Time-resolved Fluorescence Technique Research Articles

Dynamics of Bound Dyes in a Nonpolymeric Aqueous Gel Derived from a Tripodal Bile Salt

Rotational dynamics of polarity sensitive fluorescent dyes (ANS and DPH) in a nonpolymertic aqueous gel derived from tripodal cholamide I was studied using ultrafast time-resolved fluorescence technique. Results were compared with that of naturally occurring di- and trihydroxy bile salts. ANS in the gel showed two rotational correlation time (phi) components, 13.2 ns (bound to the hydrophobic region of the gel) and 1.0 ns (free aqueous ANS), whereas DPH showed only one component (4.8 ns). In the sol state, faster rotational motion was observed, both for ANS and DPH. Our data revealed that dyes get encapsulated more tightly in the gel network when compared to the micellar aggregates. ANS has more restrained rotation compared to DPH. This was attributed to the interaction of the sulfonate group of ANS with water molecules and hydrophilic parts of the gelator molecule. No restricted rotation was observed for DPH in the gel state unlike when it is in the gel phase of lipid bilayer.

Radiative lifetime and oscillator strength determinations in Sm III

Radiative lifetimes of 6 levels belonging to the 4f5(6H°, 6F°)5d configuration of Sm III have been measured for the first time using a time-resolved laser-induced fluorescence technique. Experimental data have been compared with semi-empirical calculations. The agreement is excellent for 4 levels with 6F° as a parent term. Larger discrepancies (53 and 37%) are observed for 2 levels with 6H° and 6F°, respectively, as a parent term and are explainable by strong mixing and cancellation effects occurring in the line strength calculations which are responsible of “instabilities” occurring in the lifetimes considered as a function of J value within the term. Branching ratios for the lines depopulating the levels of interest have been measured by Fourier transform spectroscopy. A first set of oscillator strengths has been deduced for this ion.

Monitoring Diffusion of Reptating Polymer Chains by a Direct Energy Transfer Method: A Monte Carlo Simulation

Full Paper: A kinetic Monte Carlo method was used to simulate the diffusion of reptating polymer chains across an interface. A time-resolved fluorescence technique in conjunction with a direct energy transfer method was used to measure the extent of diffusion of dye-labeled reptating polymer chains. The diffusion of donor- and acceptorlabeled polymer chains between adjacent compartments was randomly generated. The fluorescence decay profiles of donor molecules were simulated at several diffusion steps to produce mixing of the polymer chains. Mixing ratios of donor- and acceptor-labeled polymer chains in compartments were measured at various stages (snapshots) of diffusion. It was observed that for a given molecular weight, the average interpenetration contour length was found to be proportional to the mixing ratio. Monte Carlo analysis showed that the curvilinear diffusion coefficient is inversely proportional to the weight of polymer chains during diffusion.

Time-resolved laser-induced fluorescence lifetime measurements and theoretical transition probabilities in Tb III

Lifetimes of four short-lived levels belonging to the ${4f}^{8}{(}^{7}F)6p$ configuration of Tb III have been measured using a two-step excitation time-resolved laser-induced fluorescence technique. They agree quite well with multiconfigurational relativistic Hartree-Fock calculations performed with inclusion of core-polarization effects. Using the experimental lifetimes and the theoretical branching fractions, a set of transition probabilities has been deduced for this ion.

Radiative lifetime measurements and oscillator strengths of astrophysical interest in Ho III

Radiative lifetimes of three long-lived levels belonging to the 4f(10)5d configuration of Ho III have been measured, for the first time, using the time-resolved laser-induced fluorescence technique. A good agreement between the experimental lifetimes and theoretical results obtained within a multiconfigurational pseudo-relativistic Hartree-Fock (HFR) approach including core-polarization (CP) effects is observed provided an adequate scaling factor is applied to the transition matrix element. From the comparison theory-experiment, it has been possible to assess the reliability of the oscillator strengths of the 4f-5d transitions of astrophysical interest, particularly for the lines depopulating the levels considered in the present work. The present results fill in a gap concerning the f-values of doubly ionized holmium and are needed for a quantitative evaluation of the holmium abundance in some peculiar stars. (Less)

Time-resolved laser spectroscopy of multiply ionized atoms: natural radiative lifetimes in Ce IV.

Radiative lifetimes have been measured for two excited levels of Ce IV using the time-resolved laser-induced fluorescence technique. Ce3+ ions were produced in a laser-induced plasma. In the measurements, a suitable magnetic field was applied to reduce the recombination between electrons and the ions and thus the background light from the recombination, and special care was exercised to avoid flight-out-of-view effects on the lifetime measurements for the high-velocity ions. The experimental lifetime results, tau = 30(2) ns for the level 49 737 cm(-1) and tau = 30(3) ns for the level 52 226 cm(-1), were compared with relativistic Hartree-Fock calculations (tau = 30.5 and 30.0 ns) indicating a particularly excellent agreement.

