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At least 469 records · Page 26

Joint reconstruction of x-ray fluorescence and transmission tomography

X-ray fluorescence tomography is based on the detection of fluorescence x-ray photons produced following x-ray absorption while a specimen is rotated; it provides information on the 3D distribution of selected elements within a sample. One limitation in the quality of sample recovery is the separation of elemental signals due to the finite energy resolution of the detector. Another limitation is the effect of self-absorption, which can lead to inaccurate results with dense samples. To recover a higher quality elemental map, we combine x-ray fluorescence detection with a second data modality: conventional x-ray transmission tomography using absorption. By using these combined signals in a nonlinear optimization-based approach, we demonstrate the benefit of our algorithm on real experimental data and obtain an improved quantitative reconstruction of the spatial distribution of dominant elements in the sample. Furthermore, compared with single-modality inversion based on x-ray fluorescence alone, this joint inversion approach reduces ill-posedness and should result in improved elemental quantification and better correction of self-absorption.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

Real-time image denoising of mixed Poisson–Gaussian noise in fluorescence microscopy images using ImageJ

Fluorescence microscopy imaging speed is fundamentally limited by the measurement signal-to-noise ratio (SNR). To improve image SNR for a given image acquisition rate, computational denoising techniques can be used to suppress noise. However, common techniques to estimate a denoised image from a single frame either are computationally expensive or rely on simple noise statistical models. These models assume Poisson or Gaussian noise statistics, which are not appropriate for many fluorescence microscopy applications that contain quantum shot noise and electronic Johnson–Nyquist noise, therefore a mixture of Poisson and Gaussian noise. In this paper, we show convolutional neural networks (CNNs) trained on mixed Poisson and Gaussian noise images to overcome the limitations of existing image denoising methods. The trained CNN is presented as an open-source ImageJ plugin that performs real-time image denoising (within tens of milliseconds) with superior performance (SNR improvement) compared to conventional fluorescence microscopy denoising methods. The method is validated on external datasets with out-of-distribution noise, contrast, structure, and imaging modalities from the training data and consistently achieves high-performance ( > <#comment/> 8 d B ) denoising in less time than other fluorescence microscopy denoising methods.

47 OTHER INSTRUMENTATION↗

Direct selection of functional fluorescent-protein antibody fusions by yeast display

Antibodies are important reagents for research, diagnostics, and therapeutics. Many examples of chimeric proteins combining the specific target recognition of antibodies with complementing functionalities such as fluorescence, toxicity or enzymatic activity have been described. However, antibodies selected solely on the basis of their binding specificities are not necessarily ideal candidates for the construction of chimeras. Here, we describe a high throughput method based on yeast display to directly select antibodies most suitable for conversion to fluorescent chimera. A library of scFv binders was converted to a fluorescent chimeric form, by cloning thermal green protein into the linker between VH and VL, and directly selecting for both binding and fluorescent functionality. This allowed us to directly identify antibodies functional in the single chain TGP format, that manifest higher protein expression, easier protein purification, and one-step binding assays.

59 BASIC BIOLOGICAL SCIENCES↗

Time-resolved and steady-state studies of biologically and chemically relevant systems using laser, absorption, and fluorescence spectroscopy

In Chapter 2 several experimental and data analysis methods used in this thesis are described. In Chapter 3 steady-state fluorescence spectroscopy was used to determine the concentration of the efflux pump inhibitors (EPIs), pheophorbide a and pyropheophorbide a, in the feces of animals and it was found that their levels far exceed those reported to be inhibitory to efflux pumps. In Chapter 4 the solvation dynamics of 6-Propionyl-2-(N,Ndimethyl) aminonaphthalene (PRODAN) was studied in reverse micelles. The two fluorescent states of PRODAN solvate on different time scales and as such care must be exercised in solvation dynamic studies involving it and its analogs. In Chapter 5 we studied the experimental and theoretical solvation dynamics of coumarin 153 (C153) in wild-type (WT) and modified myoglobins. Based on the nuclear magnetic resonance (NMR) spectroscopy and time-resolved fluorescence studies, we have concluded that it is important to thoroughly characterize the structure of a protein and probe system before comparing the theoretical and experimental results. In Chapter 6 the photophysical and spectral properties of a derivative of the medically relevant compound curcumin called cyclocurcumin was studied. Based on NMR, fluorescence, and absorption studies, the ground- and excited-states of cyclocurcumin are complicated by the existence of multiple structural isomers. In Chapter 7 the hydrolysis of cellulose by a pure form of cellulase in an ionic liquid, HEMA, and its aqueous mixtures at various temperatures were studied with the goal of increasing the cellulose to glucose conversion for biofuel production. It was found that HEMA imparts an additional stability to cellulase and can allow for faster conversion of cellulose to glucose using a pre-treatment step in comparison to only buffer.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Analysis of Environmental Swipes Using Fluorescence Microscopy - Summary Report

