitroxide radicals were linked either via an ester-bond to one or two N-alkyl carboxyl-terminated groups of Cy5, or via two amido-bonds (aminocarbonyl or carbonylamino group) to the 5-position of the indolenine moieties of Cy5 and Cy3. Changes in fluorescence and ESR intensities of the hybrid compounds were measured before and after addition of Na ascorbate in PBS (pH 7.0) to reduce the radicals. Among the hybrid compounds synthesized, those that linked the nitroxide radicals via an aminocarbonyl residue at the 5-position of the indolenine moieties on Cy5 and Cy3 exhibited a 1.8- and 5.1-fold increase in fluorescence intensity with the reduction of the nitroxide segment by the addition of Na ascorbate, respectively. In contrast, fluorescence intensity was not enhanced in the other hybrid compounds. Thus, the hybrid compounds which exhibited an increase in fluorescent intensity with radical reduction can be used in the quantitative measurement of reducing species such as Fe 2+ and ascorbic acid, and hydroxyl radicals. Because these hybrid compounds have the advantage of fluorescing at longer wavelengths—661 (Cy5) or 568 (Cy3) nm, respectively, they can be used to measure radical-reducing species or radicals either in solution or in vivo.

Simultaneous three-dimensional velocity and mixing measurements by use of laser Doppler velocimetry and fluorescence probes in a water tunnel

NASA Technical Reports Server (NTRS)

Neuhart, Dan H.; Wing, David J.; Henderson, Uleses C., Jr.

1994-01-01

A water tunnel investigation was conducted to demonstrate the capabilities of a laser-based instrument that can measure velocity and fluorescence intensity simultaneously. Fluorescence intensity of an excited fluorescent dye is directly related to concentration level and is used to indicate the extent of mixing in flow. This instrument is a three-dimensional laser Doppler velocimeter (LDV) in combination with a fluorometer for measuring fluorescence intensity variations. This capability allows simultaneous flow measurements of the three orthogonal velocity components and mixing within the same region. Two different flows which were generated by two models were studied: a generic nonaxisymmetric nozzle propulsion simulation model with an auxiliary internal water source that generated a jet flow and an axisymmetric forebody model with a circular sector strake that generated a vortex flow. The off-body flow fields around these models were investigated in the Langley 16- by 24-Inch Water Tunnel. The experimental results were used to calculate 17 quantities that included mean and fluctuating velocities, Reynolds stresses, mean and fluctuating dye fluorescence intensities (proportional to concentration), and fluctuating velocity and dye concentration correlations. An uncertainty analysis was performed to establish confidence levels in the experimental results. In general, uncertainties in mean velocities varied between 1 and 7 percent of free-stream velocity; uncertainties in fluctuating velocities varied between 1 and 5 percent of reference values. The results show characteristics that are unique to each type of flow.

Aspect ratio dependence of the enhancement of fluorescence intensity by gold nanobipyramids for cancer cell imaging and photodynamic therapy

NASA Astrophysics Data System (ADS)

Wang, Jing; Wang, Shimiao; Mi, Lan; Liu, Jun

2018-07-01

Enhancement of dye fluorescence intensity was studied by modifying the aspect ratio of gold nanobipyramids (AuBPs) from 3.2 to 6.6. The emission fluorescence intensity of sulfonated aluminum phthalocyanine (AlPcS) was strongly dependent on the aspect ratio of AuBPs. Furthermore, we found that the energy transfer from excited AlPcS to AuBPs was a key determinant of the efficacy of metal-enhanced fluorescence. By means of AuBPs with a higher aspect ratio, such that the surface plasmon resonance band does not overlap with the energy level of excited AlPcS, metal-enhanced fluorescence of various AlPcS–AuBP conjugates was determined, and the maximal enhancement factor was found to be 14. The enhanced fluorescence intensity of AlPcS conjugated with AuBPs indicates promising plasmonic properties. An apoptosis assay of HeLa cells revealed that AlPcS–AuBPs, when used as a drug, can enhance the effectiveness of photodynamic therapy (PDT). Furthermore, AuBPs with the longitudinal absorption peak wavelength of 1050 nm had optimal proapoptotic effects. HeLa cells treated with AlPcS–AuBPs (ratio 0.42 µM to 0.01 nM) had viability as low as 29.31% after 32 J cm‑2 ultraviolet light exposure, indicating the strong potential of AlPcS–AuBPs to improve the efficacy of PDT.

Luminescent carbon quantum dots with high quantum yield as a single white converter for white light emitting diodes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Feng, X. T.; Zhang, Y.; Liu, X. G., E-mail: liuxuguang@tyut.edu.cn

Carbon quantum dots (CQDs) with high quantum yield (51.4%) were synthesized by a one-step hydrothermal method using thiosalicylic acid and ethylenediamine as precursor. The CQDs have the average diameter of 2.3 nm and possess excitation-independent emission wavelength in the range from 320 to 440 nm excitation. Under an ultraviolet (UV) excitation, the CQDs aqueous solutions emit bright blue fluorescence directly and exhibit broad emission with a high spectral component ratio of 67.4% (blue to red intensity to total intensity). We applied the CQDs as a single white-light converter for white light emitting diodes (WLEDs) using a UV-LED chip as the excitation lightmore » source. The resulted WLED shows superior performance with corresponding color temperature of 5227 K and the color coordinates of (0.34, 0.38) belonging to the white gamut.« less

Development of a fluorescent chelating ligand for scandium ion having a Schiff base moiety

NASA Astrophysics Data System (ADS)

Yamada, Hiroshi; Kojo, Masahito; Nakahara, Tomomi; Murakami, Kumi; Kakima, Takashi; Ichiba, Hideaki; Yajima, Takehiko; Fukushima, Takeshi

2012-05-01

A fluorescent ligand, 1-(2-hydroxy-3-methoxybenzaldehyde)-4-aminosalicylhydrazone (HMB-ASH), was newly designed and synthesized, and its fluorescence characteristics for metal ions were investigated in the pH range 3.0-10.5 (at a difference of 0.5 for each metal). After testing 31 different metal ions, it was found that HMB-ASH was able to emit fluorescence intensely at 512 nm with an excitation wavelength of 353 nm in the presence of Sc3+, one of the rare earth metals, at pH values around 3.5 and 8.0. The other metal ions hardly showed fluorescence with HMB-ASH. The fluorescence was more intense at pH 8.0, and the detection limit of Sc3+ in a buffer solution (pH 8.0) was approximately 18.8 nmol L-1 (0.85 ppb).

Chemical reactivation of resin-embedded pHuji adds red for simultaneous two-color imaging with EGFP

PubMed Central

Guo, Wenyan; Liu, Xiuli; Liu, Yurong; Gang, Yadong; He, Xiaobin; Jia, Yao; Yin, Fangfang; Li, Pei; Huang, Fei; Zhou, Hongfu; Wang, Xiaojun; Gong, Hui; Luo, Qingming; Xu, Fuqiang; Zeng, Shaoqun

2017-01-01

The pH-sensitive fluorescent proteins enabling chemical reactivation in resin are useful tools for fluorescence microimaging. EGFP or EYFP is good for such applications. For simultaneous two-color imaging, a suitable red fluorescent protein is an urgent need. Here a pH-sensitive red fluorescent protein, pHuji, is selected and verified to remain pH-sensitive in HM20 resin. We observe 183% fluorescence intensity of pHuji in resin-embeded mouse brain and 29.08-fold fluorescence intensity of reactivated pHuji compared to the quenched state. pHuji and EGFP can be quenched and chemically reactivated simultaneously in resin, thus enabling simultaneous two-color micro-optical sectioning tomography of resin-embedded mouse brain. This method may greatly facilitate the visualization of neuronal morphology and neural circuits to promote understanding of the structure and function of the brain. PMID:28717566

Optical properties of flexible fluorescent films prepared by screen printing technology

NASA Astrophysics Data System (ADS)

Chen, Yan; Ke, Taiyan; Chen, Shuijin; He, Xin; Zhang, Mei; Li, Dong; Deng, Jinfeng; Zeng, Qingguang

2018-05-01

In this work, we prepared a fluorescent film comprised phosphors and silicone on flexible polyethylene terephthalate (PET) substrate using a screen printing technology. The effects of mesh number and weight ratio of phosphors to silicone on the optical properties of the flexible films were investigated. The results indicate that the emission intensity of the film increase as the mesh decreased from 400 to 200, but the film surface gradually becomes uneven. The fluorescent film with high emission intensity and smooth surface can be obtained when the weight ratio of phosphor to gel is 2:1, and mesh number is 300. The luminous efficiency of the fabricated LEDs combined the fluorescent films with 460 nm Ga(In)N chip module can reach 75 lm/W. The investigation indicates that the approach can be applied in the remote fluorescent film conversion and decreases the requirements of the particle size and the dispersion state of fluorescent materials.

Chemical reactivation of resin-embedded pHuji adds red for simultaneous two-color imaging with EGFP.

PubMed

Guo, Wenyan; Liu, Xiuli; Liu, Yurong; Gang, Yadong; He, Xiaobin; Jia, Yao; Yin, Fangfang; Li, Pei; Huang, Fei; Zhou, Hongfu; Wang, Xiaojun; Gong, Hui; Luo, Qingming; Xu, Fuqiang; Zeng, Shaoqun

2017-07-01

The pH-sensitive fluorescent proteins enabling chemical reactivation in resin are useful tools for fluorescence microimaging. EGFP or EYFP is good for such applications. For simultaneous two-color imaging, a suitable red fluorescent protein is an urgent need. Here a pH-sensitive red fluorescent protein, pHuji, is selected and verified to remain pH-sensitive in HM20 resin. We observe 183% fluorescence intensity of pHuji in resin-embeded mouse brain and 29.08-fold fluorescence intensity of reactivated pHuji compared to the quenched state. pHuji and EGFP can be quenched and chemically reactivated simultaneously in resin, thus enabling simultaneous two-color micro-optical sectioning tomography of resin-embedded mouse brain. This method may greatly facilitate the visualization of neuronal morphology and neural circuits to promote understanding of the structure and function of the brain.

Image recovery from defocused 2D fluorescent images in multimodal digital holographic microscopy.

PubMed

Quan, Xiangyu; Matoba, Osamu; Awatsuji, Yasuhiro

2017-05-01

A technique of three-dimensional (3D) intensity retrieval from defocused, two-dimensional (2D) fluorescent images in the multimodal digital holographic microscopy (DHM) is proposed. In the multimodal DHM, 3D phase and 2D fluorescence distributions are obtained simultaneously by an integrated system of an off-axis DHM and a conventional epifluorescence microscopy, respectively. This gives us more information of the target; however, defocused fluorescent images are observed due to the short depth of field. In this Letter, we propose a method to recover the defocused images based on the phase compensation and backpropagation from the defocused plane to the focused plane using the distance information that is obtained from a 3D phase distribution. By applying Zernike polynomial phase correction, we brought back the fluorescence intensity to the focused imaging planes. The experimental demonstration using fluorescent beads is presented, and the expected applications are suggested.

A fiber optic biosensor for fluorimetric detection of triple-helical DNA.

PubMed

Uddin, A H; Piunno, P A; Hudson, R H; Damha, M J; Krull, U J

1997-10-15

A fiber optic biosensor was used for the fluorimetric detection of T/AT triple-helical DNA formation. The surfaces of two sets of fused silica optical fibers were functionalized with hexaethylene oxide linkers from which decaadenylic acid oligonucleotides were grown in the 3'to 5'and 5'to 3'direction, respectively, using a DNA synthesizer. Fluorescence studies of hybridization showed unequivocal hybridization between oligomers immobilized on the fibers and complementary oligonucleotides from the solution phase, as detected by fluorescence from intercalated ethidium bromide. The complementary oligonucleotide, dT10, which was expected to Watson-Crick hybridize upon cooling the system below the duplex melting temperature ( T m), provided a fluorescence intensity with a negative temperature coefficient. Upon further cooling, to the point where the pyrimidine motif T*AT triple-helix formation occurred, a fluorescence intensity change with a positive temperature coefficient was observed. The reverse-Hoogsteen T.AT triplex, which is known to form with branched nucleic acids, provided a corresponding decrease in fluorescence intensity with decreasing temperature. Full analytical signal evolution was attainable in minutes.

Biological 2-Input Decoder Circuit in Human Cells

PubMed Central

2015-01-01

Decoders are combinational circuits that convert information from n inputs to a maximum of 2n outputs. This operation is of major importance in computing systems yet it is vastly underexplored in synthetic biology. Here, we present a synthetic gene network architecture that operates as a biological decoder in human cells, converting 2 inputs to 4 outputs. As a proof-of-principle, we use small molecules to emulate the two inputs and fluorescent reporters as the corresponding four outputs. The experiments are performed using transient transfections in human kidney embryonic cells and the characterization by fluorescence microscopy and flow cytometry. We show a clear separation between the ON and OFF mean fluorescent intensity states. Additionally, we adopt the integrated mean fluorescence intensity for the characterization of the circuit and show that this metric is more robust to transfection conditions when compared to the mean fluorescent intensity. To conclude, we present the first implementation of a genetic decoder. This combinational system can be valuable toward engineering higher-order circuits as well as accommodate a multiplexed interface with endogenous cellular functions. PMID:24694115

Biological 2-input decoder circuit in human cells.

PubMed

Guinn, Michael; Bleris, Leonidas

2014-08-15

Decoders are combinational circuits that convert information from n inputs to a maximum of 2(n) outputs. This operation is of major importance in computing systems yet it is vastly underexplored in synthetic biology. Here, we present a synthetic gene network architecture that operates as a biological decoder in human cells, converting 2 inputs to 4 outputs. As a proof-of-principle, we use small molecules to emulate the two inputs and fluorescent reporters as the corresponding four outputs. The experiments are performed using transient transfections in human kidney embryonic cells and the characterization by fluorescence microscopy and flow cytometry. We show a clear separation between the ON and OFF mean fluorescent intensity states. Additionally, we adopt the integrated mean fluorescence intensity for the characterization of the circuit and show that this metric is more robust to transfection conditions when compared to the mean fluorescent intensity. To conclude, we present the first implementation of a genetic decoder. This combinational system can be valuable toward engineering higher-order circuits as well as accommodate a multiplexed interface with endogenous cellular functions.

The fluorescent photobleaching properties of GFP expressed in human lung cancer cells

NASA Astrophysics Data System (ADS)

Jin, Ying; Xing, Da

2003-12-01

The characteristic properties of GFP make this protein a good candidate for use as a molecular reporter to monitor patterns of protein localization, gene expression, and intracellular protein trafficking in living cells. In this study, the dicistronic expression vector (pEGFP-C1) was used to transfected into human lung cancer cell line (ASTC-a-1) and a positive clone which stably expressed GFP in high level was obtained. After more than three months' passengers, the cells were also remained the strong fluorescence under fluorescent microscope. The results showed that the green fluorescent protein expressed in tumor cells was also photobleached under intense irradiation (approximately 488 nm) and the degree of photobleaching varied with the difference of the intensity of the excitation. Using different interdiction parcel (None, ND4, ND8, ND16), there were significant differences in photobleaching among the different excitation. The photobleaching was also affected by the time length of excitation, and the intensity of fluorescence was obviously decreased along with the increasing of excitation time, especially to stronger excitation.

Rapid and prodium iodide-compatible optical clearing method for brain tissue based on sugar/sugar-alcohol

NASA Astrophysics Data System (ADS)

Yu, Tingting; Qi, Yisong; Wang, Jianru; Feng, Wei; Xu, Jianyi; Zhu, Jingtan; Yao, Yingtao; Gong, Hui; Luo, Qingming; Zhu, Dan

2016-08-01

The developed optical clearing methods show great potential for imaging of large-volume tissues, but these methods present some nonnegligible limitations such as complexity of implementation and long incubation times. In this study, we tried to screen out rapid optical clearing agents by means of molecular dynamical simulation and experimental demonstration. According to the optical clearing potential of sugar and sugar-alcohol, we further evaluated the improvement in the optical clearing efficacy of mouse brain samples, imaging depth, fluorescence preservation, and linear deformation. The results showed that drops of sorbitol, sucrose, and fructose could quickly make the mouse brain sample transparent within 1 to 2 min, and induce about threefold enhancement in imaging depth. The former two could evidently enhance the fluorescence intensity of green fluorescent protein (GFP) and prodium iodide (PI) nuclear dye. Fructose could significantly increase the fluorescence intensity of PI, but slightly decrease the fluorescence intensity of GFP. Even though the three agents caused some shrinkage in samples, the contraction in horizontal and longitudinal directions are almost the same.

A Label-Free Fluorescent Array Sensor Utilizing Liposome Encapsulating Calcein for Discriminating Target Proteins by Principal Component Analysis

PubMed Central

Imamura, Ryota; Murata, Naoki; Shimanouchi, Toshinori; Yamashita, Kaoru; Fukuzawa, Masayuki; Noda, Minoru

2017-01-01

A new fluorescent arrayed biosensor has been developed to discriminate species and concentrations of target proteins by using plural different phospholipid liposome species encapsulating fluorescent molecules, utilizing differences in permeation of the fluorescent molecules through the membrane to modulate liposome-target protein interactions. This approach proposes a basically new label-free fluorescent sensor, compared with the common technique of developed fluorescent array sensors with labeling. We have confirmed a high output intensity of fluorescence emission related to characteristics of the fluorescent molecules dependent on their concentrations when they leak from inside the liposomes through the perturbed lipid membrane. After taking an array image of the fluorescence emission from the sensor using a CMOS imager, the output intensities of the fluorescence were analyzed by a principal component analysis (PCA) statistical method. It is found from PCA plots that different protein species with several concentrations were successfully discriminated by using the different lipid membranes with high cumulative contribution ratio. We also confirmed that the accuracy of the discrimination by the array sensor with a single shot is higher than that of a single sensor with multiple shots. PMID:28714873

A Label-Free Fluorescent Array Sensor Utilizing Liposome Encapsulating Calcein for Discriminating Target Proteins by Principal Component Analysis.

PubMed

Imamura, Ryota; Murata, Naoki; Shimanouchi, Toshinori; Yamashita, Kaoru; Fukuzawa, Masayuki; Noda, Minoru

2017-07-15

A new fluorescent arrayed biosensor has been developed to discriminate species and concentrations of target proteins by using plural different phospholipid liposome species encapsulating fluorescent molecules, utilizing differences in permeation of the fluorescent molecules through the membrane to modulate liposome-target protein interactions. This approach proposes a basically new label-free fluorescent sensor, compared with the common technique of developed fluorescent array sensors with labeling. We have confirmed a high output intensity of fluorescence emission related to characteristics of the fluorescent molecules dependent on their concentrations when they leak from inside the liposomes through the perturbed lipid membrane. After taking an array image of the fluorescence emission from the sensor using a CMOS imager, the output intensities of the fluorescence were analyzed by a principal component analysis (PCA) statistical method. It is found from PCA plots that different protein species with several concentrations were successfully discriminated by using the different lipid membranes with high cumulative contribution ratio. We also confirmed that the accuracy of the discrimination by the array sensor with a single shot is higher than that of a single sensor with multiple shots.

Effect of silver nanoparticles on the 1.53 μm fluorescence in Er3+/Yb3+ codoped tellurite glasses

NASA Astrophysics Data System (ADS)

Wu, Libo; Zhou, Yaxun; Zhou, Zizhong; Cheng, Pan; Huang, Bo; Yang, Fengjing; Li, Jun

2016-07-01

Improving the spectroscopic properties of rare earth (RE) doped glass materials is a challenging task. In the present work the metallic silver nanoparticles (Ag NPs) were embedded into Er3+/Yb3+ codoped tellurite glasses with composition TeO2-Bi2O3-TiO2, prepared using melt-quenching and subsequent heat-treated techniques, and the improved effect of Ag NPs on the 1.53 μm band fluorescence of Er3+ ions was investigated. About 24 h heat-treatment of Er3+/Yb3+ codoped tellurite glass containing 1 mol % amount of AgNO3 at the temperature 370 °C yielded the well-dispersed and near-spherical Ag NPs with ∼11.4 nm average diameter as evidenced by transmission electron microscopy (TEM) image. The intense 1.53 μm band fluorescence was observed in the prepared Er3+/Yb3+ codoped tellurite glasses under the excitation of 980 nm and was further improved with the presence of Ag NPs in the glass matrix, which is attributed to the enhanced local electric field around doped RE ions induced by Ag NPs and the possible energy transfer from Ag NPs to Er3+ ions. The enhanced local electric field was well demonstrated by comparing the variation of emission spectra of hypersensitive probe Eu3+ ions in tellurite glasses with and without Ag NPs. From the Judd-Ofelt analysis, it was also found that the value of Ω6 intensity parameter increased slightly with the increase of Ag NPs concentration in a certain range, also confirming the possibility of realizing strong fluorescence emission. In addition, the amorphous structural nature was demonstrated by the measured X-ray diffraction (XRD) patterns with no sharp diffraction peak. The enhanced 1.53 μm band fluorescence indicates that the Er3+/Yb3+ codoped tellurite glass with an appropriate amount of Ag NPs is a promising candidate for the development of Er3+-doped fiber amplifiers (EDFAs) applied in the WDM systems.

Anomalous fluorescence enhancement and fluorescence quenching of graphene quantum dots by single walled carbon nanotubes.

PubMed

Das, Ruma; Rajender, Gone; Giri, P K

2018-02-07

We explore the mechanism of the fluorescence enhancement and fluorescence quenching effect of single walled carbon nanotubes (SWCNTs) on highly fluorescent graphene quantum dots (GQDs) over a wide range of concentrations of SWCNTs. At very low concentrations of SWCNTs, the fluorescence intensity of the GQDs is enhanced, while at higher concentrations, systematic quenching of fluorescence is observed. The nature of the Stern-Volmer plot for the latter case was found to be non-linear indicating a combined effect of dynamic and static quenching. The contribution of the dynamic quenching component was assessed through the fluorescence lifetime measurements. The contribution of static quenching is confirmed from the red shift of the fluorescence spectra of the GQDs after addition of SWCNTs. The fluorescence intensity is first enhanced at very low concentration due to improved dispersion and higher absorption by GQDs, while at higher concentration, the fluorescence of GQDs is quenched due to the complex formation and associated reduction of the radiative sites of the GQDs, which is confirmed from time-resolved fluorescence measurements. Laser confocal microscopy imaging provides direct evidence of the enhancement and quenching of fluorescence at low and high concentrations of SWCNTs, respectively. This study provides an important insight into tuning the fluorescence of GQDs and understanding the interaction between GQDs and different CNTs, which is important for bio-imaging and drug delivery applications.

Motor Oil Classification Based on Time-Resolved Fluorescence

PubMed Central

Mu, Taotao; Chen, Siying; Zhang, Yinchao; Guo, Pan; Chen, He; Meng, Fandong

2014-01-01

A time-resolved fluorescence (TRF) technique is presented for classifying motor oils. The system is constructed with a third harmonic Nd:YAG laser, a spectrometer, and an intensified charge coupled device (ICCD) camera. Steady-state and time-resolved fluorescence (TRF) measurements are reported for several motor oils. It is found that steady-state fluorescence is insufficient to distinguish the motor oil samples. Then contour diagrams of TRF intensities (CDTRFIs) are acquired to serve as unique fingerprints to identify motor oils by using the distinct TRF of motor oils. CDTRFIs are preferable to steady-state fluorescence spectra for classifying different motor oils, making CDTRFIs a particularly choice for the development of fluorescence-based methods for the discrimination and characterization of motor oils. The two-dimensional fluorescence contour diagrams contain more information, not only the changing shapes of the LIF spectra but also the relative intensity. The results indicate that motor oils can be differentiated based on the new proposed method, which provides reliable methods for analyzing and classifying motor oils. PMID:24988439

Red-emitting fluorescent probe for detecting hypochlorite acid in vitro and in vivo.

PubMed

Chen, Hong; Sun, Tao; Qiao, Xiao-Guang; Tang, Qian-Oian; Zhao, Shan-Chao; Zhou, Zhan

2018-06-12

Due to the importance of hypochlorous acid (HClO) in biological and industrial, development of fluorescent probes for HClO has been an active research area. Here, a new red-emitting ratiometric fluorescent probe (P) was synthesized and well defined characterization via NMR, HR-MS, and fluorescence spectrum, which serves as a selective and sensitive probe for ClO - group. The probe showed a ratiometric fluorescent response to hypochlorite at the emission intensities ratio (I 480 /I 612 ) increasing from 0.28 to 27.46. The emission intensities ratio (I 480 /I 612 ) was linearly enhanced (I 480 /I 612  = 0.064 X + 0.096) with the ClO - concentration range from 1 to 30 μM. The detection limitation for ClO - in aqueous solution is 0.47 μM. Moreover, this biocompatible red-emitting ratiometric fluorescent probe was utilized to the fluorescence imaging of ClO - in living cells and Zebrafish. Copyright © 2018. Published by Elsevier B.V.

[Resolving characteristic of CDOM by excitation-emission matrix spectroscopy combined with parallel factor analysis in the seawater of outer Yangtze Estuary in Autumn in 2010].

PubMed

Yan, Li-Hong; Chen, Xue-Jun; Su, Rong-Guo; Han, Xiu-Rong; Zhang, Chuan-Song; Shi, Xiao-Yong

2013-01-01

The distribution and estuarine behavior of fluorescent components of chromophoric dissolved organic matter in the seawater of outer Yangtze Estuary were determined by fluorescence excitation emission matrix spectra combined with parallel factor analysis. Six individual fluorescent components were identified by PARAFAC models, including three terrestrial humic-like components C1 [330 nm/390(430) nm], C2 (390 nm/480 nm), C3 (360 nm/440 nm), marine biological production component C5 (300 nm/400 nm) and protein-like components C4 (290 nm/350 nm) and C6 (275 nm/300 nm). The results indicated that C1, C2, and C3 showed a conservative mixing behavior in the whole estuarine region, especially in high-salinity region. And the fluorescence intensity proportion of C1 and C3 decreased with increase of salinity and fluorescence intensity proportion of C2 kept constant with increase of salinity in the whole estuarine region. While C4 showed conservative mixing behavior in low-salinity region and non-conservative mixing behavior in high-salinity region, and fluorescence intensity proportion of C4 increased with increase of salinity. However, C5 and C6 showed a non-conservative mixing behavior and fluorescence intensity proportion increased with increase of salinity in high-salinity region. Significantly spatial difference was recorded for CDOM absorption coefficient in the coastal region and in the open water areas with the highest value in coastal region and the lowest value in the open water areas. The scope of absorption coefficient and absorption slope was higher in coastal region than that in the open water areas. Significantly positive correlations were found between CDOM absorption coefficient and the fluorescence intensities of C1, C2, C3, and C4, but no significant correlation was found between C5 and C6, suggesting that the river inputs contributed to the coastal areas, while CDOM in the open water areas was affected by terrestrial inputs and phytoplankton degradation.

Red emissive cross-linked chitosan and their nanoparticles for imaging the nucleoli of living cells.

PubMed

Wang, Ke; Yuan, Xun; Guo, Zhenpeng; Xu, Jiying; Chen, Yi

2014-02-15

Biocompatible glutaraldehyde-cross-linked chitosan with new red fluorescence were prepared for the first time and were shaped into nanoparticles via inverse-microemulsion method. They could luminesce at ca. 670 nm either as powders and nanoparticles or in real and gelling solutions or suspensions, having a lifetime of 1.353 ns and a quantum yield of 0.08 in solution or 0.01 in solid state. The new-formed pyridinium structures and the intramolecular charge transfer effect are considered to be responsible for the new red emission, which have been proved by FTIR, (13)C NMR, and some calculation using Gaussian 09, respectively. Strikingly, they are quite inert and anti-photobleaching, with only <3% loss of fluorescent intensity per minute in average under a continuous laser illumination at 633 nm and 50 μW. Especially, their nanoparticles (5.6 nm) could enter into the negative nucleoli of living HeLa cells with low cytotoxicity for high contrast imaging inspections. Copyright © 2013 Elsevier Ltd. All rights reserved.

Spectrofluorometric determination of methocarbamol in pharmaceutical preparations and human plasma.

PubMed

Walash, Mohamed; Belal, Fathalla; Eid, Manal; EL Abass, Samah Abo

2011-03-01

A simple, sensitive and rapid spectrofluorometric method for determination of methocarbamol in pharmaceutical formulations and spiked human plasma has been developed. The proposed method is based on the measurement of the native fluorescence of methocarbamol in methanol at 313 nm after excitation at 277 nm. The relative fluorescence intensity-concentration plot was rectilinear over the range of 0.05-2.0 μg/mL, with good correlation (r=0.9999), limit of detection of 0.007 μg/ mL and a lower limit of quantification of 0.022 μg/ mL. The described method was successfully applied for the determination of methocarbamol in its tablets without interference from co-formulated drugs, such as aspirin, diclofenac, paracetamol and ibuprofen, The results obtained were in good agreement with those obtained using the official method (USP 30).The high sensitivity of the method allowed the determination of the studied drug in spiked human plasma with average percentage recovery of 99.42 ± 3.84. © Springer Science+Business Media, LLC 2010

Identifying Mechanisms of Interfacial Dynamics Using Single-Molecule Tracking

PubMed Central

Kastantin, Mark; Walder, Robert; Schwartz, Daniel K.

2012-01-01

The “soft” (i.e. non-covalent) interactions between molecules and surfaces are complex and highly-varied (e.g. hydrophobic, hydrogen bonding, ionic) often leading to heterogeneous interfacial behavior. Heterogeneity can arise either from spatial variation of the surface/interface itself or from molecular configurations (i.e. conformation, orientation, aggregation state, etc.). By observing adsorption, diffusion, and desorption of individual fluorescent molecules, single-molecule tracking can characterize these types of heterogeneous interfacial behavior in ways that are inaccessible to traditional ensemble-averaged methods. Moreover, the fluorescence intensity or emission wavelength (in resonance energy transfer experiments) can be used to simultaneously track molecular configuration and directly relate this to the resulting interfacial mobility or affinity. In this feature article, we review recent advances involving the use of single-molecule tracking to characterize heterogeneous molecule-surface interactions including: multiple modes of diffusion and desorption associated with both internal and external molecular configuration, Arrhenius activated interfacial transport, spatially dependent interactions, and many more. PMID:22716995

Synthesis and spectral properties of preorganized BODIPYs in solutions and Langmuir-Schaefer films

NASA Astrophysics Data System (ADS)

Marfin, Yuriy S.; Usoltsev, Sergey D.; Kazak, Alexandr V.; Smirnova, Antonina I.; Rumyantsev, Evgeniy V.; Molchanov, Evgeniy E.; Kuznetsov, Vladimir V.; Chumakov, Alexey S.; Glukhovskoy, Evgeny G.

2017-12-01

In order to investigate the influence of molecular structure peculiarities of boron-dipyrrine dyes (BODIPYs) on their properties in solutions and supramolecular organization in Langmuir-Schaefer (LS) films, four new BODIPY dyes with various aliphatic, aromatic or mixed nature meso-subtituents were synthesized and investigated. Spectral characteristics (electronic absorption and fluorescence) of the synthesized compounds in organic solvents and LS-films were studied. Floating monolayers of the BODIPYs were formed from chloroform solutions placed onto water subphase in Langmuir-Blodgett through. Thin films were prepared using the Langmuir-Schaefer technique by the transfer of floating monolayers onto standard polished glass, ITO covered glass or pure silicon substrate. The variation of the dye structure we consider as a preorganization aiming to influence the structure of LS-films. The morphology and structure of the LS-films was examined by fluorescent microscopy, scanning electron microscopy, atomic force microscopy and small angle X-ray diffraction analysis. It was found that the introduced substituents have no substantial influence on the position of the absorption and fluorescence bands in dilute solutions. In contrast, the fluorescent characteristics of the LS-films significantly depend on the substituent nature. Therefore, this strategy could be used for the direct tuning of compounds fluorescent properties in LS-films. Concerning the LS-film surface characteristics it was proved that the films are homogeneous, without disruptions and only some widely-spaced microcrystals could be observed. With respect to the LS-film structure, the change of the substituents introduced to the BODIPY molecule did not influence the average given periodicity of layers (d = 0.3-0.4 nm). This value corresponds to a single-layer arrangement of BODIPY molecules located parallel to the substrate surface. Nevertheless, the diffraction peak intensities depended on the molecular structure of BODIPYs and therefore the structurization in thin films. Moreover, the combination of the rigid phenyl moiety with long alkyl chains in one compound completely suppresses the aggregation of molecules maintaining the intense fluorescence in thin films. On the basis of used range of experimental and calculation methods the intralayer and interlayer structures were proposed. Intermolecular hydrogen bond formation and π-π staking of the BODIPY cores were found to be the structure forming forces during the films manufacturing, resulting the differences in crystallinity of the materials. While the alkyl-substituents prevent the type of interactions and suppress the association of the dyes and formation of excimers. Compounds under investigation show a manifestation of the intense solvatochromic properties which allow their application as sensors, including naked eye sensorics for solution polarity. Besides, the obtained results broaden prospective of functional materials usage based on BODIPY thin films as components of optoelectronics.

