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1

256 × 2 SPAD line sensor for time resolved fluorescence spectroscopy.  

PubMed

We present a CMOS chip 256 × 2 single photon avalanche diode (SPAD) line sensor, 23.78 µm pitch, 43.7% fill factor, custom designed for time resolved emission spectroscopy (TRES). Integrating time-to-digital converters (TDCs) implement on-chip mono-exponential fluorescence lifetime pre-calculation allowing timing of 65k photons/pixel at 200 Hz line rate at 40 ps resolution using centre-of-mass method (CMM). Per pixel time-correlated single-photon counting (TCSPC) histograms can also be generated with 320 ps bin resolution. We characterize performance in terms of dark count rate, instrument response function and lifetime uniformity for a set of fluorophores with lifetimes ranging from 4 ns to 6 ns. Lastly, we present fluorescence lifetime spectra of multicolor microspheres and skin autofluorescence acquired using a custom built spectrometer. In TCSPC mode, time-resolved spectra are acquired within 5 minutes whilst in CMM mode spectral lifetime signatures are acquired within 2 ms for fluorophore in cuvette and 200 ms for skin autofluorescence. We demonstrate CMOS line sensors to be a versatile tool for time-resolved fluorescence spectroscopy by providing parallelized and flexible spectral detection of fluorescence decay. PMID:25836796

Krstaji?, Nikola; Levitt, James; Poland, Simon; Ameer-Beg, Simon; Henderson, Robert

2015-03-01

2

Diagnosis of meningioma by time-resolved fluorescence spectroscopy  

PubMed Central

We investigate the use of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) as an adjunctive tool for the intraoperative rapid evaluation of tumor specimens and delineation of tumor from surrounding normal tissue. Tissue autofluorescence is induced with a pulsed nitrogen laser (337 nm, 1.2 ns) and the intensity decay profiles are recorded in the 370 to 500 nm spectral range with a fast digitizer (0.2 ns resolution). Experiments are conducted on excised specimens (meningioma, dura mater, cerebral cortex) from 26 patients (97 sites). Spectral intensities and time-dependent parameters derived from the time-resolved spectra of each site are used for tissue characterization. A linear discriminant analysis algorithm is used for tissue classification. Our results reveal that meningioma is characterized by unique fluorescence characteristics that enable discrimination of tumor from normal tissue with high sensitivity (>89%) and specificity (100%). The accuracy of classification is found to increase (92.8% cases in the training set and 91.8% in the cross-validated set correctly classified) when parameters from both the spectral and the time domain are used for discrimination. Our findings establish the feasibility of using TR-LIFS as a tool for the identification of meningiomas and enables further development of real-time diagnostic tools for analyzing surgical tissue specimens of meningioma or other brain tumors. PMID:16409091

Butte, Pramod V.; Pikul, Brian K.; Hever, Aviv; Yong, William H.; Black, Keith L.; Marcu, Laura

2010-01-01

3

Time-resolved Hyperspectral Fluorescence Spectroscopy using Frequency Modulated Excitation  

SciTech Connect

An intensity-modulated excitation light source is used together with a micro channel plate intensified CCD (ICCD) detector gated at a slightly different frequency to generate a beat frequency from a fluorescent sample. The addition of a spectrograph produces a hyperspectral time-resolved data product where the resulting beat frequency is detected with a low frame rate camera. Measuring the beat frequency of the spectrum as a function of time allows separation of the excited fluorescence from ambient constant light sources. The excitation and detector repetition rates are varied over a range of discrete frequencies, and the phase shift of the beat wave maps out the emission decay rate(s).

,; Neill, M

2012-07-01

4

Picosecond time-resolved fluorescence spectroscopy of phytochrome and stentorin  

NASA Astrophysics Data System (ADS)

Phytochrome is a tetrapyrrole chromoprotein. It serves as a sensitive photosensor for red lightmediated gene expression and other developmental/morphological responses in plants. In this paper photochemical dynamics of the phytochrome molecule have been described in terms of photoisomerization of the tetrapyrrole chromophore in its singlet excited state and subsequent thermal processes in the Pr Pfr phototransformation of phytochrome. Stentorin acts as the photosensor molecule in the ciliate Stentor coeruleus. This unicellular protozoan is most sensitive to red light (610-620 urn). Stentor also senses the direction of light propagation as evidenced by their light-avoiding and negative phototactic swimming behaviors. This aneural photosensory phenomenon is triggered by the photoreceptor stentorin. The possible involvement of a light-induced transient proton release from the photoreceptor as the primary mechanism of light-signal processing has been discussed on the basis of picosecond fluorescence decays and time-resolved fluorescence spectra of stentorin in solution. An initial sensory signal generated by the primary photoprocess of stentorin then triggers subsequent transduction steps that include calcium ion influx from the extracellular medium. Calcium ion influx from the extracellular medium to the cytosol causes the Stentor cell to reverse its ciliary beating and subsequently steer away from the light trap. II.

Song, Pill-Soon

1991-05-01

5

Conformational Analysis of DNA Repair Intermediates by Time-Resolved Fluorescence Spectroscopy  

E-print Network

to their DNA modification. This included a nicked DNA backbone (NICK), a backbone with a gap of one and twoConformational Analysis of DNA Repair Intermediates by Time-Resolved Fluorescence Spectroscopy Su General Hospital, Boston, Massachusetts 02114 ReceiVed: July 16, 2009 DNA repair enzymes are essential

Heller, Eric

6

Multispectral scanning time-resolved fluorescence spectroscopy (TRFS) technique for intravascular diagnosis  

PubMed Central

This study describes a scanning time-resolved fluorescence spectroscopy (TRFS) system designed to continuously acquire fluorescence emission and to reconstruct fluorescence lifetime images (FLIM) from a luminal surface by using a catheter-based optical probe with rotary joint and pull-back device. The ability of the system to temporally and spectrally resolve the fluorescence emission from tissue was validated using standard dyes and tissue phantoms (e.g., ex vivo pig aorta phantom). Current results demonstrate that this system is capable to reliably resolve the fluorescence emission of multiple fluorophores located in the lumen; and suggest its potential for intravascular detection of distinct biochemical features of atherosclerotic plaques. PMID:22808425

Xie, Hongtao; Bec, Julien; Liu, Jing; Sun, Yang; Lam, Matthew; Yankelevich, Diego R.; Marcu, Laura

2012-01-01

7

Fluorescence imaging and time-resolved spectroscopy of steroid using confocal synchrotron radiation microscopy  

NASA Astrophysics Data System (ADS)

The Confocal Synchrotron Radiation Microscope at Daresbury was used in a study of the transport and distribution of the steroid Coumestrol in single Leydig cells. The broad spectrum of synchrotron radiation in combination with UV compatible microscope optics affords the extension of confocal microscopy from the visible to the UV region down to about 200 nm. Consequently fluorescent molecules with absorption bands in the UV can be imaged. In addition the pulsed nature of the light source allows us to perform time-resolved fluorescence spectroscopy experiments on microscopic volumes. Coumestrol is a naturally fluorescing plant steroid exhibiting estrogenic activity. In physiological environments it has an absorption peak in the UV at 340 nm and it emits around 440 nm. First results indicate that the Coumestrol transport through the cell membrane is diffusion limited. The weak fluorescence observed in the nuclei of the Leydig cells may be due to fluorescence quenching arising from the interaction of the Coumesterol with nuclear components. However, micro-volume time-resolved fluorescence spectroscopy experiments on cell nuclei have revealed the same decay behavior for Coumesterol in both the cytoplasm and nucleus of the cells.

Gerritsen, Hans C.; van der Oord, C. J. R.; Levine, Yehudi K.; Munro, Ian H.; Jones, Gareth R.; Shaw, D. A.; Rommerts, Fokko F.

1994-08-01

8

Tubulin equilibrium unfolding followed by time-resolved fluorescence and fluorescence correlation spectroscopy  

PubMed Central

The pathway for the in vitro equilibrium unfolding of the tubulin heterodimer by guanidinium chloride (GdmCl) has been studied using several spectroscopic techniques, specifically circular dichroism (CD), two-photon Fluorescence Correlation Spectroscopy (FCS), and time-resolved fluorescence, including lifetime and dynamic polarization. The results show that tubulin unfolding is characterized by distinct processes that occur in different GdmCl concentration ranges. From 0 to 0.5 M GdmCl, a slight alteration of the tubulin heterodimer occurs, as evidenced by a small, but reproducible increase in the rotational correlation time of the protein and a sharp decrease in the secondary structure monitored by CD. In the range 0.5–1.5 M GdmCl, significant decreases in the steady-state anisotropy and average lifetime of the intrinsic tryptophan fluorescence occur, as well as a decrease in the rotational correlation time, from 48 to 26 nsec. In the same GdmCl range, the number of protein molecules (labeled with Alexa 488), as determined by two-photon FCS measurements, increases by a factor of two, indicating dissociation of the tubulin dimer into monomers. From 1.5 to 4 M GdmCl, these monomers unfold, as evidenced by the continual decrease in the tryptophan steady-state anisotropy, average lifetime, and rotational correlation time, concomitant with secondary structural changes. These results help to elucidate the unfolding pathway of the tubulin heterodimer and demonstrate the value of FCS measurements in studies on oligomeric protein systems. PMID:14691224

Sánchez, Susana A.; Brunet, Juan E.; Jameson, David M.; Lagos, Rosalba; Monasterio, Octavio

2004-01-01

9

Time-Resolved Fluorescence Spectroscopy as a Diagnostic Technique of Oral Carcinoma  

PubMed Central

Objective To investigate the benefit of using time-resolved, laser-induced fluorescence spectroscopy for diagnosing malignant and premalignant lesions of the oral cavity. Design The carcinogen 7,12-dimethylbenz[a]anthracene (DMBA) was applied to 1 cheek pouch of 19 hamsters. The contralateral pouch and the cheek pouches of 3 hamsters without DMBA exposure served as controls. Setting University of California, Davis. Participants Twenty-two golden/Syrian hamsters. Intervention A nitrogen pulse laser was used to induce tissue autofluorescence between the wavelengths of 360 and 650 nm. Main Outcome Measures Spectral intensities and time-domain measurements were obtained and compared with the histopathologic findings at each corresponding site. Results Spectral intensities and lifetime values at 3 spectral bands (SBs; SB1=380±10 nm; SB2=460±10 nm, and SB3 = 635 ± 10 nm) allowed for discrimination among healthy epithelium, dysplasia, carcinoma in situ, and invasive carcinoma. The lifetime values at SB2 were the most important when distinguishing the lesions using only time-resolved parameters. An algorithm combining spectral fluorescence parameters derived from both spectral and time-domain parameters (peak intensities, average fluorescence lifetimes, and the Laguerre coefficient [zero-order]) for healthy epithelium, dysplasia, carcinoma in situ, and invasive carcinoma provided the best diagnostic discrimination, with 100%, 100%, 69.2%, and 76.5% sensitivity and 100%, 92.2%, 97.1%, and 96.2% specificity, respectively. Conclusions The addition of time-resolved fluorescence-derived parameters significantly improves the capability of fluorescence spectroscopy–based diagnostics in the hamster buccal pouch. This technique provides a potential non-invasive diagnostic instrument for head and neck cancer. PMID:20157056