APPLICATIONS OF TIME‐RESOLVED FLUOROIMMUNOASSAY TO DETECT MAGNETIC BEAD CAPTURED ESCHERICHIA COLI O157:H7

A time‐resolved fluorescence technique was developed to detect Escherichia coli O157:H7 in ground beef burger. After a 4.5 h enrichment period, streptavidin coated magnetic beads conjugated with biotin‐labeled anti E. coli O157:H7 were used to capture the bacteria. The bacteria were, at the same time, also labeled by a nonfluorescent, europium (Eu)‐tagged anti‐E. coli O157:H7 antibody. The sandwiched bacterial complexes were then concentrated using a magnetic particle concentrator and washed to remove other solution components. Upon addition of an enhancement buffer, the Eu‐labels were then released from the antibodies and chelated to nitrilo‐triacetic acid (NTA) and trioctylphosphine oxide (TOPO) to form highly fluorescent Eu‐(2‐NTA)3(TOPO)2–3 micellar complexes. Delayed fluorescence associated with these complexes was measured and its intensity was used to estimate the original bacterial concentration spiked in hamburger. This approach was applied to detect E. coli O157:H7 spiked in hamburgers. The results indicated this method is able to detect 1 CFU/g of the bacteria after a brief enrichment for four and half hours at 37C. Specificity studies indicated that the approach exhibited no or limited cross reactivity to Salmonella typhimurium, E. coli K‐12 or Shigella dysenteriae spiked in hamburgers. Thus, the developed approach may be used as a rapid screening procedure for E. coli O157 bacteria in foods.

Lifetimes, Branching Fractions, and Oscillator Strengths of Doubly Ionized Tungsten

A first small set of W III oscillator strengths has been obtained from combined lifetime and branching fraction measurements. The branching fractions in the wavelength region of 154–334 nm were measured with a Penning discharge and a Fourier transform spectrometer. Three levels have been calibrated on absolute scales with lifetimes measured with the time-resolved laser-induced fluorescence technique. The f-values derived have uncertainties of about 8% at best. A comparison with Cowan-code calculations is given since no other data are available in the literature.

Radiative lifetimemeasurements in Tm III with time-resolved laser spectroscopyand comparisons with HFR calculations

Natural radiative lifetimes of eight levels in Tm III weremeasured with the time-resolved laser-induced fluorescence (LIF)technique. Free doubly ionized thulium ions were obtained in alaser-produced plasma. Three even-parity levels were excitedfrom the ground state with one-photon excitation, using a lasersystem generating pulses of 1 ns duration. Five odd-paritylevels were excited from the ground state with two-photonexcitation, where a picosecond laser system was employed. Inboth cases, the lifetimes were evaluated from transient LIFsignals detected with a fast-detection system. The experimentallifetime results were compared with Hartree-Fock calculationsincluding relativistic corrections. Good agreement was achievedfor the 4f126p levels while largerdiscrepancies are noted for some 4f125dlevels.

Time-resolved nonlinear laser-induced fluorescence technique for a combined lifetime and branching-fraction measurement

The single-shot time-resolved laser-induced fluorescence (LIF) technique is well established for measuring radiative lifetimes of atomic states. On top of that, we show that the branching fraction of the pumped transition can be determined and hence also the absolute transition probability, provided that the LIF signal has been sufficiently saturated by the laser pulse. This additional information is solely deduced from the temporal nonlinear behavior of the LIF signal during laser pumping. The alternative procedure is based on the fact that atoms that decay spontaneously into the initial state can be re-excited during the same laser pulse, whereas atoms that decay into third levels cannot. First measurements of this kind have been performed on singly ionized iron. Lifetimes, branching fractions, and resulting transition probabilities are presented and compared with literature data.