The Effect of Intense Laser Radiation on Atomic Collisions

NASA Astrophysics Data System (ADS)

Young, Stephen Michael Radley

1991-02-01

Available from UMI in association with The British Library. Requires signed TDF. We have carried out theoretical and experimental studies into the effect of intense laser radiation on atomic collisions. The first experiment used neon. Excitation by electron impact in a gas discharge demanded a pressure of at least 0.075 Torr. Measurement of the intensity of 3^1S_0to 3^1P_1 fluorescence has been made for the case where high intensity ASE wings in the laser profile and background laser scatter are unimportant, with the laser tuned to resonance. The field intensity required to produce strong field fluorescence (exemplified by the Mollow triplet) was found to give rise to complications capable of screening the effects sought. Our theoretical model has suggested that at finite detunings, line-centre fluorescence will dominate Rayleigh scatter and omega_3 fluorescence. Our measurements provide information on the saturation of neon fluorescence but not of the variation of the intense field collision rate. Absorption of weak field 253.7 nm laser photons by ground state mercury atoms yielded a high 6 ^3P_1 population at a lower pressure of 0.02 Torr. The Mollow triplet has been observed in the self-broadened mercury system. Dressing of the upper transition (6^3P_1rightarrow 7^3S_1) by an intense laser close to 435.8 nm yielded the strong field signal. Polarisation studies were made possible by the 3-level mercury system (radiation trapping in a 2-level system would depolarise fluorescence) perturbed by argon. The studies yielded results that were explainable in terms of the selective population of Stark shifted dressed states by a detuned, weak probe field. Use has been made of the electric-dipole radiation selection rule m_{J}=0 rightarrow m_{J^' } = 0 unless J=J^' to devise a 'Stark shift collision switch'. The competition between collision and radiation induced transitions within the mercury atom has then been studied. The resonant, strong lambda 435.8 nm field was used in conjunction with the weak lambda 253.7 nm field detuned by 0 to 6 cm^ {-1}. Measurement of fluorescence intensity in two perpendicular planes of polarisation has revealed the dominant | e_1> to | e_2> excitation channel as a function of the Stark shift by way of the U.V. detuning. Competition between the channels was dependent on the generalised Rabi frequency. However, we could only monitor the relative strength of the channels and were thus unable to say that the Stark shift switched collisions off. (Abstract shortened by UMI.).

Combination of hand-held probe and microscopy for fluorescence guided surgery in the brain tumor marginal zone.

PubMed

Richter, Johan C O; Haj-Hosseini, Neda; Hallbeck, Martin; Wårdell, Karin

2017-06-01

Visualization of the tumor is crucial for differentiating malignant tissue from healthy brain during surgery, especially in the tumor marginal zone. The aim of the study was to introduce a fluorescence spectroscopy-based hand-held probe (HHF-probe) for tumor identification in combination with the fluorescence guided resection surgical microscope (FGR-microscope), and evaluate them in terms of diagnostic performance and practical aspects of fluorescence detection. Eighteen operations were performed on 16 patients with suspected high-grade glioma. The HHF-probe and the FGR-microscope were used for detection of protoporphyrin (PpIX) fluorescence induced by 5-aminolevulinic acid (5-ALA) and evaluated against histopathological analysis and visual grading done through the FGR-microscope by the surgeon. A ratio of PpIX fluorescence intensity to the autofluorescence intensity (fluorescence ratio) was used to quantify the spectra detected by the probe. Fluorescence ratio medians (range 0 - 40) measured by the probe were related to the intensity of the fluorescence in the FGR-microscope, categorized as "none" (0.3, n=131), "weak" (1.6, n=34) and "strong" (5.4, n=28). Of 131 "none" points in the FGR-microscope, 88 (67%) exhibited fluorescence with the HHF-probe. For the tumor marginal zone, the area under the receiver operator characteristics (ROC) curve was 0.49 for the FGR-microscope and 0.65 for the HHF-probe. The probe was integrated in the established routine of tumor resection using the FGR-microscope. The HHF-probe was superior to the FGR-microscope in sensitivity; it detected tumor remnants after debulking under the FGR-microscope. The combination of the HHF-probe and the FGR-microscope was beneficial especially in the tumor marginal zone. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Laser-excited fluorescence for measuring atmospheric pollution

NASA Technical Reports Server (NTRS)

Menzies, R. T.

1975-01-01

System measures amount of given pollutant at specific location. Infrared laser aimed at location has wavelength that will cause molecules of pollutant to fluoresce. Detector separates fluorescence from other radiation and measures its intensity to indicate concentration of pollutant.

Palus Somni - Anomalies in the correlation of Al/Si X-ray fluorescence intensity ratios and broad-spectrum visible albedos. [lunar surface mineralogy

NASA Technical Reports Server (NTRS)

Clark, P. E.; Andre, C. G.; Adler, I.; Weidner, J.; Podwysocki, M.

1976-01-01

The positive correlation between Al/Si X-ray fluorescence intensity ratios determined during the Apollo 15 lunar mission and a broad-spectrum visible albedo of the moon is quantitatively established. Linear regression analysis performed on 246 1 degree geographic cells of X-ray fluorescence intensity and visible albedo data points produced a statistically significant correlation coefficient of .78. Three distinct distributions of data were identified as (1) within one standard deviation of the regression line, (2) greater than one standard deviation below the line, and (3) greater than one standard deviation above the line. The latter two distributions of data were found to occupy distinct geographic areas in the Palus Somni region.

Diurnal Variability in Chlorophyll-a, Carotenoids, CDOM and SO₄(2-) Intensity of Offshore Seawater Detected by an Underwater Fluorescence-Raman Spectral System.

PubMed

Chen, Jing; Ye, Wangquan; Guo, Jinjia; Luo, Zhao; Li, Ying

2016-07-13

A newly developed integrated fluorescence-Raman spectral system (λex = 532 nm) for detecting Chlorophyll-a (chl-a), Chromophoric Dissolved Organic Matter (CDOM), carotenoids and SO₄(2-) in situ was used to successfully investigate the diurnal variability of all above. Simultaneously using the integration of fluorescence spectroscopy and Raman spectroscopy techniques provided comprehensive marine information due to the complementarity between the different excitation mechanisms and different selection rules. The investigation took place in offshore seawater of the Yellow Sea (36°05'40'' N, 120°31'32'' E) in October 2014. To detect chl-a, CDOM, carotenoids and SO₄(2-), the fluorescence-Raman spectral system was deployed. It was found that troughs of chl-a and CDOM fluorescence signal intensity were observed during high tides, while the signal intensity showed high values with larger fluctuations during ebb-tide. Chl-a and carotenoids were influenced by solar radiation within a day cycle by different detection techniques, as well as displaying similar and synchronous tendency. CDOM fluorescence cause interference to the measurement of SO₄(2-). To avoid such interference, the backup Raman spectroscopy system with λex = 785 nm was employed to detect SO₄(2-) concentration on the following day. The results demonstrated that the fluorescence-Raman spectral system has great potential in detection of chl-a, carotenoids, CDOM and SO₄(2-) in the ocean.

In-Vivo Fluorescence Spectroscopy Of Normal And Atherosclerotic Arteries

NASA Astrophysics Data System (ADS)

Deckelbaum, Lawrence I.; Sarembock, Ian J.; Stetz, Mark L.; O'Brien, Kenneth M.; Cutruzzola, Francis W.; Gmitro, Arthur F.; Ezekowitz, Michael D.

1988-06-01

Laser-induced fluorescence spectroscopy can discriminate atherosclerotic from normal arteries in-vitro and may thus potentially guide laser angioplasty. To evaluate the feasibility of laser-induced fluorescence spectroscopy in a living blood-filled arterial system we performed fiberoptic laser-induced fluorescence spectroscopy in a rabbit model of focal femoral atherosclerosis. A laser-induced fluorescence spectroscopy score was derived from stepwise linear regression analysis of in-vitro spectra to distinguish normal aorta (score>0) from atherosclerotic femoral artery (score<0). A 400 u silica fiber, coupled to a helium cadmium laser and optical multichannel analyzer, was inserted through a 5F catheter to induce and record in-vivo fluorescence from femoral and aortoiliac arteries. Arterial spectra could be recorded in all animals (n=10: 5 occlusions, 5 stenoses). Blood spectra were of low intensity and were easily distinguished from arterial spectra. The scores (mean ± SEM) for the in-vivo spectra were -0.69 +/- 0.29 for artherosclerotic femoral, and +0.54 ±. 0.15 for normal aorta (p<.01 p=NS compared to in-vitro spectra). In-vitro, a fiber tip to tissue distance <50 u was necessary for adequate arterial LIFS in blood. At larger distances low intensity blood spectra were recorded (1/20 the intensity of tissue spectra). Thus, fiberoptic laser-induced fluorescence spectroscopy can be sucessfully performed in a blood filled artery provided the fiber tip is approximated to the tissue.

Calcium Sensitive Fluorescent Dyes Fluo-4 and Fura Red under Pressure: Behaviour of Fluorescence and Buffer Properties under Hydrostatic Pressures up to 200 MPa.

PubMed

Schneidereit, D; Vass, H; Reischl, B; Allen, R J; Friedrich, O

2016-01-01

The fluorescent Ca2+ sensitive dyes Fura Red (ratiometric) and Fluo-4 (non-ratiometric) are widely utilized for the optical assessment of Ca2+ fluctuations in vitro as well as in situ. The fluorescent behavior of these dyes is strongly depends on temperature, pH, ionic strength and pressure. It is crucial to understand the response of these dyes to pressure when applying calcium imaging technologies in the field of high pressure bioscience. Therefore, we use an optically accessible pressure vessel to pressurize physiological Ca2+-buffered solutions at different fixed concentrations of free Ca2+ (1 nM to 25.6 μM) and a specified dye concentration (12 μM) to pressures of 200 MPa, and record dye fluorescence intensity. Our results show that Fluo-4 fluorescence intensity is reduced by 31% per 100 MPa, the intensity of Fura Red is reduced by 10% per 100 MPa. The mean reaction volume for the dissociation of calcium from the dye molecules [Formula: see text] is determined to -17.8 ml mol-1 for Fluo-4 and -21.3 ml mol-1 for Fura Red. Additionally, a model is presented that is used to correct for pressure-dependent changes in pH and binding affinity of Ca2+ to EGTA, as well as to determine the influence of these changes on dye fluorescence.

Use of Panitumumab-IRDye800 to Image Microscopic Head and Neck Cancer in an Orthotopic Surgical Model

PubMed Central

Heath, C. Hope; Deep, Nicholas L.; Sweeny, Larissa; Zinn, Kurt R; Rosenthal, Eben L.

2013-01-01

Background Fluorescence imaging hardware (SPY) has recently been developed for intraoperative assessment of blood flow via detection of probes emitting in the near-infrared (NIR) spectrum. This study sought to determine if this imaging system was capable of detecting micrometastatic head and neck squamous cell carcinoma (HNSCC) in preclinical models. Methods A NIR fluorescent probe (IRDye800CW) was covalently linked to a monoclonal antibody targeting EGFR (panitumumab) or non-specific IgG. HNSCC flank (SCC-1) and orthotopic (FADU and OSC19) xenografts were imaged 48-96hrs following systemic injection of labeled panitumumab or IgG. The primary tumor and regional lymph nodes were dissected using fluorescence guidance with the SPY system and grossly assessed with a charge-coupled NIR system (Pearl). Histologic slides were also imaged with a NIR charged-coupled device (Odyssey) and fluorescence intensity was correlated with pathologic confirmation of disease. Results Orthotopic tongue tumors were clearly delineated from normal tissue with tumor-to-background ratios of 2.9(Pearl) and 2.3(SPY). Disease detection was significantly improved with panitumumab-IRDye compared to IgG-IRDye800 (P<0.05). Tissue biopsies (average size=3.7mm) positive for fluorescence were confirmed for pathologic disease by histology and immunohistochemistry (n=25/25). Biopsies of non-fluorescent tissue were proven to be negative for malignancy (n=28/28). The SPY was able to detect regional lymph node metastasis (<1.0mm) and microscopic areas of disease. Standard histological assessment in both frozen and paraffin-embedded histologic specimens was augmented using the Odyssey. Conclusions Panitumumab-IRDye800 may have clinical utility in detection and removal of microscopic HNSCC using existing intraoperative optical imaging hardware and may augment analysis of frozen and permanent pathology. PMID:22669455

Enhanced photoluminescence and characterization of multicolor carbon dots using plant soot as a carbon source.

PubMed

Tan, Mingqian; Zhang, Lingxin; Tang, Rong; Song, Xiaojie; Li, Yimin; Wu, Hao; Wang, Yanfang; Lv, Guojun; Liu, Wanfa; Ma, Xiaojun

2013-10-15

Carbon dots (C-dots) are a class of novel fluorescent nanomaterials, which have drawn great attention for their potential applications in bio-nanotechnology. Multicolor C-dots have been synthesized by chemical nitric acid oxidation using the reproducible plant soot as raw material. TEM analysis reveals that the prepared C-dots have an average size of 3.1 nm. The C-dots are well dispersed in aqueous solution and are strongly fluorescent under the irradiation of ultra-violet light. X-ray photoelectron spectroscopy characterization demonstrates that the O/C atomic ratio for C-dots change to from 0.207 to 0.436 due to the chemical oxidation process. The photo bleaching experiment reveals that the C-dots show excellent photostability as compared with the conventional organic dyes, fluorescein and rhodamine B. The fluorescence intensity of the C-dots did not change significantly in the pH range of 3-10. To further enhance the fluorescence quantum yield, the C-dots were surface modified with four types of passivation ligands, 4,7,10-trioxa-1,13-tridecanediamine (TTDDA), poly-L-lysine (PLL), cysteine and chitosan and the fluorescence quantum yields of the TTDDA, PLL, cysteine and chitosan passivated C-dots were improved 1.53-, 5.94-, 2.00- and 3.68-fold, respectively. Fourier-transform infrared (FTIR) spectra were employed to characterize the surface groups of the C-dots. The bio-application of the C-dots as fluorescent bio-probes was evaluated in cell imaging and ex vivo fish imaging, which suggests that the C-dots may have potential applications in biolabeling and bioimaging. Copyright © 2013 Elsevier B.V. All rights reserved.

Evaluation of slide based cytometry (SBC) for concentration measurements of fluorescent dyes in solution

NASA Astrophysics Data System (ADS)

Pierzchalski, Arkadiusz; Marecka, Monika; Müller, Hans-Willy; Bocsi, József; Tárnok, Attila

2009-02-01

Flow cytometers (FCM) are built for particle measurements. In principle, concentration measurement of a homogeneous solution is not possible with FCM due to the lack of a trigger signal. In contrast to FCM slide based cytometry systems could act as tools for the measurement of concentrations using volume defined cell counting chambers. These chambers enable to analyze a well defined volume. Sensovation AG (Stockach, Germany) introduced an automated imaging system that combines imaging with cytometric features analysis. Aim of this study was to apply this imaging system to quantify the fluorescent molecule concentrations. The Lumisens (Sensovation AG) slide-based technology based on fluorescence digital imaging microscopy was used. The instrument is equipped with an inverted microscope, blue and red LEDs, double band-pass filters and a high-resolution cooled 16-bit digital camera. The instrument was focussed on the bottom of 400μm deep 6 chamber slides (IBIDI GmbH, Martinsried, Germany) or flat bottom 96 well plates (Greiner Bio One GmbH, Frickenhausen, Germany). Fluorescent solutions were imaged under 90% pixel saturation in a broad concentration range (FITC: 0.0002-250 μg/ml, methylene blue (MethB): 0.0002-250 μg/ml). Exposition times were recorded. Images were analysed by the iCys (CompuCyte Corp., Cambridge, MA, USA) image analysis software with the phantom contour function. Relative fluorescence intensities were calculated from mean fluorescence intensities per phantom contours divided by the exposition time. Solution concentrations could be distinguished over a broad dynamic range of 3.5 to 5.5 decades log (range FITC: 0.0002-31.25μg/ml, MethB: 0.0076-31.25μg/ml) with a good linear relationship between dye concentration and relative fluorescence intensity. The minimal number of fluorescent molecules per pixel as determined by the mean fluorescence intensity and the molecular weight of the fluorochrome were about 800 molecules FITC and ~2.000 MethB. The novel slide-based imaging system is suitable for detection of fluorescence differences over a broad range of concentrations. This approach may lead to novel assays for measuring concentration differences in cell free solutions and cell cultures e.g. in secretion assays.

Method and apparatus for optical temperature measurements

DOEpatents

Angel, S.M.; Hirschfeld, T.B.

1986-04-22

A method and apparatus are provided for remotely monitoring temperature. Both method and apparatus employ a temperature probe material having an excitation-dependent emission line whose fluorescence intensity varies directly with temperature whenever excited by light having a first wavelength and whose fluorescence intensity varies inversely with temperature whenever excited by light having a second wavelength. Temperature is measured by alternatively illiminating the temperature probe material with light having the first wavelength and light having the second wavelength, monitoring the intensity of the successive emissions of the excitation-dependent emission line, and relating the intensity ratio of successive emissions to temperature. 3 figs.

Method and apparatus for optical temperature measurements

DOEpatents

Angel, S. Michael; Hirschfeld, Tomas B.

1988-01-01

A method and apparatus are provided for remotely monitoring temperature. Both method and apparatus employ a temperature probe material having an excitation-dependent emission line whose fluorescence intensity varies directly with temperature whenever excited by light having a first wavelength and whose fluorescence intensity varies inversely with temperature whenever excited by light having a second wavelength. Temperature is measured by alternatively illuminating the temperature probe material with light having the first wavelength and light having the second wavelength, monitoring the intensity of the successive emissions of the excitation-dependent emission line, and relating the intensity ratio of successive emissions to temperature.

Measurement of the fluorescence of crop residues: A tool for controlling soil erosion

NASA Technical Reports Server (NTRS)

Daughtry, C. S. T.; Mcmurtrey, J. E., III; Chappelle, E. W.; Hunter, W. J.

1994-01-01

Management of crop residues, the portion of a crop left in the field after harvest, is an important conservation practice for minimizing soil erosion and for improving water quality. Quantification of crop residue cover is required to evaluate the effectiveness of conservation tillage practices. Methods are needed to quantify residue cover that are rapid, accurate, and objective. The fluorescence of crop residue was found to be a broadband phenomenon with emission maxima at 420 to 495 nm for excitations of 350 to 420 nm. Soils had low intensity broadband emissions over the 400 to 690 nm region for excitations of 300 to 600 nm. The range of relative fluorescence intensities for the crop residues was much greater than the fluorescence observed of the soils. As the crop residues decompose their blue fluorescence values approach the fluorescence of the soil. Fluorescence techniques are concluded to be less ambiguous and better suited for discriminating crop residues and soils than reflectance methods. If properly implemented, fluorescence techniques can be used to quantify, not only crop residue cover, but also photosynthetic efficiency in the field.

Laser-saturated fluorescence measurements in laminar sooting diffusion flames

NASA Technical Reports Server (NTRS)

Wey, Changlie

1993-01-01

The hydroxyl radical is known to be one of the most important intermediate species in the combustion processes. The hydroxyl radical has also been considered a dominant oxidizer of soot particles in flames. In this investigation the hydroxyl concentration profiles in sooting diffusion flames were measured by the laser-saturated fluorescence (LSF) method. The temperature distributions in the flames were measured by the two-line LSF technique and by thermocouple. In the sooting region the OH fluorescence was too weak to make accurate temperature measurements. The hydroxyl fluorescence profiles for all four flames presented herein show that the OH fluorescence intensities peaked near the flame front. The OH fluorescence intensity dropped sharply toward the dark region of the flame and continued declining to the sooting region. The OH fluorescence profiles also indicate that the OH fluorescence decreased with increasing height in the flames for all flames investigated. Varying the oxidizer composition resulted in a corresponding variation in the maximum OH concentration and the flame temperature. Furthermore, it appears that the maximum OH concentration for each flame increased with increasing flame temperature.

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

PubMed

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

2015-01-25

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

Preliminary Results on Luminaire Designs for Hybrid Solar Lighting Systems

DOE Office of Scientific and Technical Information (OSTI.GOV)

Earl, D.D.

2001-06-15

We report on the design of two hybrid lighting luminaires that blend light from a fiber optic end-emitted solar source with electric T8 fluorescent lamps. Both designs involve the retrofit of a commercially-available recessed fluorescent luminaire with minimal reductions in the original luminaire's optical efficiency. Two methods for high-angle dispersion of fiber optic end-emitted solar light are described and the resulting spatial intensity distributions, simulated using ZEMAX, are compared with standard cylindrical fluorescent tubes. Differences in spatial intensity distribution are qualitatively characterized and potential design improvements discussed.

Fluorescence decay time imaging using an imaging photon detector with a radio frequency photon correlation system

NASA Astrophysics Data System (ADS)

Morgan, Christopher G.; Mitchell, A. C.; Murray, J. G.

1990-05-01

An imaging photon detector has been modified to incorporate fast timing electronics coupled to a custom built photon correlator interfaced to a RISC computer. Using excitation with intensity- muodulated light, fluorescence images can be readily obtained where contrast is determined by the decay time of emission, rather than by intensity. This technology is readily extended to multifrequency phase/demodulation fluorescence imaging or to differential polarised phase fluorometry. The potential use of the correlator for confocal imaging with a laser scanner is also briefly discussed.

Application of Time-Resolved Spectroscopies to the Study of Energetic Materials - 1982

DTIC Science & Technology

1983-05-24

fluores- cence intensity as a function of UV pulse energy, for individual laser shots. The lower curve shows the UV + probe induced fluorescence... intensity as a function of UV pulse energy, for individual laser shots. The lower curve shows the UV + probe Induced fluorescence, at 1 ns delay...locked Nd:YAG Laser Pulse ", Appl. Phys. Lett 26, 501-503 (1975). 97 43. A. J. Campillo, V. H. Kollman and S. L. Shapiro, " Intensity Dependence of

Characterizing chlorine oxidation of dissolved organic matter and disinfection by-product formation with fluorescence spectroscopy and parallel factor analysis

NASA Astrophysics Data System (ADS)

Beggs, Katherine M. H.; Summers, R. Scott; McKnight, Diane M.

2009-12-01

Relationships between chlorine demand and disinfection by-product (DBP) formation during chlorination and fluorescence of dissolved organic matter (DOM) were developed. Fluorescence excitation and emission (EEM) spectroscopy was employed, and parameters including fluorescence index, redox index, and overall fluorescence intensity (OFI) were correlated to chlorine demand and DBP formation. The EEMs were also analyzed using a well established global parallel factor analysis (PARAFAC) model which resolves the fluorescence signal into 13 components, including quinone-like and protein-like components. Over an 8-day chlorination period the OFI and sum of the 13 PARAFAC loadings decreased by more than 70%. The remaining identified quinone-like compounds within the DOM were shifted to a more oxidized state. Quinone fluorescence was strongly correlated to both reduced fluorescence intensity and to chlorine demand which indicates that fluorescence may be used to track the chlorine oxidation of DOM. Quinone fluorescence was also correlated strongly with both classes of regulated DBPs: total trihalomethanes and haloacetic acids. Quinone-like components were found to be strongly correlated to overall, short-term, and long-term specific DBP formation. The results of this study show that fluorescence is a useful tool in tracking both DOM oxidation and DBP formation during chlorination.

Aggressive resection at the infiltrative margins of glioblastoma facilitated by intraoperative fluorescein guidance.

PubMed

Neira, Justin A; Ung, Timothy H; Sims, Jennifer S; Malone, Hani R; Chow, Daniel S; Samanamud, Jorge L; Zanazzi, George J; Guo, Xiaotao; Bowden, Stephen G; Zhao, Binsheng; Sheth, Sameer A; McKhann, Guy M; Sisti, Michael B; Canoll, Peter; D'Amico, Randy S; Bruce, Jeffrey N

2017-07-01

OBJECTIVE Extent of resection is an important prognostic factor in patients undergoing surgery for glioblastoma (GBM). Recent evidence suggests that intravenously administered fluorescein sodium associates with tumor tissue, facilitating safe maximal resection of GBM. In this study, the authors evaluate the safety and utility of intraoperative fluorescein guidance for the prediction of histopathological alteration both in the contrast-enhancing (CE) regions, where this relationship has been established, and into the non-CE (NCE), diffusely infiltrated margins. METHODS Thirty-two patients received fluorescein sodium (3 mg/kg) intravenously prior to resection. Fluorescence was intraoperatively visualized using a Zeiss Pentero surgical microscope equipped with a YELLOW 560 filter. Stereotactically localized biopsy specimens were acquired from CE and NCE regions based on preoperative MRI in conjunction with neuronavigation. The fluorescence intensity of these specimens was subjectively classified in real time with subsequent quantitative image analysis, histopathological evaluation of localized biopsy specimens, and radiological volumetric assessment of the extent of resection. RESULTS Bright fluorescence was observed in all GBMs and localized to the CE regions and portions of the NCE margins of the tumors, thus serving as a visual guide during resection. Gross-total resection (GTR) was achieved in 84% of the patients with an average resected volume of 95%, and this rate was higher among patients for whom GTR was the surgical goal (GTR achieved in 93.1% of patients, average resected volume of 99.7%). Intraoperative fluorescein staining correlated with histopathological alteration in both CE and NCE regions, with positive predictive values by subjective fluorescence evaluation greater than 96% in NCE regions. CONCLUSIONS Intraoperative administration of fluorescein provides an easily visualized marker for glioma pathology in both CE and NCE regions of GBM. These findings support the use of fluorescein as a microsurgical adjunct for guiding GBM resection to facilitate safe maximal removal.

Interpretative Guidelines and Possible Indications for Indocyanine Green Fluorescence Imaging in Robot-Assisted Sphincter-Saving Operations.

PubMed

Kim, Jin Cheon; Lee, Jong Lyul; Park, Seong Ho

2017-04-01

Since the introduction of indocyanine green angiography more than 25 years ago, few studies have presented interpretative guidelines for indocyanine green fluorescent imaging. We aimed to provide interpretative guidelines for indocyanine green fluorescent imaging through quantitative analysis and to suggest possible indications for indocyanine green fluorescent imaging during robot-assisted sphincter-saving operations. This is a retrospective observational study. This study was conducted at a single center. A cohort of 657 patients with rectal cancer who consecutively underwent curative robot-assisted sphincter-saving operations was enrolled between 2010 and 2016, including 310 patients with indocyanine green imaging (indocyanine green fluorescent imaging+ group) and 347 patients without indocyanine green imaging (indocyanine green fluorescent imaging- group). We tried to quantitatively define the indocyanine green fluorescent imaging findings based on perfusion (mesocolic and colic) time and perfusion intensity (5 grades) to provide probable indications. The anastomotic leakage rate was significantly lower in the indocyanine green fluorescent imaging+ group than in the indocyanine green fluorescent imaging- group (0.6% vs 5.2%) (OR, 0.123; 95% CI, 0.028-0.544; p = 0.006). Anastomotic stricture was closely correlated with anastomotic leakage (p = 0.002) and a short descending mesocolon (p = 0.003). Delayed perfusion (>60 s) and low perfusion intensity (1-2) were more frequently detected in patients with anastomotic stricture and marginal artery defects than in those without these factors (p ≤ 0.001). In addition, perfusion times greater than the mean were more frequently observed in patients aged >58 years, whereas low perfusion intensity was seen more in patients with short descending mesocolon and high ASA classes (≥3). The 300 patients in the indocyanine green fluorescent imaging- group underwent operations 3 years before indocyanine green fluorescent imaging. Quantitative analysis of indocyanine green fluorescent imaging may help prevent anastomotic complications during robot-assisted sphincter-saving operations, and may be of particular value in high-class ASA patients, older patients, and patients with a short descending mesocolon.

Complete suppression of the fluorophore fluorescence by combined effect of multiple fluorescence quenching groups: A fluorescent sensor for Cu²⁺ with zero background signals.

PubMed

Long, Lingliang; Wu, Yanjun; Wang, Lin; Gong, Aihua; Hu, Rongfeng; Zhang, Chi

2016-02-18

The reaction-based fluorescent sensors have attracted increasing attention in the past decades. However, the application of these sensors for accurate sensing was significantly retarded by the background fluorescence from the sensors themselves. In this work, we demonstrated a novel strategy that the background fluorescence of the sensor could be completely eliminated by the combined effect of multiple fluorescence quenching groups. Based on this new strategy, as proof-of-principle study, a fluorescent sensor (CuFS) for Cu(2+) was judiciously developed. In CuFS, three types of fluorescence quenching groups were directly tethered to a commonly used coumarin fluorophore. The fluorescence of coumarin fluorophore in CuFS was completely suppressed by the combined effect of these fluorescence quenching groups. Upon treatment with 22 μM Cu(2+), sensor CuFS achieved a dramatic fluorescence enhancement (fluorescence intensity enhanced up to 811-fold) centered at 469 nm. The detection limits was determined to be 12.3 nM. The fluorescence intensity enhancement also showed a good linearity with the Cu(2+) concentration in the range of 12.3 nM to 2 μM. By fabricating test strips, sensor CuFS can be utilized as a simple tool to detect Cu(2+) in water samples. Furthermore, the fluorescent sensor was successfully applied in detecting different concentration of Cu(2+) in living cells. Copyright © 2015 Elsevier B.V. All rights reserved.

Polythiophene nanofilms for sensitive fluorescence detection of viruses in drinking water.

PubMed

Wankar, Shashwati; Turner, Nicholas W; Krupadam, Reddithota J

2016-08-15

Molecular imprints of the tobacco necrosis virus (TNV) have been formed within polythiophene nanofilms with an approximate thickness of 200nm. These films have been electrochemically deposited onto conducting Au surfaces. Upon rebinding, the TNV-polythiophene complex changes the fluorescence intensity of the nanofilm. The fluorescence intensity at 410nm was observed to be proportional to the concentration of viruses in the range of 0.1-10ngL(-1) (0.15-15pg) with the lower calculated detection limit of 2.29ngL(-1) (3.4pg). The intensity of the fluorescence emission is not affected by the thickness of the polythiophene film and the nature of TNV specific binding sites. Kinetic data analyses showed that the nanofilm responds to TNV within 2min; and cross-selectivity studies with tobacco mosaic virus (TMV) showed an excellent specificity for the targeted TNV. These binding experiments demonstrate the potential of fluorescence emission for the specific, label free and rapid detection of viruses using nanofilm sensors. Taking into account the lower limit of detection, the fluorescence sensing reported here is reliable, simple to perform, rapid, cost-effective and offers a sensitive analytical method for virus detection in water resources. Copyright © 2016 Elsevier B.V. All rights reserved.

Resolving environmental microheterogeneity and dielectric relaxation in fluorescence kinetics of protein

NASA Astrophysics Data System (ADS)

Rolinski, Olaf J.; McLaughlin, Damien; Birch, David J. S.; Vyshemirsky, Vladislav

2016-09-01

The fluorescence intensity decay of protein is easily measurable and reports on the intrinsic fluorophore-local environment interactions on the sub-nm spatial and sub-ns temporal scales, which are consistent with protein activity in numerous biomedical and industrial processes. This makes time-resolved fluorescence a perfect tool for understanding, monitoring and controlling these processes at the molecular level, but the complexity of the decay, which has been traditionally fitted to multi-exponential functions, has hampered the development of this technique over the last few decades. Using the example of tryptophan in HSA we present the alternative to the conventional approach to modelling intrinsic florescence intensity decay in protein where the key factors determining fluorescence decay, i.e. the excited-state depopulation and the dielectric relaxation (Toptygin and Brand 2000 Chem. Phys. Lett. 322 496-502), are represented by the individual relaxation functions. This allows quantification of both effects separately by determining their parameters from the global analysis of a series of fluorescence intensity decays measured at different detection wavelengths. Moreover, certain pairs of the recovered parameters of tryptophan were found to be correlated, indicating the influence of the dielectric relaxation on the transient rate of the electronic transitions. In this context the potential for the dual excited state depopulation /dielectric relaxation fluorescence lifetime sensing is discussed.