Farwell, D. Gregory; Meier, Jeremy D.; Park, Jesung; Sun, Yang; Coffman, Heather; Poirier, Brian; Phipps, Jennifer; Tinling, Steve; Enepekides, Danny J.; Marcu, Laura

2014-01-01

10

Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging  

SciTech Connect

The application of time-resolved fluorescence spectroscopy (TRFS) to in vivo tissue diagnosis requires a method for fast acquisition of fluorescence decay profiles in multiple spectral bands. This study focusses on development of a clinically compatible fiber-optic based multispectral TRFS (ms-TRFS) system together with validation of its accuracy and precision for fluorescence lifetime measurements. It also presents the expansion of this technique into an imaging spectroscopy method. A tandem array of dichroic beamsplitters and filters was used to record TRFS decay profiles at four distinct spectral bands where biological tissue typically presents fluorescence emission maxima, namely, 390, 452, 542, and 629 nm. Each emission channel was temporally separated by using transmission delays through 200 ?m diameter multimode optical fibers of 1, 10, 19, and 28 m lengths. A Laguerre-expansion deconvolution algorithm was used to compensate for modal dispersion inherent to large diameter optical fibers and the finite bandwidth of detectors and digitizers. The system was found to be highly efficient and fast requiring a few nano-Joule of laser pulse energy and <1 ms per point measurement, respectively, for the detection of tissue autofluorescent components. Organic and biological chromophores with lifetimes that spanned a 0.8–7 ns range were used for system validation, and the measured lifetimes from the organic fluorophores deviated by less than 10% from values reported in the literature. Multi-spectral lifetime images of organic dye solutions contained in glass capillary tubes were recorded by raster scanning the single fiber probe in a 2D plane to validate the system as an imaging tool. The lifetime measurement variability was measured indicating that the system provides reproducible results with a standard deviation smaller than 50 ps. The ms-TRFS is a compact apparatus that makes possible the fast, accurate, and precise multispectral time-resolved fluorescence lifetime measurements of low quantum efficiency sub-nanosecond fluorophores.

Yankelevich, Diego R. [Department of Electrical and Computer Engineering, University of California, 3101 Kemper Hall, Davis, California 95616 (United States) [Department of Electrical and Computer Engineering, University of California, 3101 Kemper Hall, Davis, California 95616 (United States); Department of Biomedical Engineering, University of California, 451 Health Sciences Drive, Davis, California 95616 (United States); Ma, Dinglong; Liu, Jing; Sun, Yang; Sun, Yinghua; Bec, Julien; Marcu, Laura, E-mail: lmarcu@ucdavis.edu [Department of Biomedical Engineering, University of California, 451 Health Sciences Drive, Davis, California 95616 (United States)] [Department of Biomedical Engineering, University of California, 451 Health Sciences Drive, Davis, California 95616 (United States); Elson, Daniel S. [Hamlyn Centre for Robotic Surgery, Department of Surgery and Cancer, Imperial College London, Exhibition Road, London SW7 2AZ (United Kingdom)] [Hamlyn Centre for Robotic Surgery, Department of Surgery and Cancer, Imperial College London, Exhibition Road, London SW7 2AZ (United Kingdom)

2014-03-15

11

Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging  

NASA Astrophysics Data System (ADS)

The application of time-resolved fluorescence spectroscopy (TRFS) to in vivo tissue diagnosis requires a method for fast acquisition of fluorescence decay profiles in multiple spectral bands. This study focusses on development of a clinically compatible fiber-optic based multispectral TRFS (ms-TRFS) system together with validation of its accuracy and precision for fluorescence lifetime measurements. It also presents the expansion of this technique into an imaging spectroscopy method. A tandem array of dichroic beamsplitters and filters was used to record TRFS decay profiles at four distinct spectral bands where biological tissue typically presents fluorescence emission maxima, namely, 390, 452, 542, and 629 nm. Each emission channel was temporally separated by using transmission delays through 200 ?m diameter multimode optical fibers of 1, 10, 19, and 28 m lengths. A Laguerre-expansion deconvolution algorithm was used to compensate for modal dispersion inherent to large diameter optical fibers and the finite bandwidth of detectors and digitizers. The system was found to be highly efficient and fast requiring a few nano-Joule of laser pulse energy and <1 ms per point measurement, respectively, for the detection of tissue autofluorescent components. Organic and biological chromophores with lifetimes that spanned a 0.8-7 ns range were used for system validation, and the measured lifetimes from the organic fluorophores deviated by less than 10% from values reported in the literature. Multi-spectral lifetime images of organic dye solutions contained in glass capillary tubes were recorded by raster scanning the single fiber probe in a 2D plane to validate the system as an imaging tool. The lifetime measurement variability was measured indicating that the system provides reproducible results with a standard deviation smaller than 50 ps. The ms-TRFS is a compact apparatus that makes possible the fast, accurate, and precise multispectral time-resolved fluorescence lifetime measurements of low quantum efficiency sub-nanosecond fluorophores.

Yankelevich, Diego R.; Ma, Dinglong; Liu, Jing; Sun, Yang; Sun, Yinghua; Bec, Julien; Elson, Daniel S.; Marcu, Laura

2014-03-01

12

Structure and dynamics of a DNA: polymerase complex by time-resolved fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

The interaction of a fluorescent DNA primer:template with the Klenow fragment of DNA polymerase I has been studied in solution using time-resolved fluorescence spectroscopy. The excited-state decay behavior and internal reorientation dynamics of a dansyl sulfonamide probe connected by a propyl chain to a modified uridine base in the primer strand were very sensitive to the local probe environment and exhibited characteristic changes upon binding of Kienow fragment to the DNA and elongation of the primer strand. Between 5 and 7 bases of duplex DNA upstream of the 3' primer terminus were protected from the solvent by the Kienow fragment and the strength of DNA:protein contacts varied within this region, being strongest at the 3' primer terminus. About 5% of the substrates were bound in a second spatially distinct site on the enzyme. Site-directed mutagenesis of the Kienow fragment was consistent with this being the active site for 3'->5' exonuclease activity.

Millar, David P.; Benkovic, Stephen J.

1990-05-01

13

Distinction of brain tissue, low grade and high grade glioma with time-resolved fluorescence spectroscopy.  

PubMed

Neuropathology frozen section diagnoses are difficult in part because of the small tissue samples and the paucity of adjunctive rapid intraoperative stains. This study aims to explore the use of time-resolved laser-induced fluorescence spectroscopy as a rapid adjunctive tool for the diagnosis of glioma specimens and for distinction of glioma from normal tissues intraoperatively. Ten low grade gliomas, 15 high grade gliomas without necrosis, 6 high grade gliomas with necrosis and/or radiation effect, and 14 histologically uninvolved "normal" brain specimens are spectroscopicaly analyzed and contrasted. Tissue autofluorescence was induced with a pulsed Nitrogen laser (337 nm, 1.2 ns) and the transient intensity decay profiles were recorded in the 370-500 nm spectral range with a fast digitized (0.2 ns time resolution). Spectral intensities and time-dependent parameters derived from the time-resolved spectra of each site were used for tissue characterization. A linear discriminant analysis diagnostic algorithm was used for tissue classification. Both low and high grade gliomas can be distinguished from histologically uninvolved cerebral cortex and white matter with high accuracy (above 90%). In addition, the presence or absence of treatment effect and/or necrosis can be identified in high grade gliomas. Taking advantage of tissue autofluorescence, this technique facilitates a direct and rapid investigation of surgically obtained tissue. PMID:16368511

Yong, William H; Butte, Pramod V; Pikul, Brian K; Jo, Javier A; Fang, Qiyin; Papaioannou, Thanassis; Black, Keith; Marcu, Laura

2006-01-01

14

Distinction of brain tissue, low grade and high grade glioma with time-resolved fluorescence spectroscopy  

PubMed Central

Neuropathology frozen section diagnoses are difficult in part because of the small tissue samples and the paucity of adjunctive rapid intraoperative stains. This study aims to explore the use of time-resolved laser-induced fluorescence spectroscopy as a rapid adjunctive tool for the diagnosis of glioma specimens and for distinction of glioma from normal tissues intraoperatively. Ten low grade gliomas, 15 high grade gliomas without necrosis, 6 high grade gliomas with necrosis and/or radiation effect, and 14 histologically uninvolved “normal” brain specimens are spectroscopicaly analyzed and contrasted. Tissue autofluorescence was induced with a pulsed Nitrogen laser (337 nm, 1.2 ns) and the transient intensity decay profiles were recorded in the 370-500 nm spectral range with a fast digitized (0.2 ns time resolution). Spectral intensities and time-dependent parameters derived from the time-resolved spectra of each site were used for tissue characterization. A linear discriminant analysis diagnostic algorithm was used for tissue classification. Both low and high grade gliomas can be distinguished from histologically uninvolved cerebral cortex and white matter with high accuracy (above 90%). In addition, the presence or absence of treatment effect and/or necrosis can be identified in high grade gliomas. Taking advantage of tissue autofluorescence, this technique facilitates a direct and rapid investigation of surgically obtained tissue. PMID:16368511

Yong, William H.; Butte, Pramod V.; Pikul, Brian K.; Jo, Javier A.; Fang, Qiyin; Papaioannou, Thanassis; Black, Keith L.; Marcu, Laura

2010-01-01

15

Intraoperative delineation of primary brain tumors using time-resolved fluorescence spectroscopy  

PubMed Central

The goal of this study is to determine the potential of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) as an adjunctive tool for delineation of brain tumor from surrounding normal tissue in order to assist the neurosurgeon in near-complete tumor excision. A time-domain TR-LIFS prototype apparatus (gated photomultiplier detection, fast digitizer) was used for recording tissue autofluorescence in normal cortex (NC), normal white matter (NWM), and various grades of gliomas intraoperatively. Tissue fluorescence was induced with a pulsed nitrogen laser (337nm, 700ps), and the intensity decay profiles were recorded in the 360-to550-nm spectral range (10-nm interval). Histopathological analysis (hematoxylin & eosin) of the biopsy samples taken from the site of TR-LIFS measurements was used for validation of spectroscopic results. Preliminary results on 17 patients demonstrate that normal cortex (N=16) and normal white matter (N=3) show two peaks of fluorescence emission at 390nm(lifetime=1.8±0.3ns) and 460nm(lifetime=0.8±0.1ns). The 390-nm emission peak is absent in low-grade glioma (N=5; lifetime=1.1ns) and reduced in high-grade glioma (N=9; lifetime=1.7±0.4ns). The emission characteristics at 460nm in all tissues correlated with the nicotinamide adenine dinucleotide fluorescence (peak: 440to460nm; lifetime: 0.8to1.0ns). These findings demonstrate the potential of using TR-LIFS as a tool for enhanced delineation of brain tumors during surgery. In addition, this study evaluates similarities and differences between TR-LIFS signatures of brain tumors obtained in vivo and those previously reported in ex vivo brain tumor specimens. PMID:20459282