Europium Cryptate-Tethered Ribonucleotide for the Labeling of RNA and Its Detection by Time-Resolved Amplification of Cryptate Emission

TRACE (time-resolved amplification of cryptate emission), also called HTRF for pharmaceutical applications, is a homogeneous time-resolved fluorescence technique well adapted for the study of molecular interactions. It is based on fluorescence resonance energy transfer (FRET) between europium trisbipyridine cryptate (TBPEu3+) as energy donor and cross-linked allophycocyanin, symbolized by XL665, as acceptor, leading to a long-lived FRET signal. TBPEu3+-labeled uridine triphosphate (UTP), referred to as K-11-UTP in the text, was obtained by coupling TBPEu3+ moiety to a C-5 functionalized UTP analog. K-11-UTP can be directly incorporated in RNA strands during enzymatic synthesis. This was demonstrated in an in vitro transcription reaction promoted by T7 RNA polymerase. The reaction was performed in the presence of K-11-UTP and biotin-labeled cytidine triphosphate (biotin-16-CTP) in admixture with natural ribonucleotides. After the addition of streptavidin–XL665 conjugate (SA-XL665), which binds on biotinylated cytidine residues, a long-lived FRET signal was obtained. This proved that both europium cryptate and biotin were incorporated into the same RNA strand and are close enough to generate a FRET signal. The study of this FRET detection assay format showed that such doubly labeled RNA can be easily detected even when a very low percentage of K-11-UTP is used (less than 1% of total UTP concentration). Europium-cryptate-labeled RNA can also be monitored using a homogeneous hybridization assay format involving a biotinylated probe. After the addition of SA-XL665, the FRET signal generated demonstrates the formation of RNA:DNA hybrids. Europium-cryptate-labeled nucleotide thus gives access to a new type of RNA nonisotopic labeling and homogeneous detection assays.

Lifetime measurements in Ce I, Ce II, and Ce III using time-resolved laser spectroscopy with application to stellar abundance determinations of cerium

Radiative lifetimes of two levels in Ce I, eight levels in Ce II, and nine levels in Ce III have been measured using the time-resolved laser-induced fluorescence technique. Free cerium atoms and singly and doubly ionized ions were obtained in a laser-produced plasma. A narrow bandwidth UV laser pulse was employed to selectively populate the short-lived upper levels and the lifetime values were evaluated from the time-resolved fluorescence signals recorded by a fast detection system. Transition probabilities for Ce III were obtained from branching fractions calculated by the Cowan code and the experimental lifetimes. The results are compared with previous measurements and calculations. Spectral lines of Ce III were identified in the spectrum of the magnetic chemically peculiar star ${\ensuremath{\alpha}}^{2}\mathrm{CVn}$ and the abundance of cerium was determined from synthetic spectrum fitting to be 800 times greater than the solar abundance.

Reaction Kinetics at Dispersed-Colloid/Solution Interfaces: Benzophenone Triplet-State Quenching by Methylated Silica Particles

Dispersed silica colloids are used as model surfaces for studying triplet-state reaction kinetics at liquid/solid interfaces. Phosphorescence from the triplet state of benzophenone is quenched by a hydrogen-atom abstraction from methyl groups immobilized on the surface of a silica colloid. The production of diphenylketyl radicals from this Norrish Type II reaction and their decay by recombination are also monitored using a time-resolved laser-induced fluorescence technique. Aggregation of the colloids results in a dispersion of benzophenone-triplet quenching rates; the fractions of excited-triplet states quenched by small clusters and larger aggregates were estimated and used to determine rate constants for reaction with the two populations of surface methyl groups. The rates of the interfacial reactions of triplet benzophenone were enhanced compared to reactions in free solution, which is likely due to weak adsorption of triplet benzophenone to the methylated silica surface.

Detection of Metastable Chlorine Ions in Time-Modulated Plasma by Time Resolved Laser-Induced Fluorescence

Metastable chlorine ions (Cl+*) were detected in time-modulated inductively coupled plasma by a time-resolved laser-induced fluorescence technique. By varying the rf power, gas pressure and modulation frequency, the Cl+* density was measured in detail. In the discharge-on period, the Cl+* density increased as the rf power increased. In the discharge-off period, the rf power decreased within 1 µs, however, the Cl+* density decreased more slowly and persisted for a while. The decay constant was evaluated to be around 10 µs by fitting the decay curve under our experimental conditions. In addition, the diffusion coefficient of Cl+* was also estimated.

A linear Paul trap for radiative lifetime measurements on ions

A linear ion trap has been designed for collimation and storage of ions from an effusive particle beam generated in a hollow cathode discharge. The motivation is to measure radiative lifetimes on multiply ionized atoms by using the time-resolved laser-induced fluorescence technique. The trap has been tested on iron and tungsten ions both as an ion guide and also as a storing device. Radiative lifetimes have been measured in the trap with a remarkably higher signal-to-noise ratio than in the original effusive atom beam.