Determination of dopamine hydrochloride by host-guest interaction based on water-soluble pillar[5]arene

NASA Astrophysics Data System (ADS)

Xiao, Xue-Dong; Shi, Lin; Guo, Li-Hui; Wang, Jun-Wen; Zhang, Xiang

2017-02-01

The supramolecular interaction between the water-soluble pillar[5]arene (WP[5]) as host and dopamine hydrochloride (DH) as guest was studied by spectrofluorometry. The fluorescence intensity of DH gradually decreased with increasing WP[5] concentration, and the possible interaction mechanism between WP[5] and DH was confirmed by 1H NMR, 2D NOESY, and molecular modelling. Based on significant DH fluorescence, a highly sensitive and selective method for DH determination was developed for the first time. The fluorescence intensity was measured at 312 nm, with excitation at 285 nm. The effects of pH, temperature, and reaction time on the fluorescence spectra of the WP[5]-DH complex were investigated. A linear relationship between fluorescence intensity and DH concentration in the range of 0.07-6.2 μg mL- 1 was obtained. The corresponding linear regression equation is ΔF = 25.76 C + 13.56 (where C denotes the concentration in μg mL- 1), with the limit of detection equal to 0.03 μg mL- 1 and the correlation coefficient equal to 0.9996. This method can be used for the determination of dopamine in injection and urine samples. In addition, the WP[5]-DH complex has potential applications in fluorescent sensing and pharmacokinetics studies of DH.

Specific detection of Vibrio parahaemolyticus by fluorescence quenching immunoassay based on quantum dots.

PubMed

Wang, Ling; Zhang, Junxian; Bai, Haili; Li, Xuan; Lv, Pintian; Guo, Ailing

2014-07-01

In this study, anti-Vibrio parahaemolyticus polyclonal and monoclonal antibodies were prepared through intradermal injection immune and lymphocyte hybridoma technique respectively. CdTe quantum dots (QDs) were synthesized at pH 9.3, 98 °C for 1 h with stabilizer of 2.7:1. The fluorescence intensity was 586.499, and the yield was 62.43%. QD probes were successfully prepared under the optimized conditions of pH 7.4, 37 °C for 1 h, 250 μL of 50 mg/mL EDC · HCl, 150 μL of 4 mg/mL NHS, buffer system of Na2HPO4-citric acid, and 8 μL of 2.48 mg/mL polyclonal antibodies. As gold nanoparticles could quench fluorescence of quantum dots, the concentration of V. parahaemolyticus could be detected through measuring the reduction of fluorescence intensity in immune sandwich reaction composed of quantum dot probe, gold-labeled antibody, and the sample. For pure culture, fluorescence intensity of the system was proportional with logarithm concentration of antigen, and the correlation coefficient was 99.764%. The fluorescence quenching immunoassay based on quantum dots is established for the first time to detect Vibrio parahaemolyticus. This method may be used as rapid testing procedure due to its high simplicity and sensitivity.

Fluorescence recognition of chiral amino alcohols by using a novel ionic liquid sensor.

PubMed

Cai, Pengfei; Wu, Datong; Zhao, Xiaoyong; Pan, Yuanjiang

2017-08-07

A novel task-specific ionic liquid derived from l-phenylalaninol was prepared as an enantioselective fluorescent sensor for the first time. Fluorescent chiral ionic liquid 1 (FCIL1) is found to exhibit highly enantioselective fluorescence enhancements toward both aromatic and non-aromatic chiral amino alcohols. When (S)-FCIL1 was treated with the enantiomers of phenylalaninol, a great fluorescence enhancement at 349 nm could be observed and the value of the enantiomeric fluorescence difference (ef) is 5.92. This demonstrated that the chiral sensor (S)-FCIL1 exhibited an excellent enantioselective response behaviour to d-phenylalaninol. Besides that, both the fluorescence intensity at 349 nm (I 349 ) and the ratio of I 349 to I 282 depend linearly on the concentration of amino alcohols. Both the concentration and the enantiomeric composition could be determined by using the chiral ionic liquid. Differently, the sensor treated with the enantiomers of 2-amino-1-butanol showed an opposite result: the fluorescence intensity of the S-enantiomer is higher than that of the R-enantiomer. Furthermore, the size of the substituents on the chiral carbon might be important for the enantioselective fluorescent response.

Comparative study of the fluorescence intensity of dental composites and human teeth submitted to artificial aging.

PubMed

Jablonski, Tatiana; Takahashi, Marcos Kenzo; Brum, Rafeal Torres; Rached, Rodrigo Nunes; Souza, Evelise M

2014-01-01

The aim of this study was to evaluate quantitatively the fluorescence of resin composites and human teeth, and to determine the stability of fluorescence after aging. Ten specimens were built using a 1 mm thick increment of dentin composite overlapped by a 0.5 mm thick increment of enamel composite. Ten sound human molars were sectioned and silicon carbide-polished to obtain enamel and dentin slabs 1.5 mm in thickness. Fluorescence measurements were carried out by a fluorescence spectrophotometer before and after thermocycling (2000 cycles, 5°C and 55°C). One-way analysis of variance (ANOVA) with repeated measures and Tukey's test were performed at a significance level of 5%. Most of the tested composites showed significant differences in fluorescence both before and after aging (P < 0.05). Opallis was the only composite whose fluorescence was similar to that of human teeth at both periods of evaluation (P > 0.05), and was the only composite that showed comparable results of fluorescence to the tooth structure before and after thermocycling. With the exception of Filtek Supreme, there were significant reductions in fluorescence intensity for all the tested composites (P < 0.05).

Detection and evaluation of normal and malignant cells using laser-induced fluorescence spectroscopy.

PubMed

Khosroshahi, Mohamad E; Rahmani, Mahya

2012-01-01

The aim of this research is to study the normalized fluorescence spectra (intensity variations and area under the fluorescence signal), relative quantum yield, extinction coefficient and intracellular properties of normal and malignant human bone cells. Using Laser-Induced Fluorescence Spectroscopy (LIFS) upon excitation of 405 nm, the comparison of emission spectra of bone cells revealed that fluorescence intensity and the area under the spectra of malignant bone cells was less than that of normal. In addition, the area ratio and shape factor were changed. We obtained two emission bands in spectra of normal cells centered at about 486 and 575 nm and for malignant cells about 482 and 586 nm respectively, which are most likely attributed to NADH and riboflavins. Using fluorescein sodium emission spectrum, the relative quantum yield of bone cells is numerically determined.

pH-responsive fluorescence chemical sensor constituted by conjugated polymers containing pyridine rings.

PubMed

Adachi, Naoya; Kaneko, Yuki; Sekiguchi, Kazuki; Sugiyama, Hiroki; Sugeno, Masafumi

2015-12-01

Poly(p-pyridinium phenylene ethynylene)s (PPyPE) functionalized with alternating donor-acceptor repeat units were synthesized by a Pd-catalyzed Sonogashira coupling reaction between diethynyl monomer and di-iodopyridine for use as a pH-responsive fluorescence chemical sensor. The synthesized PPyPE, containing pyridine units, was characterized by FT-IR, (1)H and (13)C NMR, UV-visible and fluorescence spectroscopies. We investigated the relationship between changes of optical properties and protonation/deprotonation of PPyPE containing pyridine units in solution. Addition of HCl decreased and red-shifted the fluorescence intensity of the conjugated polymers that contained pyridine rings; fluorescence intensity of the polymers increased upon addition of NaOH solution. The synthesized PPyPE was found to be an effective and reusable chemical sensor for pH sensing. Copyright © 2015 John Wiley & Sons, Ltd.

Micro-Raman spectroscopy of algae: composition analysis and fluorescence background behavior.

PubMed

Huang, Y Y; Beal, C M; Cai, W W; Ruoff, R S; Terentjev, E M

2010-04-01

Preliminary feasibility studies were performed using Stokes Raman scattering for compositional analysis of algae. Two algal species, Chlorella sorokiniana (UTEX #1230) and Neochloris oleoabundans (UTEX #1185), were chosen for this study. Both species were considered to be candidates for biofuel production. Raman signals due to storage lipids (specifically triglycerides) were clearly identified in the nitrogen-starved C. sorokiniana and N. oleoabundans, but not in their healthy counterparts. On the other hand, signals resulting from the carotenoids were found to be present in all of the samples. Composition mapping was conducted in which Raman spectra were acquired from a dense sequence of locations over a small region of interest. The spectra obtained for the mapping images were filtered for the wavelengths of characteristic peaks that correspond to components of interest (i.e., triglyceride or carotenoid). The locations of the components of interest could be identified by the high intensity areas in the composition maps. Finally, the time evolution of fluorescence background was observed while acquiring Raman signals from the algae. The time dependence of fluorescence background is characterized by a general power law decay interrupted by sudden high intensity fluorescence events. The decreasing trend is likely a result of photo-bleaching of cell pigments due to prolonged intense laser exposure, while the sudden high intensity fluorescence events are not understood. (c) 2009 Wiley Periodicals, Inc.

Sensitive and background-free determination of thiols from wastewater samples by MOF-5 extraction coupled with high-performance liquid chromatography with fluorescence detection using a novel fluorescence probe of carbazole-9-ethyl-2-maleimide.

PubMed

Lv, Zhengxian; Sun, Zhiwei; Song, Cuihua; Lu, Shuaimin; Chen, Guang; You, Jinmao

2016-12-01

A sensitive and background-free pre-column derivatization method for the determination of thiol compounds using metal-organic framework material (MOF-5) as dispersive solid-phase extraction (DSPE) adsorbent followed by high-performance liquid chromatography fluorescence detection (HPLC-FLD) has been developed. In this paper, a novel labeling reagent, carbazole-9-ethyl-2-maleimide(CAEM), was synthesized and reacted with thiols at 40°C for 10min in the presence of PBS buffer (0.02mol/L, pH 7.5). Interestingly, CAEM itself had no fluorescence, while its derivatives exhibited intense fluorescence with an excitation maximum at λ ex 274nm and an emission maximum at λ em 363nm, which greatly reduced the background interference and improved the sensitivity of the method. Furthermore, the MOF-5 was prepared and used as DSPE adsorbent for the selective adsorption of thiols from wastewater sample. Under the optimized experimental conditions, an excellent linearity for all analytes over their concentration ranges of 0.01-1.0μmol/L (R 2 >0.9986)were obtained with the limit of detection (LOD) ranging from 8 to 17.1pmol/L for nine tested thiols. The feasibility of this method for the determination of thiols in wastewater samples had been evaluated and satisfactory average recoveries (n=3) were achieved with the range of 86.6-98.5%. Copyright © 2016 Elsevier B.V. All rights reserved.

Understanding the fundamental mechanisms of biofilms development and dispersal: BIAM (Biofilm Intensity and Architecture Measurement), a new tool for studying biofilms as a function of their architecture and fluorescence intensity.

PubMed

Baudin, Marine; Cinquin, Bertrand; Sclavi, Bianca; Pareau, Dominique; Lopes, Filipa

2017-09-01

Confocal laser scanning microscopy (CLSM) is one of the most relevant technologies for studying biofilms in situ. Several tools have been developed to investigate and quantify the architecture of biofilms. However, an approach to quantify correctly the evolution of intensity of a fluorescent signal as a function of the structural parameters of a biofilm is still lacking. Here we present a tool developed in the ImageJ open source software that can be used to extract both structural and fluorescence intensity from CLSM data: BIAM (Biofilm Intensity and Architecture Measurement). This is of utmost significance when studying the fundamental mechanisms of biofilm growth, differentiation and development or when aiming to understand the effect of external molecules on biofilm phenotypes. In order to provide an example of the potential of such a tool in this study we focused on biofilm dispersion. cis-2-Decenoic acid (CDA) is a molecule known to induce biofilm dispersion of multiple bacterial species. The mechanisms by which CDA induces dispersion are still poorly understood. To investigate the effects of CDA on biofilms, we used a reporter strain of Escherichia coli (E. coli) that expresses the GFPmut2 protein under control of the rrnBP1 promoter. Experiments were done in flow cells and image acquisition was made with CLSM. Analysis carried out using the new tool, BIAM, indicates that CDA affects the fluorescence intensity of the biofilm structures as well as biofilm architectures. Indeed, our results demonstrate that CDA removes more than 35% of biofilm biovolume and suggest that it results in an increase of the biofilm's mean fluorescence intensity (MFI) by more than 26% compared to the control biofilm in the absence of CDA. Copyright © 2017. Published by Elsevier B.V.

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

PubMed Central

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

2018-01-01

Background Glypican-3 (GPC3) is highly expressed in most of the hepatocellular carcinomas (HCCs), even in small HCCs. It may be used as a potential biomarker for early detection of HCC. The aptamer is a promising targeting agent with unique advantages over antibody. This study was to introduce a novel GPC3 specific aptamer (AP613-1), to verify its specific binding property in vitro, and to evaluate its targeting efficiency in vivo by performing near-infrared (NIR) fluorescence imaging on an HCC xenograft model. Methods AP613-1 was generated from the systematic evolution of ligands by exponential enrichment. Flow cytometry and aptamer-based immunofluorescence imaging were performed to verify the binding affinity of AP613-1 to GPC3 in vitro. NIR Fluorescence images of nude mice with unilateral (n=12) and bilateral (n=4) subcutaneous xenograft tumors were obtained. Correlation between the tumor fluorescence intensities in vivo and ex vivo was analyzed. Results AP613-1 could specifically bind to GPC3 in vitro. In vivo and ex vivo tumors, fluorescence intensities were in excellent correlation (P<0.001, r=0.968). The fluorescence intensity is significantly higher in tumors given Alexa Fluor 750 (AF750) labeled AP613-1 than in those given AF750 labeled initial ssDNA library both in vivo (P<0.001) and ex vivo (P=0.022). In the mice with bilateral subcutaneous tumors injected with AF750 labeled AP613-1, Huh-7 tumors showed significantly higher fluorescence intensities than A549 tumors both in vivo (P=0.016) and ex vivo (P=0.004). Conclusions AP613-1 displays a specific binding affinity to GPC3 positive HCC. Fluorescently labeled AP613-1 could be used as an imaging probe to subcutaneous HCC in xenograft models. PMID:29675356

Strong signal increase in STED fluorescence microscopy by imaging regions of subdiffraction extent

PubMed Central

Göttfert, Fabian; Pleiner, Tino; Heine, Jörn; Westphal, Volker; Görlich, Dirk; Sahl, Steffen J.; Hell, Stefan W.

2017-01-01

Photobleaching remains a limiting factor in superresolution fluorescence microscopy. This is particularly true for stimulated emission depletion (STED) and reversible saturable/switchable optical fluorescence transitions (RESOLFT) microscopy, where adjacent fluorescent molecules are distinguished by sequentially turning them off (or on) using a pattern of light formed as a doughnut or a standing wave. In sample regions where the pattern intensity reaches or exceeds a certain threshold, the molecules are essentially off (or on), whereas in areas where the intensity is lower, that is, around the intensity minima, the molecules remain in the initial state. Unfortunately, the creation of on/off state differences on subdiffraction scales requires the maxima of the intensity pattern to exceed the threshold intensity by a large factor that scales with the resolution. Hence, when recording an image by scanning the pattern across the sample, each molecule in the sample is repeatedly exposed to the maxima, which exacerbates bleaching. Here, we introduce MINFIELD, a strategy for fundamentally reducing bleaching in STED/RESOLFT nanoscopy through restricting the scanning to subdiffraction-sized regions. By safeguarding the molecules from the intensity of the maxima and exposing them only to the lower intensities (around the minima) needed for the off-switching (on-switching), MINFIELD largely avoids detrimental transitions to higher molecular states. A bleaching reduction by up to 100-fold is demonstrated. Recording nanobody-labeled nuclear pore complexes in Xenopus laevis cells showed that MINFIELD-STED microscopy resolved details separated by <25 nm where conventional scanning failed to acquire sufficient signal. PMID:28193881

DNA Hairpins Containing the Cytidine Analog Pyrrolo-dC: Structural, Thermodynamic, and Spectroscopic Studies

PubMed Central

Zhang, Xu; Wadkins, Randy M.

2009-01-01

Structures formed by single-strand DNA have become increasingly interesting because of their roles in a number of biological processes, particularly transcription and its regulation. Of particular importance is the fact that antitumor drugs such as Actinomycin D can selectively bind DNA hairpins over fully paired, double-strand DNA. A new fluorescent base analog, pyrrolo-deoxycytidine (PdC), can now be routinely incorporated into single-strand DNA. The fluorescence of PdC is particularly useful for studying the formation of single-strand DNA in regions of double-strand DNA. The fluorescence is quenched when PdC is paired with a complementary guanine residue, and thus is greatly enhanced upon formation of single-strand DNA. Hence, any process that results in melting or opening of DNA strands produces an increase in the fluorescence intensity of this base analog. In this study we measured the structural effects of incorporating PdC into DNA hairpins, and the effect of this incorporation on the binding of the hairpins by a fluorescent analog of the drug Actinomycin D. Two hairpin DNAs were used: one with PdC in the stem (basepaired) and one with PdC in the loop (unpaired). The thermal stability, 7-aminoactinomycin D binding, and three-dimensional structures of PdC incorporated into these DNA hairpins were all quite similar as compared to the hairpins containing an unmodified dC residue. Fluorescence lifetime measurements indicate that two lifetimes are present in PdC, and that the increase in fluorescence of the unpaired PdC residue compared to the basepaired PdC is due to an increase in the contribution of the longer lifetime to the average fluorescence lifetime. Our data indicate that PdC can be used effectively to differentiate paired and unpaired bases in DNA hairpin secondary structures, and should be similarly applicable for related structures such as cruciforms and quadruplexes. Further, our data indicate that PdC can act as a fluorescence resonance energy transfer donor for the fluorescent drug 7-aminoactinomycin D. PMID:19254547

DNA hairpins containing the cytidine analog pyrrolo-dC: structural, thermodynamic, and spectroscopic studies.

PubMed

Zhang, Xu; Wadkins, Randy M

2009-03-04

Structures formed by single-strand DNA have become increasingly interesting because of their roles in a number of biological processes, particularly transcription and its regulation. Of particular importance is the fact that antitumor drugs such as Actinomycin D can selectively bind DNA hairpins over fully paired, double-strand DNA. A new fluorescent base analog, pyrrolo-deoxycytidine (PdC), can now be routinely incorporated into single-strand DNA. The fluorescence of PdC is particularly useful for studying the formation of single-strand DNA in regions of double-strand DNA. The fluorescence is quenched when PdC is paired with a complementary guanine residue, and thus is greatly enhanced upon formation of single-strand DNA. Hence, any process that results in melting or opening of DNA strands produces an increase in the fluorescence intensity of this base analog. In this study we measured the structural effects of incorporating PdC into DNA hairpins, and the effect of this incorporation on the binding of the hairpins by a fluorescent analog of the drug Actinomycin D. Two hairpin DNAs were used: one with PdC in the stem (basepaired) and one with PdC in the loop (unpaired). The thermal stability, 7-aminoactinomycin D binding, and three-dimensional structures of PdC incorporated into these DNA hairpins were all quite similar as compared to the hairpins containing an unmodified dC residue. Fluorescence lifetime measurements indicate that two lifetimes are present in PdC, and that the increase in fluorescence of the unpaired PdC residue compared to the basepaired PdC is due to an increase in the contribution of the longer lifetime to the average fluorescence lifetime. Our data indicate that PdC can be used effectively to differentiate paired and unpaired bases in DNA hairpin secondary structures, and should be similarly applicable for related structures such as cruciforms and quadruplexes. Further, our data indicate that PdC can act as a fluorescence resonance energy transfer donor for the fluorescent drug 7-aminoactinomycin D.

Photoinhibition of Photosystems I and II Using Chlorophyll Fluorescence Measurements

ERIC Educational Resources Information Center

Quiles, Maria Jose

2005-01-01

In this study the photoinhibition of photosystems (PS) I and II caused by exposure to high intensity light in oat ("Avena sativa," var Prevision) is measured by the emission of chlorophyll fluorescence in intact leaves adapted to darkness. The maximal quantum yield of PS II was lower in plants grown under high light intensity than in plants grown…

Rapid ELISA Using a Film-Stack Reaction Field with Micropillar Arrays

PubMed Central

Suzuki, Yuma; Morioka, Kazuhiro; Ohata, Soichiro; Nakajima, Hizuru; Uchiyama, Katsumi; Yang, Ming

2017-01-01

A film-stack reaction field with a micropillar array using a motor stirrer was developed for the high sensitivity and rapid enzyme-linked immunosorbent assay (ELISA) reaction. The effects of the incubation time of a protein (30 s, 5 min, and 10 min) on the fluorescence intensity in ELISAs were investigated using a reaction field with different micropillar array dimensions (5-µm, 10-µm and 50-µm gaps between the micropillars). The difference in fluorescence intensity between the well with the reaction field of 50-µm gap for the incubation time of 30 s and the well without the reaction field with for incubation time of 10 min was 6%. The trend of the fluorescence intensity in the gap between the micro pillars in the film-stack reaction field was different between the short incubation time and the long incubation time. The theoretical analysis of the physical parameters related with the biomolecule transport indicated that the reaction efficiency defined in this study was the dominant factor determining the fluorescence intensity for the short incubation time, whereas the volumetric rate of the circulating flow through the space between films and the specific surface area were the dominant factors for the long incubation time. PMID:28696378

Rapid ELISA Using a Film-Stack Reaction Field with Micropillar Arrays.

PubMed

Suzuki, Yuma; Morioka, Kazuhiro; Ohata, Soichiro; Shimizu, Tetsuhide; Nakajima, Hizuru; Uchiyama, Katsumi; Yang, Ming

2017-07-11

A film-stack reaction field with a micropillar array using a motor stirrer was developed for the high sensitivity and rapid enzyme-linked immunosorbent assay (ELISA) reaction. The effects of the incubation time of a protein (30 s, 5 min, and 10 min) on the fluorescence intensity in ELISAs were investigated using a reaction field with different micropillar array dimensions (5-µm, 10-µm and 50-µm gaps between the micropillars). The difference in fluorescence intensity between the well with the reaction field of 50-µm gap for the incubation time of 30 s and the well without the reaction field with for incubation time of 10 min was 6%. The trend of the fluorescence intensity in the gap between the micro pillars in the film-stack reaction field was different between the short incubation time and the long incubation time. The theoretical analysis of the physical parameters related with the biomolecule transport indicated that the reaction efficiency defined in this study was the dominant factor determining the fluorescence intensity for the short incubation time, whereas the volumetric rate of the circulating flow through the space between films and the specific surface area were the dominant factors for the long incubation time.

Spectral and Temporal Properties of the Alpha and Beta Subunits and (alpha Beta) Monomer Isolated from Nostoc SP. Using Picosecond Laser Spectroscopy.

NASA Astrophysics Data System (ADS)

Dagen, Aaron J.

1985-12-01

The fluorescence decay profiles, relative quantum yield and transmission of the (alpha), (beta) and ((alpha)(beta)) complexes from phycoerythrin isolated from the photosynthetic antenna system of Nostoc sp. and measured by single picosecond laser spectroscopic techniques is studied. The fluorescence decay profiles of all three complexes are found to be intensity independent for the intensity range investigated ((TURN)4 x 10('13) to (TURN)4 x 10('15) photons-cm('-2) per pulse). The apparent decrease in the relative quantum yield of all three complexes as intensity increases is offset by a corresponding increase in the relative transmission. This evidence, along with the intensity independent fluorescence kinetics, suggests that exciton annihilation is absent in these complexes. The decay profiles are fit to models assuming energy transfer amongst fluorescing chromophores. The intraprotein transfer rate is found to be 100 ps in the (alpha) subunit, 666 ps in the (beta) subunit. Constraining these rates to be identical in the monomer results in explaining the monomer kinetics by an increase in the nonradiative rate of the f(,(beta)) chromophore, an apparent result of aggregation effects.

Spectral and temporal properties of the alpha and beta subunits and (alpha beta) monomer isolated from Nostoc sp. using picosecond laser spectroscopy

NASA Astrophysics Data System (ADS)

Dagen, A. J.

1985-12-01

The fluorescence decay profiles, relative quantum yield and transmission of the alpha, beta and (alpha beta) complexes from phycoerythrin isolated from the photosynthetic antenna system of Nostoc sp. and measured by single picosecond laser spectroscopic techniques is studied. The fluorescence decay profiles of all three complexes are found to be intensity independent for the intensity range investigated (approx. 4x10 to the 13th power to 4x10 to the 15th power photons/sq cm per pulse). The apparent decrease in the relative quantum yield of all three complexes as intensity increases is offset by a corresponding increase in the relative transmission. This evidence, along with the intensity independent fluorescence kinetics, suggests that exciton annihilation is absent in these complexes. The decay profiles are fit to models assuming energy transfer amongst fluorescing chromophores. The intraprotein transfer rate is found to be 100 ps in the alpha subunit, 666 ps in the beta subunit. Constraining these rates to be identical in the monomer results in explaining the monomer kinetics by an increase in the nonradiative rate of the f beta chromophore, an apparent result of aggregation effects.

Characterization of dissolved organic matter in a submerged membrane bioreactor by using three-dimensional excitation and emission matrix fluorescence spectroscopy.

PubMed

Wang, Zhiwei; Wu, Zhichao; Tang, Shujuan

2009-04-01

Three-dimensional excitation-emission matrix (EEM) fluorescence spectroscopy was employed to characterize dissolved organic matter (DOM) in a submerged membrane bioreactor (MBR). Three fluorescence peaks could be identified from the EEM fluorescence spectra of the DOM samples in the MBR. Two peaks were associated with the protein-like fluorophores, and the third was related to the visible humic acid-like fluorophores. Only two main peaks were observed in the EEM fluorescence spectra of the extracellular polymeric substance (EPS) samples, which were due to the fluorescence of protein-like and humic acid-like matters, respectively. However, the EEM fluorescence spectra of membrane foulants were observed to have three peaks. It was also found that the dominant fluorescence substances in membrane foulants were protein-like substances, which might be due to the retention of proteins in the DOM and/or EPS in the MBR by the fine pores of the membrane. Quantitative analysis of the fluorescence spectra including peak locations, fluorescence intensity, and different peak intensity ratios and the fluorescence regional integration (FRI) analysis were also carried out in order to better understand the similarities and differences among the EEM spectra of the DOM, EPS, and membrane foulant samples and to further provide an insight into membrane fouling caused by the fluorescence substances in the DOM in submerged MBRs.

Analysis of the 3D structure og agglutinated erythrocyte using CellScan and confocal microscopy: characterization by FLIM-FRET

NASA Astrophysics Data System (ADS)

Riquelme, Bibiana D.; Dumas, Dominique; Valverde de Rasia, Juana; Rasia, Rodolfo J.; Stoltz, Jean Francois

2003-10-01

We report the adhesion of human erythrocyte membranes mediated by monoclonal antibodies anti-glycophorin. The distribution of the linked antibodies on membrane was identified with selective fluorescence labels. To analyze the antibody distribution on interfacial region between two cells agglutinated and on its surface, three types of fluorescence marked strategy were evaluated. The 3D images were obtained in a CellScan and Confocal Laser Scanning Microscopy CLSM. We considered the FRET signal to characterize the agglutination of Red Blood Cells (RBC) by specific monoclonal antibodies (anti-glycophorin A or B). The fluorescence labeling demonstrated that distribution of antibody on erythrocyte membranes is not homogeneous. The fluorescence intensity on contact region in the agglutinated is bigger than the intensity on exterior surface. Tentatively, we interpreted these intensity differences in terms of the mobility of antibody linked to the glycocalix on cell surface. Such mobility has a large consequence in the morphology of cellular agglutinated.

Highly sensitive spectrofluorimetric determination of trace amounts NADP using Europium ion-doxycycline complex probe

NASA Astrophysics Data System (ADS)

Peng, Qian; Hou, Faju; Jiang, Chongqiu

2006-09-01

A new spectrofluorimetric method was developed for determination of trace amount of Coenzyme II (NADP). Using europium ion-doxycycline (DC) as a fluorescent probe, in the buffer solution of pH 8.44, NADP can remarkably enhance the fluorescence intensity of the Eu 3+-DC complex at λ = 612 nm and the enhanced fluorescence intensity is in proportion to the concentration of NADP. Optimum conditions for the determination of NADP were also investigated. The dynamic range for the determination of NADP is 3.3 × 10 -7 to 6.1 × 10 -6 mol l -1 with detection limit of 6.8 × 10 -8 mol l -1. This method is simple, practical and relatively free interference from coexisting substances and can be successfully applied to determination of NADP in synthetic water samples and in serum samples. Moreover, the enhancement mechanisms of the fluorescence intensity in the Eu 3+-DC system and the Eu 3+-DC-NADP system have been also discussed.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rout, Dipak; Vijaya, R.; Centre for Lasers and Photonics, Indian Institute of Technology Kanpur, Kanpur 208016

Well-ordered opaline photonic crystals are grown by inward growing self-assembly method from Rhodamine B dye-doped polystyrene colloids. Subsequent to self-assembly, the crystals are infiltrated with gold nanoparticles of 40 nm diameter. Measurements of the stopband features and photoluminescence intensity from these crystals are supplemented by fluorescence decay time analysis. The fluorescence decay times from the dye-doped photonic crystals before and after the infiltration are dramatically different from each other. A lowered fluorescence decay time was observed for the case of gold infiltrated crystal along with an enhanced emission intensity. Double-exponential decay nature of the fluorescence from the dye-doped crystal gets convertedmore » into single-exponential decay upon the infiltration of gold nanoparticles due to the resonant radiative process resulting from the overlap of the surface plasmon resonance with the emission spectrum. The influence of localized surface plasmon due to gold nanoparticles on the increase in emission intensity and decrease in decay time of the emitters is established.« less

[The spectroscopic studies on the binding of Al(III) to EHPG].

PubMed

Li, Ying-qi; Bai, Hai-jing; Yang, Bin-sheng

2002-06-01

Interaction of ethylene-N,N'-bis(o-hydioxyphenylglycine) (EHPG) with Al3+ has been investigated by both UV difference and fluorescent spectra. Both results show that the molar ration of the complex is most likely 1:1. Aluminum binding produces peaks at 235 and 291 nm. The molar absorptivity of aluminum ions to EHPG at 235 nm is 1.27 x 10(4) cm-1.mol-1.L. The conditional stability constant for Al3+ binding to EHPG is determined to be IgK = 14.20 +/- 0.03 in 0.1 mol.L-1 Hepes buffer at room temperature, pH 7.4 by UV difference spectra. At the same condition, the fluorescent intensity of EHPG at 310 nm has been monitored. In result, the fluorescent intensity of EHPG at 310 nm is decreased with the addition of Al3+. Then the quench of the fluorescent intensity is ascribed to deprotonated phenolic groups coordinated to aluminum ions.

Joint level-set and spatio-temporal motion detection for cell segmentation.

