Concentration Of Fluorescent Dye Research Articles

Maximising the light output of a Luminescent Solar Concentrator

Luminescent Solar Concentrators (LSC's) are currently being developed so that sunlight may be more fully utilised in place of conventional lighting sources. In order to optimise the light output of an LSC, various collector properties including fluorescent dye concentration, collector dimensions and matrix quality have been investigated. This study, involving measurement and computer modelling of the light transport in a single colour LSC, shows that luminous output is highly sensitive to weak extinction processes in the emission spectral band. These fine details in the loss spectrum play a key role in determining the output and efficiency of an LSC system. The separation of dye and matrix losses enables a quantitative study of the effect of matrix optical quality on performance.

Adsorption of Fluorescent Dyes on Oxide Nanoparticles Studied by Fluorescence Correlation Spectroscopy

In the present work, we have studied the adsorption of Rhodamine 6G on negative colloidal particles (Silica-Bindzil, Ludox-SM30 and Ludox-HS30) and Calcein on positive colloidal particles (Ludox-CL and Alumina) at very low concentrations by use of fluorescent correlation spectroscopy (FCS). In this relation we have introduced a set of equations which take into account the different quantum yield of the dye molecules and the particles as well as the adsorption of the dye molecules on the sample cell. The concentrations of fluorescent dye were varied between 2 × 10-9 and 2 × 10-7 M, and the concentrations of particles were in the range of 0.01−0.03 g/L. The adsorption data fit well with a Langmuir adsorption isotherm. The values of the adsorption constants at pH 5.5−5.7, ionic strength NaCl 1 mM, and room temperature (22 ± 0.5 °C) were in the range (0.2−2.5) × 108 L/mol, and the adsorption energy varied between −10 and 13 kJ/mol. The cooperative binding model (generalized Freundlich) was also checked but showed less satisfactory results. This work demonstrates the high performance of the FCS method for investigation of the adsorption equilibrium at solid−liquid interface. In particular the technique does not required any separation step of the adsorbed and nonadsorbed species and is not influenced by the adsorption on the sample cell walls which is often a source of artifacts when working with very low concentrations.

A comparison of imaging methods for use in an array biosensor

An array biosensor has been developed which uses an actively-cooled, charge-coupled device (CCD) imager. In an effort to save money and space, a complementary metal-oxide semiconductor (CMOS) camera and photodiode were tested as replacements for the cooled CCD imager. Different concentrations of CY5 fluorescent dye in glycerol were imaged using the three different detection systems with the same imaging optics. Signal discrimination above noise was compared for each of the three systems.

Imaging Fluorescent Dye Concentrations on Soil Surfaces

To investigate transport processes in soils, detailed information about the spatial distribution of solutes is required. We describe a method to obtain concentration maps of fluorescent tracers on cross sections of soil cores with a high spatial resolution. The fluorescence signal of two dye tracers, brilliant sulfaflavine (BF; 1H‐Benz(de)isoquinoline‐5‐sulfonic acid, 2,3‐dihydro‐6‐amino‐1,3‐dioxo‐2‐(p‐tolyl)‐, monosodium salt) and sulforhodamine B (SB; xanthylium, 3,6‐bis(diethylamino)‐9‐(2,4‐disulfophenyl)‐, inner salt, sodium salt), was imaged on the exposed cross sections. The fluorescence signal was corrected for variable illumination light intensity and optical properties of the soil across the exposed surface. Correction factors for varying optical soil properties were derived from the image of the reflected excitation light at the exposed surface. Linear calibration relations related the corrected fluorescence image to the total tracer concentration (Ct) map, that is, mass of dye dissolved in the soil solution and sorbed to the soil particles per unit volume bulk soil. Corrections for varying optical properties of the soil surface were important to reduce the uncertainty of the concentration that was estimated from the fluorescence signal. For BF, the calibration relations were different for different soil materials and a soil specific calibration had to be used. Variations in background fluorescence were an important source of uncertainty of the BF concentration estimates but can be overcome by applying higher concentrations. For SB, variations in calibration relations and in the background fluorescence were considerably smaller, and so is the uncertainty of the estimated SB concentrations.