Butte, Pramod V.; Fang, Qiyin; Jo, Javier A.; Yong, William H.; Pikul, Brian K.; Black, Keith L.; Marcu, Laura

2010-01-01

16

Conformational Analysis of DNA Repair Intermediates by Time-Resolved Fluorescence Spectroscopy  

PubMed Central

DNA repair enzymes are essential for maintaining the integrity of the DNA sequence. Unfortunately, very little is known about how these enzymes recognize damaged regions along the helix. Structural analysis of cellular repair enzymes bound to DNA reveals that these enzymes are able to recognize DNA in a variety of conformations. However, the prevalence of these deformations in the absence of enzymes remains unclear, as small populations of DNA conformations are often difficult to detect by NMR and X-ray crystallography. Here, we used time-resolved fluorescence spectroscopy to examine the conformational dynamics of linear, nicked, gapped, and bulged DNA in the absence of protein enzymes. This analysis reveals that damaged DNA is polymorphic in nature and able to adopt multiple individual conformations. We show that DNA repair intermediates that contain a one-nucleotide gap and bulge have a significant propensity to adopt conformations in which the orphan base resides outside the DNA helix, while DNA structures damaged by a nick or two-nucleotide gap favor intrahelical conformations. Because changes in DNA conformation appear to guide the recognition of DNA repair enzymes, we suggest that the current approach could be used to study the mechanism of DNA repair. PMID:19673467

2009-01-01

17

Dynamics and Flexibility of Human Aromatase Probed by FTIR and Time Resolved Fluorescence Spectroscopy  

PubMed Central

Human aromatase (CYP19A1) is a steroidogenic cytochrome P450 converting androgens into estrogens. No ligand-free crystal structure of the enzyme is available to date. The crystal structure in complex with the substrate androstenedione and the steroidal inhibitor exemestane shows a very compact conformation of the enzyme, leaving unanswered questions on the conformational changes that must occur to allow access of the ligand to the active site. As H/D exchange kinetics followed by FTIR spectroscopy can provide information on the conformational changes in proteins where solvent accessibility is affected, here the amide I region was used to measure the exchange rates of the different elements of the secondary structure for aromatase in the ligand-free form and in the presence of the substrate androstenedione and the inhibitor anastrozole. Biphasic exponential functions were found to fit the H/D exchange data collected as a function of time. Two exchange rates were assigned to two populations of protons present in different flexible regions of the protein. The addition of the substrate androstenedione and the inhibitor anastrozole lowers the H/D exchange rates of the ?-helices of the enzyme when compared to the ligand-free form. Furthermore, the presence of the inhibitor anastrozole lowers exchange rate constant (k1) for ?-sheets from 0.22±0.06 min?1 for the inhibitor-bound enzyme to 0.12±0.02 min?1 for the free protein. Dynamics effects localised in helix F were studied by time resolved fluorescence. The data demonstrate that the fluorescence lifetime component associated to Trp224 emission undergoes a shift toward longer lifetimes (from ?5.0 to ?5.5 ns) when the substrate or the inhibitor are present, suggesting slower dynamics in the presence of ligands. Together the results are consistent with different degrees of flexibility of the access channel and therefore different conformations adopted by the enzyme in the free, substrate- and inhibitor-bound forms. PMID:24349198

Sadeghi, Sheila J.; Castrignanò, Silvia; Mei, Giampiero; Di Venere, Almerinda; Nicolai, Eleonora; Allegra, Paola; Gilardi, Gianfranco

2013-01-01

18

Development of a Time Resolved Fluorescence Spectroscopy System for Near Real-Time Clinical Diagnostic Applications  

E-print Network

tool to characterize lifetimes and autofluorescence of normal brain tissue and glioma. They also differentiated between normal cerebral cortex regions and normal white matter, and could categorize gliomas into low grade glioma, high grade glioma... and high grade glioma with necrotic change with reasonable accuracy using time resolved fluorescence parameters. Lifetime Kinetics and Analysis Data analysis techniques for time-domain TRFS methods will be reviewed and discussed here as the work...

Trivedi, Chintan A.

2010-07-14

19

Time-resolved hyperspectral fluorescence spectroscopy using frequency-modulated excitation  

NASA Astrophysics Data System (ADS)

An intensity-modulated excitation light source is used together with a micro-channel plate intensified CCD detector gated at a slightly different frequency to generate a beat frequency from a fluorescent sample. The addition of a spectrograph produces a hyperspectral time-resolved data product where the resulting beat frequency is detected with a low frame rate camera. Measuring the beat frequency of the spectrum as a function of time allows separation of the excited fluorescence from ambient constant light sources. The excitation and detector repetition rates are varied over a range of discrete frequencies, and the phase shift of the beat wave maps out the emission decay rate(s).

DiBenedetto, John; Capelle, Gene A.; O'Neill, Mary

2012-07-01

20

Time-resolved and steady-state fluorescence spectroscopy from bacteria subjected to bactericidal agents  

NASA Astrophysics Data System (ADS)

The time-resolved and steady-state changes in fluorescence were investigated from one spore-forming (Bacillus subtilis) and four non-spore forming (Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, and Pseudomonas aeruginosa) bacteria subjected to different bactericidal agents. The bactericidal agents were sodium hypochlorite (bleach) hydrogen peroxide, formaldehyde, and UV light exposure. Application of sodium hypochlorite resulted in an almost total lose of fluorescence signal and large decrease in the optical density of the bacterial suspension. Addition of hydrogen peroxide resulted in a 35% decrease in emission intensity fom the Sa and an 85-95% decrease for the other bacteria. Ultraviolet light exposure resulted in a 5-35% decrease in the emission intensity of the tryptophan band. The addition of formaldehyde to the bacteria did not result in significant changes in the steady-state emission intensity, but did shift the tryptophan emission peak position to shorter wavelengths by 3 to 5 nm. Time-resolved fluorescence measurements showed that the fluorescence lifetime of tryptophan in the bacteria could not be described by a single exponential decay, and was similar to that of tryptophan in neutral aqueous solution. Upon addition of formaldehyde to the Gram positive bacteria (Bs and Sa) the strength of the short lifetime component increased dramatically, while for the Gram negative bacteria, a smaller increase was observed. These fluorescence changes reflect the different mechanisms of the bactericidal agents and may provide a useful tool to monitor the effectiveness of disinfectants.

Katz, Alvin; Alimova, Alexandra; Siddique, Masood; Savage, Howard E.; Shah, Mahendra; Rosen, Richard; Alfano, Robert

2004-03-01

21

Pyrene binary probes for unambiguous detection of mRNA using time-resolved fluorescence spectroscopy  

PubMed Central

We report here the design, synthesis and application of pyrene binary oligonucleotide probes for selective detection of cellular mRNA. The detection strategy is based on the formation of a fluorescent excimer when two pyrene groups are brought into close proximity upon hybridization of the probes with the target mRNA. The pyrene excimer has a long fluorescence lifetime (>40 ns) compared with that of cellular extracts (?7 ns), allowing selective detection of the excimer using time-resolved emission spectra (TRES). Optimized probes were used to target a specific region of sensorin mRNA yielding a strong excimer emission peak at 485 nm in the presence of the target and no excimer emission in the absence of the target in buffer solution. While direct fluorescence measurement of neuronal extracts showed a strong fluorescent background, obscuring the detection of the excimer signal, time-resolved emission measurements indicated that the emission decay of the cellular extracts is ?8 times faster than that of the pyrene excimer probes. Thus, using TRES of the pyrene probes, we are able to selectively detect mRNA in the presence of cellular extracts, demonstrating the potential for application of pyrene excimer probes for imaging mRNAs in cellular environments that have background fluorescence. PMID:16769776

Martí, Angel A.; Li, Xiaoxu; Jockusch, Steffen; Li, Zengmin; Raveendra, Bindu; Kalachikov, Sergey; Russo, James J.; Morozova, Irina; Puthanveettil, Sathyanarayanan V.; Ju, Jingyue; Turro, Nicholas J.

2006-01-01

22

Detection of Rupture-Prone Atherosclerotic Plaques by Time-Resolved Laser Induced Fluorescence Spectroscopy  

PubMed Central

Objective Plaque with dense inflammatory cells, including macrophages, thin fibrous cap and superficial necrotic/lipid core is thought to be prone-to-rupture. We report a time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) technique for detection of such markers of plaque vulnerability in human plaques. Methods The autofluorescence of carotid plaques (65 endarterectomy patients) induced by a pulsed laser (337 nm, 0.7 ns) was measured from 831 distinct areas. The emission was resolved spectrally- (360–550 nm range) and temporally- (0.3 ns resolution) using a prototype fiber-optic TR-LIFS apparatus. Lesions were evaluated microscopically and quantified as to the % of different components (fibrous cap, necrotic core, inflammatory cells, foam cells, mature and degraded collagen, elastic fibers, calcification, and smooth muscle cell of the vessel wall). Results We determined that the spectral intensities and time-dependent parameters at discrete emission wavelengths 1) allow for discrimination (sensitivity >81%, specificity >94%) of various compositional and pathological features associated with plaque vulnerability including infiltration of macrophages into intima and necrotic/lipid core under a thin fibrous cap, and 2) show a linear correlation with plaque biochemical content: elastin (P<0.008), collagen (P<0.02), inflammatory cells (P<0.003), necrosis (P<0.004). Conclusion Our results demonstrate the feasibility of TR-LIFS as a method for the identification of markers of plaque vulnerability. Current findings enable future development of TR-LIFS based clinical devices for rapid investigation of atherosclerotic plaques and detection of those at high-risk. PMID:18926540

Marcu, Laura; Jo, Javier A.; Fang, Qiyin; Papaioannou, Thanassis; Reil, Todd; Qiao, Jian-Hua; Baker, J. Dennis; Freischlag, Julie A.; Fishbein, Michael C.

2009-01-01

23

Drug/protein interactions studied by time-resolved fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

We report here on a recent time-resolved fluorescence study [1] of the interaction between flurbiprofen (FBP), a chiral non-steroidal anti-inflammatory drug, and human serum albumin (HSA), the main transport protein in the human body. We compare the results obtained for the drug-protein complex with those of various covalently linked flurbiprofentryptophan dyads having well-defined geometries. In all cases stereoselective dynamic fluorescence quenching is observed, varying greatly from one system to another. In addition, the fluorescence anisotropy decays also display a clear stereoselectivity. For the drug-protein complexes, this can be interpreted in terms of the protein microenvironment playing a significant role in the conformational relaxation of FBP, which is more restricted in the case of the (R)- enantiomer.

Gustavsson, Thomas; Markovitsi, Dimitra; Vayá, Ignacio; Bonancía, Paula; Jiménez, M. C.; Miranda, Miguel A.