This report summarizes efforts as part of a collaboration between Oak Ridge National Laboratory (ORNL) and the University of Kansas (KU) during fiscal year 2016 for the purpose of screening environmental swipes (ESs) using fluorescence microscopy. The overall goal of this work is to image actinide-bearing species by exploiting the fluorescence properties of the uranyl moiety. If successfully developed, this capability would permit automated screening of actinide species in environmental swipes to prioritize samples for destructive analysis. The evaluation of efforts to date has shown that uranyl fluoride can be imaged using fluorescence microscopy and the vibronic structure can be resolved; however, difficulties arise when attempting to image these materials on ESs because of spectral interference. Due to the fluorescence decay differences between cellulosic fibers and uranyl-containing materials, time-gating approaches were investigated to minimize this interference. This approach shows promise; however, the technique requires additional developmental work before it can be transitioned to a robust method for laboratory use.

54 ENVIRONMENTAL SCIENCES↗

Synthesis and preliminary biological evaluations of fluorescent or 149 Promethium labeled Trastuzumab-polyethylenimine

Radioimmunotherapy utilize a targeting antibody coupled to a therapeutic isotope to target and treat a tumor or disease. In this study we examine the synthesis and cell binding of a polymer scaffold containing a radiotherapeutic isotope and a targeting antibody. Methods: The multistep synthesis of a fluorescent or 149 Promethium-labeled Trastuzumab-polyethyleneimine (PEI), Trastuzumab, or PEI is described. In vitro uptake, internalization and/or the binding affinity to the Her2/neu expressing human breast adenocarcinoma SKBr3 cells was investigated with the labeled compounds. Fluorescent-labeled Trastuzumab-PEI was internalized more into cells at 2 and 18 h than fluorescent-labeled Trastuzumab or PEI. The fluorescent-labeled Trastuzumab was concentrated on the cell surface at 2 and 18 h and the labeled PEI had minimal uptake. DOTA-PEI was prepared and contained an average of 16 chelates per PEI; the compound was radio-labeled with 149 Promethium and conjugated to Trastuzumab. The purified 149 Pm-DOTA-PEI-Trastuzumab had a radiochemical purity of 96.7% and a specific activity of 0.118 TBq/g. The compound demonstrated a dissociation constant for the Her2/neu receptor of 20.30 ± 6.91 nM. In conclusion, the results indicate the DOTA-PEI-Trastuzumab compound has potential as a targeted therapeutic carrier, and future in vivo studies should be performed.

07 ISOTOPE AND RADIATION SOURCES↗

Abundance of fluorescent biological aerosol particles at temperatures conducive to the formation of mixed-phase and cirrus clouds

Some types of biological particles are known to nucleate ice at warmer temperatures than mineral dust, with the potential to influence cloud microphysical properties and climate. However, the prevalence of these particle types above the atmospheric boundary layer is not well known. Many types of biological particles fluoresce when exposed to ultraviolet light, and the Wideband Integrated Bioaerosol Sensor takes advantage of this characteristic to perform real-time measurements of fluorescent biological aerosol particles (FBAPs). This instrument was flown on the National Center for Atmospheric Research Gulfstream V aircraft to measure concentrations of fluorescent biological particles from different potential sources and at various altitudes over the US western plains in early autumn. Clear-air number concentrations of FBAPs between 0.8 and 12 µm diameter usually decreased with height and generally were about 10–100 L -1 in the continental boundary layer but always much lower at temperatures colder than 255 K in the free troposphere. At intermediate temperatures where biological ice-nucleating particles may influence mixed-phase cloud formation (255 K ≤ T ≤ 270 K), concentrations of fluorescent particles were the most variable and were occasionally near boundary-layer concentrations. Predicted vertical distributions of ice-nucleating particle concentrations based on FBAP measurements in this temperature regime sometimes reached typical concentrations of primary ice in clouds but were often much lower. If convection was assumed to lift boundary-layer FBAPs without losses to the free troposphere, better agreement between predicted ice-nucleating particle concentrations and typical ice crystal concentrations was achieved. Ice-nucleating particle concentrations were also measured during one flight and showed a decrease with height, and concentrations were consistent with a relationship to FBAPs established previously at the forested surface site below. The vertical distributions of FBAPs measured on five flights were also compared with those for bacteria, fungal spores, and pollen predicted from the EMAC global chemistry–climate model for the same geographic region.