PubMed

Boukari, Fatima; Makrogiannis, Sokratis

2016-08-10

Cell segmentation is a critical step for quantification and monitoring of cell cycle progression, cell migration, and growth control to investigate cellular immune response, embryonic development, tumorigenesis, and drug effects on live cells in time-lapse microscopy images. In this study, we propose a joint spatio-temporal diffusion and region-based level-set optimization approach for moving cell segmentation. Moving regions are initially detected in each set of three consecutive sequence images by numerically solving a system of coupled spatio-temporal partial differential equations. In order to standardize intensities of each frame, we apply a histogram transformation approach to match the pixel intensities of each processed frame with an intensity distribution model learned from all frames of the sequence during the training stage. After the spatio-temporal diffusion stage is completed, we compute the edge map by nonparametric density estimation using Parzen kernels. This process is followed by watershed-based segmentation and moving cell detection. We use this result as an initial level-set function to evolve the cell boundaries, refine the delineation, and optimize the final segmentation result. We applied this method to several datasets of fluorescence microscopy images with varying levels of difficulty with respect to cell density, resolution, contrast, and signal-to-noise ratio. We compared the results with those produced by Chan and Vese segmentation, a temporally linked level-set technique, and nonlinear diffusion-based segmentation. We validated all segmentation techniques against reference masks provided by the international Cell Tracking Challenge consortium. The proposed approach delineated cells with an average Dice similarity coefficient of 89 % over a variety of simulated and real fluorescent image sequences. It yielded average improvements of 11 % in segmentation accuracy compared to both strictly spatial and temporally linked Chan-Vese techniques, and 4 % compared to the nonlinear spatio-temporal diffusion method. Despite the wide variation in cell shape, density, mitotic events, and image quality among the datasets, our proposed method produced promising segmentation results. These results indicate the efficiency and robustness of this method especially for mitotic events and low SNR imaging, enabling the application of subsequent quantification tasks.

Detecting peanuts inoculated with toxigenic and atoxienic Aspergillus flavus strains with fluorescence hyperspectral imagery

NASA Astrophysics Data System (ADS)

Xing, Fuguo; Yao, Haibo; Hruska, Zuzana; Kincaid, Russell; Zhu, Fengle; Brown, Robert L.; Bhatnagar, Deepak; Liu, Yang

2017-05-01

Aflatoxin contamination in peanut products has been an important and long-standing problem around the world. Produced mainly by Aspergillus flavus and Aspergillus parasiticus, aflatoxins are the most toxic and carcinogenic compounds among toxins. This study investigated the application of fluorescence visible near-infrared (VNIR) hyperspectral images to assess the spectral difference between peanut kernels inoculated with toxigenic and atoxigenic inocula of A. flavus and healthy kernels. Peanut kernels were inoculated with NRRL3357, a toxigenic strain of A. flavus, and AF36, an atoxigenic strain of A. flavus, respectively. Fluorescence hyperspectral images under ultraviolet (UV) excitation were recorded on peanut kernels with and without skin. Contaminated kernels exhibited different fluorescence features compared with healthy kernels. For the kernels without skin, the inoculated kernels had a fluorescence peaks shifted to longer wavelengths with lower intensity than healthy kernels. In addition, the fluorescence intensity of peanuts without skin was higher than that of peanuts with skin (10 times). The fluorescence spectra of kernels with skin are significantly different from that of the control group (p<0.001). Furthermore, the fluorescence intensity of the toxigenic, AF3357 peanuts with skin was lower than that of the atoxigenic AF36 group. Discriminate analysis showed that the inoculation group can be separated from the controls with 100% accuracy. However, the two inoculation groups (AF3357 vis AF36) can be separated with only ∼80% accuracy. This study demonstrated the potential of fluorescence hyperspectral imaging techniques for screening of peanut kernels contaminated with A. flavus, which could potentially lead to the production of rapid and non-destructive scanning-based detection technology for the peanut industry.

Effects of Anisotropic Excitation in Laser-Induced Fluorescence Spectroscopy (LIFS)

NASA Astrophysics Data System (ADS)

Fujimoto, Takashi; Goto, Chiaki; Uetani, Yasunori; Fukuda, Kuniya

1985-07-01

Various features of the effect of alignment in the upper-level population on the observed emission-line intensity, i.e., the spatially-anisotropic intensity distribution and polarization, are demonstrated using laser-induced fluorescence spectroscopy on the neon 2p53s-2p53p transitions in a plasma. Disalignment by atomic collision is observed on the 2p2 level, and its rate coefficient is determined as (1.70± 0.03)× 10-10 cm3s-1. The case of hyperfine-structure lines is discussed. Polarization is observed in the hydrogen Balmer α line fluorescence following the laser excitation of the same transition. Conditions are given under which the alignment effect is eliminated or can be neglected. Cases of unpolarized-light excitation and high-intensity excitation are discussed.

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

PubMed

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.

The impact of fluorescent dyes on the performances of polystyrene-based plastic scintillators

NASA Astrophysics Data System (ADS)

Zhu, Jun; Deng, Cheng; Jiang, Huimin; Zheng, Zhanlong; Gong, Rui; Bi, Yutie; Zhang, Lin; Lin, Runxiong

2016-11-01

To investigate the influence of both the first luminescent additive and the wavelength-shifter on the performance of plastic scintillator, a series of polystyrene-based scintillator had been prepared by thermal polymerization. Three first luminescent additives (PPO, p-TP and b-PBD) and four wavelength-shifters (POPOP, Bis-MSB, Me-MSB and DPA) were added to the scintillators respectively. The comparison results showed that PPO and POPOP were the most adequate fluorescent dyes for the polystyrene-based plastic scintillator. Moreover, with the increase of the concentration of PPO and POPOP, the fluorescence intensity and light yield were increased firstly and then decreased. The plastic scintillator containing 2% PPO and 0.02% POPOP had the highest fluorescence intensity and light yield.

Development of a TiO2/SiO2 waveguide-mode chip for an ultraviolet near-field fluorescence sensor.

PubMed

Kuroda, Chiaki; Nakai, Midori; Fujimaki, Makoto; Ohki, Yoshimichi

2018-03-19

Aimed at detecting fluorescent-labeled biological substances sensitively, a sensor that utilizes near-field light has attracted much attention. According to our calculations, a planar structure composed of two dielectric layers can enhance the electric field of UV near-field light effectively by inducing waveguide-mode (WM) resonance. The fluorescence intensity obtainable by a WM chip with an optimized structure is 5.5 times that obtainable by an optimized surface plasmon resonance chip. We confirmed the above by making a WM chip consisting of TiO 2 and SiO 2 layers on a silica glass substrate and by measuring the fluorescence intensity of a solution of quantum dots dropped on the chip.

Recovering fluorophore concentration profiles from confocal images near lateral refractive index step changes.

PubMed

Jonášová, Eleonóra Parelius; Bjørkøy, Astrid; Stokke, Bjørn Torger

2016-12-01

Optical aberrations due to refractive index mismatches occur in various types of microscopy due to refractive differences between the sample and the immersion fluid or within the sample. We study the effects of lateral refractive index differences by fluorescence confocal laser scanning microscopy due to glass or polydimethylsiloxane cuboids and glass cylinders immersed in aqueous fluorescent solution, thereby mimicking realistic imaging situations in the proximity of these materials. The reduction in fluorescence intensity near the embedded objects was found to depend on the geometry and the refractive index difference between the object and the surrounding solution. The observed fluorescence intensity gradients do not reflect the fluorophore concentration in the solution. It is suggested to apply a Gaussian fit or smoothing to the observed fluorescence intensity gradient and use this as a basis to recover the fluorophore concentration in the proximity of the refractive index step change. The method requires that the reference and sample objects have the same geometry and refractive index. The best results were obtained when the sample objects were also used for reference since small differences such as uneven surfaces will result in a different extent of aberration.

The ratioed image film thickness meter

NASA Astrophysics Data System (ADS)

Husen, Nicholas M.; Liu, Tianshu; Sullivan, John P.

2018-06-01

A technique for measuring the thickness of a fluorescent oil film is presented. Incident light is cast upon the oil film and the intensity of the luminescent signal from the fluorescent dye is ratioed with the intensity of the incident light which is scattered from the surface of the model. The quotient is independent of the intensity of the incident light and proportional to the film thickness. Experiments are presented supporting that for sufficiently thin films the ratio is independent of the intensity of the incident light as well as independent of the angle from which the experiment is imaged and the angle from which the incident light is cast.

Fluorescence Approaches to Growing Macromolecule Crystals

NASA Technical Reports Server (NTRS)

Pusey, Marc; Forsythe, Elizabeth; Achari, Aniruddha

2006-01-01

Trace fluorescent labeling, typically < 1%, can be a powerful aid in macromolecule crystallization. Precipitation concentrates a solute, and crystals are the most densely packed solid form. The more densely packed the fluorescing material, the more brightly the emission from it, and thus fluorescence intensity of a solid phase is a good indication of whether one has crystals or not. The more brightly fluorescing crystalline phase is easily distinguishable, even when embedded in an amorphous precipitate. This approach conveys several distinct advantages: one can see what the protein is doing in response to the imposed conditions, and distinguishing between amorphous and microcrystalline precipitated phases are considerably simpler. The higher fluorescence intensity of the crystalline phase led us to test if we could derive crystallization conditions from screen outcomes which had no obvious crystalline material, but simply "bright spots" in the precipitated phase. Preliminary results show that the presence of these bright spots, not observable under white light, is indeed a good indicator of potential crystallization conditions.

Determination of adenosine disodium triphosphate (ATP) using oxytetracycline-Eu 3+ as a fluorescence probe by spectrofluorimetry

NASA Astrophysics Data System (ADS)

Hou, Faju; Miao, Yanhong; Jiang, Chongqiu

2005-10-01

A new spectrofluorimetric method was developed for determination of adenosine disodium triphosphate (ATP). We studied the interactions between oxytetracycline (OTC)-Eu 3+ complex and adenosine disodium triphosphate (ATP) by using UV-vis absorption and fluorescence spectra. Using oxytetracycline (OTC)-Eu 3+ as a fluorescence probe, under the optimum conditions, ATP can remarkably enhance the fluorescence intensity of the OTC-Eu 3+ complex at λ = 612 nm and the enhanced fluorescence intensity of Eu 3+ ion is in proportion to the concentration of ATP. Optimum conditions for the determination of ATP were also investigated. The linear ranges for ATP are 8.00 × 10 -8-1.50 × 10 -6 mol L -1 with detection limits of 2.67 × 10 -9 mol L -1. This method is simple, practical and relatively free interference from coexisting substances and can be successfully applied to determination of ATP in samples. The mechanism of fluorescence enhancement between oxytetracycline (OTC)-Eu 3+ complex and ATP was also studied.

In vivo optical imaging of dihydroethidium oxidation in the mouse brain employing fluorescence intensity and lifetime contrast

NASA Astrophysics Data System (ADS)

Hall, David J.; Han, Sung-Ho; Dugan, Laura

2009-02-01

Reactive oxygen species (ROS) are believed to be involved in many diseases and injuries to the brain, but the molecular processes are not well understood due to a lack of in vivo imaging techniques to evaluate ROS. The fluorescent oxidation products of dihydroethidium (DHE) can monitor ROS production in vivo. Here we demonstrate the novel optical imaging of brain in live mice to measure ROS production via generation of fluorescent DHE oxidation products (ox-DHE) by ROS. We show that in Sod2+/- mice, which have partial loss of a key antioxidant enzyme, superoxide dismutase-2, that ox-DHE fluorescence intensity was significantly higher than in hSOD1 mice, which have four-fold overexpression of superoxide dismutase-1 activity, which had almost no ox-DHE fluorescence, confirming specificity of ox-DHE to ROS production. The DHE oxidation products were also confirmed by detecting a characteristic fluorescence lifetime of the oxidation product, which was validated with ex vivo measurements.

Generation of fluorescent silver nanoclusters in reverse micelles using gamma irradiation: low vs. high dosages and spectral evolution with time

NASA Astrophysics Data System (ADS)

Martin, Brett D.; Fontana, Jake; Wang, Zheng; Trammell, Scott A.

2015-04-01

Reverse micelles (RMs) containing aqueous solutions of Ag+ ions in their core produce fluorescent Ag nanoclusters (NCs), upon exposure to gamma irradiation. The fluorescence spectra of the NCs evolve over days to weeks after the exposure, and usually show large increases in intensity. Responses of as high as 2.8 × 104 CPS/Gy were reached. A dosage as low as 0.5 Gy (10 % of the lethal dosage for humans) produces NCs having fluorescence intensities higher than background. The RMs can be employed in novel gamma radiation detectors with appearance of fluorescence indicating that radiation was once present. In applications involving detection and tracking of fissile materials, the evolution of the fluorescence spectra over time may provide additional information about the radiation source. A two-phase liquid system is used for RM formation in a simple procedure. It is likely that this synthesis method may be adapted to produce NCs from other metal ions.

Computer Modeling of the Structure and Spectra of Fluorescent Proteins

PubMed Central

Grigorenko, B.L.; Savitsky, A.P.

2009-01-01

Fluorescent proteins from the family of green fluorescent proteins are intensively used as biomarkers in living systems. The chromophore group based on the hydroxybenzylidene-imidazoline molecule, which is formed in nature from three amino-acid residues inside the protein globule and well shielded from external media, is responsible for light absorption and fluorescence. Along with the intense experimental studies of the properties of fluorescent proteins and their chromophores by biochemical, X-ray, and spectroscopic tools, in recent years, computer modeling has been used to characterize their properties and spectra. We present in this review the most interesting results of the molecular modeling of the structural parameters and optical and vibrational spectra of the chromophorecontaining domains of fluorescent proteins by methods of quantum chemistry, molecular dynamics, and combined quantum-mechanical-molecular-mechanical approaches. The main emphasis is on the correlation of theoretical and experimental data and on the predictive power of modeling, which may be useful for creating new, efficient biomarkers. PMID:22649601

Novel strategy combining SYBR Green I with carbon nanotubes for highly sensitive detection of Salmonella typhimurium DNA.

PubMed

Mao, Pingdao; Ning, Yi; Li, Wenkai; Peng, Zhihui; Chen, Yongzhe; Deng, Le

2014-01-10

A simple, selective, sensitive and label-free fluorescent method for detecting trpS-harboring Salmonella typhimurium was developed in this study. This assay used the non-covalent interaction of single-stranded DNA (ssDNA) probes with SWNTs, since SWNTs can quench fluorescence. Fluorescence recovery (78% with 1.8 nM target DNA) was detected in the presence of target DNA as ssDNA probes detached from SWNTs hybridized with target DNA, and the resulting double-stranded DNA (dsDNA) intercalated with SYBR Green I (SG) dyes. The increasing fluorescence intensity reached 4.54-fold. In contrast, mismatched oligonucleotides (1- or 3-nt difference to the target DNA) did not contribute to significant fluorescent recovery, which demonstrated the specificity of the assay. The increasing fluorescence intensity increased 3.15-fold when purified PCR products containing complementary sequences of trpS gene were detected. These results confirmed the ability to use this assay for detecting real samples. Copyright © 2013 Elsevier Inc. All rights reserved.

Plasmonic enhancement of cyanine dyes for near-infrared light-triggered photodynamic/photothermal therapy and fluorescent imaging

NASA Astrophysics Data System (ADS)

Lu, Mindan; Kang, Ning; Chen, Chuan; Yang, Liuqing; Li, Yang; Hong, Minghui; Luo, Xiangang; Ren, Lei; Wang, Xiumin

2017-11-01

Near-infrared (NIR) triggered cyanine dyes have attracted considerable attention in multimodal tumor theranostics. However, NIR cyanine dyes used in tumor treatment often suffer from low fluorescence intensity and weak singlet oxygen generation efficiency, resulting in inadequate diagnostic and therapy efficacy for tumors. It is still a great challenge to improve both the photodynamic therapy (PDT) and fluorescent imaging (FLI) efficacy of cyanine dyes in tumor applications. Herein, a novel multifunctional nanoagent AuNRs@SiO2-IR795 was developed to realize the integrated photothermal/photodynamic therapy (PTT/PDT) and FLI at a very low dosage of IR795 (0.4 μM) based on metal-enhanced fluorescence (MEF) effects. In our design, both the fluorescence intensity and reactive oxygen species of AuNRs@SiO2-IR795 nanocomposites were significantly enhanced up to 51.7 and 6.3 folds compared with free IR795, owing to the localized surface plasmon resonance band of AuNRs overlapping with the absorption or fluorescence emission band of the IR795 dye. Under NIR laser irradiation, the cancer cell inhibition efficiency in vitro with synergetic PDT/PTT was up to 82.3%, compared with 10.3% for free IR795. Moreover, the enhanced fluorescence intensity of our designed nanocomposites was helpful to track their behavior in tumor cells. Therefore, our designed nanoagents highlight the applications of multimodal diagnostics and therapy in tumors based on MEF.

Calcium Sensitive Fluorescent Dyes Fluo-4 and Fura Red under Pressure: Behaviour of Fluorescence and Buffer Properties under Hydrostatic Pressures up to 200 MPa

PubMed Central

Vass, H.; Reischl, B.; Allen, R. J.; Friedrich, O.

2016-01-01

The fluorescent Ca2+ sensitive dyes Fura Red (ratiometric) and Fluo-4 (non-ratiometric) are widely utilized for the optical assessment of Ca2+ fluctuations in vitro as well as in situ. The fluorescent behavior of these dyes is strongly depends on temperature, pH, ionic strength and pressure. It is crucial to understand the response of these dyes to pressure when applying calcium imaging technologies in the field of high pressure bioscience. Therefore, we use an optically accessible pressure vessel to pressurize physiological Ca2+-buffered solutions at different fixed concentrations of free Ca2+ (1 nM to 25.6 μM) and a specified dye concentration (12 μM) to pressures of 200 MPa, and record dye fluorescence intensity. Our results show that Fluo-4 fluorescence intensity is reduced by 31% per 100 MPa, the intensity of Fura Red is reduced by 10% per 100 MPa. The mean reaction volume for the dissociation of calcium from the dye molecules Δdv¯ is determined to -17.8 ml mol-1 for Fluo-4 and -21.3 ml mol-1 for Fura Red. Additionally, a model is presented that is used to correct for pressure-dependent changes in pH and binding affinity of Ca2+ to EGTA, as well as to determine the influence of these changes on dye fluorescence. PMID:27764134

Diurnal Variability in Chlorophyll-a, Carotenoids, CDOM and SO42− Intensity of Offshore Seawater Detected by an Underwater Fluorescence-Raman Spectral System

PubMed Central

Chen, Jing; Ye, Wangquan; Guo, Jinjia; Luo, Zhao; Li, Ying

2016-01-01

A newly developed integrated fluorescence-Raman spectral system (λex = 532 nm) for detecting Chlorophyll-a (chl-a), Chromophoric Dissolved Organic Matter (CDOM), carotenoids and SO42− in situ was used to successfully investigate the diurnal variability of all above. Simultaneously using the integration of fluorescence spectroscopy and Raman spectroscopy techniques provided comprehensive marine information due to the complementarity between the different excitation mechanisms and different selection rules. The investigation took place in offshore seawater of the Yellow Sea (36°05′40′′ N, 120°31′32′′ E) in October 2014. To detect chl-a, CDOM, carotenoids and SO42−, the fluorescence-Raman spectral system was deployed. It was found that troughs of chl-a and CDOM fluorescence signal intensity were observed during high tides, while the signal intensity showed high values with larger fluctuations during ebb-tide. Chl-a and carotenoids were influenced by solar radiation within a day cycle by different detection techniques, as well as displaying similar and synchronous tendency. CDOM fluorescence cause interference to the measurement of SO42−. To avoid such interference, the backup Raman spectroscopy system with λex = 785 nm was employed to detect SO42− concentration on the following day. The results demonstrated that the fluorescence-Raman spectral system has great potential in detection of chl-a, carotenoids, CDOM and SO42− in the ocean. PMID:27420071

Novel single-stranded DNA binding protein-assisted fluorescence aptamer switch based on FRET for homogeneous detection of antibiotics.

PubMed

Wang, Ye; Gan, Ning; Zhou, You; Li, Tianhua; Cao, Yuting; Chen, Yinji

2017-01-15

Herein, a smart single-stranded DNA binding protein (SSB)-assisted fluorescence aptamer switch based on fluorescence resonance energy transfer (FRET) was designed. The FRET switch was synthesized by connecting SSB labeled quantum dots (QDs@SSB) as donor with aptamer (apt) labeled gold nanoparticles (AuNPs@apt) as acceptor, and it was employed for detecting chloramphenicol (CAP) in a homogenous solution. In the assay, the interaction between core-shell QDs@SSB and AuNPs@apt leads to a dramatic quenching (turning off). After adding CAP in the detection system, AuNPs@apt can bind the target specifically then separate QDs@SSB with AuNPs@apt-target, resulting in restoring the fluorescence intensity of QDs (turning on). Consequently, the fluorescence intensity recovers and the recovery extent can be used for detection of CAP in homogenous phase via optical responses. Under optimal conditions, the fluorescence intensity increased linearly with increasing concentrations of CAP from 0.005 to 100ngmL -1 . The limit of this fluorescence aptamer switch was around 3pgmL -1 for CAP detection. When the analyte is changed, the assay can be applied to detect other targets only by changing relative aptamer in AuNPs@apt probe. Furthermore, it has potential to be served as a simple, sensitive and portable platform for antibiotic contaminants detection in biological and environmental samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Fluorescence lifetime as a new parameter in analytical cytology measurements

NASA Astrophysics Data System (ADS)

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

1996-05-01

A phase-sensitive flow cytometer has been developed to quantify fluorescence decay lifetimes on fluorochrome-labeled cells/particles. This instrument combines flow cytometry (FCM) and frequency-domain fluorescence spectroscopy measurement principles to provide unique capabilities for making phase-resolved lifetime measurements, while preserving conventional FCM capabilities. Cells are analyzed as they intersect a high-frequency, intensity-modulated (sine wave) laser excitation beam. Fluorescence signals are processed by conventional and phase-sensitive signal detection electronics and displayed as frequency distribution histograms. In this study we describe results of fluorescence intensity and lifetime measurements on fluorescently labeled particles, cells, and chromosomes. Examples of measurements on intrinsic cellular autofluorescence, cells labeled with immunofluorescence markers for cell- surface antigens, mitochondria stains, and on cellular DNA and protein binding fluorochromes will be presented to illustrate unique differences in measured lifetimes and changes caused by fluorescence quenching. This innovative technology will be used to probe fluorochrome/molecular interactions in the microenvironment of cells/chromosomes as a new parameter and thus expand the researchers' understanding of biochemical processes and structural features at the cellular and molecular level.

Fluorescence detection of trace PCB101 based on PITC immobilized on porous AAO membrane.

PubMed

Wang, Meiling; Meng, Guowen; Huang, Qing; Li, Mingtao; Li, Zhongbo; Tang, Chaolong

2011-01-21

A sensitive and selective fluorescent membrane for rapid detection of trace 2,2',4,5,5'-pentachlorinated biphenyl (PCB101) has been achieved by immobilizing the fluorophore phenyl isothiocyanate (PITC) onto porous anodic aluminium oxide (AAO) membrane (denoted as PITC@AAO). The fluorescence of the PITC@AAO membrane is obviously enhanced after titrating the analyte PCB101 into the membrane, being ascribed to the halogen-bonding interaction between the fluorophore PITC and the analyte PCB101. The fluorescence intensity increases with the PCB101 concentration in the low range below 1 ppm, and there exists an approximate linear relationship between the relative fluorescence intensity and the PCB101 concentration in the low range of 1-6 ppb. Moreover, the PITC@AAO membrane shows good selectivity; for example, it is insensitive to common structural analogs (polychlorinated aromatics). The mechanisms of the fluorescence enhancement and the better sensitivity and selectivity of the PITC@AAO membrane to PCB101 than that of PITC/n-hexane solution are also discussed. This work demonstrates that trace (in ppb range) PCBs can be detected by simple fluorescence measurement.

Detection of large deletion in human BRCA1 gene in human breast carcinoma MCF-7 cells by using DNA-Silver Nanoclusters

NASA Astrophysics Data System (ADS)

Borghei, Yasaman-Sadat; Hosseini, Morteza; Ganjali, Mohammad Reza

2018-01-01

Here we describe a label-free detection strategy for large deletion mutation in breast cancer (BC) related gene BRCA1 based on a DNA-silver nanocluster (NC) fluorescence upon recognition-induced hybridization. The specific hybridization of DNA templated silver NCs fluorescent probe to target DNAs can act as effective templates for enhancement of AgNCs fluorescence, which can be used to distinguish the deletion of BRCA1 due to different fluorescence intensities. Under the optimal conditions, the fluorescence intensity of the DNA-AgNCs at emission peaks around 440 nm (upon excitation at 350 nm) increased with the increasing deletion type within a dynamic range from 1.0 × 10-10 to 2.4 × 10-6 M with a detection limit (LOD) of 6.4 × 10-11 M. In this sensing system, the normal type shows no significant fluorescence; on the other hand, the deletion type emits higher fluorescence than normal type. Using this nanobiosensor, we successfully determined mutation using the non-amplified genomic DNAs that were isolated from the BC cell line.

Shedding quantitative fluorescence light on novel regulatory mechanisms in skeletal biomedicine and biodentistry.

PubMed

Lee, Ji-Won; Iimura, Tadahiro

2017-02-01

Digitalized fluorescence images contain numerical information such as color (wavelength), fluorescence intensity and spatial position. However, quantitative analyses of acquired data and their validation remained to be established. Our research group has applied quantitative fluorescence imaging on tissue sections and uncovered novel findings in skeletal biomedicine and biodentistry. This review paper includes a brief background of quantitative fluorescence imaging and discusses practical applications by introducing our previous research. Finally, the future perspectives of quantitative fluorescence imaging are discussed.

Fluorescence spectra of atmospheric aerosol at Adelphi, Maryland, USA: measurement and classification of single particles containing organic carbon

NASA Astrophysics Data System (ADS)

Pinnick, Ronald G.; Hill, Steven C.; Pan, Yong-Le; Chang, Richard K.

We measured laser-induced fluorescence spectra from individual supermicron-sized atmospheric particles drawn into our laboratory at Adelphi, MD, an urban site in the Washington, DC metroplex. A virtural impactor concentrator is used along with an aerodynamic-focusing-nozzle which forms, within an optical chamber, a focused aerosol jet where single aerosol particles can be interrogated on-the-fly with a pulsed 266-nm-wavelength laser. Sample rates are a few liter per minute, and are size dependent. Crossed-diode laser beams indicate when a particle is traversing the sample region and are used to trigger the UV laser to fire and the gated intensified CCD to record the fluorescence spectrum. Our breadboard fluorescence particle spectrometer measures particles in the 3-10 μm diameter size range. Typical trigger rates are a few per second. The usable spectral range is from about 295 to 605 nm. The majority of the particles have very weak fluorescence (on average 8 percent of particles have fluorescence signals above noise). The spectra were grouped using a heirarchical cluster analysis, with parameters chosen so that spectra typically cluster into 4-12 main categories. From the set of all cluster spectra we chose 8 template spectra for reanalyzing all the data. On average, 92 percent (81-94 percent) of the spectra were similar to these templates (using the same thresholds used for the cluster analysis). The major emission bands of the most commonly occurring spectra have peaks: near 460 nm (28 percent of fluorescent particles on average), a very broad hump, and may be humic acids or humic like substances; near 317 nm (on average 24 percent of fluorescent particles); near 321 and 460 nm (a double hump, 12 percent of fluorescent particles); and near 341 nm (8 percent of fluorescent particles). Some of the fluorescence in spectra peaking in the 317-341 nm range is probably from dicyclic aromatics and heterocyclics, including the amino acid tryptophan in biological particles such as bacteria and spores.

Novel 1:1 labeling and purification process for C-terminal thioester and single cysteine recombinant proteins using generic peptidic toolbox reagents.

PubMed

Portal, Christophe F; Seifert, Jan-Marcus; Buehler, Christof; Meisner-Kober, Nicole-Claudia; Auer, Manfred

2014-07-16

We developed a versatile set of chemical labeling reagents which allow dye ligation to the C-terminus of a protein or a single internal cysteine and target purification in a simple two-step process. This simple process results in a fully 1:1 labeled conjugate suitable for all quantitative fluorescence spectroscopy and imaging experiments. We refer to a "generic labeling toolbox" because of the flexibility to choose one of many available dyes, spacers of different lengths and compositions which increase the target solubility, a variety of affinity purification tags, and different cleavage chemistries to release the 1:1 labeled proteins. Studying protein function in vitro or in the context of live cells and organisms is of vital importance in biological research. Although label free detection technologies gain increasing interest in molecular recognition science, fluorescence spectroscopy is still the most often used detection technique for assays and screens both in academic as well as in industrial groups. For generations, fluorescence spectroscopists have labeled their proteins of interest with small fluorescent dyes by random chemical linking on the proteins' exposed lysines and cysteines. Chemical reactions with a certain excess of activated esters or maleimides of longer wavelength dyes hardly ever result in quantitative labeling of the target protein. Most of the time, more than one exposed amino acid side chain reacts. This results in a mixture of dye-protein complexes of different labeling stoichiometries and labeling sites. Only mass spectrometry allows resolving the precise chemical composition of the conjugates. In "classical" ensemble averaging fluorescent experiments, these labeled proteins are still useful, and quantification of, e.g., ligand binding experiments, is achieved via knowledge of the overall protein concentration and a fluorescent signal change which is proportional to the amount of complex formed. With the development of fluorescence fluctuation analysis techniques working at single molecule resolution, like fluorescence correlation spectroscopy (FCS), fluorescence cross correlation spectroscopy (FCCS), fluorescence intensity diffusion analysis (FIDA), etc., it became important to work with homogeneously labeled target proteins. Each molecule participating in a binding equilibrium should be detectable when it freely fluctuates through the confocal focus of a microscope. The measured photon burst for each transition contains information about the size and the stoichiometry of a protein complex. Therefore, it is important to work with reagents that contain an exact number of tracers per protein at identical positions. The ideal fluorescent tracer-protein complex stoichiometry is 1:1. While genetic tags such as fluorescent proteins (FPs) are widely used to detect proteins, FPs have several limitations compared to chemical tags. For example, FPs cannot easily compete with organic dyes in the flexibility of modification and spectral range; moreover, FPs have disadvantages in brightness and photostability and are therefore not ideal for most biochemical single molecule studies. We present the synthesis of a series of exemplaric toolbox reagents and labeling results on three target proteins which were needed for high throughput screening experiments using fluorescence fluctuation analysis at single molecule resolution. On one target, Hu-antigen R (HuR), we demonstrated the activity of the 1:1 labeled protein in ribonucleic acid (RNA) binding, and the ease of resolving the stoichiometry of an RNA-HuR complex using the same dye on protein and RNA by Fluorescence Intensity Multiple Distribution Analysis (FIMDA) detection.

Development of dansyl-modified oligonucleotide probes responding to structural changes in a duplex.

PubMed

Suzuki, Yoshio; Kowata, Keiko; Komatsu, Yasuo

2013-11-15

We have synthesized a nonnucleoside amidite block of dansyl fluorophore to prepare dansyl-modified oligonucleotides (ONTs). The fluorescence intensities of dansyl-ONT specifically increased by the presence of adjacent guanosine residues but, significantly reduced in a dansyl-flipping duplex. These changes were caused by solvatochromism effect due to the number of guanine which is hydrophobic functional group and the external environment of dansyl group. The fluorescence intensities could be plotted as a function of the ONTs concentrations and the increase in the fluorescence was observed to equimolar concentrations of target DNA. This duplex exhibited higher melting temperature relative to the corresponding duplexes containing other base pairs. Similar changes in fluorescence could be detected upon hybridization with complementary RNAs. Thus, the dansyl-modified ONTs provide sequence specific fluorescent probe of DNA and RNA. Copyright © 2013 Elsevier Ltd. All rights reserved.

Plasmonic platforms of self-assembled silver nanostructures in application to fluorescence

PubMed Central

Luchowski, Rafal; Calander, Nils; Shtoyko, Tanya; Apicella, Elisa; Borejdo, Julian; Gryczynski, Zygmunt; Gryczynski, Ignacy

2011-01-01

Fluorescence intensity changes were investigated theoretically and experimentally using self-assembled colloidal structures on silver semitransparent mirrors. Using a simplified quasi-static model and finite element method, we demonstrate that near-field interactions of metallic nanostructures with a continuous metallic surface create conditions that produce enormously enhanced surface plasmon resonances. The results were used to explain the observed enhancements and determine the optimal conditions for the experiment. The theoretical parts of the studies are supported with reports on detailed emission intensity changes which provided multiple fluorescence hot spots with 2–3 orders of enhancements. We study two kinds of the fluorophores: dye molecules and fluorescent nanospheres characterized with similar spectral emission regions. Using a lifetime-resolved fluorescence/reflection confocal microscopy technique, we find that the largest rate for enhancement (~1000-fold) comes from localized areas of silver nanostructures. PMID:21403765

Laser-induced fluorescence imaging of acetone inside evaporating and burning fuel droplets

NASA Astrophysics Data System (ADS)

Shringi, D. S.; Shaw, B. D.; Dwyer, H. A.