2014-09-01

24

Structural incorporation of Cm(III) in trioctahedral smectite hectorite: A time-resolved laser fluorescence spectroscopy (TRLFS) study  

NASA Astrophysics Data System (ADS)

Structural incorporation of the actinide curium in the octahedral layer of the trioctahedral Mg-rich smectite hectorite was studied using time-resolved laser fluorescence spectroscopy. Organo-hectorite nanoparticles were synthesized at 90 °C in the presence of Cm(III). Within 120 h, hectorite particles were formed. Time-resolved laser fluorescence spectroscopy was used to identify Cm(III) species during various synthesis steps and to characterize the structural incorporation mechanism. The formation of a Cm-containing Mg hydroxide precursor and the reaction with aqueous silica in a pH range of 9-10 to form TOT layers were identified to be key steps for trivalent actinide incorporation in hectorite via coprecipitation.

Brandt, Heike; Bosbach, Dirk; Panak, Petra J.; Fanghänel, Thomas

2007-01-01

25

In vivo detection of macrophages in a rabbit atherosclerotic model by time-resolved laser-induced fluorescence spectroscopy  

Microsoft Academic Search

Accumulation of numerous macrophages in the fibrous cap is a key identifying feature of plaque inflammation and vulnerability. This study investigates the use of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) as a potential tool for detection of macrophage foam cells in the intima of atherosclerotic plaques. Experiments were conducted in vivo on 14 New Zealand rabbits (6 control, 8 hypercholesterolemic) following

Laura Marcu; Qiyin Fang; Javier A. Jo; Thanassis Papaioannou; Amir Dorafshar; Todd Reil; Jian-Hua Qiao; J. Dennis Baker; Julie A. Freischlag; Michael C. Fishbein

2005-01-01

26

In vivo detection of macrophages in a rabbit atherosclerotic model by time-resolved laser-induced fluorescence spectroscopy  

PubMed Central

Accumulation of numerous macrophages in the fibrous cap is a key identifying feature of plaque inflammation and vulnerability. This study investigates the use of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) as a potential tool for detection of macrophage foam cells in the intima of atherosclerotic plaques. Experiments were conducted in vivo on 14 New Zealand rabbits (6 control, 8 hypercholesterolemic) following aortotomy to expose the intimal luminal surface of the aorta. Tissue autofluorescence was induced with a nitrogen pulse laser (337 nm, 1 ns). Lesions were histologically classified by the percent of collagen or macrophage foam cells as well as thickness of the intima. Using parameters derived from the time-resolved fluorescence emission of plaques, we determined that intima rich in macrophage foam cells can be distinguished from intima rich in collagen with high sensitivity (>85%) and specificity (>95%). This study demonstrates, for the first time, that a time-resolved fluorescence-based technique can differentiate and demark macrophage content versus collagen content in vivo. Our results suggest that TR-LIFS technique can be used in clinical applications for identification of inflammatory cells important in plaque formation and rupture. PMID:16039283

Marcu, Laura; Fang, Qiyin; Jo, Javier A.; Papaioannou, Thanassis; Dorafshar, Amir; Reil, Todd; Qiao, Jian-Hua; Baker, J. Dennis; Freischlag, Julie A.; Fishbein, Michael C.

2007-01-01

27

Fluorescence Instrument Response Standards in Two-Photon Time-Resolved Spectroscopy  

PubMed Central

We studied the fluorescence properties of several potential picosecond lifetime standards suitable for two-photon excitation from a Ti : sapphire femtosecond laser. The fluorescence emission of the selected fluorophores (rose bengal, pyridine 1, and LDS 798) covered the visible to near-infrared wavelength range from 550 to 850 nm. We suggest that these compounds can be used to measure the appropriate instrument response functions needed for accurate deconvolution of fluorescence lifetime data. Lifetime measurements with multiphoton excitation that use scatterers as a reference may fail to properly resolve fluorescence intensity decays. This is because of the different sensitivities of photodetectors in different spectral regions. Also, detectors often lose sensitivity in the near-infrared region. We demonstrate that the proposed references allow a proper reconvolution of measured lifetimes. We believe that picosecond lifetime standards for two-photon excitation will find broad applications in multiphoton spectroscopy and in fluorescence lifetime imaging microscopy (FLIM). PMID:20719056

LUCHOWSKI, RAFAL; SZABELSKI, MARIUSZ; SARKAR, PABAK; APICELLA, ELISA; MIDDE, KRISHNA; RAUT, SANGRAM; BOREJDO, JULIAN; GRYCZYNSKI, ZYGMUNT; GRYCZYNSKI, IGNACY

2011-01-01

28

Fluorescence instrument response standards in two-photon time-resolved spectroscopy.  

PubMed

We studied the fluorescence properties of several potential picosecond lifetime standards suitable for two-photon excitation from a Ti:sapphire femtosecond laser. The fluorescence emission of the selected fluorophores (rose bengal, pyridine 1, and LDS 798) covered the visible to near-infrared wavelength range from 550 to 850 nm. We suggest that these compounds can be used to measure the appropriate instrument response functions needed for accurate deconvolution of fluorescence lifetime data. Lifetime measurements with multiphoton excitation that use scatterers as a reference may fail to properly resolve fluorescence intensity decays. This is because of the different sensitivities of photodetectors in different spectral regions. Also, detectors often lose sensitivity in the near-infrared region. We demonstrate that the proposed references allow a proper reconvolution of measured lifetimes. We believe that picosecond lifetime standards for two-photon excitation will find broad applications in multiphoton spectroscopy and in fluorescence lifetime imaging microscopy (FLIM). PMID:20719056

Luchowski, Rafal; Szabelski, Mariusz; Sarkar, Pabak; Apicella, Elisa; Midde, Krishna; Raut, Sangram; Borejdo, Julian; Gryczynski, Zygmunt; Gryczynski, Ignacy

2010-08-01

29

TIME-RESOLVED VIBRATIONAL SPECTROSCOPY  

SciTech Connect

This document contains the Proceedings from the 14th International Conference on Time-Resolved Vibrational Spectroscopy, which was held in Meredith, NH from May 9-14, 2009. The study of molecular dynamics in chemical reaction and biological processes using time-resolved spectroscopy plays an important role in our understanding of energy conversion, storage, and utilization problems. Fundamental studies of chemical reactivity, molecular rearrangements, and charge transport are broadly supported by the DOE�s Office of Science because of their role in the development of alternative energy sources, the understanding of biological energy conversion processes, the efficient utilization of existing energy resources, and the mitigation of reactive intermediates in radiation chemistry. In addition, time-resolved spectroscopy is central to all five of DOE�s grand challenges for fundamental energy science. The Time-Resolved Vibrational Spectroscopy conference is organized biennially to bring the leaders in this field from around the globe together with young scientists to discuss the most recent scientific and technological advances. The latest technology in ultrafast infrared, Raman, and terahertz spectroscopy and the scientific advances that these methods enable were covered. Particular emphasis was placed on new experimental methods used to probe molecular dynamics in liquids, solids, interfaces, nanostructured materials, and biomolecules.

Andrei Tokmakoff, MIT (Conference Chair) [Conference Chair; Paul Champion, Northeastern University; Edwin J. Heilweil, NIST; Keith A. Nelson, MIT; Larry Ziegler, Boston University

2009-05-14

30

Solvation dynamics of coumarin 153 embedded in AOT + phenol organogels studied by time-resolved fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

We investigate solvation dynamics of organogel utilizing ps-ns fluorescence spectroscopy. The organogel studied in this Letter comprises bis(2-ethylhexyl) sulfosuccinate (AOT) and p-chlorophenol in the m-xylene solvent, that produce an organogel architecture with self-assembly. Within the organogel, an emitting probe, coumarin 153 (C153), is embedded. We then obtain dynamic response functions of solvation derived from the time-resolved fluorescence spectra of C153. We propose that total energy of the C153-organogel system relaxes with a relaxation time of 3.9 ns, whereas the entire rearrangement of the organogel structure around C153 is achieved with that of 6.1 ns, respectively.

Nishiyama, Katsura; Takata, Kei; Watanabe, Keiichi; Shigematsu, Hirotake

2012-03-01

31

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

PubMed Central

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

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

2011-01-01

32

The study of polyplex formation and stability by time-resolved fluorescence spectroscopy of SYBR Green I-stained DNA.  

PubMed

Polyplexes are nanoparticles formed by the self-assembly of DNA/RNA and cationic polymers specifically designed to deliver exogenous genetic material to cells by a process called transfection. There is a general consensus that a subtle balance between sufficient extracellular protection and intracellular release of nucleic acids is a key factor for successful gene delivery. Therefore, there is a strong need to develop suitable tools and techniques for enabling the monitoring of the stability of polyplexes in the biological environment they face during transfection. In this work we propose time-resolved fluorescence spectroscopy in combination with SYBR Green I-DNA dye as a reliable tool for the in-depth characterization of the DNA/vector complexation state. As a proof of concept, we provide essential information on the assembly and disassembly of complexes formed between DNA and each of three cationic polymers, namely a novel promising chitosan-graft-branched polyethylenimine copolymer (Chi-g-bPEI), one of its building block 2 kDa bPEI and the gold standard transfectant 25 kDa bPEI. Our results highlight the higher information content provided by the time-resolved studies of SYBR Green I/DNA, as compared to conventional steady state measurements of ethidium bromide/DNA that enabled us to draw relationships among fluorescence lifetime, polyplex structural changes and transfection efficiency. PMID:25308511

D'Andrea, Cosimo; Pezzoli, Daniele; Malloggi, Chiara; Candeo, Alessia; Capelli, Giulio; Bassi, Andrea; Volonterio, Alessandro; Taroni, Paola; Candiani, Gabriele

2014-12-01

33

Energy transfer processes in chlorophyll f-containing cyanobacteria using time-resolved fluorescence spectroscopy on intact cells.  

PubMed

We examined energy transfer dynamics in the unique chlorophyll (Chl) f-containing cyanobacterium Halomicronema hongdechloris. The absorption band of Chl f appeared during cultivation of this organism under far-red light. The absorption maximum of Chl f in organic solvents occurs at a wavelength of approximately 40 nm longer than that of Chl a. In vivo, the cells display a new absorption band at approximately 730 nm at 298 K, which is at a significantly longer wavelength than that of Chl a. We primarily assigned this band to a long wavelength form of Chl a. The function of Chl f is currently unknown. We measured the fluorescence of cells using time-resolved fluorescence spectroscopy in the picosecond-to-nanosecond time range and found clear differences in fluorescence properties between the cells that contained Chl f and the cells that did not. After excitation, the fluorescence peaks of photosystem I and photosystem II appeared quickly but diminished immediately. A unique fluorescence peak located at 748 nm subsequently appeared in cells containing Chl f. This finding strongly suggests that the Chl f in this alga exists in photosystem I and II complexes and is located close to each molecule of Chl a. This article is part of a special issue entitled: photosynthesis research for sustainability: keys to produce clean energy. PMID:24792349

Tomo, Tatsuya; Shinoda, Toshiyuki; Chen, Min; Allakhverdiev, Suleyman I; Akimoto, Seiji

2014-09-01

34

Coherent photon interference elimination and spectral correction in femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy  

NASA Astrophysics Data System (ADS)

We report an improved setup of femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy (FNOPAS) with a 210 fs temporal response. The system employs a Cassegrain objective to collect and focus fluorescence photons, which eliminates the interference from the coherent photons in the fluorescence amplification by temporal separation of the coherent photons and the fluorescence photons. The gain factor of the Cassegrain objective-assisted FNOPAS is characterized as 1.24 × 105 for Rhodamine 6G. Spectral corrections have been performed on the transient fluorescence spectra of Rhodamine 6G and Rhodamine 640 in ethanol by using an intrinsic calibration curve derived from the spectrum of superfluorescence, which is generated from the amplification of the vacuum quantum noise. The validity of spectral correction is illustrated by comparisons of spectral shape and peak wavelength between the corrected transient fluorescence spectra of these two dyes acquired by FNOPAS and their corresponding standard reference spectra collected by the commercial streak camera. The transient fluorescence spectra of the Rhodamine 6G were acquired in an optimized phase match condition, which gives a deviation in the peak wavelength between the retrieved spectrum and the reference spectrum of 1.0 nm, while those of Rhodamine 640 were collected in a non-optimized phase match condition, leading to a deviation in a range of 1.0-3.0 nm. Our results indicate that the improved FNOPAS can be a reliable tool in the measurement of transient fluorescence spectrum for its high temporal resolution and faithfully corrected spectrum.