54 ENVIRONMENTAL SCIENCES↗

Viscosities, diffusion coefficients, and mixing times of intrinsic fluorescent organic molecules in brown limonene secondary organic aerosol and tests of the Stokes–Einstein equation

Viscosities and diffusion rates of organics within secondary organic aerosol (SOA) remain uncertain. Using the bead-mobility technique, we measured viscosities as a function of water activity ( a w ) of SOA generated by the ozonolysis of limonene followed by browning by exposure to NH 3 (referred to as brown limonene SOA or brown LSOA). These measurements together with viscosity measurements reported in the literature show that the viscosity of brown LSOA increases by 3–5 orders of magnitude as the a w decreases from 0.9 to approximately 0.05. In addition, we measured diffusion coefficients of intrinsic fluorescent organic molecules within brown LSOA matrices using rectangular area fluorescence recovery after photobleaching. Based on the diffusion measurements, as the a w decreases from 0.9 to 0.33, the average diffusion coefficient of the intrinsic fluorescent organic molecules decreases from 5.5 × 10 - 9 to 7.1 × 10 - 13 cm 2 s -1 and the mixing times of intrinsic fluorescent organic molecules within 200 nm brown LSOA particles increases from 0.002 to 14 s. These results suggest that the mixing times of large organics in the brown LSOA studied here are short ( < 1 h) for a w and temperatures often found in the planetary boundary layer (PBL). Since the diffusion coefficients and mixing times reported here correspond to SOA generated using a high mass loading ( ~1000 µ g m -3 ), biogenic SOA particles found in the atmosphere with mass loadings ≤10 µ g m -3 are likely to have higher viscosities and longer mixing times (possibly 3 orders of magnitude longer). These new measurements of viscosity and diffusion were used to test the accuracy of the Stokes–Einstein relation for predicting diffusion rates of organics within brown LSOA matrices. The results show that the Stokes–Einstein equation gives accurate predictions of diffusion coefficients of large organics within brown LSOA matrices when the viscosity of the matrix is as high as 10 2 to 10 4 Pa s. These results have important implications for predicting diffusion and mixing within SOA particles in the atmosphere.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Measurement report: A comparative analysis of an intensive incursion of fluorescing African dust particles over Puerto Rico and another over Spain

Measurements during episodes of African dust, made with two wideband integrated bioaerosol spectrometers (WIBSs), one on the northeastern coast of Puerto Rico and the other in the city of León, Spain, show unmistakable, bioaerosol-like fluorescing aerosol particles (FAPs) that can be associated with these dust episodes. The Puerto Rico event occurred during a major incursion of African dust during June 2020. The León event occurred in the late winter and spring of 2022, when widespread, elevated layers of dust inundated the Iberian Peninsula. Satellite and back-trajectory analyses confirm that dust from northern Africa was the source of the particles during both events. The WIBSs measure the size of individual particles in the range from 0.5 to 30 µm, derive a shape factor, and classify seven types of fluorescence from the FAPs. In general, it is not possible to directly determine the specific biological identity from fluorescence signatures; however, measurements of these types of bioaerosols in laboratory studies allow us to compare ambient fluorescence patterns with whole microbial cells measured under controlled conditions. Here we introduce some new metrics that offer a more quantitative approach for comparing FAP characteristics derived from particles measured under different environmental conditions. The analysis highlights the similarities and differences at the two locations and reveals differences that can be attributed to the age and history of the dust plumes, e.g., the amount of time that the air masses were in the mixed layer and the frequency of precipitation along the air mass trajectory.

54 ENVIRONMENTAL SCIENCES↗

Multiplet effects on the L/sub 2,3/ fluorescence yield of multiply ionized Ar

The 2p fluorescence yield of Ar in the presence of zero to six 3p holes has been calculated by statistically averaging the fluorescence yields of initial states that consist of individual multiplet configurations, formed by coupling the 2p vacancy to the partially filled 3p shell. The L(sub 2,3) fluorescence yields for the (2p)-1 (3p)-n configurations of Ar are found to be 1.48, 17.97, 24.83, 37.84, 79.61, 112.16, and 121.48 x .0001 for n = 0, 1, 2, 3, 4, 5, and 6, respectively. Results agree reasonably well with experimental fluorescence yields deduced from ion-atom collision measurements.