2009-01-01

Laser-induced fluorescence was used to visualize acetone fields inside individual droplets of pure acetone as well as droplets composed of methanol or 1-propanol initially mixed with acetone. Droplets were supported on a horizontal wire and two vaporization conditions were investigated: (1) slow evaporation in room air and (2) droplet combustion, which leads to substantially faster droplet surface regression rates. Acetone was preferentially gasified, causing its concentration in droplets to drop in time with resultant decreases in acetone fluorescence intensities. Slowly vaporizing droplets did not exhibit large spatial variations of fluorescence within droplets, indicating that these droplets were relatively well mixed. Ignition of droplets led to significant variations in fluorescence intensities within droplets, indicating that these droplets were not well mixed. Ignited droplets composed of mixtures of 1-propanol and acetone showed large time-varying changes in shapes for higher acetone concentrations, suggesting that bubble formation was occurring in these droplets.

DNA nanostructure-based fluorescence thermometer with silver nanoclusters

NASA Astrophysics Data System (ADS)

Bu, Congcong; Mu, Lixuan; Cao, Xingxing; Chen, Min; She, Guangwei; Shi, Wensheng

2018-07-01

DNA nanostructure-based fluorescence thermometers were fabricated by linking fluorescent silver nanoclusters (AgNCs) and guanine-rich（G-rich）DNA chains via a thermally sensitive DNA stem-loop at terminals 5‧ and 3‧. Variations of temperature alter the distance between the AgNCs and G-rich DNA chain, affecting the interaction between them. As a result, the intensity of fluorescence emission from the AgNCs at 636 nm can be sensitively modulated. It was found that the intensity of such red emission is more temperature sensitive than the equivalent green emission at 543 nm; sensitivity of ‑3.6%/°C was achieved. Through variation of the melting temperature of the DNA stem-loop, the response temperature range of the thermometers could be readily adjusted. Novel DNA nanostructure-based fluorescence thermometers as described in this work are anticipated to be able to measure the temperature of biological systems at small scales—even a single cell.

Contribution of glue layer into epidermis sample fluorescence dynamics

NASA Astrophysics Data System (ADS)

Salomatina, Elena V.; Chernova, Svetlana P.; Pravdin, Alexander B.

2000-04-01

In this work, the temporal behavior of autofluorescence of epidermis samples under UV-irradiation has ben studied. The samples were prepared using surface epidermis stripping technique. Fluorescence spectra and kinetic curves of fluorescence intensity have been obtained. It has been concluded that the glue composition used allows the measurement of epidermis fluorescence dynamics with the first 60 min of experiment.

Evaluation of CDOM sources and their links with water quality in the lakes of Northeast China using fluorescence spectroscopy

NASA Astrophysics Data System (ADS)

Zhao, Ying; Song, Kaishan; Wen, Zhidan; Fang, Chong; Shang, Yingxin; Lv, Lili

2017-07-01

The spatial distributions of the fluorescence intensities Fmax for chromophoric dissolved organic matter (CDOM) components, the fluorescence indices (FI370 and FI310) and their correlations with water quality of 19 lakes in the Songhua River Basin (SHRB) across semiarid regions of Northeast China were examined with the data collected in September 2012 and 2015. The 19 lakes were divided into two groups according to EC (threshold value = 800 μS cm-1): fresh water (N = 13) and brackish water lakes (N = 6). The fluorescent characteristics of CDOM in the 19 lakes were investigated using excitation-emission matrix fluorescence spectroscopy (EEM) coupled with parallel factor (PARAFAC) and multivariate analysis. Two humic-like components (C1 and C3), one tryptophan-like component (C2), and one tyrosine-like component (C4) were identified by PARAFAC. The component C4 was not included in subsequent analyses due to the strong scatter in some colloidal water samples from brackish water lakes. The correlations between Fmax for the three EEM-PARAFAC extracted CDOM components C1-C3, the fluorescence indices (FI370 and FI310) and the water quality parameters (i.e., TN, TP, Chl-a, pH, EC, turbidity (Turb) and dissolved organic carbon (DOC)) were determined by redundancy analysis (RDA). The results of RDA analysis showed that spatial variation in land cover, pollution sources, and salinity/EC gradients in water quality affected Fmax for the fluorescent components C1-C3 and the fluorescence indices (FI370 and FI310). Further examination indicated that the CDOM fluorescent components and the fluorescence indices (FI370 and FI310) did not significantly differ (t-test, p > 0.05) in fresh water (N = 13) and brackish water lakes (N = 6). There was a difference in the distribution of the average Fmax for the CDOM fluorescent components between C1 to C3 from agricultural sources and urban wastewater sources in hypereutrophic brackish water lakes. The Fmax for humic-like components C1 and C3 spatially varied with land cover among the 19 lakes. Our results indicated that the spatial distributions of Fmax for CDOM fluorescent components and their correlations with water quality can be evaluated by EEM-PARAFAC and multivariate analysis among the 19 lakes across semiarid regions of Northeast China, which has potential implication for lakes with similar genesis.

Single-Step Assembly of Multi-Modal Imaging Nanocarriers: MRI and Long-Wavelength Fluorescence Imaging

PubMed Central

Pinkerton, Nathalie M.; Gindy, Marian E.; Calero-DdelC, Victoria L.; Wolfson, Theodore; Pagels, Robert F.; Adler, Derek; Gao, Dayuan; Li, Shike; Wang, Ruobing; Zevon, Margot; Yao, Nan; Pacheco, Carlos; Therien, Michael J.; Rinaldi, Carlos; Sinko, Patrick J.

2015-01-01

MRI and NIR-active, multi-modal Composite NanoCarriers (CNCs) are prepared using a simple, one-step process, Flash NanoPrecipitation (FNP). The FNP process allows for the independent control of the hydrodynamic diameter, co-core excipient and NIR dye loading, and iron oxide-based nanocrystal (IONC) content of the CNCs. In the controlled precipitation process, 10 nm IONCs are encapsulated into poly(ethylene glycol) stabilized CNCs to make biocompatible T2 contrast agents. By adjusting the formulation, CNC size is tuned between 80 and 360 nm. Holding the CNC size constant at an intensity weighted average diameter of 99 ± 3 nm (PDI width 28 nm), the particle relaxivity varies linearly with encapsulated IONC content ranging from 66 to 533 mM-1s-1 for CNCs formulated with 4 to 16 wt% IONC. To demonstrate the use of CNCs as in vivo MRI contrast agents, CNCs are surface functionalized with liver targeting hydroxyl groups. The CNCs enable the detection of 0.8 mm3 non-small cell lung cancer metastases in mice livers via MRI. Incorporating the hydrophobic, NIR dye PZn3 into CNCs enables complementary visualization with long-wavelength fluorescence at 800 nm. In vivo imaging demonstrates the ability of CNCs to act both as MRI and fluorescent imaging agents. PMID:25925128

Visualization of oxygen transfer across the air-water interface using a fluorescence oxygen visualization method.

PubMed

Lee, Minhee

2002-04-01

Oxygen concentration fields in a water body were visualized by the fluorescence oxygen visualization (FOV) method. Pyrenebutyric acid (PBA) was used as a fluorescent indicator of oxygen, and an intensive charge coupled-device (ICCD) camera as an image detector. Sequential images (over 2000 images) of the oxygen concentration field around the surface water of the tank (1 x 1 x 0.75 m3) were produced during the 3 h experiment. From image processing, the accurate pathway of oxygen-rich, cold water at the water surface was also visualized. The amount of oxygen transferred through the air-water interface during the experiment was measured and the oxygen transfer coefficient (K(L)) was determined as 0.22 m/d, which was much higher than that is expected in molecular diffusion. Results suggest that vertical penetration of cold water was the main pathway of oxygen in the water body in the tank. The average velocity of cold water penetrating downward in water body was also measured from consecutive images and the value was 0.3-0.6 mm/s. The FOV method used in this research should have wide application in experimental fluid mechanics and can also provide a phenomenological description of oxygen transfer under physically realizable natural conditions in lakes and reservoirs.

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

NASA Astrophysics Data System (ADS)

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

2015-06-01

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

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

PubMed

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

2018-05-15

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

ANAlyte: A modular image analysis tool for ANA testing with indirect immunofluorescence.

PubMed

Di Cataldo, Santa; Tonti, Simone; Bottino, Andrea; Ficarra, Elisa

2016-05-01

The automated analysis of indirect immunofluorescence images for Anti-Nuclear Autoantibody (ANA) testing is a fairly recent field that is receiving ever-growing interest from the research community. ANA testing leverages on the categorization of intensity level and fluorescent pattern of IIF images of HEp-2 cells to perform a differential diagnosis of important autoimmune diseases. Nevertheless, it suffers from tremendous lack of repeatability due to subjectivity in the visual interpretation of the images. The automatization of the analysis is seen as the only valid solution to this problem. Several works in literature address individual steps of the work-flow, nonetheless integrating such steps and assessing their effectiveness as a whole is still an open challenge. We present a modular tool, ANAlyte, able to characterize a IIF image in terms of fluorescent intensity level and fluorescent pattern without any user-interactions. For this purpose, ANAlyte integrates the following: (i) Intensity Classifier module, that categorizes the intensity level of the input slide based on multi-scale contrast assessment; (ii) Cell Segmenter module, that splits the input slide into individual HEp-2 cells; (iii) Pattern Classifier module, that determines the fluorescent pattern of the slide based on the pattern of the individual cells. To demonstrate the accuracy and robustness of our tool, we experimentally validated ANAlyte on two different public benchmarks of IIF HEp-2 images with rigorous leave-one-out cross-validation strategy. We obtained overall accuracy of fluorescent intensity and pattern classification respectively around 85% and above 90%. We assessed all results by comparisons with some of the most representative state of the art works. Unlike most of the other works in the recent literature, ANAlyte aims at the automatization of all the major steps of ANA image analysis. Results on public benchmarks demonstrate that the tool can characterize HEp-2 slides in terms of intensity and fluorescent pattern with accuracy better or comparable with the state of the art techniques, even when such techniques are run on manually segmented cells. Hence, ANAlyte can be proposed as a valid solution to the problem of ANA testing automatization. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

3D Actin Network Centerline Extraction with Multiple Active Contours

PubMed Central

Xu, Ting; Vavylonis, Dimitrios; Huang, Xiaolei

2013-01-01

Fluorescence microscopy is frequently used to study two and three dimensional network structures formed by cytoskeletal polymer fibers such as actin filaments and actin cables. While these cytoskeletal structures are often dilute enough to allow imaging of individual filaments or bundles of them, quantitative analysis of these images is challenging. To facilitate quantitative, reproducible and objective analysis of the image data, we propose a semi-automated method to extract actin networks and retrieve their topology in 3D. Our method uses multiple Stretching Open Active Contours (SOACs) that are automatically initialized at image intensity ridges and then evolve along the centerlines of filaments in the network. SOACs can merge, stop at junctions, and reconfigure with others to allow smooth crossing at junctions of filaments. The proposed approach is generally applicable to images of curvilinear networks with low SNR. We demonstrate its potential by extracting the centerlines of synthetic meshwork images, actin networks in 2D Total Internal Reflection Fluorescence Microscopy images, and 3D actin cable meshworks of live fission yeast cells imaged by spinning disk confocal microscopy. Quantitative evaluation of the method using synthetic images shows that for images with SNR above 5.0, the average vertex error measured by the distance between our result and ground truth is 1 voxel, and the average Hausdorff distance is below 10 voxels. PMID:24316442

Near-Infrared Fluorescence Imaging of Liver Metastases in Rats using Indocyanine Green

PubMed Central

van der Vorst, Joost R.; Hutteman, Merlijn; Mieog, Sven D.; de Rooij, Karien E.; Kaijzel, Eric L.; Löwik, Clemens W.G.M.; Putter, Hein; Kuppen, Peter J.K.; Frangioni, John V.; van de Velde, Cornelis J.H.; Vahrmeijer, Alexander L.

2011-01-01

Background Near-infrared (NIR) fluorescence imaging using indocyanine green (ICG) is a promising technique to obtain real-time assessment of the extent and number of colorectal liver metastases during surgery. The current study aims to optimize dosage and timing of ICG administration. Materials and methods Liver tumors were induced in 18 male WAG/Rij rats by subcapsular inoculation of CC531 rat colorectal cancer cells into three distinct liver lobes. Rats were divided in 2 groups: imaging after 24 and 48 hours or 72 and 96 hours after intravenous ICG administration. In each time group, rats were allocated to three dose groups: 0.04, 0.08, or 0.16 mg ICG. Intraoperative imaging and ex vivo measurements were performed using Mini-FLARE™ and confirmed by fluorescence microscopy. Fluorescence intensity was quantified using the Mini-FLARE software and the difference between tumor signal and liver signal (tumor-to-liver ratio; TLR) was calculated. Results In all 18 rats, all colorectal liver metastases (N = 34), some as small as 1.2 mm, were identified using ICG and the Mini-FLARE™ imaging system. Average tumor-to-liver ratio (TLR) over all groups was 3.0 ± 1.2. TLR was significantly higher in the 72 h time group compared to other time points. ICG dose did not significantly influence TLR, but a trend was found favoring the 0.08 mg dose group. Fluorescence microscopy demonstrated a clear fluorescent rim around the tumor. Conclusions This study demonstrates that colorectal cancer liver metastases can be clearly identified during surgery using ICG and the Mini-FLARE™ imaging system, with optimal timing of 72 h post-injection and an optimal dose of 0.08 mg (0.25 mg/kg) ICG. NIR fluorescence imaging has the potential to improve intraoperative detection of micrometastases and thus the completeness of resection. PMID:21396660

Seasonal characterization of CDOM for lakes in semiarid regions of Northeast China using excitation-emission matrix fluorescence and parallel factor analysis (EEM-PARAFAC)

NASA Astrophysics Data System (ADS)

Zhao, Ying; Song, Kaishan; Wen, Zhidan; Li, Lin; Zang, Shuying; Shao, Tiantian; Li, Sijia; Du, Jia

2016-03-01

The seasonal characteristics of fluorescent components in chromophoric dissolved organic matter (CDOM) for lakes in the semiarid region of Northeast China were examined by excitation-emission matrix (EEM) spectra and parallel factor analysis (PARAFAC). Two humic-like (C1 and C2) and protein-like (C3 and C4) components were identified using PARAFAC. The average fluorescence intensity of the four components differed under seasonal variation from June and August 2013 to February and April 2014. Components 1 and 2 exhibited a strong linear correlation (R2 = 0.628). Significantly positive linear relationships between CDOM absorption coefficients a(254) (R2 = 0.72, 0.46, p < 0.01), a(280) (R2 = 0.77, 0.47, p < 0.01), a(350) (R2 = 0.76, 0.78, p < 0.01) and Fmax for two humic-like components (C1 and C2) were exhibited, respectively. A significant relationship (R2 = 0.930) was found between salinity and dissolved organic carbon (DOC). However, almost no obvious correlation was found between salinity and EEM-PARAFAC-extracted components except for C3 (R2 = 0.469). Results from this investigation demonstrate that the EEM-PARAFAC technique can be used to evaluate the seasonal dynamics of CDOM fluorescent components for inland waters in the semiarid regions of Northeast China, and to quantify CDOM components for other waters with similar environmental conditions.

Enhanced broadband near-infrared luminescence from Pr3+-doped tellurite glass with silver nanoparticles

NASA Astrophysics Data System (ADS)

Cheng, Pan; Zhou, Yaxun; Zhou, Minghan; Su, Xiue; Zhou, Zizhong; Yang, Gaobo

2017-11-01

Pr3+-doped tellurite glasses containing metallic silver NPs were synthesized by the conventional melt-quenching technique. Structural, thermal and optical properties of the synthesized glass samples were characterized by X-Ray diffraction (XRD) curves, Raman spectra, differential scanning calorimeter (DSC) curves, transmission electron microscopy (TEM) images, UV/Vis/NIR absorption and near-infrared fluorescence emission spectra. The XRD curves confirmed the amorphous structural nature of the synthesized glasses, the Raman spectra identified the presence of different vibrational groups, the DSC curves verified the good thermal stability, and the TEM images revealed the nucleated silver NPs with average diameter about 10 nm dispersed in the glass matrix and its surface Plasmon resonance (SPR) absorption band was located at around 510 nm. Besides, Judd-Ofelt intensity parameters Ωt (t = 2, 4, 6) and other important spectroscopic parameters like transition probability, radiative lifetime, branching ratio were calculated to evaluate the radiative properties of Pr3+ levels from the measured optical absorption spectra. It was found that Pr3+-doped tellurite glasses could emit an ultra-broadband fluorescence extending from 1250 to 1650 nm under the 488 nm excitation, and this fluorescence emission increased further with the introduction of silver NPs. The enhanced fluorescence was mainly attributed to the increased local electric field around Pr3+ induced by silver NPs. The present results demonstrate that Pr3+-Ag codoped tellurite glass is a promising candidate for the near-infrared band ultra-broadband fiber amplifiers covering the expanded low-loss communication window.

Toxic Effects of Ethyl Cinnamate on the Photosynthesis and Physiological Characteristics of Chlorella vulgaris Based on Chlorophyll Fluorescence and Flow Cytometry Analysis

PubMed Central

Jiao, Yang; Ouyang, Hui-Ling; Jiang, Yu-Jiao; Kong, Xiang-Zhen; He, Wei; Liu, Wen-Xiu; Yang, Bin; Xu, Fu-Liu

2015-01-01

The toxic effects of ethyl cinnamate on the photosynthetic and physiological characteristics of Chlorella vulgaris were studied based on chlorophyll fluorescence and flow cytometry analysis. Parameters, including biomass, F v/F m (maximal photochemical efficiency of PSII), ФPSII (actual photochemical efficiency of PSII in the light), FDA, and PI staining fluorescence, were measured. The results showed the following: (1) The inhibition on biomass increased as the exposure concentration increased. 1 mg/L ethyl cinnamate was sufficient to reduce the total biomass of C. vulgaris. The 48-h and 72-h EC50 values were 2.07 mg/L (1.94–2.20) and 1.89 mg/L (1.82–1.97). (2) After 24 h of exposure to 2–4 mg/L ethyl cinnamate, the photosynthesis of C. vulgaris almost ceased, manifesting in ФPSII being close to zero. After 72 h of exposure to 4 mg/L ethyl cinnamate, the F v/F m of C. vulgaris dropped to zero. (3) Ethyl cinnamate also affected the cellular physiology of C. vulgaris, but these effects resulted in the inhibition of cell yield rather than cell death. Exposure to ethyl cinnamate resulted in decreased esterase activities in C. vulgaris, increased average cell size, and altered intensities of chlorophyll a fluorescence. Overall, esterase activity was the most sensitive variable. PMID:26101784

Effect of light intensity on the degree of ammonia toxicity on PSII activity of Arthrospira platensis and Chlorella vulgaris.

PubMed

Markou, Giorgos; Muylaert, Koenraad

2016-09-01

Herein the effect of increasing light intensity on the degree of ammonia toxicity and its impact on the photosynthetic performance of Arthrospira and Chlorella was investigated using Chl fluorescence as a technique to characterize their photosystem II (PSII) activity. The results revealed that the increase of light intensity amplifies the ammonia toxicity on PSII. Chl fluorescence transients shown that at a given free ammonia (FA) concentration (100mg-N/L), the photochemistry potential decreased by increasing light intensity. The inhibition of the PSII was not reversible either by re-incubating the cells under dark or under decreased FA concentration. Moreover, the decrease of photochemical and non-photochemical quenching (NPQ) of fluorescence suggest that ammonia toxicity decreases the open available PSII centers, as well the inability of PSII to transfer the generated electrons beyond QA. The collapse of NPQ suggests that ammonia toxicity inhibits the photoprotection mechanism(s) and hence renders PSII more sensitive to photoinhibition. Copyright © 2016 Elsevier Ltd. All rights reserved.

Green Tea Catechins Quench the Fluorescence of Bacteria-Conjugated Alexa Fluor Dyes

PubMed Central

Zhao, Lin; Li, Wei; Zhu, Shu; Tsai, Sheena; Li, Jianhua; Tracey, Kevin J.; Wang, Ping; Fan, Saijun; Sama, Andrew E.; Wang, Haichao

2013-01-01

Accumulating evidence suggests that Green tea polyphenolic catechins, especially the (-)-epigallocatechin gallate (EGCG), can be cross-linked to many proteins, and confer a wide range of anti-bacterial activities possibly by damaging microbial cytoplasmic lipids and proteins. At the doses that conferred protection against lethal polymicrobial infection (induced by cecal ligation and puncture), EGCG significantly reduced bacterial loads particularly in the liver and lung. To elucidate its bactericidal mechanisms, we determined whether EGCG affected the fluorescence intensities of bacteria-conjugated Alexa Fluor 488 or 594 dyes. When mixed with unconjugated Alexa Fluor 488 or 594 dyes, EGCG or analogs did not affect the fluorescence intensity of these dyes. In a sharp contrast, EGCG and some analogs (e.g., Catechin Gallate, CG), markedly reduced the fluorescence intensity of Gram-positive Staphylococcus aureus-conjugated Alexa 594 and Gram-negative Escherichia coli-conjugated Alexa 488. Interestingly, co-treatment with ethanol impaired the EGCG-mediated fluorescence quenching of the G+ S. aureus, but not of the G- E. coli-conjugated Alexa Flour dyes. In light of the notion that Alexa Fluor dyes can be quenched by aromatic amino acids, it is plausible that EGCG exerts anti-microbial activities possibly by altering microbial protein conformations and functions. This possibility can now be explored by screening other fluorescence-quenching agents for possible antimicrobial activities. PMID:24011199

Photoimmunotherapy of Gastric Cancer Peritoneal Carcinomatosis in a Mouse Model

PubMed Central

Sato, Kazuhide; Choyke, Peter L.; Kobayashi, Hisataka

2014-01-01

Photoimmunotherapy (PIT) is a new cancer treatment that combines the specificity of antibodies for targeting tumors with the toxicity induced by photosensitizers after exposure to near infrared (NIR) light. We performed PIT in a model of disseminated gastric cancer peritoneal carcinomatosis and monitored efficacy with in vivo GFP fluorescence imaging. In vitro and in vivo experiments were conducted with a HER2-expressing, GFP-expressing, gastric cancer cell line (N87-GFP). A conjugate comprised of a photosensitizer, IR-700, conjugated to trastuzumab (tra-IR700), followed by NIR light was used for PIT. In vitro PIT was evaluated by measuring cytotoxicity with dead staining and a decrease in GFP fluorescence. In vivo PIT was evaluated in a disseminated peritoneal carcinomatosis model and a flank xenograft using tumor volume measurements and GFP fluorescence intensity. In vivo anti-tumor effects of PIT were confirmed by significant reductions in tumor volume (at day 15, p<0.0001 vs. control) and GFP fluorescence intensity (flank model: at day 3, PIT treated vs. control p<0.01 and peritoneal disseminated model: at day 3 PIT treated vs. control, p<0.05). Cytotoxic effects in vitro were shown to be dependent on the light dose and caused necrotic cell rupture leading to GFP release and a decrease in fluorescence intensity in vitro. Thus, loss of GFP fluorescence served as a useful biomarker of cell necrosis after PIT. PMID:25401794

Use of an automated fluorescent microsphere method to measure regional blood flow in the fetal lamb.

PubMed

Tan, W; Riggs, K W; Thies, R L; Rurak, D W

1997-08-01

We have developed a method for measuring regional blood flow by means of fluorescent microspheres in all organs and tissues of the fetal lamb, including brain, heart, lung, liver, gut, spleen, kidney, adrenal, brown fat, skin, muscle, bone, and placenta. Five different fluorescent-labeled microspheres were used: blue (B), yellow-green (Y), orange (O), red (R), and crimson (C). An automated, 96-well microplate fluorescent reader (bottom reading) was chosen for the assay because of the rapidity and high throughput that it offers. Tissue samples were digested by 4 M ethanolic KOH. The sedimentation method and dye extraction with Cellosolve acetate, as previously reported by others, were used for the sample processing. The bones were crushed and allowed to directly soak in Cellosolve acetate to extract the dye. The relationship between microsphere number and fluorescent intensity was linear over a broad range of microsphere numbers (80-20,000/mL). The coefficients of variation of within-run and between-run precision were 3.39 +/- 1.10% and 4.54 +/- 1.10%, respectively. Recovery of microspheres from tissues and blood averaged 94.3 +/- 2.5% and was not dependent on microsphere number. The spillover of the fluorescent signals into adjacent colors was 4.0 +/- 0.1% for O to Y, 8.1 +/- 0.4% for O to R, and 9.1 +/- 0.5% for R to C, and these values were constant over a wide range in concentrations of the microsphere pairs. No evidence was obtained for quenching of the emission of one fluorophore via photon absorption by another fluorophore. The measurements of regional blood flow obtained with fluorescent microspheres in three chronically instrumented fetal lambs at approximately 140 days gestation were similar to the flow estimates obtained using radioactive microspheres in four other fetal lambs at the same gestational age. The fluorescent method is thus a viable alternative to the radioactive technique for the measurement of regional blood flow to all fetal organs and tissues, particularly when an automated fluorescent microplate reader is employed to reduce analysis time.

Noninvasive detection of diabetes mellitus

NASA Astrophysics Data System (ADS)

Eppstein, Jonathan A.; Bursell, Sven-Erik

1992-05-01

Recent advances in fluorescence spectroscopy of the lens reveal the potential of a non-invasive device and methodology to sensitively measure changes in the lens of the eye associated with diabetes mellitus. The system relies on the detection of the spectrum of fluorescence emitted from a selected volume (approximately 1/10 mm3) of the lens of living human subjects using low power excitation illumination from monochromatic light sources. The sensitivity of this technique is based on the measurement of the fluorescence intensity in a selected region of the fluorescence spectrum and normalization of this fluorescence with respect to attenuation (scattering and absorption) of the incident excitation light. The amplitude of the unshifted Rayleigh line, measured as part of the fluorescence spectrum, is used as a measure of the attenuation of the excitation light in the lens. Using this methodology we have demonstrated that the normalized lens fluorescence provides a more sensitive discrimination between diabetic and non-diabetic lenses than more conventional measurements of fluorescence intensity from the lens. The existing instrumentation will be described as well as the proposed design for a commercial version of the instrument expected to be ready for FDA trials by late 1992. The results from clinical measurements are used to describe a relationship between normalized lens fluorescence and hemoglobin A1c levels in diabetic patients.

Intrinsic photosensitizer fluorescence measured using multi-diameter single-fiber spectroscopy in vivo

NASA Astrophysics Data System (ADS)

van Leeuwen-van Zaane, Floor; Gamm, Ute A.; van Driel, Pieter B. A. A.; Snoeks, Thomas J.; de Bruijn, Henriette S.; van der Ploeg-van den Heuvel, Angelique; Sterenborg, Henricus J. C. M.; Löwik, Clemens W.; Amelink, Arjen; Robinson, Dominic J.

2014-01-01

Quantification of fluorescence in vivo is complicated by the influence of tissue optical properties on the collected fluorescence signal. When tissue optical properties in the measurement volume are quantified, one can obtain the intrinsic fluorescence, which equals the product of fluorophore absorption coefficient and quantum yield. We applied this method to in vivo single-fiber fluorescence spectroscopy measurements on mouse tongue, skin, liver, and oral squamous cell carcinoma, where we detected intrinsic fluorescence spectra of the photosensitizers chlorin e6 and Bremachlorin at t=[3,4.5,6,24,48] h incubation time. We observed a tissue-dependent maximum of 35% variation in the total correction factor over the visible wavelength range. Significant differences in spectral shape over time between sensitizers were observed. Although the wavelength position of the fluorescence intensity maximum for ce6 shifted to the red, Bremachlorin showed a blue shift. Furthermore, the Bremachlorin peak appeared to be broader than the ce6 fluorescence peak. Intrinsic fluorescence intensity, which can be related to photosensitizer concentration, was decreasing for all time points but showed significantly more Bremachlorin present compared to ce6 at long incubation times. Results from this study can be used to define an optimal treatment protocol for Bremachlorin-based photodynamic therapy.

A ratiometric threshold for determining presence of cancer during fluorescence-guided surgery.

PubMed

Warram, Jason M; de Boer, Esther; Moore, Lindsay S; Schmalbach, Cecelia E; Withrow, Kirk P; Carroll, William R; Richman, Joshua S; Morlandt, Anthony B; Brandwein-Gensler, Margaret; Rosenthal, Eben L

2015-07-01

Fluorescence-guided imaging to assist in identification of malignant margins has the potential to dramatically improve oncologic surgery. However, a standardized method for quantitative assessment of disease-specific fluorescence has not been investigated. Introduced here is a ratiometric threshold derived from mean fluorescent tissue intensity that can be used to semi-quantitatively delineate tumor from normal tissue. Open-field and a closed-field imaging devices were used to quantify fluorescence in punch biopsy tissues sampled from primary tumors collected during a phase 1 trial evaluating the safety of cetuximab-IRDye800 in patients (n = 11) undergoing surgical intervention for head and neck cancer. Fluorescence ratios were calculated using mean fluorescence intensity (MFI) from punch biopsy normalized by MFI of patient-matched tissues. Ratios were compared to pathological assessment and a ratiometric threshold was established to predict presence of cancer. During open-field imaging using an intraoperative device, the threshold for muscle normalized tumor fluorescence was found to be 2.7, which produced a sensitivity of 90.5% and specificity of 78.6% for delineating disease tissue. The skin-normalized threshold generated greater sensitivity (92.9%) and specificity (81.0%). Successful implementation of a semi-quantitative threshold can provide a scientific methodology for delineating disease from normal tissue during fluorescence-guided resection of cancer. © 2015 Wiley Periodicals, Inc.

Evaluating the effect of local pH on fluorescence emissions from oral bacteria of the genus Prevotella

NASA Astrophysics Data System (ADS)

Hope, Christopher K.; Higham, Susan M.

2016-08-01

A number of anaerobic oral bacteria, notably Prevotellaceae, exhibit red fluorescence when excited by short-wavelength visible light due to their accumulation of porphyrins, particularly protoporphyrin IX. pH affects the fluorescence of abiotic preparations of porphyrins due to transformations in speciation between monomers, higher aggregates, and dimers. To elucidate whether the porphyrin speciation phenomenon could be manifested within a microbiological system, suspensions of Prevotella intermedia and Prevotella nigrescens were examined by fluorescence spectrophotometry while being titrated against NaOH. The initial pH of the samples was <6, which was then raised toward the maximum found within a diseased periodontal pocket, being ˜pH 8.7. The intensity of the fluorescence emissions increased between 600 and 650 nm with increasing pH. Peak fluorescence emissions occurred at 635±1 nm with a second emission peak developing with increasing pH at 622 nm. A linear relationship was demonstrated between pH and the log10 ratio of 635:622 nm excitation fluorescence intensities. These findings suggest that the pH range found within the oral cavity could affect the fluorescence of oral bacteria in vivo, which may in turn have connotations for any clinical diagnoses that may be inferred from dental plaque fluorescence.

Binding of naproxen enantiomers to human serum albumin studied by fluorescence and room-temperature phosphorescence

NASA Astrophysics Data System (ADS)

Lammers, Ivonne; Lhiaubet-Vallet, Virginie; Ariese, Freek; Miranda, Miguel A.; Gooijer, Cees

2013-03-01

The interaction of the enantiomers of the non-steroidal anti-inflammatory drug naproxen (NPX) with human serum albumin (HSA) has been investigated using fluorescence and phosphorescence spectroscopy in the steady-state and time-resolved mode. The absorption, fluorescence excitation, and fluorescence emission spectra of (S)-NPX and (R)-NPX differ in shape in the presence of HSA, indicating that these enantiomers experience a different environment when bound. In solutions containing 0.2 M KI, complexation with HSA results in a strongly increased NPX fluorescence intensity and a decreased NPX phosphorescence intensity due to the inhibition of the collisional interaction with the heavy atom iodide. Fluorescence intensity curves obtained upon selective excitation of NPX show 8-fold different slopes for bound and free NPX. No significant difference in the binding constants of (3.8 ± 0.6) × 105 M-1 for (S)-NPX and (3.9 ± 0.6) × 105 M-1 for (R)-NPX was found. Furthermore, the addition of NPX quenches the phosphorescence of the single tryptophan in HSA (Trp-214) based on Dexter energy transfer. The short-range nature of this mechanism explains the upward curvature of the Stern-Volmer plot observed for HSA: At low concentrations NPX binds to HSA at a distance from Trp-214 and no quenching occurs, whereas at high NPX concentrations the phosphorescence intensity decreases due to dynamic quenching by NPX diffusing into site I from the bulk solution. The dynamic quenching observed in the Stern-Volmer plots based on the longest phosphorescence lifetime indicates an overall binding constant to HSA of about 3 × 105 M-1 for both enantiomers.