Dang, Wei; Mao, Pengcheng; Weng, Yuxiang

2013-07-01

35

Development of a dual-modal tissue diagnostic system combining time-resolved fluorescence spectroscopy and ultrasonic backscatter microscopy  

PubMed Central

We report a tissue diagnostic system which combines two complementary techniques of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) and ultrasonic backscatter microscopy (UBM). TR-LIFS evaluates the biochemical composition of tissue, while UBM provides tissue microanatomy and enables localization of the region of diagnostic interest. The TR-LIFS component consists of an optical fiber-based time-domain apparatus including a spectrometer, gated multichannel plate photomultiplier, and fast digitizer. It records the fluorescence with high sensitivity (nM concentration range) and time resolution as low as 300 ps. The UBM system consists of a transducer, pulser, receiving circuit, and positioning stage. The transducer used here is 45 MHz, unfocused, with axial and lateral resolutions 38 and 200 ?m. Validation of the hybrid system and ultrasonic and spectroscopic data coregistration were conducted both in vitro (tissue phantom) and ex vivo (atherosclerotic tissue specimens of human aorta). Standard histopathological analysis of tissue samples was used to validate the UBM-TRLIFS data. Current results have demonstrated that spatially correlated UBM and TR-LIFS data provide complementary characterization of both morphology (necrotic core and calcium deposits) and biochemistry (collagen, elastin, and lipid features) of the atherosclerotic plaques at the same location. Thus, a combination of fluorescence spectroscopy with ultrasound imaging would allow for better identification of features associated with tissue pathologies. Current design and performance of the hybrid system suggests potential applications in clinical diagnosis of atherosclerotic plaque. PMID:19566223

Sun, Yang; Park, Jesung; Stephens, Douglas N.; Jo, Javier A.; Sun, Lei; Cannata, Jonathan M.; Saroufeem, Ramez M. G.; Shung, K. Kirk; Marcu, Laura

2009-01-01

36

Modified diglycol-amides for actinide separation: solvent extraction and time-resolved laser fluorescence spectroscopy complexation studies  

SciTech Connect

In this work, the back-bone of the diglycolamide-structure of the TODGA extractant was modified by adding one or two methyl groups to the central methylene carbon-atoms. The influence of these structural modifications on the extraction behavior of trivalent actinides and lanthanides and other fission products was studied in solvent extraction experiments. The addition of methyl groups to the central methylene carbon atoms leads to reduced distribution ratios, also for Sr(II). This reduced extraction efficiency for Sr(II) is beneficial for process applications, as the co-extraction of Sr(II) can be avoided, resulting in an easier process design. The use of these modified diglycol-amides in solvent extraction processes is discussed. Furthermore, the complexation of Cm(III) and Eu(III) to the ligands was studied using Time-Resolved-Laser-Fluorescence-Spectroscopy (TRLFS). The complexes were characterized by slope analysis and conditional stability constants were determined.

Wilden, A.; Modolo, G.; Lange, S.; Sadowski, F.; Bosbach, D. [Foschungszentrum Juelich GmbH, IEK-6, Juelich (Germany); Beele, B.B.; Panak, P.J. [Ruprecht-Karls-Universitaet Heidelberg, Physikalisch Chemisces Institut - PCI, Heidelberg (Germany); Karlsruher Institut fuer Technologie - INE, Karlsruhe (Germany); Skerencak-Frech, A.; Geist, A. [Karlsruher Institut fuer Technologie - INE, Karlsruhe (Germany); Iqbal, M. [University of Twente, Laboratory of Molecular Nanofabrication, Enschede (Netherlands); Department of Chemistry, University of Sargodha, Sargodha 40100 (Pakistan); Verboom, W. [University of Twente, Laboratory of Molecular Nanofabrication, Enschede (Netherlands)

2013-07-01

37

Short-term light adaptation of a cyanobacterium, Synechocystis sp. PCC 6803, probed by time-resolved fluorescence spectroscopy.  

PubMed

In photosynthetic organisms, the interactions among pigment-protein complexes change in response to light conditions. In the present study, we analyzed the transfer of excitation energy from the phycobilisome (PBS) and photosystem (PS) II to PSI in the cyanobacterium Synechocystis sp. PCC 6803. After 20 min of dark adaptation, Synechocystis cells were illuminated for 5 min with strong light with different spectral profiles, blue, green, two kinds of red, and white light. After illumination, the energy-transfer characteristics were evaluated using steady-state fluorescence and picosecond time-resolved fluorescence spectroscopy techniques. The fluorescence rise and decay curves were analyzed by global analysis to obtain fluorescence decay-associated spectra, followed by spectral component analysis. Under illumination with strong light, the contribution of the energy transfer from the PSII to PSI (spillover) became greater, and that of the energy transfer from the PBS to PSI decreased; the former change was larger than the latter. The energy transfer pathway to PSI was sensitive to red light. We discuss the short-term adaptation of energy-transfer processes in Synechocystis under strong-light conditions. PMID:24495908

Akimoto, Seiji; Yokono, Makio; Yokono, Erina; Aikawa, Shimpei; Kondo, Akihiko

2014-08-01

38

Steady state and time resolved effects of guanidine hydrochloride on the structure of Humicola lanuginosa lipase revealed by fluorescence spectroscopy.  

PubMed Central

Effects of guanidine hydrochloride (GdnHCl) on the structure and dynamics of wild-type Humicola lanuginosa lipase (HLL) and its two mutants were studied. The latter were S146A (with the active site Ser replaced by Ala) and the single Trp mutant W89m, with substitutions W117F, W221H, and W260H. Steady-state, stopped-flow, and time-resolved laser-induced fluorescence spectroscopy were carried out as a function of [GdnHCl]. The maximum emission wavelength and fluorescence lifetimes revealed the microenvironment of the tryptophan(s) in these lipases to become more polar upon increasing [GdnHCl]. However, significant extent of tertiary structure in GdnHCl is suggested by the observation that both wild-type HLL and W89m remain catalytically active at rather high GdnHCl concentrations of >6 and 4.0 M, respectively. Changes in steady-state emission anisotropy, as well as variation in rotational correlation times and residual anisotropy values, demonstrate that upon increasing [GdnHCl] the structure of the lipases became more loose, with increasing amplitude of structural fluctuations. Finally, intermediate states in the course of exposure of the proteins to GdnHCl were revealed by stopped-flow fluorescence measurements. PMID:10752622

Zhu, K.; Jutila, A.; Kinnunen, P. K.

2000-01-01

39

Time resolved fluorescence of CdSe nanocrystals using single molecule spectroscopy  

E-print Network

A wide variety of spectroscopic studies of CdSe nanocrystals (NCs) are presented in this thesis, all studying some aspect of the temporal evolution of NC fluorescence tinder different conditions. In particular the methods ...

Fisher, Brent R

2005-01-01

40

An ultrafast time-resolved fluorescence spectroscopy system for metal ion complexation studies with organic ligands  

Microsoft Academic Search

A dedicated spectrofluorimeter using ultrashort laser pulses as an excitation source was developed to measure the fluorescence properties of organic ligands for metal ion complexation with organic ligands. The laser system consists of an oscillator system for generation of femtosecond laser pulses, an amplifier system to increase the pulse energy of the generated pulses to about 2 mJ and an

G. Geipel; M Acker; D Vulpius; G Bernhard; H Nitsche; Th Fanghänel

2004-01-01

41

Nondestructive Evaluation of Tissue Engineered Articular Cartilage Using Time-Resolved Fluorescence Spectroscopy and Ultrasound Backscatter Microscopy  

PubMed Central

The goal of this study is to evaluate the ability of a bimodal technique integrating time-resolved fluorescence spectroscopy (TRFS) and ultrasound backscatter microscopy (UBM) for nondestructive detection of changes in the biochemical, structural, and mechanical properties of self-assembled engineered articular cartilage constructs. The cartilage constructs were treated with three chemical agents (collagenase, chondroitinase-ABC, and ribose) to induce changes in biochemical content (collagen and glycosaminoglycan [GAG]) of matured constructs (4 weeks); and to subsequently alter the mechanical properties of the construct. The biochemical changes were evaluated using TRFS. The microstructure and the thickness of the engineered cartilage samples were characterized by UBM. The optical and ultrasound results were validated against those acquired via conventional techniques including collagen and GAG quantification and measurement of construct stiffness. Current results demonstrated that a set of optical parameters (e.g., average fluorescence lifetime and decay constants) showed significant correlation (p<0.05) with biochemical and mechanical data. The high-resolution ultrasound images provided complementary cross-section information of the cartilage samples morphology. Therefore, the technique was capable of nondestructively evaluating the composition of extracellular matrix and the microstructure of engineered tissue, demonstrating great potential as an alternative to traditional destructive assays. PMID:22010819

Responte, Donald; Xie, Hongtao; Liu, Jing; Fatakdawala, Hussain; Hu, Jerry; Athanasiou, Kyriacos A.

2012-01-01

42

Interaction of europium and nickel with calcite studied by Rutherford Backscattering Spectrometry and Time-Resolved Laser Fluorescence Spectroscopy  

NASA Astrophysics Data System (ADS)

This study aims at elucidating the mechanisms regulating the interaction of Eu and Ni with calcite (CaCO3). Calcite powders or single crystals (some mm sized) were put into contact with Eu or Ni solutions at concentrations ranging from 10-3 to 10-5 mol L-1 for Eu and 10-3 mol L-1 for Ni. The sorption durations ranged from 1 week to 1 month. Rutherford Backscattering Spectrometry (RBS) well adapted to discriminate incorporation processes such as: (i) adsorption or co precipitation at the mineral surfaces or, (ii) incorporation into the mineral structure (through diffusion for instance), has been carried out. Moreover, using the fluorescence properties of europium, the results have been compared to those obtained by Time-Resolved Laser Fluorescence Spectroscopy (TRLFS) on calcite powders. For the single crystals, complementary SEM observations of the mineral surfaces at low voltage were also performed. Results showed that Ni accumulates at the calcite surface whereas Eu is also incorporated at a greater depth. Eu seems therefore to be incorporated into two different states in calcite: (i) heterogeneous surface accumulation and (ii) incorporation at depth greater than 160 nm after 1 month of sorption. Ni was found to accumulate at the surface of calcite without incorporation.