Imaging Fluorescent Dye Concentrations on Soil Surfaces

To investigate transport processes in soils, detailed information about the spatial distribution of solutes is required. We describe a method to obtain concentration maps of fluorescent tracers on cross sections of soil cores with a high spatial resolution. The fluorescence signal of two dye tracers, brilliant sulfaflavine (BF; 1H-Benz(de)isoquinoline-5-sulfonic acid, 2,3-dihydro-6-amino-1,3-dioxo-2-(p-tolyl)-, monosodium salt) and sulforhodamine B (SB; xanthylium, 3,6-bis(diethylamino)-9-(2,4-disulfophenyl)-, inner salt, sodium salt), was imaged on the exposed cross sections. The fluorescence signal was corrected for variable illumination light intensity and optical properties of the soil across the exposed surface. Correction factors for varying optical soil properties were derived from the image of the reflected excitation light at the exposed surface. Linear calibration relations related the corrected fluorescence image to the total tracer concentration (Ct) map, that is, mass of dye dissolved in the soil solution and sorbed to the soil particles per unit volume bulk soil. Corrections for varying optical properties of the soil surface were important to reduce the uncertainty of the concentration that was estimated from the fluorescence signal. For BF, the calibration relations were different for different soil materials and a soil specific calibration had to be used. Variations in background fluorescence were an important source of uncertainty of the BF concentration estimates but can be overcome by applying higher concentrations. For SB, variations in calibration relations and in the background fluorescence were considerably smaller, and so is the uncertainty of the estimated SB concentrations.

Assessment of Membrane Potentials of Mitochondrial Populations in Living Cells

Mitochondrial membrane potentials (MMP) reflect the functional status of mitochondria within cells. Fluorescent probes to estimate these potentials within cells have been available for some time, but measurements of populations of mitochondria are not possible by existing methods. Therefore, comparisons between different cell types (e.g., fibroblasts and neuroblastoma), fibroblast cell lines from different patients, or even the same cell following various experimental paradigms are not feasible. The current approach estimates populations of MMP within living cells at 37°C using the combination of conventional fluorescence microscopy and three-dimensional deconvolution by exhaustive photon reassignment. With this method, raw images are acquired rapidly with low-intensity (nonlaser) light with minimal concentrations of fluorescent dye. The method uses the fluorescent dye tetramethylrhodamine methyl ester, which equilibrates in cells according to the Nernst equation and provides a numerical, replicable estimate of MMP for populations of cellular mitochondria. This method can detect either increases or decreases in MMP as small as 5%. Furthermore, MMP in different cell types appear distinct. Values in fibroblasts (−105 ± 0.9 mV) and N2a cells (−81 ± 0.7 mV) were very different by this method. This approach bridges investigations of individual mitochondria to those that assess MMP by examining global fluorescence from cells.

Fluorescence anisotropy sensor and its application to polymer processing and characterization

An optical sensor containing polarizing optical components has been constructed to monitor fluorescence anisotropy during polymer processing and to carry out remote sensing of polymer products doped with fluorescent dyes. The sensor is a compact unit that is used to polarize incident excitation light as well as to analyze the polarization of generated fluorescent light. Optical fibers are used to carry light between the sensor head and the light source and detecting equipment. The anisotropy measurement yields information about the orientation of a fluorescent dye molecule that has been doped into polymer matrix. Fluorescent dyes that have geometrical asymmetry in their molecular structure are used. Experiments are described for which the sensor is positioned in line during extrusion, during specimen extension, and where the sensor is used to carry out area scans of films and sheets. Measurements were made on polyethylene, polyethylene terephthalate, and polybutadiene resins that contained a low concentration of fluorescent dye.

Simultaneous detection of cell volume and intracellular pH in isolated rat duodenal cells by confocal microscopy and BCECF.

The combination of confocal laser scan microscopy and the pH-sensitive fluorescent dye BCECF allowed us to record simultaneously intracellular pH, cell viability and relative cell volume. pH was measured by using the pH-sensitive excitation wavelength at 488 nm and the pH-independent excitation wavelength at 442 nm to obtain ratio images. Cell volume was traced by measuring fluorescence dye concentration at 442 nm. Isolated villus tip rat duodenal enterocytes were exposed to 20 mM NH4Cl, sodium free, or 1 mM amiloride buffer. Sodium free buffer and amiloride buffer acidified the cells. Cell volume did not change in sodium free buffer, or NH4Cl exposure, but amiloride led to an increase in cell volume of 20%. After acidification of the duodenal cells, amiloride buffer increased cell volume by almost 50%. These studies revealed that cell volume regulation during pH changes in short-living cells could easily be detected by confocal microscopy and BCECF.