Chen, M. H.↗

Photomultiplier window materials under electron irradiation - Fluorescence and phosphorescence

The fluorescence and phosphorescence of photomultiplier window materials under electron irradiation have been investigated using a Sr-90/Y-90 beta emitter as the electron source. Spectral emission curves of UV-grade, optical-grade, and electron-irradiated samples of MgF2 and LiF, and of CaF2, BaF2, sapphire, fused silica, and UV-transmitting glasses were obtained over the 200-650-nm spectral range. Fluorescence yields were determined on these materials utilizing photomultiplier tubes with cesium telluride, bialkali, and trialkali (S-20) photocathodes, respectively. Optical-grade MgF2 and LiF, as well as electron-irradiated UV-grade samples of these two materials, show enhanced fluorescence due to color-center formation and associated emission bands in the blue and red wavelength regions. Large variations in fluorescence intensities were found in UV-grade sapphire samples of different origins, particularly in the red end of the spectrum, presumably due to various amounts of chromium-ion content. Phosphorescence decay with time is best described by a sum of exponential terms, with time constants ranging from a few minutes to several days.

Viehmann, W.↗

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

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.

Killinger, D. K.↗

Saturation of fluorescence in flames with a Gaussian laser beam

The method of saturated fluorescence for measuring species concentrations in flames is usually performed with laser beams that do not provide a constant intensity distribution across the focal volume. Because of the intensity distribution across the beam, the fluorescence signal does not depend on laser power or intensity in the same manner as for uniform illumination. This leads to anomolous apparent saturation intensities. In the following, the effect is considered for atomic fluorescence. Relations for the fluorescence signal under two common excitation geometries are derived and uncertainty relations used to consider the benefits of high laser intensity.

Daily, J. W.↗

Detectability limit and uncertainty considerations for laser induced fluorescence spectroscopy in flames

Laser induced fluorescence spectroscopy of flames is discussed, and derived uncertainty relations are used to calculate detectability limits due to statistical errors. Interferences due to Rayleigh scattering from molecules as well as Mie scattering and incandescence from particles have been examined for their effect on detectability limits. Fluorescence trapping is studied, and some methods for reducing the effect are considered. Fluorescence trapping places an upper limit on the number density of the fluorescing species that can be measured without signal loss.

Daily, J. W.↗

Laser excited fluorescence in the cesium-xenon excimer and the cesium dimer

Argon ion laser lines are used to excite fluorescence in a mixture of cesium and xenon. Excimer band fluorescence is observed at higher pressures (about 1 atm) while at lower pressures (several torr) a diffuse fluorescence due to the cesium dimer is observed whose character changes with exciting wavelength. The excimer fluorescence is shown to be directly related to the location of the exciting wavelength within previously measured Cs/Xe line shapes. This fact suggests that the excimer systems may be efficiently pumped through these line shapes. Qualitative energy-level schemes are proposed to explain the observations in both the excimer and dimer systems.

Exton, R. J.↗

Quantitative absorption and fluorescence studies of NO between 1060 and 2000 A

Synchrotron radiation in the 1060 to 2000 A region was used to measure the average absorption and fluorescence cross sections of NO and to determine approximate photodissociation quantum yields. Several vibrational levels of the D(2) sigma(+), E(2) sigma(+), and B(2) delta states have high fluorescence quantum yields. The C(2) and B(2) states do not fluoresce when the excitation energies are above the first dissociation limit, in accord with previous experiments. In general, the fluorescence yields decrease with increasing photon energy. The quantitative measurements are compared with spectroscopic observations and are found to be reasonably consistent.

Guest, J. A.↗

Fluorescent radiation converter

A fluorescence radiation converter is described which includes a substantially undoped optically transparent substrate and a waveshifter coating deposited on at least one portion of the substrate for absorption of radiation and conversion of fluorescent radiation. The coating is formed to substantially 1000 g/liter of a solvent, 70 to 200 g/liter of an organic polymer, and 0.2 to 25 g/liter of at least one organic fluorescent dye. The incoming incident radiation impinges on the coating. Radiation is absorbed by the fluorescent dye and is re-emitted as a longer wavelength radiation. Radiation is trapped within the substrate and is totally internally reflected by the boundary surface. Emitted radiation leaves the substrate ends to be detected.

Viehmann, W.↗

Laser measurement of the spectral extinction coefficients of fluorescent, highly absorbing liquids

A conceptual method is developed to deduce rapidly the spectral extinction coefficient of fluorescent, highly absorbing liquids, such as crude or refined petroleum oils. The technique offers the advantage of only requiring one laser wavelength and a single experimental assembly and execution for any specific fluorescent liquid. The liquid is inserted into an extremely thin wedge-shaped cavity for stimulation by a laser from one side and flurescence measurement on the other side by a monochromator system. For each arbitrarily selected extinction wavelength, the wedge is driven slowly to increasing thicknesses until the fluorescence extinguishes. The fluorescence as a function of wedge thickness permits a determination of the extinction coefficient using an included theoretical model. When the monochromator is set to the laser emission wavelength, the extinction coefficient is determined using the usual on-wavelength signal extinction procedure.

Hoge, F. E.↗