Sabau, A.; Pipon, Y.; Toulhoat, N.; Lomenech, C.; Jordan, N.; Moncoffre, N.; Barkleit, A.; Marmier, N.; Brendler, V.; Surblé, S.; Giffaut, E.

2014-08-01

43

A space-and time-resolved single-photon counting detector for fluorescence microscopy and spectroscopy  

E-print Network

A space- and time-resolved single-photon counting detector for fluorescence microscopy compare them to those of standard single-molecule detectors such as single-photon avalanche photodiode for single-molecule imaging and high-throughput study of biomolecular interactions. Keywords: single

Michalet, Xavier

44

Comparison of beetroot extracts originating from several sites using time-resolved laser-induced fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Beetroot (Beta vulgaris) juice contains a large number of fluorophores which can fluoresce. There is a growing interest in beetroot extracts analysis. In contrast, there is only limited information about beetroot obtained without sample preparation and/or extraction of components from the sample. In this work, we continue our previous study (Rabasovi? et al 2009 Acta Phys. Pol. A 116 570-2), analyzing and comparing beetroot extracts from several sites, using the time-resolved laser-induced fluorescence technique to measure the fluorescence of samples at different excitation wavelengths (340-470?nm) and for different sample dilutions.

Rabasovi?, M. S.; Ševi?, D.; Terzi?, M.; Marinkovi?, B. P.

2012-05-01

45

Effect of ouabain on metabolic oxidative state in living cardiomyocytes evaluated by time-resolved spectroscopy of endogenous NAD(P)H fluorescence  

NASA Astrophysics Data System (ADS)

Time-resolved spectrometry of endogenous nicotinamide dinucleotide phosphate [NAD(P)H] fluorescence is a useful method to evaluate metabolic oxidative state in living cells. Ouabain is a well-known pharmaceutical drug used in the treatment of cardiovascular disease, the effects of which on myocardial metabolism were recently demonstrated. Mechanisms implicated in these actions are still poorly understood. We investigate the effect of ouabain on the metabolic oxidative state of living cardiac cells identified by time-resolved fluorescence spectroscopy of mitochondrial NAD(P)H. Spectral unmixing is used to resolve individual NAD(P)H fluorescence components. Ouabain decreased the integral intensity of NAD(P)H fluorescence, leading to a reduced component amplitudes ratio corresponding to a change in metabolic state. We also noted that lactate/pyruvate, affecting the cytosolic NADH gradient, increased the effect of ouabain on the component amplitudes ratio. Cell oxidation levels, evaluated as the percentage of oxidized NAD(P)H, decreased exponentially with rising concentrations of the cardiac glycoside. Ouabain also stimulated the mitochondrial NADH production. Our study sheds a new light on the role that ouabain plays in the regulation of metabolic state, and presents perspective on a noninvasive, pharmaceutical approach for testing the effect of drugs on the mitochondrial metabolism by means of time-resolved fluorescence spectroscopy in living cells.

Chorvatova, Alzbeta; Elzwiei, Fathia; Mateasik, Anton; Chorvat, Dusan

2012-10-01

46

Differences in excitation energy transfer of Arthrospira platensis cells grown in seawater medium and freshwater medium, probed by time-resolved fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Excitation energy transfer of Arthrospira platensis cells grown in f/2 medium (a high salinity medium) and SOT medium (a control) was investigated by steady-state and time-resolved spectroscopies. Growth in f/2 medium induced changes in absorption and fluorescence spectra as well as in the energy transfer pathways. Excitation energy captured by phycobilisome (PBS) was transferred directly to photosystem (PS) I, instead of being first transferred to an intermediate (PBS ? PSII ? PSI), as observed in SOT medium. The respiration rate increased while photosynthetic rate reduced in f/2 medium. Possible causes of the differences in light-harvesting and energy-transfer processes between the two media are discussed.

Arba, Muhammad; Aikawa, Shimpei; Niki, Kenta; Yokono, Makio; Kondo, Akihiko; Akimoto, Seiji

2013-11-01

47

Time-resolved fluorescence decay measurements for flowing particles  

DOEpatents

Time-resolved fluorescence decay measurements for flowing particles. An apparatus and method for the measurement and analysis of fluorescence for individual cells and particles in flow are described, wherein the rapid measurement capabilities of flow cytometry and the robust measurement and analysis procedures of time-domain fluorescence lifetime spectroscopy are combined. A pulse-modulated cw laser is employed for excitation of the particles. The characteristics and the repetition rate of the excitation pulses can be readily adjusted to accommodate for fluorescence decays having a wide range of lifetimes.

Deka, Chiranjit (Miami, FL); Steinkamp, John A. (Los Alamos, NM)

1999-01-01

48

Time-resolved fluorescence decay measurements for flowing particles  

DOEpatents

Time-resolved fluorescence decay measurements are disclosed for flowing particles. An apparatus and method for the measurement and analysis of fluorescence for individual cells and particles in flow are described, wherein the rapid measurement capabilities of flow cytometry and the robust measurement and analysis procedures of time-domain fluorescence lifetime spectroscopy are combined. A pulse-modulated CW laser is employed for excitation of the particles. The characteristics and the repetition rate of the excitation pulses can be readily adjusted to accommodate for fluorescence decays having a wide range of lifetimes. 12 figs.

Deka, C.; Steinkamp, J.A.

1999-06-01

49

Nanosecond Time-Resolved Fluorescence Spectroscopy in the Physical Chemistry Laboratory: Formation of the Pyrene Excimer in Solution  

NASA Astrophysics Data System (ADS)

The fluorescence lifetime and spectrum of pyrene in fluid solution depend on experimental conditions, including sample concentration, solvent viscosity, and the presence of dissolved oxygen. Students are introduced to nanosecond-domain transient spectroscopy in fluid solution by using a pulsed-laser apparatus to make direct measurements of the effects of these variables. They become familiar with the operation of a nitrogen laser and an associated apparatus for fluorescence spectra and lifetime measurements, including the optical and electronic hardware and the computer software used to acquire and analyze the data. The experiment specifically illustrates the role of sample concentration and solvent viscosity on fluorescence lifetimes and spectra, and the effects of argon purging to remove dissolved oxygen.

van Dyke, David A.; Pryor, Brian A.; Smith, Philip G.; Topp, Michael R.

1998-05-01

50

Interaction of Cm(III) and Am(III) with human serum transferrin studied by time-resolved laser fluorescence and EXAFS spectroscopy.  

PubMed

The complexation of Cm(III) with human serum transferrin was investigated in a pH range from 3.5 to 11.0 using time-resolved laser fluorescence spectroscopy (TRLFS). At pH ? 7.4 Cm(III) is incorporated at the Fe(III) binding site of transferrin whereas at lower pH a partially bound Cm(III) transferrin species is formed. At physiological temperature (310 K) at pH 7.4, about 70% of the partially bound and 30% of the incorporated Cm(III) transferrin species are present in solution. The Cm(III) results obtained by TRLFS are in very good agreement with Am(III) EXAFS results, confirming the incorporation of Am(III) at the Fe(III) binding site at pH 8.5. PMID:24626477

Bauer, Nicole; Fröhlich, Daniel R; Panak, Petra J

2014-05-14

51

Time Resolved Raman and Fluorescence Spectrometer for Planetary Mineralogy  

NASA Astrophysics Data System (ADS)

Raman spectroscopy is a prime candidate for the next generation of planetary instruments, as it addresses the primary goal of mineralogical analysis which is structure and composition. It does not require sample preparation and provides unique mineral fingerprints, even for mixed phase samples. However, large fluorescence return from many mineral samples under visible light excitation can seriously compromise the quality of the spectra or even render Raman spectra unattainable. Fluorescence interference is likely to be a problem on Mars and is evident in Raman spectra of Martian Meteorites[1]. Our approach uses time resolution for elimination of fluorescence from Raman spectra, allowing for traditional visible laser excitation (532 nm). Since Raman occurs instantaneously with the laser pulse and fluorescence lifetimes vary from nsec to msec depending on the mineral, it is possible to separate them out in time. Complementary information can also be obtained simultaneously using the time resolved fluorescence data. The Simultaneous Spectral Temporal Adaptive Raman Spectrometer (SSTARS) is a planetary instrument under development at the Jet Propulsion Laboratory, capable of time-resolved in situ Raman and fluorescence spectroscopy. A streak camera and pulsed miniature microchip laser provide psec scale time resolution. Our ability to observe the complete time evolution of Raman and fluorescence in minerals provides a foundation for design of pulsed Raman and fluorescence spectrometers in diverse planetary environments. We will discuss the SSTARS instrument design and performance capability. We will also present time-resolved pulsed Raman spectra collected from a relevant set of minerals selected using available data on Mars mineralogy[2]. Of particular interest are minerals resulting from aqueous alteration on Mars. For comparison, we will present Raman spectra obtained using a commercial continuous wave (CW) green (514 nm) Raman system. In many cases using a CW laser the strong mineral fluorescence saturates the detector and Raman spectra are unattainable. This problem is overcome by using time resolved Raman where fluorescence is eliminated. [1]Frosch et al., Anal. Chem. 2007, 79, 1101-1108 [2]Bell, J.,ed, The Martian Surface: Composition, Mineralogy, and physical Properties, Cambridge University Press, 2008

Blacksberg, Jordana; Rossman, George

2010-05-01

52

Time-resolved laser-induced fluorescence spectroscopy of Nd 3+ in molten LiCl-KCl eutectic  

NASA Astrophysics Data System (ADS)

The characteristics of the laser-induced fluorescence of Nd3+ in LiCl-KCl eutectic in the wavelength region of 360-900 nm were investigated for information concerning the chemical speciation of Nd-chloride complexes. When pumped at either 355 or 532 nm, Nd3+ in molten salt emits visible and near-IR fluorescence. The fluorescence peaks at 750 nm (4F7/2 + 4S3/2 ? 4I9/2) and 810 nm (4F5/2 + 2H9/2 ? 4I9/2) were particularly prominent at temperatures above the melting point. The fluorescence decay of these transition lines showed a bi-exponential behaviour of the fluorescence lifetime. These results provide evidence that two different chemical species of Nd3+ coexist in this system.

Jung, E. C.; Bae, S.-E.; Park, Y. J.; Song, K.