Dual-Color Fluorescence Imaging to Monitor CYP3A4 and CYP3A7 Expression in Human Hepatic Carcinoma HepG2 and HepaRG Cells

PubMed Central

Kubiura, Musashi; Hayashi, Ayaka; Ohbayashi, Tetsuya; Kazuki, Yasuhiro; Chesné, Christophe; Oshimura, Mitsuo; Tada, Masako

2014-01-01

Human adult hepatocytes expressing CYP3A4, a major cytochrome P450 enzyme, are required for cell-based assays to evaluate the potential risk of drug-drug interactions caused by transcriptional induction of P450 enzymes in early-phase drug discovery and development. However, CYP3A7 is preferentially expressed in premature hepatoblasts and major hepatic carcinoma cell lines. The human hepatocellular carcinoma cell line HepaRG possesses a high self-renewal capacity and can differentiate into hepatic cells similar to human adult hepatocytes in vitro. Transgenic HepaRG cells, in which the expression of fluorescent reporters is regulated by 35 kb regulatory elements of CYP3A4, have a distinct advantage over human hepatocytes isolated by collagenase perfusion, which are unstable in culture. Thus, we created transgenic HepaRG and HepG2 cells by replacing the protein-coding regions of human CYP3A4 and CYP3A7 with enhanced green fluorescent protein (EGFP) and DsRed reporters, respectively, in a bacterial artificial chromosome vector that included whole regulatory elements. The intensity of DsRed fluorescence was initially high during the proliferation of transgenic HepaRG cells. However, most EGFP-positive cells were derived from those in which DsRed fluorescence was extinguished. Comparative analyses in these transgenic clones showed that changes in the total fluorescence intensity of EGFP reflected fold changes in the mRNA level of endogenous CYP3A4. Moreover, CYP3A4 induction was monitored by the increase in EGFP fluorescence. Thus, this assay provides a real-time evaluation system for quality assurance of hepatic differentiation into CYP3A4-expressing cells, unfavourable CYP3A4 induction, and fluorescence-activated cell sorting-mediated enrichment of CYP3A4-expressing hepatocytes based on the total fluorescence intensities of fluorescent reporters, without the need for many time-consuming steps. PMID:25101946

Dual-color fluorescence imaging to monitor CYP3A4 and CYP3A7 expression in human hepatic carcinoma HepG2 and HepaRG cells.

PubMed

Tsuji, Saori; Kawamura, Fumihiko; Kubiura, Musashi; Hayashi, Ayaka; Ohbayashi, Tetsuya; Kazuki, Yasuhiro; Chesné, Christophe; Oshimura, Mitsuo; Tada, Masako

2014-01-01

Human adult hepatocytes expressing CYP3A4, a major cytochrome P450 enzyme, are required for cell-based assays to evaluate the potential risk of drug-drug interactions caused by transcriptional induction of P450 enzymes in early-phase drug discovery and development. However, CYP3A7 is preferentially expressed in premature hepatoblasts and major hepatic carcinoma cell lines. The human hepatocellular carcinoma cell line HepaRG possesses a high self-renewal capacity and can differentiate into hepatic cells similar to human adult hepatocytes in vitro. Transgenic HepaRG cells, in which the expression of fluorescent reporters is regulated by 35 kb regulatory elements of CYP3A4, have a distinct advantage over human hepatocytes isolated by collagenase perfusion, which are unstable in culture. Thus, we created transgenic HepaRG and HepG2 cells by replacing the protein-coding regions of human CYP3A4 and CYP3A7 with enhanced green fluorescent protein (EGFP) and DsRed reporters, respectively, in a bacterial artificial chromosome vector that included whole regulatory elements. The intensity of DsRed fluorescence was initially high during the proliferation of transgenic HepaRG cells. However, most EGFP-positive cells were derived from those in which DsRed fluorescence was extinguished. Comparative analyses in these transgenic clones showed that changes in the total fluorescence intensity of EGFP reflected fold changes in the mRNA level of endogenous CYP3A4. Moreover, CYP3A4 induction was monitored by the increase in EGFP fluorescence. Thus, this assay provides a real-time evaluation system for quality assurance of hepatic differentiation into CYP3A4-expressing cells, unfavourable CYP3A4 induction, and fluorescence-activated cell sorting-mediated enrichment of CYP3A4-expressing hepatocytes based on the total fluorescence intensities of fluorescent reporters, without the need for many time-consuming steps.

Multispectral fluorescence image algorithms for detection of frass on mature tomatoes

USDA-ARS?s Scientific Manuscript database

A multispectral algorithm derived from hyperspectral line-scan fluorescence imaging under violet LED excitation was developed for the detection of frass contamination on mature tomatoes. The algorithm utilized the fluorescence intensities at five wavebands, 515 nm, 640 nm, 664 nm, 690 nm, and 724 nm...

Temperature dependence of the ratio of intensities of up-conversion fluorescence bands of YVO4 and YGdVO4 crystals and lead fluoride nano glass ceramics activated with erbium ions

NASA Astrophysics Data System (ADS)

Varaksa, Yu. A.; Sinitsyn, G. V.; Khodasevich, M. A.; Aseev, V. A.; Kolobkova, E. V.; Yasyukevich, A. S.

2015-01-01

Up-conversion fluorescence spectra of YVO4 and YGdVO4 crystals and lead fluoride nano glass ceramics coactivated with erbium and ytterbium ions have been studied in the wavelength range of 520-560 nm under 967-nm pumping. The ratio of intensities of fluorescence bands in the ranges of 520-530 and 540-550 nm has been measured in the temperature range of from room temperature to 150°C. It is shown that the considered materials can be used for preparing a sensing element of optical fluorescent temperature sensors; the sensitivity of measuring the temperature of nano glass-ceramics can be close to that of crystal samples.

Spectrally selective fluorescence imaging of Chlorobaculum tepidum reaction centers conjugated to chelator-modified silver nanowires.

PubMed

Kowalska, Dorota; Szalkowski, Marcin; Ashraf, Khuram; Grzelak, Justyna; Lokstein, Heiko; Niedziolka-Jonsson, Joanna; Cogdell, Richard; Mackowski, Sebastian

2018-03-01

A polyhistidine tag (His-tag) present on Chlorobaculum tepidum reaction centers (RCs) was used to immobilize photosynthetic complexes on a silver nanowire (AgNW) modified with nickel-chelating nitrilo-triacetic acid (Ni-NTA). The optical properties of conjugated nanostructures were studied using wide-field and confocal fluorescence microscopy. Plasmonic enhancement of RCs conjugated to AgNWs was observed as their fluorescence intensity dependence on the excitation wavelength does not follow the excitation spectrum of RC complexes in solution. The strongest effect of plasmonic interactions on the emission intensity of RCs coincides with the absorption spectrum of AgNWs and is observed for excitation into the carotenoid absorption. From the absence of fluorescence decay shortening, we attribute the emission enhancement to increase of absorption in RC complexes.

Detecting crop population growth using chlorophyll fluorescence imaging.

PubMed

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

2017-12-10

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

Synthesis of novel fluorescently labeled water-soluble fullerenes and their application to its cellar uptake and distribution properties

NASA Astrophysics Data System (ADS)

Hashimoto, Akiko; Yamanaka, Takehiro; Takamura-Enya, Takeji

2017-12-01

Fullerene is a well-known carbon nanomaterial, which can be potentially used for drug manufacture or delivery. Despite several successful examples of utilizing fullerene derivatives as drug candidate materials, their low water solubility under physiological conditions negatively affects the cell penetration efficiency after treatment. In this work, we successfully synthesized two fullerene derivatives with covalently attached fluorescein and boron dipyrromethene (BODIPY) fluorophore moieties, which exhibited cellular uptake and intracellular localization. While both fluorophores decreased their fluorescence intensity in the vicinity of fullerene, the cellar uptake of the fluorescein-modified fullerene was detected via fluorescence microscopy observations. Moreover, decreases in the fluorescence intensities of the intact fluorescein and BODIPY species were observed when both fluorophores and fullerene coexisted in aqueous media.

Evaluation of automated threshold selection methods for accurately sizing microscopic fluorescent cells by image analysis.

PubMed Central

Sieracki, M E; Reichenbach, S E; Webb, K L

1989-01-01

The accurate measurement of bacterial and protistan cell biomass is necessary for understanding their population and trophic dynamics in nature. Direct measurement of fluorescently stained cells is often the method of choice. The tedium of making such measurements visually on the large numbers of cells required has prompted the use of automatic image analysis for this purpose. Accurate measurements by image analysis require an accurate, reliable method of segmenting the image, that is, distinguishing the brightly fluorescing cells from a dark background. This is commonly done by visually choosing a threshold intensity value which most closely coincides with the outline of the cells as perceived by the operator. Ideally, an automated method based on the cell image characteristics should be used. Since the optical nature of edges in images of light-emitting, microscopic fluorescent objects is different from that of images generated by transmitted or reflected light, it seemed that automatic segmentation of such images may require special considerations. We tested nine automated threshold selection methods using standard fluorescent microspheres ranging in size and fluorescence intensity and fluorochrome-stained samples of cells from cultures of cyanobacteria, flagellates, and ciliates. The methods included several variations based on the maximum intensity gradient of the sphere profile (first derivative), the minimum in the second derivative of the sphere profile, the minimum of the image histogram, and the midpoint intensity. Our results indicated that thresholds determined visually and by first-derivative methods tended to overestimate the threshold, causing an underestimation of microsphere size. The method based on the minimum of the second derivative of the profile yielded the most accurate area estimates for spheres of different sizes and brightnesses and for four of the five cell types tested. A simple model of the optical properties of fluorescing objects and the video acquisition system is described which explains how the second derivative best approximates the position of the edge. Images PMID:2516431

Dual-detector X-ray fluorescence imaging of ancient artifacts with surface relief

PubMed Central

Smilgies, Detlef-M.; Powers, Judson A.; Bilderback, Donald H.; Thorne, Robert E.

2012-01-01

Interpretation of X-ray fluorescence images of archeological artifacts is complicated by the presence of surface relief and roughness. Using two symmetrically arranged fluorescence detectors in a back-reflection geometry, the proper X-ray fluorescence yield can be distinguished from intensity variations caused by surface topography. This technique has been applied to the study of Roman inscriptions on marble. PMID:22713888

Multi-channel lock-in amplifier assisted femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy with efficient rejection of superfluorescence background.

PubMed

Mao, Pengcheng; Wang, Zhuan; Dang, Wei; Weng, Yuxiang

2015-12-01

Superfluorescence appears as an intense background in femtosecond time-resolved fluorescence noncollinear optical parametric amplification spectroscopy, which severely interferes the reliable acquisition of the time-resolved fluorescence spectra especially for an optically dilute sample. Superfluorescence originates from the optical amplification of the vacuum quantum noise, which would be inevitably concomitant with the amplified fluorescence photons during the optical parametric amplification process. Here, we report the development of a femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectrometer assisted with a 32-channel lock-in amplifier for efficient rejection of the superfluorescence background. With this spectrometer, the superfluorescence background signal can be significantly reduced to 1/300-1/100 when the seeding fluorescence is modulated. An integrated 32-bundle optical fiber is used as a linear array light receiver connected to 32 photodiodes in one-to-one mode, and the photodiodes are further coupled to a home-built 32-channel synchronous digital lock-in amplifier. As an implementation, time-resolved fluorescence spectra for rhodamine 6G dye in ethanol solution at an optically dilute concentration of 10(-5)M excited at 510 nm with an excitation intensity of 70 nJ/pulse have been successfully recorded, and the detection limit at a pump intensity of 60 μJ/pulse was determined as about 13 photons/pulse. Concentration dependent redshift starting at 30 ps after the excitation in time-resolved fluorescence spectra of this dye has also been observed, which can be attributed to the formation of the excimer at a higher concentration, while the blueshift in the earlier time within 10 ps is attributed to the solvation process.

Multi-channel lock-in amplifier assisted femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy with efficient rejection of superfluorescence background

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mao, Pengcheng; Wang, Zhuan; Dang, Wei

Superfluorescence appears as an intense background in femtosecond time-resolved fluorescence noncollinear optical parametric amplification spectroscopy, which severely interferes the reliable acquisition of the time-resolved fluorescence spectra especially for an optically dilute sample. Superfluorescence originates from the optical amplification of the vacuum quantum noise, which would be inevitably concomitant with the amplified fluorescence photons during the optical parametric amplification process. Here, we report the development of a femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectrometer assisted with a 32-channel lock-in amplifier for efficient rejection of the superfluorescence background. With this spectrometer, the superfluorescence background signal can be significantly reduced to 1/300–1/100more » when the seeding fluorescence is modulated. An integrated 32-bundle optical fiber is used as a linear array light receiver connected to 32 photodiodes in one-to-one mode, and the photodiodes are further coupled to a home-built 32-channel synchronous digital lock-in amplifier. As an implementation, time-resolved fluorescence spectra for rhodamine 6G dye in ethanol solution at an optically dilute concentration of 10{sup −5}M excited at 510 nm with an excitation intensity of 70 nJ/pulse have been successfully recorded, and the detection limit at a pump intensity of 60 μJ/pulse was determined as about 13 photons/pulse. Concentration dependent redshift starting at 30 ps after the excitation in time-resolved fluorescence spectra of this dye has also been observed, which can be attributed to the formation of the excimer at a higher concentration, while the blueshift in the earlier time within 10 ps is attributed to the solvation process.« less

Improving confocal microscopy with solid-state semiconductor excitation sources

NASA Astrophysics Data System (ADS)

Sivers, Nelson L.

To efficiently excite the fluorescent dyes used in imaging biological samples with a confocal microscope, the wavelengths of the exciting laser must be near the fluorochrome absorption peak. However, this causes imaging problems when the fluorochrome absorption and emission spectra overlap significantly, i.e. have small Stokes shifts, which is the case for most fluorochromes that emit in the red to infrared. As a result, the reflected laser excitation cannot be distinguished from the information-containing fluorescence signal. However, cryogenically cooling the exciting laser diode enabled the laser emission wavelengths to be tuned to shorter wavelengths, decreasing the interference between the laser and the fluorochrome's fluorescence. This reduced the amount of reflected laser light in the confocal image. However, the cooled laser diode's shorter wavelength signal resulted in slightly less efficient fluorochrome excitation. Spectrophotometric analysis showed that as the laser diodes were cooled, their output power increased, which more than compensated for the lower fluorochrome excitation and resulted in significantly more intense fluorescence. Thus, by tuning the laser diode emission wavelengths away from the fluorescence signal, less reflected laser light and more fluorescence information reached the detector, creating images with better signal to noise ratios. Additionally, new, high, luminous flux, light-emitting diodes (LEDs) are now powerful enough to create confocal fluorescence signals comparable to those produced by the traditional laser excitation sources in fluorescence confocal microscopes. The broader LED spectral response effectively excited the fluorochrome, yet was spectrally limited enough for standard filter sets to separate the LED excitation from the fluorochrome fluorescence signal. Spectrophotometric analysis of the excitation and fluorescence spectra of several fluorochromes showed that high-powered, LED-induced fluorescence contained the same spectral information and could be more intense than that produced by lasers. An alternative, LED-based, confocal microscope is proposed in this thesis that would be capable of exciting multiple fluorochromes in a single specimen, producing images of several distinct cellular components simultaneously. The inexpensive, LED-based, confocal microscope would require lower peak excitation intensities to produce fluorescence signals equal to those produced by laser excitation, reducing cellular damage and slowing fluorochrome photobleaching.

Damage Precursor Detection in Polymer Matrix Composites Using Novel Smart Composite Particles

DTIC Science & Technology

2016-09-20

during the deformation test. Good agreement was observed with experimental results : the intensity of fluorescence was found to be directly proportional to...agreement is observed with experimental results , for which the intensity of fluorescence was found to be directly proportional to the deformation. Epoxy...the estimated Tgs of both neat epoxy and the smart polymer were compared with the experimental results obtained by DSC. Unit cell preparation

Chemical Environment Effects on K[beta]/K[alpha] Intensity Ratio: An X-Ray Fluorescence Experiment on Periodic Trends

ERIC Educational Resources Information Center

Durham, Chaney R.; Chase, Jeffery M.; Nivens, Delana A.; Baird, William H.; Padgett, Clifford W.

2011-01-01

X-ray fluorescence (XRF) data from an energy-dispersive XRF instrument were used to investigate the chlorine K[alpha] and K[beta] peaks in several group 1 salts. The ratio of the peak intensity is sensitive to the local chemical environment of the chlorine atoms studied in this experiment and it shows a periodic trend for these salts. (Contains 1…

Rapid Optical Detection and Classification of Microbes in Suspicious Powders

DTIC Science & Technology

2018-06-01

per kilogram (J kg–1) [sievert (Sv)] * Specific details regarding the implementation of SI units may be viewed at http://www.bipm.org/en/si...amino acids, peptides, and proteins. Above about 340 nm fluorescence occurs from multi -ring organic materials and their variants. Therefore, to...emission wavelength, and fluorescence intensity, represented as iso-intensity contours in the EEM diagram. The two different fluorophors shown with

Optical Characteristics and Distribution of Chromophoric Dissolved Organic Matter in Onega Bay (White Sea) during the Summer Season (Findings from an Expedition from June 22 to 26, 2015)

NASA Astrophysics Data System (ADS)

Zaitseva, A. F.; Konyukhov, I. V.; Kazimirko, Yu. V.; Pogosyan, S. I.

2018-03-01

Onega Bay waters are characterized by a high content of chromophoric dissolved organic matter (CDOM). The absorbance spectra and fluorescence intensity (excitation wavelength 455 nm, emission wavelength >680 nm) were used to assess the distribution of CDOM content in water filtered through a GF/F filter. The CDOM content at different points in Onega Bay showed more than a fourfold difference, as inferred from the measured values. The CDOM content in surface waters was, as a rule, higher than in the deeper horizons. A higher CDOM content was measured near the Onega River, near the middle part of the Onega shore, and near the Pomor shore opposite the town of Belomorsk. River runoff is the major source of CDOM in Onega Bay water. The CDOM chemical composition in Onega Bay waters was heterogeneous. The ratio of the fluorescence intensity to the absorbance value was higher near the mouths of rivers and in intensive mixing zones than in water characterized by high salinity. A highly significant linear correlation ( R 2 = 0.7825) between water salinity and CDOM fluorescence intensity was demonstrated. The contribution of fluorescent compounds to river runoff CDOM is substantially higher than the contribution to the composition marine CDOM.

Visible photoluminescence of color centers in LiF crystals for absorbed dose evaluation in clinical dosimetry

NASA Astrophysics Data System (ADS)

Villarreal-Barajas, J. E.; Piccinini, M.; Vincenti, M. A.; Bonfigli, F.; Khan, R. F.; Montereali, R. M.

2015-04-01

Among insulating materials, lithium fluoride (LiF) has been successfully used as ionizing radiation dosemeter for more than 60 years. Thermoluminescence (TL) has been the most commonly used reading technique to evaluate the absorbed dose. Lately, optically stimulated luminescence (OSL) of visible emitting color centers (CCs) has also been explored in pure and doped LiF. This work focuses on the experimental behaviour of nominally pure LiF crystals dosemeters for 6 MV x rays at low doses based on photoluminescence (PL) of radiation induced CCs. Polished LiF crystals were irradiated using 6 MV x rays produced by a clinical linear accelerator. The doses (absorbed dose to water) covered the 1-100 Gy range. Optical absorption spectra show stable formation of primary F defects up to a maximum concentration of 2×1016 cm-3, while no significant M absorption band at around 450 nm was detected. On the other hand, under Argon laser excitation at 458 nm, PL spectra of the irradiated LiF crystals clearly exhibited the characteristic F2 and F+3 visible broad emission bands. Their sum intensity is linearly proportional to the absorbed dose in the investigated range. PL integrated intensity was also measured using a conventional fluorescence optical microscope under blue lamp illumination. The relationship between the absorbed dose and the integrated F2 and F+3 PL intensities, represented by the net average pixel number in the optical fluorescence images, is also fairly linear. Even at the low point defect densities obtained at the investigated doses, these preliminary experimental results are encouraging for further investigation of CCs PL in LiF crystals for clinical dosimetry.

Character and chlorine reactivity of dissolved organic matter from a mountain pine beetle impacted watershed.

PubMed

Beggs, Katherine M H; Summers, R Scott

2011-07-01

Lodgepole pine needle leachates from trees killed by the mountain pine beetle epidemic in Colorado were evaluated for dissolved organic matter (DOM) character, biodegradation, treatability by coagulation and disinfection byproduct (DBP) formation. An average of 8.0 (±0.62) mg-DOC/g-dry weight of litter was leached from three sets of needle samples representing different levels of forest floor degradation. Fluorescence analysis included collection of excitation and emission matrices, examination of peak intensities and development of a 4-component parallel factor (PARAFAC) analysis model. Peak intensity and PARAFAC analyses provided complementary results showing that fresh leachates were initially dominated by polyphenolic/protein-like components (60-70%) and humic-like fluorescence increased (40-70%) after biodegradation. Humic-like components were removed by coagulation (20-64%), while polyphenolic/protein-like components were not, which may create challenges for utilities required to meet OM removal regulations. DBP formation yields after 24 h chlorination were 20.5-26.4 μg/mg-DOC for trihalomethanes and 9.0-14.5 μg/mg-DOC for haloacetic acids for fresh leachates; increased after biodegradation to 19.2-64.2 and 7.1-30.9 μg/mg-DOC, respectively; and decreased after coagulation (fresh: 11.3-17.7;5.7-7.6 μg/mg-DOC, respectively; biodegraded: 12.0-27.3 and 2.9-7.2 μg/mg-DOC, respectively), reflective of changes in concentration of humic material. Humic-like PARAFAC components and peak intensities were positively correlated (R(2) ≥ 0.45) to DBP concentrations, while polyphenolic/protein-like components were not (R(2) ≤ 0.17).

2-(Benzothiazol-2-yl)-phenyl-β-d-galactopyranoside derivatives as fluorescent pigment dyeing substrates and their application for the assay of β-d-galactosidase activities.

PubMed

Otsubo, Tadamune; Minami, Akira; Fujii, Haruna; Taguchi, Risa; Takahashi, Tadanobu; Suzuki, Takashi; Teraoka, Fumiteru; Ikeda, Kiyoshi

2013-04-01

2-(Benzothiazol-2-yl)-phenyl-β-d-galactopyranoside derivatives were synthesized as novel artificial fluorescent pigment dyeing substrates for β-d-galactosidase. The substrates, which exhibited non-fluorescence or weak fluorescence in solution phase, were smoothly hydrolyzed by β-d-galactosidase from Aspergillus oryzae and yielded a water-insoluble strong fluorescent pigment. The difference of fluorescent intensity exhibited a linear relationship with the amount of enzyme. Copyright © 2013 Elsevier Ltd. All rights reserved.

Fluorescence detection of dental calculus

NASA Astrophysics Data System (ADS)

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

2010-11-01

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

Taking into Account Interelement Interference in X-Ray Fluorescence Analysis of Thin Two-Layer Ti/V Systems

NASA Astrophysics Data System (ADS)

Mashin, N. I.; Razuvaev, A. G.; Cherniaeva, E. A.; Gafarova, L. M.; Ershov, A. V.

2018-03-01

We propose a new method for determining the thickness of layers in x-ray fluorescence analysis of two-layer Ti/V systems, using easily fabricated standardized film layers obtained by sputter deposition of titanium on a polymer film substrate. We have calculated correction factors taking into account the level of attenuation for the intensity of the primary emission from the x-ray tube and the analytical line for the element of the bottom layer in the top layer, and the enhancement of the fluorescence intensity for the top layer by the emission of atoms in the bottom layer.

Development and characterisation of a brain tumour mimicking protoporphyrin IX fluorescence phantom (Conference Presentation)

NASA Astrophysics Data System (ADS)

Xie, Yijing; Tisca, Cristiana; Peveler, William; Noimark, Sacha; Desjardins, Adrien E.; Parkin, Ivan P.; Ourselin, Sebastien; Vercauteren, Tom

2017-02-01

5-ALA-PpIX fluorescence-guided brain tumour resection can increase the accuracy at which cancerous tissue is removed and thereby improve patient outcomes, as compared with standard white light imaging. Novel optical devices that aim to increase the specificity and sensitivity of PpIX detection are typically assessed by measurements in tissue-mimicking optical phantoms of which all optical properties are defined. Current existing optical phantoms specified for PpIX lack consistency in their optical properties, and stability with respect to photobleaching, thus yielding an unstable correspondence between PpIX concentration and the fluorescence intensity. In this study, we developed a set of aqueous-based phantoms with different compositions, using deionised water or PBS buffer as background medium, intralipid as scattering material, bovine haemoglobin as background absorber, and either PpIX dissolved in DMSO or a novel nanoparticle with similar absorption and emission spectrum to PpIX as the fluorophore. We investigated the phantom stability in terms of aggregation and photobleaching by comparing with different background medium and fluorophores, respectively. We characterised the fluorescence intensity of the fluorescent nanoparticle in different concentration of intralipid and haemoglobin and its time-dependent stability, as compared to the PpIX-induced fluorescence. We corroborated that the background medium was essential to prepare a stable aqueous phantom. The novel fluorescent nanoparticle used as surrogate fluorophore of PpIX presented an improved temporal stability and a reliable correspondence between concentration and emission intensity. We proposed an optimised phantom composition and recipe to produce reliable and repeatable phantom for validation of imaging device.

Fluorescence evolution of leachates during treatment processes from two contrasting landfills.

PubMed

Sun, W L; Liu, T T; Cui, F; Ni, J R

2008-10-01

Landfill leachates are composed of a complex mixture of organic matter, including a wide range of potentially fluorescent organic compounds. The fluorescence excitation-emission matrix (FEEM) of leachates during treatment processes is investigated. Particular attention is paid to the fluorescence evolution of leachates during treatment processes. Two typical types of landfill, landfill A (a direct municipal solid waste (MSW) landfill) and landfill B (disposal of bottom ashes from MSW incinerators), in a city in Southern China were selected. The results show that two characteristic and intense excitation-emission peaks located at Ex/Em = 310-330 nm/395-410 nm (peak alpha) and Ex/Em = 250-260 nm/450-460 nm (peak alpha') are observed. As the aromatic chemicals, capable of emitting fluorescence, are more recalcitrant to biodegradation than aliphatic chemicals, enhancement of the dissolved organic carbon normalized fluorescence intensities is demonstrated during treatment processes of leachate A and leachate B. This is confirmed by the variation of ultraviolet absorptivity of leachates at 254 nm. Peak alpha' and peak alpha are attributed to a mixture of xenobiotic organic compounds with low molecular weight and relatively stable aromatic fulvic-like matters with high molecular weight, respectively. Humic substances are more resistant to biodegradation than xenobiotic organic compounds, so a significant reduction in the Ialpha'/Ialpha values (fluorescence intensity ratios of peak alpha' and peak a) of leachate A was observed during treatment processes. However, no evident variation for the Ialpha/Ialpha values of leachate B was found during treatment processes owing to the low concentrations of xenobiotic organic compounds in leachate B after incineration.

The synthesis and characterization of 7-hydroxy-4-methylcoumarin and the investigation of the fluorescence properties of its 7-hydroxy-4-methylcoumarin-chitosan films

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wahyuningrum, Deana, E-mail: deana@chem.itb.ac.id; Zulqarnaen, Muhammad; Suendo, Veinardi

Chitosan fluorescent films containing 7-hydroxy-4-methylcoumarin (7H4MC) have been successfully prepared. Used chitosan was obtained from chitin isolated from skin of tiger prawns (Penaeus monodon) through the deproteination, demineralization, and deacetylation process. The yields of chitin and chitosan are 10.66% and 23.83%, respectively. The chitosan has 55.00% degree of deacetylation based on FTIR spectroscopy. Average molecular mass of chitosan which was determined by Ostwald viscometry method is 8.55 × 10{sup 6} g/mol. The 7H4MC was synthesized from resorcinol and ethyl acetoacetate using amberlyst-15 as catalyst based on Pechmann reaction with chemical yields of 90.01% and the melting point of 189–190°C. Themore » FTIR, {sup 1}H–NMR, and {sup 13}C–NMR spectroscopies confirmed the structure which corresponds to the structure of 7H4MC. The films of chitosan containing 7H4MC were prepared by solvent evaporation method in 2% (v/v) acetic acid. The 7H4MC content in each film was 0% (blank), 0.2%, 0.4%, 0.6%, and 0.8% (w/w). The UV-Vis spectrum of 7H4MC in methanol showed λ{sub max} at 235 and 337 nm. The observed fluorescence is the fluorescence color of cyan. The excitation wavelengths are 200, 235, 275, 337, and 365 nm. The highest intensity of cyan color fluorescence of chitosan containing 7H4MC films was obtained at the concentration of 0.2% of 7-hydroxy-4-methylcoumarin at the excitation wavelength of 275 nm.« less

Fluorescent components and spatial patterns of chromophoric dissolved organic matters in Lake Taihu, a large shallow eutrophic lake in China.

PubMed

Yao, Bo; Hu, Chunming; Liu, Qingquan

2016-11-01

Water samples at both surface and bottom layers were taken from 102 sites in Lake Taihu to study the fluorescent components and spatial patterns of chromophoric dissolved organic matters (CDOM). Three-dimensional excitation-emission matrix data obtained from the samples were analyzed by parallel factor approach in which four humic-like and two protein-like fluorescent components (named C1-C6) were identified. The results showed that fluorescence intensities were higher in the northern and western lake regions, and notable declines of fluorescence maxima (F max ) were observed from the northwest to the center and then to the southeast of the lake. Calculated biological index (BIX) values ranged from 0.88 to 1.44 and humification index (HIX) values from 0.64 to 3.37 for all the samples. The spatial variations of BIX and HIX values suggested stronger allochthonous CDOM characteristics in Zhushan Bay and the western area and autochthonous characteristics in the southern and eastern areas. Vertically, the average F max value of the surface samples was about 6 % less than that of the bottom samples, but noticeable variations existed among different sampling sites and components. These distribution characteristics of CDOM were mainly attributed to the spatial heterogeneity of sources and wind-induced transportation process. Interestingly, the C6 component (Ex max /Em max  = 250/455 nm) seemed to be unique in samples from Zhushan Bay and probably resulted from the discharge of the Taige River. Therefore, it could be used as an indicator of point-source discharge and a tracer to study the fate of CDOM in the lake.

Mapping Diffusion in a Living Cell via the Phasor Approach

PubMed Central

Ranjit, Suman; Lanzano, Luca; Gratton, Enrico

2014-01-01

Diffusion of a fluorescent protein within a cell has been measured using either fluctuation-based techniques (fluorescence correlation spectroscopy (FCS) or raster-scan image correlation spectroscopy) or particle tracking. However, none of these methods enables us to measure the diffusion of the fluorescent particle at each pixel of the image. Measurement using conventional single-point FCS at every individual pixel results in continuous long exposure of the cell to the laser and eventual bleaching of the sample. To overcome this limitation, we have developed what we believe to be a new method of scanning with simultaneous construction of a fluorescent image of the cell. In this believed new method of modified raster scanning, as it acquires the image, the laser scans each individual line multiple times before moving to the next line. This continues until the entire area is scanned. This is different from the original raster-scan image correlation spectroscopy approach, where data are acquired by scanning each frame once and then scanning the image multiple times. The total time of data acquisition needed for this method is much shorter than the time required for traditional FCS analysis at each pixel. However, at a single pixel, the acquired intensity time sequence is short; requiring nonconventional analysis of the correlation function to extract information about the diffusion. These correlation data have been analyzed using the phasor approach, a fit-free method that was originally developed for analysis of FLIM images. Analysis using this method results in an estimation of the average diffusion coefficient of the fluorescent species at each pixel of an image, and thus, a detailed diffusion map of the cell can be created. PMID:25517145

Development and biodistribution of a theranostic aluminum phthalocyanine nanophotosensitizer.