On-line time-resolved fluorescence spectroscopy of emulsion polymerizations to study the local dynamics in the polymerizing particles

An on-line time-resolved fluorescence technique is applied to study local dynamics in polymer particles during an emulsifier-free heterogeneous polymerization. The polymerization of styrene in the aqueous phase, initiated with a polymeric initiator (hydrolyzed propene-maleic acid copolymer with tert-butyl perester groups) was chosen as an illustrative system. To analyze the heterogeneous reaction pyrenyl probes were attached to the backbone of the initiator. Within the end phase of the polymerization an increase of fluorescence lifetime and intensity was observed. Fluorescence quenching experiments of the photo excited pyrene with dibenzoyl peroxide in toluene proved that the perester groups act as quencher. The calculated quenching constant points out that the quenching reaction is diffusion controlled. The also investigated bulk polymerization of styrene with dibenzoyl peroxide as initiator and pyrene as fluorophore showed that a change of the quenching probability at the gel point is responsible for a fluorescence lifetime and intensity enhancement. The emulsion polymerization showed the gel effect, too, although the macro-initiator forms a shell around the polystyrene core. At the gel point the chain mobility is reduced so that the encounter probability of pyrenyl and perester groups is limited. Despite the turbidity of the reaction mixture, fluorescence techniques allow to study the dynamics in the polymerizing particles.

The high-pressure range of the reaction CH(2Π)+CO+M⇒HCCO+M

The pulsed laser photolysis (PLP), time-resolved saturated laser-induced fluorescence (SLIF) technique has been used to study the reaction CH+CO+M⇒HCCO+M (1) in the temperature range between 300–800 K and at total pressures between 4 and 160 bar helium. The CH radicals have been generated using UV-multiphoton dissociation of CHBr3 at 248 nm. In the investigated temperature and pressure range, the usual falloff behavior of the reaction has been found. Falloff curves have been constructed and a high-pressure limiting rate constant with a weak negative temperature dependence of k1,∞=1.7×10−10(T/300 K)−0.4 cm3 molecule−1 s−1 has been extracted. This high-pressure limiting rate constant has been treated in terms of the statistical adiabatic channel model. Isobar Arrhenius curves have been constructed for pressures between 0.01 and 100 bar, including the formation of C2O+H at high temperatures.

Measurement of the rate constant for the association reaction CH + N2 at 53 K and its relevance to Triton's atmosphere

The pulsed laser photolysis (PLP), time‐resolved laser‐induced fluorescence (LIF) technique has been used to study the association reaction CH + N2 in a CRESU (Cinétique de Réaction en Ecoulement Supersonique Uniforme) apparatus at 53 K and in the pressure range 0.46–6 mbar. The third order rate constant was found to be ko = (5.7±0.30) × 10−30cm6s−1. Consequences for the production of HCN in the atmosphere of Triton are discussed in light of this result.

Determination of the limiting low pressure rate constants of the reactions of CH with N2 and CO: a CRESU measurement at 53 K

The pulsed laser photolysis, time-resolved laser-induced fluorescence technique has been implemented in a CRESU (Cine′tiquedeRe′actionenEcoulementSupersoniqueUniforme) apparatus to measure the rates of the association reactions of CH radicals with N2 and CO at 53 K and five different densities of the carrier gas Ar. The pressure-dependent rate constants have been combined with those reported previously for the temperature range 202⩽T/K⩽584 (R. A. Brownsword, L. B. Herbert, I. W. M. Smith and D. W. A. Stewart, J. Chem. Soc., FaradayTrans., 1996, 92, 1087) to yield, via a global fitting procedure, the following expressions for the two rate constants in the limit of low pressure: These expressions are very similar to those deduced previously by Brownsword etal., demonstrating that the analysis of rate constants for association reactions by the methods proposed by Troe (J. Chem. Phys., 1977, 66, 4758) can be applied over a wide range of temperature. These low temperature kinetic data, especially those for CH+N2, have some relevance to the chemistry of the atmospheres of some planets and planetary moons.

Measurement of internal stresses in coatings using time resolved fluorescence

AbstractThis paper describes a new nondestructive technique for detecting internal stresses in coatings using time resolved fluorescence. The measurement principle is based upon an experimental result that the decay time of fluorescence from poly(3‐octylthiophene), P3OT, dispersed in uniaxially‐stretched polymer films decreases with increasing tensile stress acting on the films. Internal stresses in the clear coat and in the base coat of a multilayer structure, which was composed of electrodeposited coat, surface coat, base coat, and clear coat, were estimated from the decay time of fluorescence from P3OT in both coats. The order of internal stresses in the clear coat and base coat of the multilayer system was 1 MPa. When the coatings were piled up, the internal stress decreased as the distance from a metal substrate was increased. It was found that moisture and temperature influenced internal stresses in the clear coat rather than in the base coat. Internal stress in the clear coat, which was one layer coated on a metal substrate, was measured by the traditional bimetallic method or by the time resolved fluorescence technique. Comparing both methods, it was concluded that the time resolved fluorescence technique gave reliable values for internal stresses in coatings.