2011-11-01

53

In vivo validation of a bimodal technique combining time-resolved fluorescence spectroscopy and ultrasonic backscatter microscopy for diagnosis of oral carcinoma  

PubMed Central

Abstract. Tissue diagnostic features generated by a bimodal technique integrating scanning time-resolved fluorescence spectroscopy (TRFS) and ultrasonic backscatter microscopy (UBM) are investigated in an in vivo hamster oral carcinoma model. Tissue fluorescence is excited by a pulsed nitrogen laser and spectrally and temporally resolved using a set of filters/dichroic mirrors and a fast digitizer, respectively. A 41-MHz focused transducer (37-?m axial, 65-?m lateral resolution) is used for UBM scanning. Representative lesions of the different stages of carcinogenesis show that fluorescence characteristics complement ultrasonic features, and both correlate with histological findings. These results demonstrate that TRFS-UBM provide a wealth of co-registered, complementary data concerning tissue composition and structure as it relates to disease status. The direct co-registration of the TRFS data (sensitive to surface molecular changes) with the UBM data (sensitive to cross-sectional structural changes and depth of tumor invasion) is expected to play an important role in pre-operative diagnosis and intra-operative determination of tumor margins. PMID:23117798

Sun, Yang; Xie, Hongtao; Liu, Jing; Lam, Matthew; Chaudhari, Abhijit J.; Zhou, Feifei; Bec, Julien; Yankelevich, Diego R.; Dobbie, Allison; Tinling, Steven L.; Gandour-Edwards, Regina F.; Monsky, Wayne L.; Gregory Farwell, D.; Marcu, Laura

2012-01-01

54

Monitoring Local Unfolding of Bovine Serum Albumin During Denaturation Using Steady-State and Time-Resolved Fluorescence Spectroscopy  

Microsoft Academic Search

In a previous report (J. Fluoresc. 16, 153, 2006) we studied the chaotropiclly induced denaturation of Bovine Serum Albumin (BSA) using the fluorescence decay\\u000a kinetics at different stages in the denaturation of BSA by guanidinium hydrochloride (GuHCl). In this work, we gain a more\\u000a detailed insight into the BSA denaturation process by investigating the thermodynamics of the process. Structural changes

Denisio M. Togashi; Alan G. Ryder; Domhnall O’Shaughnessy

2010-01-01

55

Time resolved fluorescence tomography of turbid media based on lifetime  

E-print Network

Time resolved fluorescence tomography of turbid media based on lifetime contrast Anand T. N. Kumar-based analysis of the entire temporal fluorescence response from a turbid medium. Simulations are used to show, (170.3650) Lifetime-based sensing, (170.6920) Time-resolved imaging, (170.7050) Turbid media References

Kumar, Anand T.N.

56

Picosecond time-resolved fluorescent spectroscopy of 1-anilino-8-naphthalene sulfonate binding with staphylococcal nuclease in the native and molten globule states.  

PubMed

We studied the picosecond time-resolved fluorescent spectroscopy of 1-anilino-8-naphthalene sulfonate (ANS), which binds to the staphylococcal nuclease (SNase) of the wild-type (WT) and the molten globule (MG) state. Three ANS emission bands at ?530nm, ?495nm, and ?475nm are resolved, corresponding to three ANS states: the free ANS in solution and associated form adsorbing to surface sites and binding to active sites. The surface hydrophobicity of the WT is moderate and different from the MG state, as shown both in the position of the bands and by the concentration dependent ANS fluorescent decay. For MG, the decay of two blue bands accelerated with the increment of the ANS concentration, whereas the WT did not show this dependency. However, when pdTp, an inhibitor, was attached to the active site of the MG state, band 2 decay was also independent of the ANS concentration. These results indicate that the protein hydrophobic sites have two types of interactions with ANS. PMID:25771383

Gao, Guangyu; Li, Yu; Wang, Wei; Zhong, Dongping; Wang, Shufeng; Gong, Qihuang

2015-04-01

57

Speciation of Eu3+ bound to humic substances by time-resolved laser fluorescence spectroscopy (TRLFS) and parallel factor analysis (PARAFAC)  

NASA Astrophysics Data System (ADS)

The bioavailability and toxicity of metal ions including radionuclides in the biosphere are greatly influenced by their speciation. Humic substances (HSs) are important constituents of various soil and water systems and have significant impact on the speciation and mobility of metal ions because of their high affinity to metal ions. In this study, the speciation of europium (Eu3+), a chemical homologue of trivalent actinides, with HSs collected from various origins was investigated by time-resolved laser fluorescence spectroscopy (TRLFS). The difficulties associated with the separation of the contribution of different Eu3+ species due to overlapping spectra or similar fluorescence lifetimes were addressed and mitigated by applying a multi-mode factor analysis, parallel factor analysis (PARAFAC), which resulted in the number, spectra, decay curves and relative fluorescence intensity profiles of different Eu3+ species. Subsequently, the interpretation of the Eu3+ species, was tackled by principal component analysis (PCA) and partial linear square (PLS) regression to deduce the nature of the Eu3+ species by taking into account the physicochemical properties of the HSs. Three factors corresponding to different Eu3+ species were obtained at 70 ?M Eu3+ for all HSs investigated except for one humic acid. One of the factors corresponded to free Eu3+ ion interacting with HSs via diffusion. The remaining two factors were thought to be Eu3+ bound to HSs: one bound to acidic functional groups of HSs and the other to the sites of HSs influenced by the carbon backbone structures. It was also found that the latter factor exhibited strong energy transfer from the excited Eu3+ center to HSs. At lower Eu3+ concentration (10 ?M), two factors having similar fluorescent characteristics to those of the second and third factors were obtained.

Lukman, Steven; Saito, Takumi; Aoyagi, Noboru; Kimura, Takaumi; Nagasaki, Shinya

2012-07-01

58

Time-resolved fluorescence spectroscopy and imaging of DNA labeled with DAPI and Hoechst 33342 using three-photon excitation.  

PubMed

We examined the fluorescence spectral properties of the DNA stains DAPI (4',6-diamidino-2-phenylindole, hydrochloride) and Hoechst 33342 (bis-benzimide, or 2,5'-bi'1H-benzimidazole2'-(4-ethoxyphenyl)-5-(4-methyl-1-piperazi nyl)) with two-photon (2h nu) and three-photon (3h nu) excitation using femtosecond pulses from a Ti:sapphire laser from 830 to 885 nm. The mode of excitation of DAPI bound to DNA changed from two-photon at 830 nm to three-photon at 885 nm. In contrast, Hoechst 33342 displayed only two-photon excitation from 830 to 885 nm. DAPI-DNA displayed the same emission spectra and decay times for 2h nu and 3h nu excitation. Hoechst 33342-DNA displayed the same intensity decay for excitation at 830 and 885 nm. Both probes displayed higher anisotropies for multiphoton excitation as compared to one-photon excitation with ultraviolet wavelengths, and DAPI-DNA displays a higher anisotropy for 3h nu at 885 nm than for 2h nu at 830 nm. We used 970-nm excitation of DAPI-stained chromosomes to obtain the first three-dimensional images with three-photon excitation. Three-photon excitation of DAPI-stained chromosomes at 970 nm was demonstrated by the power dependence in the fluorescence microscope. PMID:9017187

Lakowicz, J R; Gryczynski, I; Malak, H; Schrader, M; Engelhardt, P; Kano, H; Hell, S W

1997-02-01

59

Time-resolved fluorescence lifetime for cutaneous melanoma detection  

PubMed Central

Melanoma is the most aggressive skin cancer type. It is characterized by pigmented lesions with high tissue invasion and metastatic potential. The early detection of melanoma is extremely important to improve patient prognosis and survival rate, since it can progress to the deadly metastatic stage. Presently, the melanoma diagnosis is based on the clinical analysis of the macroscopic lesion characteristics such as shape, color, borders following the ABCD rules. The aim of this study is to evaluate the time-resolved fluorescence lifetime of NADH and FAD molecules to detect cutaneous melanoma in an experimental in vivo model. Forty-two lesions were analyzed and the data was classified using linear discriminant analysis, a sensitivity of 99.4%, specificity of 97.4% and accuracy of 98.4% were achieved. These results show the potential of this fluorescence spectroscopy for melanoma detection. PMID:25401022

Pires, Layla; Nogueira, Marcelo Saito; Pratavieira, Sebastião; Moriyama, Lilian Tan; Kurachi, Cristina

2014-01-01

60

Time-resolved nanosecond fluorescence lifetime imaging and picosecond infrared spectroscopy of combretastatin A-4 in solution and in cellular systems  

NASA Astrophysics Data System (ADS)

Fluorescence lifetime images of intrinsic fluorescence obtained with two-photon excitation at 630 nm are shown following uptake of a series of E-combretastatins into live cells, including human umbilical vein endothelial cells (HUVECs) that are the target for the anticancer activity of combretastatins. Images show distribution of the compounds within the cell cytoplasm and in structures identified as lipid droplets by comparison with images obtained following Nile red staining of the same cells. The intracellular fluorescent lifetimes are generally longer than in fluid solution as a consequence of the high viscosity of the cellular environment. Following incubation, the intracellular concentrations of a fluorinated derivative of E-combretastatin A-4 in HUVECs are between two and three orders of magnitude higher than the concentration in the surrounding medium. Evidence is presented to indicate that at moderate laser powers (up to 6 mW), it is possible to isomerize up to 25% of the combretastatin within the femtolitre focal volume of the femtosecond laser beam. This suggests that it may be possible to activate the E-combretastatin (with low cellular toxicity) to the Z-isomer with high anticancer drug activity using two-photon irradiation. The isomerization of Z- and E-combretastatins by 266 nm irradiation has been probed by ultrafast time-resolved infrared spectroscopy. Results for the E-isomer show a rapid loss of excess vibrational energy in the excited state with a lifetime of 7 ps, followed by a slower process with a lifetime of 500 ps corresponding to the return to the ground state as also determined from the fluorescence lifetime. In contrast, the Z-isomer, whilst also appearing to undergo a rapid cooling of the initial excited state, has a much shorter overall excited state lifetime of 14 ps. DedicationThis paper is dedicated to the memory of Professor Christopher G Morgan (1949-2011). He was a valued colleague and friend at the University of Salford and made significant contributions to the development and applications of fluorescence lifetime imaging.