PubMed

Asem, Heba; El-Fattah, Ahmed Abd; Nafee, Noha; Zhao, Ying; Khalil, Labiba; Muhammed, Mamoun; Hassan, Moustapha; Kandil, Sherif

2016-03-01

Aluminum phthalocyanine (AlPc) is an efficient second generation photosensitizer (PS) with high fluorescence ability. Its use in photodynamic therapy (PDT) is hampered by hydrophobicity and poor biodistribution. AlPc was converted to a biocompatible nanostructure by incorporation into amphiphilic polyethylene glycol-polycaprolactone (PECL) copolymer nanoparticles, allowing efficient entrapment of the PS in the hydrophobic core, water dispersibility and biodistribution enhancement by PEG-induced surface characteristics. A series of synthesized PECL copolymers were used to prepare nanophotosensitizers with an average diameter of 66.5-99.1nm and encapsulation efficiency (EE%) of 66.4-78.0%. One formulation with favorable colloidal properties and relatively slow release over 7 days was selected for in vitro photophysical assessment and in vivo biodistribution studies in mice. The photophysical properties of AlPc were improved by encapsulating AlPc into PECL-NPs, which showed intense fluorescence emission at 687nm and no AlPc aggregation has been induced after entrapment into the nanoparticles. Biodistribution of AlPc loaded NPs (AlPc-NPs) and free AlPc drug in mice was monitored by in vivo whole body fluorescence imaging and ex vivo organ imaging, with in vivo imaging system (IVIS). Compared to a AlPc solution in aqueous TWEEN 80 (2 w/v%), the developed nanophotosensitizer showed targeted drug delivery to lungs, liver and spleen as monitored by the intrinsic fluorescence of AlPc at different time points (1h, 24h and 48h) post iv. administration. The AlPc-based copolymer nanoparticles developed offer potential as a single agent-multifunctional theranostic nanophotosensitizer for PDT coupled with imaging-guided drug delivery and biodistribution, and possibly also fluorescence diagnostics. Copyright © 2015 Elsevier B.V. All rights reserved.

[Investigation of quantitative detection of water quality using spectral fluorescence signature].

PubMed

He, Jun-hua; Cheng, Yong-jin; Han, Yan-ling; Zhang, Hao; Yang, Tao

2008-08-01

A method of spectral analysis, which can simultaneously detect dissolved organic matter (DOM) and chlorophyll a (Chl-a) in natural water, was developed in the present paper with the intention of monitoring water quality fast and quantitatively. Firstly, the total luminescence spectra (TLS) of water sample from East Lake in Wuhan city were measured by the use of laser (532 nm) induced fluorescence (LIF). There were obvious peaks of relative intensity at the wavelength value of 580, 651 and 687 nm in the TLS of the sample, which correspond respectively to spectra of DOM, and the Raman scattering of water and Chl-a in the water. Then the spectral fluorescence signature (SFS) technique was adopted to analyze and distinguish spectral characteristics of DOM and Chl-a in natural water. The calibration curves and function expressions, which indicate the relation between the normalized fluorescence intensities of DOM and Chl-a in water and their concentrations, were obtained respectively under the condition of low concentration(< 40 mg x L(-1))by using normalization of Raman scattering spectrum of water. The curves have a high linearity. When the concentration of the solution with humic acid is large (> 40 mg x L(-1)), the Raman scattering signal is totally absorbed by the molecules of humic acid being on the ground state, so the normalization technique can not be adopted. However the function expression between the concentration of the solution with humic acid and its relative fluorescence peak intensity can be acquired directly with the aid of experiment of fluorescence spectrum. It is concluded that although the expression is non-linearity as a whole, there is a excellent linear relation between the fluorescence intensity and concentration of DOM when the concentration is less than 200 mg x L(-1). The method of measurement based on spectral fluorescence signature technique and the calibration curves gained will have prospects of broad application. It can recognize fast what pollutants are and detect quantitatively their contents in water. It is realizable to monitor the quality of natural water with real time, dynamics and inlarge area.

Chlorophyll Fluorescence Analysis of Cyanobacterial Photosynthesis and Acclimation

PubMed Central

Campbell, Douglas; Hurry, Vaughan; Clarke, Adrian K.; Gustafsson, Petter; Öquist, Gunnar

1998-01-01

Cyanobacteria are ecologically important photosynthetic prokaryotes that also serve as popular model organisms for studies of photosynthesis and gene regulation. Both molecular and ecological studies of cyanobacteria benefit from real-time information on photosynthesis and acclimation. Monitoring in vivo chlorophyll fluorescence can provide noninvasive measures of photosynthetic physiology in a wide range of cyanobacteria and cyanolichens and requires only small samples. Cyanobacterial fluorescence patterns are distinct from those of plants, because of key structural and functional properties of cyanobacteria. These include significant fluorescence emission from the light-harvesting phycobiliproteins; large and rapid changes in fluorescence yield (state transitions) which depend on metabolic and environmental conditions; and flexible, overlapping respiratory and photosynthetic electron transport chains. The fluorescence parameters FV/FM, FV′/FM′,qp,qN, NPQ, and φPS II were originally developed to extract information from the fluorescence signals of higher plants. In this review, we consider how the special properties of cyanobacteria can be accommodated and used to extract biologically useful information from cyanobacterial in vivo chlorophyll fluorescence signals. We describe how the pattern of fluorescence yield versus light intensity can be used to predict the acclimated light level for a cyanobacterial population, giving information valuable for both laboratory and field studies of acclimation processes. The size of the change in fluorescence yield during dark-to-light transitions can provide information on respiration and the iron status of the cyanobacteria. Finally, fluorescence parameters can be used to estimate the electron transport rate at the acclimated growth light intensity. PMID:9729605

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

NASA Astrophysics Data System (ADS)

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.

Ternary complex formation of Eu(III) with o-phthalate in aqueous solutions.

PubMed

Park, K K; Jung, E C; Cho, H-R; Kim, W H

2009-08-15

Ternary hydroxo complex formation of Eu(III) with o-phthalate was investigated by potentiometry and fluorescence spectrophotometry. Curves of the equilibrium pH versus the amount of NaOH added showed that the pH value starting to form a Eu(III) precipitate was decreased due to the formation of a ternary hydroxo complex, EuOHL(s) (L = phthalate). The formation of EuOHL(s) was qualitatively confirmed by the enhancement of the fluorescence intensity of Eu(III) in the precipitate with the light absorbed by phthalate, and was quantitatively confirmed by the measurement of the amounts of Eu(III), OH(-) and phthalate included in the precipitate. The solubility product of EuOHL(s) was determined as pK(sp)(0) = 15.6+/-0.4. Characteristic features in the fluorescence spectra and the solubility product of the Eu(III)-phthalate complex were compared with those of the Eu(III)-PDA (PDA = pyridine-2,6-dicarboxylate) complex. The fluorescence intensity of the EuL(+) complex of L = PDA was about 11 times stronger than that of L = phthalate. The origin of the difference in the fluorescence intensity is discussed based on the intramolecular energy transfer effect from the lowest triplet energy level of the ligand to the resonance energy level of Eu(III).

Characterization of dissolved organic matter in an urbanized estuary located in Northeastern Brazil.

PubMed

Arguelho, Maria de Lara Palmeira de Macedo; Alves, José do Patrocínio Hora; Monteiro, Adnívia Santos Costa; Garcia, Carlos Alexandre Borges

2017-06-01

The Sal River estuary, which is located in the state of Sergipe, Northeastern Brazil, stands out as an urban estuary, anthropogenically impacted by untreated and treated wastewater discharge. Synchronous fluorescence spectroscopy and measurement of dissolved organic carbon (DOC) were used for characterization of dissolved organic matter (DOM) in the estuarine water. Dissolved organic carbon concentrations ranged from 7.5 to 19.0 mg L -1 and, in general, the highest values were recorded during dry season. For both seasons (dry and rainy), DOC presented an inverse linear relationship with salinity, which indicates a conservative dilution of organic matter coming into the estuary. During rainy season, anthropogenic organic constituents and humic substances from land-based sources predominated in DOM composition, carried by river flow. Whereas during the dry season, it has been observed a significant increase of products generated by microbial degradation of anthropogenic organic matter. The relationships between fluorescence intensity and salinity suggest a conservative behavior during rainy season and a non-conservative behavior during dry season, with addition of fluorescent organic matter into the intermediate zone of the estuary. Photodegradation by action of sunlight caused a decrease in fluorescence intensity of humic and tryptophan-like constituents and the release of photoproducts, resulting in an increase in fluorescence intensity of protein-like constituents.

Differentiation of cancerous and normal brain tissue using label free fluorescence and Stokes shift spectroscopy

NASA Astrophysics Data System (ADS)

Zhou, Yan; Wang, Leana; Liu, Cheng-hui; He, Yong; Yu, Xinguang; Cheng, Gangge; Wang, Peng; Shu, Cheng; Alfano, Robert R.

2016-03-01

In this report, optical biopsy was applied to diagnose human brain cancer in vitro for the identification of brain cancer from normal tissues by native fluorescence and Stokes shift spectra (SSS). 77 brain specimens including three types of human brain tissues (normal, glioma and brain metastasis of lung cancers) were studied. In order to observe spectral changes of fluorophores via fluorescence, the selected excitation wavelength of UV at 300 and 340 nm for emission spectra and a different Stokes Shift spectra with intervals Δλ = 40 nm were measured. The fluorescence spectra and SSS from multiple key native molecular markers, such as tryptophan, collagen, NADH, alanine, ceroid and lipofuscin were observed in normal and diseased brain tissues. Two diagnostic criteria were established based on the ratios of the peak intensities and peak position in both fluorescence and SSS spectra. It was observed that the ratio of the spectral peak intensity of tryptophan (340 nm) to NADH (440 nm) increased in glioma, meningioma (benign), malignant meninges tumor, and brain metastasis of lung cancer tissues in comparison with normal tissues. The ratio of the SS spectral peak (Δλ = 40 nm) intensities from 292 nm to 366 nm had risen similarly in all grades of tumors.

Ultrasensitive detection of amifostine and alkaline phosphatase based on the growth of CdS quantum dots.

PubMed

Na, Weidan; Liu, Siyu; Liu, Xiaotong; Su, Xingguang

2015-11-01

In this study, we reported a simple and sensitive fluorescence nanosensor for rapid detection of amifostine and alkaline phosphatase (ALP). The novel nanosensor was based on the fluorescence "turn on-off" of CdS quantum dots (QDs). Firstly, Cd(2+) cation could react with S(2-) anion to generate fluorescent CdS QDs in the presence of amifostine. The fluorescence (FL) intensity of amifostine-capped CdS QDs (Amifostine-CdS QDs) was increased with the increasing amounts of amifostine, and could be used for amifostine detection. However, amifostine could be converted to 2-(3-aminopropylamino) ethanethiol (WR1065) in the presence of ALP based on the dephosphorylation of ALP. Under the optimum conditions, the affinity of WR1065 to CdS QDs was weaker than that of amifostine. Therefore the new generation of WR1065-CdS QDs would reduce the FL intensity with the increase of ALP concentration, and the fluorescence of CdS QDs was turn off. The metabolic process of amifostine in the presence of alkaline phosphatase could be also studied via the change of FL intensity of CdS QDs. The present method was cost-effective, convenient, and does not require any complicated synthetic procedures. Copyright © 2015 Elsevier B.V. All rights reserved.

Simultaneous determination of micellar structure and drag reduction in a surfactant solution flow using the fluorescence probe method

NASA Astrophysics Data System (ADS)

Wakimoto, Tatsuro; Araga, Koichi; Katoh, Kenji

2018-03-01

As widely known, the addition of a specific type of surfactant to water reduces drag in a pipe flow. This effect is considered to be a result of the suppression of turbulent transition caused by the ordered structure of rod-like micelles that is referred to as a shear-induced structure (SIS). However, it is typically difficult to determine the SIS since it is necessary to noninvasively detect the SIS with several hundred nanometers in the actual moving flow. In this study, we used the fluorescence probe method to locally determine the SIS in a pipe flow. When hydrophobic fluorescence molecules are added to the surfactant solution, the fluorescence molecules are trapped in micelles. Thus, fluorescence intensity varies based on the change in the micellar structure. We verified the applicability of the fluorescence probe method to the SIS detection and determined the relationship between the micellar structure and the drag reduction in the pipe flow by simultaneously measuring the fluorescence intensity and pipe friction factor. The experimental result demonstrates that the SIS formation in the near-wall region is closely correlated with the drag reduction and suggests that the near-wall SIS suppresses the turbulent transition.

A compact multi-channel fluorescence sensor with ambient light suppression

NASA Astrophysics Data System (ADS)

Egly, Dominik; Geörg, Daniel; Rädle, Matthias; Beuermann, Thomas

2012-03-01

A multi-channel fluorescence sensor has been developed for process monitoring and fluorescence diagnostics. It comprises a fiber-optic set-up with an immersion probe and an intensity-modulated high power ultraviolet light-emitting diode as a light source for fluorescence excitation. By applying an electronic lock-in procedure, fluorescence signals are selectively detectable at ambient light levels of 1000 000 times higher intensity. The sensor was designed to be compact, low cost and easily adaptable to a wide field of application. The set-up was used to simultaneously monitor three important metabolic fluorophores: NAD(P)H, flavins and porphyrins during the cultivation of a baker's yeast. Moreover, the accumulation and degradation kinetics of protoporphyrin IX induced by 5-aminolevulinic acid on the skin could be recorded by the sensor. The detection limit for protoporphyrin IX was determined to be 4 × 10-11 mol L-1. The linear signal amplification of the sensor and time courses of fluorescence signals monitored during yeast fermentations were validated using a commercial CCD spectrometer. The robust and flexible set-up of the fiber-optic measurement system promises easy implementation of this non-invasive analytical tool to fluorescence monitoring and diagnostics in R&D and production.

Use of multiphoton tomography and fluorescence lifetime imaging to investigate skin pigmentation in vivo

NASA Astrophysics Data System (ADS)

Dancik, Yuri; Favre, Amandine; Loy, Chong Jin; Zvyagin, Andrei V.; Roberts, Michael S.

2013-02-01

There is a growing body of literature showing the usefulness of multiphoton tomography (MPT) and fluorescence lifetime imaging for in situ characterization of skin constituents and the ensuing development of noninvasive diagnostic tools against skin diseases. Melanin and pigmentation-associated skin cancers constitute some of the major applications. We show that MPT and fluorescence lifetime imaging can be used to measure changes in cutaneous melanin concentration and that these can be related to the visible skin color. Melanin in the skin of African, Indian, Caucasian, and Asian volunteers is detected on the basis of its emission wavelength and fluorescence lifetimes in solution and in a melanocyte-keratinocyte cell culture. Fluorescence intensity is used to characterize the melanin content and distribution as a function of skin type and depth into the skin (stratum granulosum and stratum basale). The measured fluorescence intensities in given skin types agree with melanin amounts reported by others using biopsies. Our results suggest that spatial distribution of melanin in skin can be studied using MPT and fluorescence lifetime imaging, but further studies are needed to ascertain that the method can resolve melanin amount in smaller depth intervals.

Selective colorimetric and fluorescent quenching determination of uranyl ion via its complexation with curcumin

NASA Astrophysics Data System (ADS)

Zhu, Jing-Hui; Zhao, Xin; Yang, Jidong; Tan, Yu-Ting; Zhang, Lei; Liu, Shao-Pu; Liu, Zhong-Fang; Hu, Xiao-Li

2016-04-01

Under pH 4.0 HAc-NaAc buffer medium, curcumin alone possesses extraordinary weak fluorescence emission. Nevertheless, the introduction of Triton X-100 micelles can largely enhance the fluorescence intensity of curcumin. Uranyl ions can complex with micelles-capped curcumin, along with the slight red shift of curcumin fluorescence (about 1-7 nm), a clear decrement of absorbance (424 nm) and fluorescence (507 nm) intensities, and a distinct color change from bright yellow to orange. The fluorescence decrements (ΔF, 507 nm) are positively correlated to the amount of uranyl ions in the concentration range of 3.7 × 10- 6-1.4 × 10- 5 mol L- 1. The detection limit of this fluorescence quenching methods is 3.7 × 10- 6 mol L- 1, which is nearly 9000 times lower than the maximum allowable level in drinking water proposed by World Health Organization. Good selectivity is achieved because of a majority of co-existing substances (such as Ce4 +, La3 +, and Th4 +) do not affect the detection. The content of uranyl ions in tap water samples was determined by the proposed method with satisfactory results.

Noncontact point spectroscopy guided by two-channel fluorescence imaging in a hamster cheek pouch model

NASA Astrophysics Data System (ADS)

Yang, Victor X.; Yeow, Jenny; Lilge, Lothar D.; Kost, James; Mang, Thomas S.; Wilson, Brian C.

1999-07-01

A system for in vivo, fluorescence image-guided, non-contact point fluorescence spectroscopy is presented. A 442 nm HeCd laser is used as the fluorescence excitation source. An intensified CCD serves as the detector for both imaging and spectroscopy, on which two regions of 300 X 300 pixels were used for green (500 +/- 18 nm) and red (630 +/- 18 nm) imaging channels, and a strip of 600 X 120 pixels are used for emission spectroscopy (450 - 750 nm). At a working distance of 40 mm, the system has a spatial resolution of 0.16 mm and a spectral resolution of 5 nm. System performance is demonstrated in a carcinogenesis model in hamsters, where tumors were induced by painting DMBA in the cheek pouch. Autofluorescence and Photofrin-induced fluorescence measurements were performed every 2 weeks during the 18 weeks of tumor induction. Punch biopsies on selected animals were taken for histological staging. The results show that autofluorescence fluorescence can distinguish dysplasia from normal mucosal tissue model, utilizing the peak red intensity (or the red-to-green intensity ratio). Photofrin-induced fluorescence was superior to autofluorescence for differentiating high grade dysplasia from invasive cancer.

Combination of confocal principle and aperture stop separation improves suppression of crystalline lens fluorescence in an eye model.

PubMed

Klemm, Matthias; Blum, Johannes; Link, Dietmar; Hammer, Martin; Haueisen, Jens; Schweitzer, Dietrich

2016-09-01

Fluorescence lifetime imaging ophthalmoscopy (FLIO) is a new technique to detect changes in the human retina. The autofluorescence decay over time, generated by endogenous fluorophores, is measured in vivo. The strong autofluorescence of the crystalline lens, however, superimposes the intensity decay of the retina fluorescence, as the confocal principle is not able to suppress it sufficiently. Thus, the crystalline lens autofluorescence causes artifacts in the retinal fluorescence lifetimes determined from the intensity decays. Here, we present a new technique to suppress the autofluorescence of the crystalline lens by introducing an annular stop into the detection light path, which we call Schweitzer's principle. The efficacy of annular stops with an outer diameter of 7 mm and inner diameters of 1 to 5 mm are analyzed in an experimental setup using a model eye based on fluorescent dyes. Compared to the confocal principle, Schweitzer's principle with an inner diameter of 3 mm is able to reduce the simulated crystalline lens fluorescence to 4%, while 42% of the simulated retina fluorescence is preserved. Thus, we recommend the implementation of Schweitzer's principle in scanning laser ophthalmoscopes used for fundus autofluorescence measurements, especially the FLIO device, for improved image quality.

Use of multiphoton tomography and fluorescence lifetime imaging to investigate skin pigmentation in vivo.

PubMed

Dancik, Yuri; Favre, Amandine; Loy, Chong Jin; Zvyagin, Andrei V; Roberts, Michael S

2013-02-01

There is a growing body of literature showing the usefulness of multiphoton tomography (MPT) and fluorescence lifetime imaging for in situ characterization of skin constituents and the ensuing development of noninvasive diagnostic tools against skin diseases. Melanin and pigmentation-associated skin cancers constitute some of the major applications. We show that MPT and fluorescence lifetime imaging can be used to measure changes in cutaneous melanin concentration and that these can be related to the visible skin color. Melanin in the skin of African, Indian, Caucasian, and Asian volunteers is detected on the basis of its emission wavelength and fluorescence lifetimes in solution and in a melanocyte-keratinocyte cell culture. Fluorescence intensity is used to characterize the melanin content and distribution as a function of skin type and depth into the skin (stratum granulosum and stratum basale). The measured fluorescence intensities in given skin types agree with melanin amounts reported by others using biopsies. Our results suggest that spatial distribution of melanin in skin can be studied using MPT and fluorescence lifetime imaging, but further studies are needed to ascertain that the method can resolve melanin amount in smaller depth intervals.

Dynamic measurement of fluorescent proteins spectral distribution on virus infected cells

NASA Astrophysics Data System (ADS)

Lee, Ja-Yun; Wu, Ming-Xiu; Kao, Chia-Yun; Wu, Tzong-Yuan; Hsu, I.-Jen

2006-09-01

We constructed a dynamic spectroscopy system that can simultaneously measure the intensity and spectral distributions of samples with multi-fluorophores in a single scan. The system was used to monitor the fluorescence distribution of cells infected by the virus, which is constructed by a recombinant baculoviruses, vAcD-Rhir-E, containing the red and green fluorescent protein gene that can simultaneously produce dual fluorescence in recombinant virus-infected Spodoptera frugiperda 21 cells (Sf21) under the control of a polyhedrin promoter. The system was composed of an excitation light source, a scanning system and a spectrometer. We also developed an algorithm and fitting process to analyze the pattern of fluorescence distribution of the dual fluorescence produced in the recombinant virus-infected cells. All the algorithm and calculation are automatically processed in a visualized scanning program and can monitor the specific region of sample by calculating its intensity distribution. The spectral measurement of each pixel was performed at millisecond range and the two dimensional distribution of full spectrum was recorded within several seconds. We have constructed a dynamic spectroscopy system to monitor the process of virus-infection of cells. The distributions of the dual fluorescence were simultaneously measured at micrometer resolution.

A theoretical investigation of single-molecule fluorescence detection on thin metallic layers.

PubMed

Enderlein, J

2000-04-01

In the present paper, the excitation and detection of single-molecule fluorescence over thin metallic films is studied theoretically within the framework of classical electrodynamics. The model takes into account the specific conditions of surface plasmon-assisted optical excitation, fluorescence quenching by the metal film, and detection geometry. Extensive numerical results are presented for gold, silver, and aluminum films, showing the detectable fluorescence intensities and their dependence on film thickness and the fluorescent molecule's position under optimal excitation conditions.

"Open-Box" Approach to Measuring Fluorescence Quenching Using an iPad Screen and Digital SLR Camera

ERIC Educational Resources Information Center

Koenig, Michael H.; Yi, Eun P.; Sandridge, Matthew J.; Mathew, Alexander S.; Demas, James N.

2015-01-01

Fluorescence quenching is an analytical technique and a common undergraduate laboratory exercise. Unfortunately, a typical quenching experiment requires the use of an expensive fluorometer that measures the relative fluorescence intensity of a single sample in a closed compartment unseen by the experimenter. To overcome these shortcomings, we…

Detection of rheumatoid arthritis in humans by fluorescence imaging

NASA Astrophysics Data System (ADS)

Ebert, Bernd; Dziekan, Thomas; Weissbach, Carmen; Mahler, Marianne; Schirner, Michael; Berliner, Birgitt; Bauer, Daniel; Voigt, Jan; Berliner, Michael; Bahner, Malte L.; Macdonald, Rainer

2010-02-01

The blood pool agent indo-cyanine green (ICG) has been investigated in a prospective clinical study for detection of rheumatoid arthritis using fluorescence imaging. Temporal behavior as well as spatial distribution of fluorescence intensity are suited to differentiate healthy and inflamed finger joints after i.v. injection of an ICG bolus.

Fluorescent carbon dots nanosensor for label-free determination of vitamin B12 based on inner filter effect

NASA Astrophysics Data System (ADS)

Ding, Longhua; Yang, Hongmei; Ge, Shenguang; Yu, Jinghua

2018-03-01

A simple and effective fluorescent assay for the determination of vitamin B12 was developed. In this study, carbon dots (CDs) were prepared by one-pot hydrothermal method and directly used as a fluorophore in the inner filter effect (IFE). Both of the maximum absorption peak of vitamin B12 and excitation maxima of CDs are located at 360 nm, hence, the excited light of CDs can be absorbed by vitamin B12, resulting in the fluorescence reduction of CDs. And the fluorescence intensity of CDs decreases with the increasing concentration of vitamin B12. This IFE-based sensing strategy shows a good linear relationship between the normalized fluorescence intensity and the concentration of vitamin B12 ranging from 0 to 60 μM, with a limit of detection (LOD) of 0.1 μM at a signal-to-noise ratio of 3. Furthermore, this proposed approach was successfully applied to vitamin B12 sensing in injections. This IFE sensing platform based on various fluorescent nanomaterials has a high promise for the detection of other biomolecules due to its inherent convenience.

A virus-MIPs fluorescent sensor based on FRET for highly sensitive detection of JEV.

PubMed

Liang, Caishuang; Wang, Huan; He, Kui; Chen, Chunyan; Chen, Xiaoming; Gong, Hang; Cai, Changqun

2016-11-01

Major stumbling blocks in the recognition and detection of virus are the unstable biological recognition element or the complex detection means. Here a fluorescent sensor based on virus-molecular imprinted polymers (virus-MIPs) was designed for specific recognition and highly sensitive detection of Japanese encephalitis virus (JEV). The virus-MIPs were anchored on the surface of silica microspheres modified by fluorescent dye, pyrene-1-carboxaldehyde (PC). The fluorescence intensity of PC can be enhanced by the principle of fluorescence resonance energy transfer (FRET), where virus acted as energy donor and PC acted as energy acceptor. The enhanced fluorescence intensity was proportional to the concentration of virus in the range of 24-960pM, with a limit of detection (LOD, 3σ) of 9.6pM, and the relative standard deviation was 1.99%. In additional, the specificity study confirmed the resultant MIPs has high-selectivity for JEV. This sensor would become a new key for the detection of virus because of its high sensitive, simple operation, high stability and low cost. Copyright © 2016. Published by Elsevier B.V.

CdSe/ZnS quantum dot fluorescence spectra shape-based thermometry via neural network reconstruction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Munro, Troy; Laboratory of Soft Matter and Biophysics, Department of Physics and Astronomy, KU Leuven, Celestijnenlaan 200D, B-3001 Heverlee; Liu, Liwang

As a system of interest gets small, due to the influence of the sensor mass and heat leaks through the sensor contacts, thermal characterization by means of contact temperature measurements becomes cumbersome. Non-contact temperature measurement offers a suitable alternative, provided a reliable relationship between the temperature and the detected signal is available. In this work, exploiting the temperature dependence of their fluorescence spectrum, the use of quantum dots as thermomarkers on the surface of a fiber of interest is demonstrated. The performance is assessed of a series of neural networks that use different spectral shape characteristics as inputs (peak-based—peak intensity,more » peak wavelength; shape-based—integrated intensity, their ratio, full-width half maximum, peak normalized intensity at certain wavelengths, and summation of intensity over several spectral bands) and that yield at their output the fiber temperature in the optically probed area on a spider silk fiber. Starting from neural networks trained on fluorescence spectra acquired in steady state temperature conditions, numerical simulations are performed to assess the quality of the reconstruction of dynamical temperature changes that are photothermally induced by illuminating the fiber with periodically intensity-modulated light. Comparison of the five neural networks investigated to multiple types of curve fits showed that using neural networks trained on a combination of the spectral characteristics improves the accuracy over use of a single independent input, with the greatest accuracy observed for inputs that included both intensity-based measurements (peak intensity) and shape-based measurements (normalized intensity at multiple wavelengths), with an ultimate accuracy of 0.29 K via numerical simulation based on experimental observations. The implications are that quantum dots can be used as a more stable and accurate fluorescence thermometer for solid materials and that use of neural networks for temperature reconstruction improves the accuracy of the measurement.« less

An indole-linked C8-deoxyguanosine nucleoside acts as a fluorescent reporter of Watson-Crick versus Hoogsteen base pairing.

PubMed

Schlitt, Katherine M; Millen, Andrea L; Wetmore, Stacey D; Manderville, Richard A

2011-03-07

Pyrrole- and indole-linked C(8)-deoxyguanosine nucleosides act as fluorescent reporters of H-bonding specificity. Their fluorescence is quenched upon Watson-Crick H-bonding to dC, while Hoogsteen H-bonding to G enhances emission intensity. The indole-linked probe is ∼ 10-fold brighter and shows promise as a fluorescent reporter of Hoogsteen base pairing.

Improved axial point spread function in a two-frequency laser scanning confocal fluorescence microscope

NASA Astrophysics Data System (ADS)

Wu, Jheng-Syong; Chung, Yung-Chin; Chien, Jun-Jei; Chou, Chien

2018-01-01

A two-frequency laser scanning confocal fluorescence microscope (TF-LSCFM) based on intensity modulated fluorescence signal detection was proposed. The specimen-induced spherical aberration and scattering effect were suppressed intrinsically, and high image contrast was presented due to heterodyne interference. An improved axial point spread function in a TF-LSCFM compared with a conventional laser scanning confocal fluorescence microscope was demonstrated and discussed.

Assessing acid rain and climate effects on the temporal variation of dissolved organic matter in the unsaturated zone of a karstic system from southern China

NASA Astrophysics Data System (ADS)

Liao, Jin; Hu, Chaoyong; Wang, Miao; Li, Xiuli; Ruan, Jiaoyang; Zhu, Ying; Fairchild, Ian J.; Hartland, Adam

2018-01-01

Acid rain has the potential to significantly impact the quantity and quality of dissolved organic matter (DOM) leached from soil to groundwater. Yet, to date, the effects of acid rain have not been investigated in karstic systems, which are expected to strongly buffer the pH of atmospheric rainfall. This study presents a nine-year DOM fluorescence dataset from a karst unsaturated zone collected from two drip sites (HS4, HS6) in Heshang Cave, southern China between 2005 and 2014. Cross-correlograms show that fluorescence intensity of both dripwaters lagged behind rainfall by ∼1 year (∼11 months lag for HS4, and ∼13 months for HS6), whereas drip rates responded quite quickly to rainfall (0 months lag for HS4, and ∼3 months for HS6), based on optimal correlation coefficients. The rapid response of drip rates to rainfall is related to the change of reservoir head pressure in summer, associated with higher rainfall. In winter, low rainfall has a limited effect on head pressure, and drip rates gradually slow to a constant value associated with base flow from the overlying reservoir- this effect being most evident on inter-annual timescales (R2 = 0.80 for HS4 and R2 = 0.86 for HS6, n = 9, p < 0.01). We ascribed the ∼1 year lag of fluorescence intensity to the effect of the soil moisture deficit and the karst process on delaying water and solute transport. After eliminating the one year lag, the congruent seasonal pacing and amplitude between fluorescence intensity and rainfall observed suggests that the seasonality of fluorescence intensity was mainly controlled by the monsoonal rains which can govern the output of DOM from the soil, as well as the residence time of water in the unsaturated zone. On inter-annual timescales, a robust linear relationship between fluorescence intensity and annual (effective) precipitation amount (R2 = 0.86 for HS4 and R2 = 0.77 for HS6, n = 9, p < 0.01) was identified, implying that annual (effective) precipitation is the main determinant of DOM concentration in the aquifer. Conversely, the insensitivity of fluorescence intensity and fluorescence wavelength maxima to variations in the pH of local rainfall suggests that acid rain over the study period (∼pH 5.6 to ∼ 4.5) had no discernable effect on the quantity and quality of DOM in karst soil and soil solution, likely being strongly buffered by soil carbonates. Therefore, despite large increases in anthropogenic acid rain in recent Chinese history, hydrologic forcing is the predominant factor driving variations in DOM in karst aquifers.

Elimination of autofluorescence background from fluorescence tissue images by use of time-gated detection and the AzaDiOxaTriAngulenium (ADOTA) fluorophore.