20] Calcium influx during an action potential

Action potentials open voltage-sensitive calcium channels in excitable cells, leading to an influx of calcium ions. Calcium ions may control, among others, cell excitability, neurotransmitter release, or gene transcription. This chapter discusses two different methods that can be used to quantify the Ca 2+ flows into a cell or cell compartment during an action potential. Quantification of Ca 2+ influx may be a first step toward the construction of a model for the calcium dynamics of cells. It may also serve as a reference to study pathological processes such as cell death during ischemia or amyotrophic lateral sclerosis, where increases in calcium influx have generally been implicated. The two methods that are compared in this chapter are a voltage-clamp method, in which cells are voltage clamped with an action potential waveform command, and a fluorescence method, in which cells are loaded with a high concentration of a calcium-sensitive fluorescent dye. Both methods are applied to a giant axosomatic terminal, called the “calyx of Held,” in a slice preparation of the rat brain stem. Each principal cell in the medial nucleus of the trapezoid body (MNTB) is contacted by a single calyx-type terminal. This synapse is part of a fast auditory pathway that is involved in the localization of sound. This chapter is interested in the relationship between calcium influx and neurotransmitter release at this glutamatergic synapse. Both methods gave similar values for the number of calcium ions that enter the presynaptic terminal during an action potential. It first discusses the action potential waveform voltage-clamp method.

Absorption and fluorescence spectroscopic investigation of indocyanine green

Absorption spectra, fluorescence quantum distributions, fluorescence quantum yields and degrees of fluorescence polarization vs. dye concentration were determined for indocyanine green in methanol, water and aqueous albumin solution. The monomer fluorescence quantum yield is limited to a few per cent by internal conversion. In water, dimerization starts at low concentrations (approximately 3 × 10 −8 m ol dm −3) and lowers the fluorescence quantum yield. The strong affinity of the dye to albumin shifts the onset of dimerization to higher concentrations. In methanol, dimerization is weak and closely spaced pair formation dominates at high concentrations. Dye adsorption to albumin and dye aggregation in the solvents were analysed experimentally and theoretically.

Spray Visualization by Laser Sheet Tomography.

Laser sheet tomography was applied to the visualization of the cross section of a spray. In this method, fluorescent dye, eosine-Y(C20H6Br4Na2O), was used at a concentration of 10 g/1 of the injectant, water, and the light source which illuminated the spray was a Nd : YAG laser (532wavelength, 20ns pulse width) in laser sheet tomography. The thickness of the laser sheet which was scattered by the spray was measured to elucidate the effect of the concentration of fluorescent dye in the injected liquid. In the case of water without eosine-Y, multiscattered light-illuminated drops were observed outside of the laser sheet ; therefore the thickness of the laser sheet increased. However, in the case of water with eosine-Y, there were few drops which were illuminated by the multiscattered light, and only drops which existed within the laser sheet scattered green laser light and emitted the yellow fluorescent light of eosine-Y.

Planar Chromatography Coupled to Mass Spectrometry

Abstract Applications of thin-layer chromatography/mass spectrometry are expanding rapidly due to commercial availability of the devices, and improved understanding of the procedures required to measure good quality mass spectrometric data. Several of the common approaches to TLC/MS coupling are pursued in our laboratory; recent focus has been on techniques of sample preparation and plate treatment that allow direct TLC/MS analysis to be completed on almost any instrument. Specific examples to be covered are the direct derivatization of alkaloid samples on Empore TLC plates for volatilization into an electron/chemical ionization source, development and concentration of thin-film fluorescent dyes for analysis by liquid secondary ion mass spectrometry (LSIMS), and the use of a CCD-based imaging system to explore the integration of optical and mass spectrometric information for the characterization of samples separated by thin layer chromatography.