PubMed

Rich, Ryan M; Stankowska, Dorota L; Maliwal, Badri P; Sørensen, Thomas Just; Laursen, Bo W; Krishnamoorthy, Raghu R; Gryczynski, Zygmunt; Borejdo, Julian; Gryczynski, Ignacy; Fudala, Rafal

2013-02-01

Sample autofluorescence (fluorescence of inherent components of tissue and fixative-induced fluorescence) is a significant problem in direct imaging of molecular processes in biological samples. A large variety of naturally occurring fluorescent components in tissue results in broad emission that overlaps the emission of typical fluorescent dyes used for tissue labeling. In addition, autofluorescence is characterized by complex fluorescence intensity decay composed of multiple components whose lifetimes range from sub-nanoseconds to a few nanoseconds. For these reasons, the real fluorescence signal of the probe is difficult to separate from the unwanted autofluorescence. Here we present a method for reducing the autofluorescence problem by utilizing an azadioxatriangulenium (ADOTA) dye with a fluorescence lifetime of approximately 15 ns, much longer than those of most of the components of autofluorescence. A probe with such a long lifetime enables us to use time-gated intensity imaging to separate the signal of the targeting dye from the autofluorescence. We have shown experimentally that by discarding photons detected within the first 20 ns of the excitation pulse, the signal-to-background ratio is improved fivefold. This time-gating eliminates over 96 % of autofluorescence. Analysis using a variable time-gate may enable quantitative determination of the bound probe without the contributions from the background.

Using silicon-coated gold nanoparticles to enhance the fluorescence of CdTe quantum dot and improve the sensing ability of mercury (II)

NASA Astrophysics Data System (ADS)

Zhu, Jian; Chang, Hui; Li, Jian-Jun; Li, Xin; Zhao, Jun-Wu

2018-01-01

The effect of silicon-coated gold nanoparticles with different gold core diameter and silica shell thickness on the fluorescence emission of CdTe quantum dots (QDs) was investigated. For gold nanoparticles with a diameter of 15 nm, silica coating can only results in fluorescence recover of the bare gold nanoparticle-induced quenching of QDs. However, when the size of gold nanoparticle is increased to 60 nm, fluorescence enhancement of the QDs could be obtained by silica coating. Because of the isolation of the silica shell-reduced quenching effect and local electric field effect, the fluorescence of QDs gets intense firstly and then decreases. The maximum fluorescence enhancement takes place as the silica shell has a thickness of 30 nm. This enhanced fluorescence from silicon-coated gold nanoparticles is demonstrated for sensing of Hg2 +. Under optimal conditions, the enhanced fluorescence intensity decreases linearly with the concentration of Hg2 + ranging from 0 to 200 ng/mL. The limit of detection for Hg2 + is 1.25 ng/mL. Interference test and real samples detection indicate that the influence from other metal ions could be neglected, and the Hg2 + could be specifically detected.

Research of the fluorescence detection apparatus for nutrients

NASA Astrophysics Data System (ADS)

Wang, Yu; Yan, Huimin; Ni, Xuxiang; Xu, Xiaoyi; Chen, Shibing

2015-10-01

The research of the multifunctional analyzer of Clinical Nutrition, which integrates the absorbance, luminescence, fluorescence and other optical detection methods, can overcome the functional limitations of a single technology on human nutrition analysis, and realize a rapid and accurate analysis of the nutrients. This article focuses on the design of fluorescence detection module that uses a photomultiplier tube(PMT) to detect weak fluorescence, and utilizes the single photon counting method to measure the fluorescence intensity, and then according to the relationship between the fluorescent marker and fluorescence intensity, the concentration of the analyte can be derived. Using fluorescein isothiocyanate(FITC, the most widely used fluorescein currently)to mark antibodies in the experiment, therefore, according to the maximum absorption wavelength and the maximum emission wavelength of the fluorescein isothiocyanate, to select the appropriate filters to set up the optical path. In addition, the fluorescence detection apparatus proposed in this paper uses an aspherical lens with large numerical aperture, in order to improve the capacity of signal acquisition more effectively, and the selective adoption of flexible optical fiber can realize a compact opto-mechanical structure, which is also conducive to the miniaturization of the device. The experimental results show that this apparatus has a high sensitivity, can be used for the detection and analysis of human nutrition.

Fluorescence lifetime measurements in flow cytometry

NASA Astrophysics Data System (ADS)

Beisker, Wolfgang; Klocke, Axel

1997-05-01

Fluorescence lifetime measurements provide insights int eh dynamic and structural properties of dyes and their micro- environment. The implementation of fluorescence lifetime measurements in flow cytometric systems allows to monitor large cell and particle populations with high statistical significance. In our system, a modulated laser beam is used for excitation and the phase shift of the fluorescence signal recorded with a fast computer controlled digital oscilloscope is processed digitally to determine the phase shift with respect to a reference beam by fast fourier transform. Total fluorescence intensity as well as other parameters can be determined simultaneously from the same fluorescence signal. We use the epi-illumination design to allow the use of high numerical apertures to collect as much light as possible to ensure detection of even weak fluorescence. Data storage and processing is done comparable to slit-scan flow cytometric data using data analysis system. The results are stored, displayed, combined with other parameters and analyzed as normal listmode data. In our report we discuss carefully the signal to noise ratio for analog and digital processed lifetime signals to evaluate the theoretical minimum fluorescence intensity for lifetime measurements. Applications to be presented include DNA staining, parameters of cell functions as well as different applications in non-mammalian cells such as algae.

Emission Behavior of Fluorescently Labeled Silver Nanoshell: Enhanced Self-Quenching by Metal Nanostructure.

PubMed

Zhang, Jian; Fu, Yi; Lakowicz, Joseph R

2007-02-08

Labeled silica beads with an average diameter of 100 nm were synthesized by incorporating with 20-600 μM Ru(bpy)(3) (2+) complexes. Silver shells were deposited on the beads layer-by-layer with the shell thickness of 5-50 nm. The emission band became narrower and the intensity was enhanced depending on the shell thickness. Self-quenching of the probe was observed at high concentration. Poisson statistics were employed to analyze self-quenching of the fluorophores. The estimated quenching distance was extended from 6 to 16 nm with shell growth from 0 to 50 nm. Moreover, the silver shells were also labeled with Rhodamine 6G. Fluorescence enhancement and reduced lifetime were also observed for silver-silica shell containing R6G. We found that by adjustment of probe concentration and silver shell thickness, a Ru(bpy)(3) (2+)-labeled particle could be 600 times brighter than an isolated Ru(bpy)(3) (2+) molecule. We expect labeled metal core-shell structures can become useful probes for high sensitivity and/or single particle assay.

BSA Au clusters as a probe for enhanced fluorescence detection using multipulse excitation scheme.

PubMed

Raut, Sangram L; Rich, Ryan; Fudala, Rafal; Kokate, R; Kimball, J D; Borejdo, Julian; Vishwanatha, Jamboor K; Gryczynski, Zygmunt; Gryczynski, Ignacy

2014-01-01

Although BSA Au clusters fluoresce in red region (λmax: 650 nm), they are of limited use due to low fluorescence quantum yield (~6%). Here we report an enhanced fluorescence imaging application of fluorescent bio-nano probe BSA Au clusters using multipulse excitation scheme. Multipulse excitation takes advantage of long fluorescence lifetime (> 1 µs) of BSA Au clusters and enhances its fluorescence intensity 15 times over short lived cellular auto-fluorescence. Moreover we have also shown that by using time gated detection strategy signal (fluorescence of BSA Au clusters) to noise (auto-fluorescence) ratio can be increased by 30 fold. Thereby with multipulse excitation long lifetime probes can be used to develop biochemical assays and perform optical imaging with zero background.

Frequency Domain Fluorimetry Using a Mercury Vapor Lamp

DTIC Science & Technology

2009-04-07

independence from light scatter and excitation/emission intensity variations in order to extract the sample’s fluorescent lifetime. Mercury vapor lamps ...the modulation amplitude of the lamp , An, via: max 0 1 ( ) sin(2 ) n fluorescence n n n I t B nf tπ θ = ∝ +∑ (8... lamp is estimated by assuming the lamp is emitting as a point source of uniform intensity into the lower hemisphere and has a reflector collecting

Measurement of cytoplasmic Ca2+ concentration in Saccharomyces cerevisiae induced by air cold plasma

NASA Astrophysics Data System (ADS)

Xiaoyu, DONG

2018-03-01

In this study, a novel approach to measure the absolute cytoplasmic Ca2+ concentration ([Ca2+]cyt) using the Ca2+ indicator fluo-3 AM was established. The parameters associated with the probe fluo-3 AM were optimized to accurately determine fluorescence intensity from the Ca2+-bound probe. Using three optimized parameters (final concentration of 6 mM probe, incubation time of 135 min, loading probe before plasma treatment), the maximum fluorescence intensity (F max = 527.8 a.u.) and the minimum fluorescence intensity (F min = 63.8 a.u.) were obtained in a saturated Ca2+ solution or a solution of lacking Ca2+. Correspondingly, the maximum [Ca2+]cyt induced by cold plasma was 1232.5 nM. Therefore, the Ca2+ indicator fluo-3 AM was successfully applied to measure the absolute [Ca2+]cyt in Saccharomyces cerevisiae stimulated by cold plasma at atmospheric air pressure.

Azimuthal phase retardation microscope for visualizing actin filaments of biological cells

NASA Astrophysics Data System (ADS)

Shin, In Hee; Shin, Sang-Mo

2011-09-01

We developed a new theory-based azimuthal phase retardation microscope to visualize distributions of actin filaments in biological cells without having them with exogenous dyes, fluorescence labels, or stains. The azimuthal phase retardation microscope visualizes distributions of actin filaments by measuring the intensity variations of each pixel of a charge coupled device camera while rotating a single linear polarizer. Azimuthal phase retardation δ between two fixed principal axes was obtained by calculating the rotation angles of the polarizer at the intensity minima from the acquired intensity data. We have acquired azimuthal phase retardation distributions of human breast cancer cell, MDA MB 231 by our microscope and compared the azimuthal phase retardation distributions with the fluorescence image of actin filaments by the commercial fluorescence microscope. Also, we have observed movement of human umbilical cord blood derived mesenchymal stem cells by measuring azimuthal phase retardation distributions.

Aqueous synthesis of L-cysteine and mercaptopropionic acid co-capped ZnS quantum dots with dual emissions

NASA Astrophysics Data System (ADS)

Ren, Yingkun; Wang, Yongbo; Yang, Min; Liu, Enzhou; Hu, Xiaoyun; Zhang, Xu; Fan, Jun

2018-07-01

In this paper, L-cysteine (L-cys) and mercaptopropionic acid (MPA) co-capped ZnS quantum dots (QDs) with dual emissions have been successfully synthesized by a one-pot aqueous-phase synthesis method. The intensities of the dual emissions could be controlled by regulating the molar ratio of L-cys to MPA, and the fluorescence color also turned from blue to yellow accordingly. The relationship between the ligands and fluorescence was investigated and the results indicated that L-cys could cause two emissions and MPA improved the emission intensity. In addition, the L-cys-MPA co-capped ZnS QDs showed high photostability under UV irradiation. Therefore, the L-cys-MPA co-capped ZnS QDs, which show the dual emissions and tunable emission intensities, have great potentials for use in ratiometric fluorescence sensors and multicolor bioimaging.

Direct chemiluminescence of carbon dots induced by potassium ferricyanide and its analytical application

NASA Astrophysics Data System (ADS)

Amjadi, Mohammad; Manzoori, Jamshid L.; Hallaj, Tooba; Sorouraddin, Mohammad H.

2014-03-01

The chemiluminescence (CL) of water-soluble fluorescent carbon dots (C-dots) induced by direct chemical oxidation was investigated. C-dots were prepared by solvothermal method and characterized by fluorescence spectra and transmission electron microscopy. It was found that K3Fe(CN)6 could directly oxidize C-dots to produce a relatively intense CL emission. The mechanism of CL generation was investigated based on the fluorescence and CL emission spectra and the effect of radical scavengers on the CL intensity. The inhibitive effect of some metal ions and biologically important molecules on the CL intensity of the system was examined and the potential of the system for the determination of these species at trace levels was studied. In order to evaluate the capability of method to real sample analysis, it was applied to the determination of Cr(VI) and adrenaline in water and injection samples, respectively.

Observing the Heterogeneous Electro-redox of Individual Single-Layer Graphene Sheets.

PubMed

Chen, Tao; Zhang, Yuwei; Xu, Weilin

2016-09-27

Electro-redox-induced heterogeneous fluorescence of an individual single-layer graphene sheet was observed in real time by a total internal reflection fluorescence microscope. It was found that the fluorescence intensity of an individual sheet can be tuned reversibly by applying periodic voltages to control the redox degree of graphene sheets. Accordingly, the oxidation and reduction kinetics of an individual single-layer graphene sheet was studied at different voltages. The electro-redox-induced reversible variation of fluorescence intensity of individual sheets indicates a reversible band gap tuning strategy. Furthermore, correlation analysis of redox rate constants on individual graphene sheets revealed a redox-induced spatiotemporal heterogeneity or dynamics of graphene sheets. The observed controllable redox kinetics can rationally guide the precise band gap tuning of individual graphene sheets and then help their extensive applications in optoelectronics and devices for renewable energy.

Effect of defocusing on laser ablation plume observed by laser-induced fluorescence imaging spectroscopy

NASA Astrophysics Data System (ADS)

Oba, Masaki; Miyabe, Masabumi; Akaoka, Katsuaki; Wakaida, Ikuo

2016-02-01

We used laser-induced fluorescence imaging with a varying beam focal point to observe ablation plumes from metal and oxide samples of gadolinium. The plumes expand vertically when the focal point is far from the sample surface. In contrast, the plume becomes hemispherical when the focal point is on the sample surface. In addition, the internal plume structure and the composition of the ablated atomic and ionic particles also vary significantly. The fluorescence intensity of a plume from a metal sample is greater than that from an oxide sample, which suggests that the number of monatomic species produced in each plume differs. For both the metal and oxide samples, the most intense fluorescence from atomic (ionic) species is observed with the beam focal point at 3-4 mm (2 mm) from the sample surface.

Development of a fluorescent x-ray source for medical imaging

NASA Astrophysics Data System (ADS)

Toyofuku, F.; Tokumori, K.; Nishimura, K.; Saito, T.; Takeda, T.; Itai, Y.; Hyodo, K.; Ando, M.; Endo, M.; Naito, H.; Uyama, C.

1995-02-01

A fluorescent x-ray source for medical imaging, such as K-edge subtraction angiography and monochromatic x-ray CT, has been developed. Using a 6.5 GeV accumulation ring in Tsukuba, fluorescent x rays, which range from about 30 to 70 keV are generated by irradiating several target materials. Measurements have been made of output intensities and energy spectra for different target angles and extraction angles. The intensities of fluorescent x rays at a 30 mA beam current are on the order of 1-3×106 photons/mm2/s at 30 cm from the local spot where the incident beam is collimated to 1 mm2. A phantom which contains three different contrast media (iodine, barium, gadolinium) was used for the K-edge energy subtraction, and element selective CT images were obtained.

Fluorometric "Switch-on" Detection of Heparin Based on a System Composed of Rhodamine-labeled Chitosan Oligosaccharide Lactate, and Graphene Oxide.

PubMed

Yun, Kyusik; Zhong, Linlin

2018-05-16

A novel fluorescence "Switch on" for the detection of heparin based on the RhB-COL/GO system was achieved. A strong fluorescence dye, Rhodamine B, was modified by chitosan oligosaccharide lactate (COL), which plays a major role in the formation of a positively charged RhB-COL complex. RhB-COL was soluble and stable in solution, which was characterized by using Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. GO sheets quenched the fluorescence intensity of RhB-COL due to electron transfer from RhB to the GO surface. The decrease in fluorescence intensity of RhB-COL with increasing GO concentration was recorded using a Cary Eclipse fluorescence spectrophotometer. On the other hand, the addition of heparin replaced GO to bind with the RhB-COL surface via an electrostatic and noncovalent bond due to the abundant negative charge, which resulted in recovery of the fluorescence intensity. This RhB-COL/GO system possessed high selectivity and good sensitivity for the detection of heparin compared to other biomolecules, such as glycine, D-glucose, hyaluronic acid, L-glutamic acid, and ascorbic acid. The linear response toward heparin was measured over the range, 0-1.8 U·mL-1, with a low detection limit of 0.04 U·mL-1. The satisfactory sensing performance of RhB-COL/GO for heparin supports new "switch-on" sensor applications in heparin-related biomedical detection. © 2018 IOP Publishing Ltd.

Fluorometric ‘switch-on’ detection of heparin based on a system composed of rhodamine-labeled chitosan oligosaccharide lactate, and graphene oxide

NASA Astrophysics Data System (ADS)

Zhong, Linlin; Yun, Kyusik

2018-07-01

A novel fluorescence ‘Switch on’ for the detection of heparin based on the RhB-COL/GO system was achieved. A strong fluorescence dye, Rhodamine B, was modified by chitosan oligosaccharide lactate (COL), which plays a major role in the formation of a positively charged RhB-COL complex. RhB-COL was soluble and stable in solution, which was characterized by using Fourier transform infrared spectroscopy and x-ray photoelectron spectroscopy. GO sheets quenched the fluorescence intensity of RhB-COL due to electron transfer from RhB to the GO surface. The decrease in fluorescence intensity of RhB-COL with increasing GO concentration was recorded using a Cary Eclipse fluorescence spectrophotometer. On the other hand, the addition of heparin replaced GO to bind with the RhB-COL surface via an electrostatic and noncovalent bond due to the abundant negative charge, which resulted in recovery of the fluorescence intensity. This RhB-COL/GO system possessed high selectivity and good sensitivity for the detection of heparin compared to other biomolecules, such as glycine, D-glucose, hyaluronic acid, L-glutamic acid, and ascorbic acid. The linear response toward heparin was measured over the range, 0–1.8 U · ml‑1, with a low detection limit of 0.04 U · ml‑1. The satisfactory sensing performance of RhB-COL/GO for heparin supports new ‘switch-on’ sensor applications in heparin-related biomedical detection.

Interactions of Indocyanine Green and Lipid in Enhancing Near-Infrared Fluorescence Properties: The Basis for Near-Infrared Imaging in Vivo

PubMed Central

2015-01-01

Indocyanine green (ICG) is a near-infrared (NIR) contrast agent commonly used for in vivo cardiovascular and eye imaging. For medical diagnosis, ICG is limited by its aqueous instability, concentration-dependent aggregation, and rapid degradation. To overcome these limitations, scientists have formulated ICG in various liposomes, which are spherical lipid membrane vesicles with an aqueous core. Some encapsulate ICG, while others mix it with liposomes. There is no clear understanding of lipid–ICG interactions. Therefore, we investigated lipid–ICG interactions by fluorescence and photon correlation spectroscopy. These data were used to design stable and maximally fluorescent liposomal ICG nanoparticles for NIR optical imaging of the lymphatic system. We found that ICG binds to and is incorporated completely and stably into the lipid membrane. At a lipid:ICG molar ratio of 250:1, the maximal fluorescence intensity was detected. ICG incorporated into liposomes enhanced the fluorescence intensity that could be detected across 1.5 cm of muscle tissue, while free ICG only allowed 0.5 cm detection. When administered subcutaneously in mice, lipid-bound ICG in liposomes exhibited a higher intensity, NIR image resolution, and enhanced lymph node and lymphatic vessel visualization. It also reduced the level of fluorescence quenching due to light exposure and degradation in storage. Lipid-bound ICG could provide additional medical diagnostic value with NIR optical imaging for early intervention in cases of lymphatic abnormalities. PMID:24512123

Arthritis imaging using a near-infrared fluorescence folate-targeted probe

PubMed Central

Chen, Wei-Tsung; Mahmood, Umar; Weissleder, Ralph; Tung, Ching-Hsuan

2005-01-01

A recently developed near-infrared fluorescence-labeled folate probe (NIR2-folate) was tested for in vivo imaging of arthritis using a lipopolysaccharide intra-articular injection model and a KRN transgenic mice serum induction mouse model. In the lipopolysaccharide injection model, the fluorescence signal intensity of NIR2-folate (n = 12) and of free NIR2 (n = 5) was compared between lipopolysaccharide-treated and control joints. The fluorescence signal intensity of the NIR2-folate probe at the inflammatory joints was found to be significantly higher than the control normal joints (up to 2.3-fold, P < 0.001). The NIR2-free dye injection group showed a persistent lower enhancement ratio than the NIR2-folate probe injection group. Excessive folic acid was also given to demonstrate a competitive effect with the NIR2-folate. In the KRN serum transfer model (n = 4), NIR2-folate was applied at different time points after serum transfer, and the inflamed joints could be detected as early as 30 hours after arthritogenic antibody transfer (1.8-fold increase in signal intensity). Fluorescence microscopy, histology, and immunohistochemistry validated the optical imaging results. We conclude that in vivo arthritis detection was feasible using a folate-targeted near-infrared fluorescence probe. This receptor-targeted imaging method may facilitate improved arthritis diagnosis and early assessment of the disease progress by providing an in vivo characterization of active macrophage status in inflammatory joint diseases. PMID:15743478

Monitoring the Wet-Heat Inactivation Dynamics of Single Spores of Bacillus Species by Using Raman Tweezers, Differential Interference Contrast Microscopy, and Nucleic Acid Dye Fluorescence Microscopy▿

PubMed Central

Zhang, Pengfei; Kong, Lingbo; Wang, Guiwen; Setlow, Peter; Li, Yong-qing

2011-01-01

Dynamic processes during wet-heat treatment of individual spores of Bacillus cereus, Bacillus megaterium, and Bacillus subtilis at 80 to 90°C were investigated using dual-trap Raman spectroscopy, differential interference contrast (DIC) microscopy, and nucleic acid stain (SYTO 16) fluorescence microscopy. During spore wet-heat treatment, while the spores' 1:1 chelate of Ca2+ with dipicolinic acid (CaDPA) was released rapidly at a highly variable time Tlag, the levels of spore nucleic acids remained nearly unchanged, and the Tlag times for individual spores from the same preparation were increased somewhat as spore levels of CaDPA increased. The brightness of the spores' DIC image decreased by ∼50% in parallel with CaDPA release, and there was no spore cortex hydrolysis observed. The lateral diameters of the spores' DIC image and SYTO 16 fluorescence image also decreased in parallel with CaDPA release. The SYTO 16 fluorescence intensity began to increase during wet-heat treatment at a time before Tlag and reached maximum at a time slightly later than Trelease. However, the fluorescence intensities of wet-heat-inactivated spores were ∼15-fold lower than those of nutrient-germinated spores, and this low SYTO 16 fluorescence intensity may be due in part to the low permeability of the dormant spores' inner membranes to SYTO 16 and in part to nucleic acid denaturation during the wet-heat treatment. PMID:21602365

Seasonal and downstream alterations of dissolved organic matter and dissolved inorganic ions in a human-impacted mountainous tributary of the Yellow River, China.

PubMed

Zhang, Shurong; Bai, Yijuan; Wen, Xin; Ding, Aizhong; Zhi, Jianhui

2018-04-22

Human activities impose important disturbances on both organic and inorganic chemistry in fluvial systems. In this study, we investigated the intra-annual and downstream variations of dissolved organic carbon (DOC), dissolved organic matter (DOM) excitation-emission matrix fluorescence (EEM) with parallel factor analysis (PARAFAC), major ions, and dissolved inorganic nitrogen (DIN) species in a mountainous tributary of the Yellow River, China. Both DOM quantity and quality, as represented by DOC and DOM fluorescence respectively, changed spatially and seasonally in the studied region. Fluorescence intensity of tryptophan-like components (C3) were found much higher at the populated downstream regions than in the undisturbed forested upstream regions. Seasonally, stronger fluorescence intensity of protein-like components (C3 and C4) was observed in the low-flow period (December) and in the medium-flow period (March) than in the high-flow period (May), particularly for the downstream reaches, reflecting the dominant impacts of wastewater pollution in the downstream regions. In contrast to the protein-like fluorescence, humic-like fluorescence components C1 and C2 exhibited distinctly higher intensity in the high-flow period with smaller spatial variation indicating strong flushing effect of increasing water discharge on terrestrial-sourced humic-like materials in the high-flow period. Pollution-affected dissolved inorganic ions, particularly Na + , Cl - , and NH 4 + -N, showed similar spatial and seasonal variations with protein-like fluorescence of DOM. The significant positive correlations between protein-like fluorescence of DOM and pollution-affected ions, particularly Na + , Cl - , and NH 4 + -N, suggested that there were similar pollution sources and transportation pathways of both inorganic and organic pollutants in the region. The combination of DOM fluorescence properties and inorganic ions could provide an important reference for the pollution source characterization and river basin management.

Tryptophan and Non-Tryptophan Fluorescence of the Eye Lens Proteins Provides Diagnostics of Cataract at the Molecular Level

PubMed Central

Gakamsky, Anna; Duncan, Rory R.; Howarth, Nicola M.; Dhillon, Baljean; Buttenschön, Kim K.; Daly, Daniel J.; Gakamsky, Dmitry

2017-01-01

The chemical nature of the non-tryptophan (non-Trp) fluorescence of porcine and human eye lens proteins was identified by Mass Spectrometry (MS) and Fluorescence Steady-State and Lifetime spectroscopy as post-translational modifications (PTM) of Trp and Arg amino acid residues. Fluorescence intensity profiles measured along the optical axis of human eye lenses with age-related nuclear cataract showed increasing concentration of fluorescent PTM towards the lens centre in accord with the increased optical density in the lens nucleolus. Significant differences between fluorescence lifetimes of “free” Trp derivatives hydroxytryptophan (OH-Trp), N-formylkynurenine (NFK), kynurenine (Kyn), hydroxykynurenine (OH-Kyn) and their residues were observed. Notably, the lifetime constants of these residues in a model peptide were considerably greater than those of their “free” counterparts. Fluorescence of Trp, its derivatives and argpyrimidine (ArgP) can be excited at the red edge of the Trp absorption band which allows normalisation of the emission spectra of these PTMs to the fluorescence intensity of Trp, to determine semi-quantitatively their concentration. We show that the cumulative fraction of OH-Trp, NFK and ArgP emission dominates the total fluorescence spectrum in both emulsified post-surgical human cataract protein samples, as well as in whole lenses and that this correlates strongly with cataract grade and age. PMID:28071717

A Ratiometric Threshold for Determining Presence of Cancer During Fluorescence-guided Surgery

PubMed Central

Warram, Jason M; de Boer, Esther; Moore, Lindsay S.; Schmalbach, Cecelia E; Withrow, Kirk P; Carroll, William R; Richman, Joshua S; Morlandt, Anthony B; Brandwein-Gensler, Margaret; Rosenthal, Eben L

2015-01-01

Background&Objective Fluorescence-guided imaging to assist in identification of malignant margins has the potential to dramatically improve oncologic surgery. However a standardized method for quantitative assessment of disease-specific fluorescence has not been investigated. Introduced here is a ratiometric threshold derived from mean fluorescent tissue intensity that can be used to semi-quantitatively delineate tumor from normal tissue. Methods Open-field and a closed-field imaging devices were used to quantify fluorescence in punch biopsy tissues sampled from primary tumors collected during a phase 1 trial evaluating the safety of cetuximab-IRDye800 in patients (n=11) undergoing surgical intervention for head and neck cancer. Fluorescence ratios were calculated using mean fluorescence intensity (MFI) from punch biopsy normalized by MFI of patient-matched tissues. Ratios were compared to pathological assessment and a ratiometric threshold was established to predict presence of cancer. Results During open-field imaging using an intraoperative device, the threshold for muscle normalized tumor fluorescence was found to be 2.7, which produced a sensitivity of 90.5% and specificity of 78.6% for delineating disease tissue. The skin-normalized threshold generated greater sensitivity (92.9%) and specificity (81.0%). Conclusion Successful implementation of a semi-quantitative threshold can provide a scientific methodology for delineating disease from normal tissue during fluorescence-guided resection of cancer. PMID:26074273

Diffusion behavior of the fluorescent proteins eGFP and Dreiklang in solvents of different viscosity monitored by fluorescence correlation spectroscopy

NASA Astrophysics Data System (ADS)

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

2016-12-01

Fluorescence correlation spectroscopy relies on temporal autocorrelation analysis of fluorescence intensity fluctuations that spontaneously arise in systems at equilibrium due to molecular motion and changes of state that cause changes in fluorescence, such as triplet state transition, photoisomerization and other photophysical transformations, to determine the rates of these processes. The stability of a fluorescent molecule against dark state conversion is of particular concern for chromophores intended to be used as reference tags for comparing diffusion processes on multiple time scales. In this work, we analyzed properties of two fluorescent proteins, the photoswitchable Dreiklang and its parental eGFP, in solvents of different viscosity to vary the diffusion time through the observation volume element by several orders of magnitude. In contrast to eGFP, Dreiklang undergoes a dark-state conversion on the time scale of tens to hundreds of microseconds under conditions of intense fluorescence excitation, which results in artificially shortened diffusion times if the diffusional motion through the observation volume is sufficiently slowed down. Such photophysical quenching processes have also been observed in FCS studies on other photoswitchable fluorescent proteins including Citrine, from which Dreiklang was derived by genetic engineering. This property readily explains the discrepancies observed previously between the diffusion times of eGFP- and Dreiklang-labeled plasma membrane protein complexes.

A novel and sensitive fluorescence sensor for glutathione detection by controlling the surface passivation degree of carbon quantum dots.

PubMed

Pan, Jiahong; Zheng, Zengyao; Yang, Jianying; Wu, Yaoyu; Lu, Fushen; Chen, Yaowen; Gao, Wenhua

2017-05-01

A novel fluorescence sensor based on controlling the surface passivation degree of carbon quantum dots (CQDs) was developed for glutathione (GSH) detection. First, we found that the fluorescence intensity of the CQDs which was obtained by directly pyrolyzing citric acid would increased largely after the surface passivation treatment by 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC). In the light of this phenomenon, we designed a simple, rapid and selective fluorescence sensor based on the surface passivated CQDs. A certain and excess amount of EDC were mixed with GSH, part of EDC would form a stable complex with GSH owing to the exposed sulfhydryl group of GSH. As the synthesized CQDs were added into the above mixture solution, the fluorescence intensity of the (EDC/GSH)/CQDs mixture solution could be directly related to the amount of GSH. Compared to other fluorescence analytical methods, the fluorescence sensor we design is neither the traditional fluorescent "turn on" probes nor "turn off" probes. It is a new fluorescence analytical method that target object indirectly control the surface passivation degree of CQDs so that it can realize the detection of the target object. Moreover, the proposed method manifested great advantages including short analysis time, low cost and ease of operation. Copyright © 2017 Elsevier B.V. All rights reserved.

Facile synthesis of iron oxide nanoparticle and synergistic effect of iron nanoparticle in the presence of sunlight for the degradation of DOM from textile wastewater

NASA Astrophysics Data System (ADS)

Parvin, Fahmida; Nayna, Omme Kulsum; Tareq, Shafi M.; Rikta, Sharmin Yousuf; Kamal, Abdul KI

2018-05-01

This study explores the capacity of synthesized Fe2O3 nanoparticles (NPs) under sunlight for the degradation of dissolved organic matter (DOM) from synthetic (Procion blue dye) solution as well as from textile wastewater (TWW). Fe2O3 NPs were properly synthesized and confirmed by UV absorbance, FTIR spectra and SEM image analysis. Photocatalytic degradation of DOM from TWW and synthetic solution was performed by catalyst Fe2O3 NPs (5 mg/L) in the presence of solar irradiation (up to 40 h). The DOM degradation of the TWW and synthetic solution has been analyzed by fluorescence 3D excitation emission matrix (3D EEM). Synergistic effect was expected and it was found that the rate of decrease of fluorescence intensity increased with time. Within 20 h, for the synthetic solution, reduction of fluorescence intensity (80%) reaches an equilibrium. In contrast, the rate of decrease in the fluorescence intensity is highest (91%) in 40 h of irradiation for TWW. This reduction of fluorescence intensity indicates the degradation of DOM and can be expressed well by second-order model kinetics. Reduction of TOC, BOD5 and COD load again validated the degradation of DOM from TWW by catalyst Fe2O3 NPs-induced solar irradiation. We applied the treated wastewater on the plant to observe the reusability of the treated TWW, and the morphological data analysis of the plant demonstrates that the catalyst Fe2O3 NPs-induced solar-irradiated wastewater exhibits less adverse impact on plant morphology.