Correlation of endothal and fluorescent dye concentrations following concurrent application to tidal canals

AbstractHerbicide residue samples are expensive and time‐consuming to collect and analyze. The potential for using ‘Rhodamine WT’ to simulate endothal dissipation in moving water was tested in the weed‐infested Three Sisters tidal canals (Citrus Co., FL). Concurrent applications were made using a variety of application techniques. Significant correlations between dye and endothal concentrations were found in each treatment, indicating that ‘Rhodamine WT’ concentrations can be used to predict endothal movement and concentrations. The ratios of endothal:dye half‐lives were similar (0.2, 0.25, 0.17) for three of the application methods tested, indicating that estimations of dye half‐lives could be used to predict those for endothal.

Study of propidium iodide binding to DNA in intact cells by flow cytometry

We studied the in situ binding of propidium iodide to DNA in fixed human lymphocytes, using flow cytometry. Experimental data of fluorescence emission vs dye concentration and vs cell concentration were obtained. Data were interpreted by means of two different mathematical models specific for the staining reaction, and the binding parameters were obtained by "best-fitting" of the data. A model based on two classes of binding sites with different affinity constants gave the most satisfactory fitting. The accessibility of the in situ chromatin turned out to be reduced with respect to the non in situ accessibility for ethidium bromide as reported in the literature. The present study shows the usefulness of the flow-cytometric technique for probing DNA structure in intact cells.

Latex nanosphere delivery system (LNDS): novel nanometer-sized carriers of fluorescent dyes and active agents selectively target neuronal subpopulations via uptake and retrograde transport

A wide range of latex particles are described which are capable of carrying high concentrations of fluorescent dyes, drugs, and photoactive agents selectively to subpopulations of neurons in vitro and in vivo. Particle size, charge, and concentration were all found to influence uptake into cultured neurons or retrograde transport in vivo. Chromophore loadings of greater than 14% (w/w) were obtained. Incorporation of a photoactivated dye (chlorinε 6) into the polymer did not compromise the ability of the dye to produce singlet oxygen following light exposure. We refer to this unique family of latex particles as the latex nanosphere delivery system (LNDS). The LNDS will be useful for studies of neuroanatomy and nervous system development, as well as more general areas of biomedical research where it is desirable to selectively label subpopulations of cells. The LNDS also offers a means of providing targeted delivery of drugs or photoactive agents to selected subpopulations of cells; this will allow experimentation not currently possible using any existent methodology.

The binding kinetics and interaction of DNA fluorochromes used in the analysis of nuclei and chromosomes by flow cytometry

The interactions and binding characteristics of DNA dyes used in the flow cytometric analysis of chromatin were studied using human chromosomes and mouse thymocyte nuclei. The kinetics of dye binding and the relationship between fluorescence intensity and dye concentration are presented. Under the conditions used, Hoechst 33258, propidium iodide and chromomycin A3 reach an equilibrium with thymocyte nuclei after approximately 5 min, 20 min and more than 1 h, respectively. The same binding kinetics are observed with Hoechst 33258 and chromomycin when nuclei are stained with a mixture of the two dyes. Sodium citrate, which improves the resolution of flow karyotypes, causes a rapid increase in Hoechst and propidium iodide fluorescence, but a decrease in the fluorescence of chromomycin. The relative peak positions of chromosomes in a flow karyotype are unaffected by sodium citrate addition. The spectral interaction between Hoechst and chromomycin is quantified. There is variation among the human chromosome types in the amount of energy transferred from Hoechst to chromomycin. By measuring the Hoechst and chromomycin fluorescence of each chromosome after Hoechst excitation, it is shown that the amount of energy transferred is correlated to the ratio of the amount of Hoechst to chromomycin bound. Although the energy transfer between the two dyes is considerable, this has little effect on the reproducibility of flow karyotype measurements. The relative peak positions of all human chromosomes in a 64 X 64 channel flow karyotype, except for the 13 and Y chromosomes, vary in the order of 0.5 channel over a 16-fold change in either Hoechst or chromomycin concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

An improved objective slit-lamp fluorophotometer using tungsten-halogen lamp excitation and synchronous detection.

A new design of fluorophotometer is described. The instrument can measure lower concentrations of fluorescent dye with greater accuracy than previous models.