Bisby, Roger H.; Botchway, Stanley W.; Greetham, Greg M.; Hadfield, John A.; McGown, Alan T.; Parker, Anthony W.; Scherer, Kathrin M.; Towrie, Mike

2012-08-01

61

TIME-RESOLVED TERAHERTZ TRANSMISSION SPECTROSCOPY OF DIELECTRICS  

E-print Network

TIME-RESOLVED TERAHERTZ TRANSMISSION SPECTROSCOPY OF DIELECTRICS PETR KUZEL AND JAN PETZELT Republic. Using the method of time-domain terahertz-transmission spectroscopy we measured the far Terahertz pulses; far infrared; time-resolved spectroscopy INTRODUCTION The time-domain terahertz-transmission

Ku?el, Petr

62

Proteins in Action Monitored by Time-Resolved FTIR Spectroscopy  

E-print Network

Proteins in Action Monitored by Time-Resolved FTIR Spectroscopy Carsten Kçtting and Klaus Gerwert*[a] Introduction Time-resolved Fourier-transform infrared (trFTIR) difference spectroscopy can reveal great the FTIR literature. For further literature the reviews of Heberle,[3] Mäntele,[4] Vogel and Siebert,[5

Gerwert, Klaus

63

Conformational States of the Rapana thomasiana Hemocyanin and Its Substructures Studied by Dynamic Light Scattering and Time-Resolved Fluorescence Spectroscopy  

PubMed Central

Hemocyanins are dioxygen-transporting proteins freely dissolved in the hemolymph of mollusks and arthropods. Dynamic light scattering and time-resolved fluorescence measurements show that the oxygenated and apo-forms of the Rapana thomasiana hemocyanin, its structural subunits RtH1 and RtH2, and those of the functional unit RtH2e, exist in different conformations. The oxygenated respiratory proteins are less compact and more asymmetric than the respective apo-forms. Different conformational states were also observed for the R. thomasiana hemocyanin in the absence and presence of an allosteric regulator. The results are in agreement with a molecular mechanism for cooperative dioxygen binding in molluscan hemocyanins including transfer of conformational changes from one functional unit to another. PMID:15533921

Georgieva, Dessislava; Schwark, Daniel; Nikolov, Peter; Idakieva, Krassimira; Parvanova, Katja; Dierks, Karsten; Genov, Nicolay; Betzel, Christian

2005-01-01

64

Time-resolved optical fluorescence spectroscopy of heterogeneous turbid media with special emphasis on brain tissue structures including diseased regions: A sensitivity analysis  

NASA Astrophysics Data System (ADS)

Fluorescence-enhanced optical imaging based on near-infrared light provides a promising tool to differentiate diseased lesions from normal tissue. However, the measurement sensitivity of the fluorescence signals acquired at the output surface of the tissue is greatly influenced by the tissue structure, the optical properties, the location and the size of the target. In this paper, we present a numerical model based on the Monte Carlo method that allows to simulate time-resolved reflectance signals acquired on the surface of the scalp of a human head model bearing a fluorescent diseased region (tumor, glioma). The influence of tumor depth, tumor size and tumor shape evolution on the computed signals are analyzed by taking into account the multi-layered tissue structure. The simulations show that the mean-time-of-flight and the difference between two mean-times acquired at two source-detector distances are both relevant to this problem type. Furthermore, the simulations suggest that the use of the difference between mean-flight-times may be interesting to probe scattering changes that occur in the cerebrospinal fluid (CSF).

Vaudelle, Fabrice; L'huillier, Jean-Pierre

2013-09-01

65

Time-Resolved Electron Energy Loss Spectroscopy  

Microsoft Academic Search

Two recent instrumental improvements in high-resolution electron energy loss spectroscopy make possible the recording of complete surface vibrational spectra on the millisecond time scale. This is the first spectroscopic probe capable of directly measuring fundamental surface rate processes in real time with a resolution less than or equal to 1 millisecond. Such measurements are the key to understanding surface kinetics

T. H. Ellis; L. H. Dubois; S. D. Kevan; M. J. Cardillo

1985-01-01

66

Research on time-resolved terahertz spectroscopy  

NASA Astrophysics Data System (ADS)

We have built a set of terahertz time-domain spectroscopy system using electro-optic crystals. Conventional terahertz time-domain spectroscopy based on Fourier-transform for spectra analysis, which mixes the frequency components of the entire temporal terahertz waveform in one frequency domain; therefore, it yields different terahertz spectra from a same terahertz pulse with different scanning lengths. We introduce a new technique for the joint time-frequency analysis of terahertz time-domain spectroscopy based on wavelet-transform technique. With this technique, the frequency components in different time locations are clearly exhibited on a two-dimensional plane; therefore, the noise in the pulse tail cannot affect the frequency in the main pulse. This technique clearly separates the frequency of terahertz from that of its echo in the time domain; therefore, the interference spectrum occur in Fourier-transform is naturally removed. By varying the shape of analysis wavelet, high time resolution and high frequency resolution are easily obtained. The absorption coefficients of envelope, plastic, foam and cotton have been measured with the wavelet technique.

Deng, Yuqiang; Sun, Qing; Liu, Feng; Wang, Changlei; Xing, Qirong

2010-10-01

67

Seventh international conference on time-resolved vibrational spectroscopy  

SciTech Connect

The International Conference on Time-Resolved Vibrational Spectroscopy (TRVS) is widely recognized as the major international forum for the discussion of advances in this rapidly growing field. The 1995 conference was the seventh in a series that began at Lake Placid, New York, 1982. Santa Fe, New Mexico, was the site of the Seventh International Conference on Time-Resolved Vibrational Spectroscopy, held from June 11 to 16, 1995. TRVS-7 was attended by 157 participants from 16 countries and 85 institutions, and research ranging across the full breadth of the field of time-resolved vibrational spectroscopy was presented. Advances in both experimental capabilities for time-resolved vibrational measurements and in theoretical descriptions of time-resolved vibrational methods continue to occur, and several sessions of the conference were devoted to discussion of these advances and the associated new directions in TRVS. Continuing the interdisciplinary tradition of the TRVS meetings, applications of time-resolved vibrational methods to problems in physics, biology, materials science, and chemistry comprised a large portion of the papers presented at the conference.

Dyer, R.B.; Martinez, M.A.D.; Shreve, A.; Woodruff, W.H. [comps.

1997-04-01

68

Motor Oil Classification Based on Time-Resolved Fluorescence  

PubMed Central

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

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

2014-01-01

69

Steady-state and time-resolved fluorometry of fluorescent pollutants and heavy metal complexes  

NASA Astrophysics Data System (ADS)

Time-resolved laser-induced fluorescence spectroscopy is one of the most sensitive optical methods which is well suited for on-line in situ analysis. Here, three examples for the steady- state and time-resolved fluorescence analysis of environmentally important analytes, the fluorescent monoaromatic hydrocarbons benzene, toluene, and xylene as well as non fluorescent heavy metal ions forming a fluorescent complex with a cation coordinating fluorescence probe, are presented and the potential of both methods is discussed. For BTX, various mixtures of the spectrally similar compounds B, T, and X showing different fluorescence lifetimes were studied with both methods. As an example for fluorometric metal ion analysis, the fluorescence probe BP(OH)2 (2,2'-bipyridyl- 3,3'-diol) was employed for the determination of d10 metal ions in water and the newly developed fluorescence probe APTA for the detection of Cu(II). Cation complexation of BP(OH2 yields spectrally very similar complexes which differ in their fluorescence lifetimes. Complexation of APTA to Cu(II) leads to small spectral changes and a strong increase in fluorescence quantum yield and lifetime. For the analytes studied, a comparison of the detection limits, standard deviations, and linear dynamic range of both methods clearly demonstrates the analytical potential of time-resolved fluorometry.

Resch, Ute; Rurack, Knut

1997-05-01

70

Phase transition dynamics of fluorescent-labeled poly( N-isopropylacrylamide) in aqueous solution as revealed by time-resolved spectroscopy combined with a laser T-jump technique  

NASA Astrophysics Data System (ADS)

In order to clarify the dynamics of phase transition from coiled states to globular states in a thermo-responsive polymer, poly( N-isopropylacrylamide) (PNIPAM), we developed a method of dynamic (time-resolved) fluorescence spectroscopy combined with a laser T-jump technique. Using this, we revealed the phase transition dynamics of fluorescent-labeled PNIPAM in aqueous solutions. The phase transition dynamics was reflected in a dynamic Stokes shift of a TICT fluorescence of the probe. The time constant of the phase transition ( ?) was determined to be 35 ?s, irrespective of the sample concentration studied (0.1-1.0 wt.%), and this was longer than that predicted from the diffusion-collision model.

Tsuboi, Yasuyuki; Yoshida, Yasuhiro; Kitamura, Noboru; Iwai, Kaoru

2009-01-01

71

Time-resolved laser-induced fluorescence system  

NASA Astrophysics Data System (ADS)

Fluorescence methods are being used increasingly in the measurement of species concentrations in gases, liquids and solids. Laser induced fluorescence is spontaneous emission from atoms or molecules that have been excited by laser radiation. Here we present a time resolved fluorescence instrument that consists of a 5 ?J Nitrogen laser (337.1 nm), a sample holder, a quartz optical fiber, a spectrometer, a PMT and a PC that allows the measurement of visible fluorescence spectra (350-750 nm). Time response of the system is approximately 5 ns. The instrument has been used in the measurement of colored bond paper, antifreeze, diesel, cochineal pigment and malignant tissues. The data acquisition was achieved through computer control of a digital oscilloscope (using General Purpose Interface Bus GPIB) and the spectrometer via serial (RS232). The instrument software provides a graphic interface that lets make some data acquisition tasks like finding fluorescence spectra, and fluorescence lifetimes. The software was developed using the Lab-View 6i graphic programming package and can be easily managed in order to add more functions to it.

Bautista, F. J.; De la Rosa, J.; Gallegos, F. J.

2006-02-01

72

Advances in ultrafast time resolved fluorescence physics for cancer detection in optical biopsy  

NASA Astrophysics Data System (ADS)

We discuss the use of time resolved fluorescence spectroscopy to extract fundamental kinetic information on molecular species in tissues. The temporal profiles reveal the lifetime and amplitudes associated with key active molecules distinguishing the local spectral environment of tissues. The femtosecond laser pulses at 310 nm excite the tissue. The emission profile at 340 nm from tryptophan is non-exponential due to the micro-environment. The slow and fast amplitudes and lifetimes of emission profiles reveal that cancer and normal states can be distinguished. Time resolved optical methods offer a new cancer diagnostic modality for the medical community.

Alfano, R. R.

2012-03-01

73

Picosecond Time-Resolved Fourier Transform Raman Spectroscopy of 9,10-Diphenylanthracene in the Excited Singlet State  

E-print Network

Time-resolved Fourier transform Raman spectroscopy of the highly fluorescent chromophore 9,10-diphenylanthracene (DPA) in cyclohexane and ethanol is described. Raman spectra of the first excited singlet state of DPA were obtained with 100-ps...

Jas, Gouri S.; Wan, Chaozhi; Johnson, Carey K.

1995-05-01

74

Time-resolved wide-field optically sectioned fluorescence microscopy  

NASA Astrophysics Data System (ADS)

We present the implementation of a fast wide-field optical sectioning technique called HiLo microscopy on a fluorescence lifetime imaging microscope. HiLo microscopy is based on the fusion of two images, one with structured illumination and another with uniform illumination. Optically sectioned images are then digitally generated thanks to a fusion algorithm. HiLo images are comparable in quality with confocal images but they can be acquired faster over larger fields of view. We obtain 4D imaging by combining HiLo optical sectioning, time-gated detection, and z-displacement. We characterize the performances of this set-up in terms of 3D spatial resolution and time-resolved capabilities in both fixed- and live-cell imaging modes.

Dupuis, Guillaume; Benabdallah, Nadia; Chopinaud, Aurélien; Mayet, Céline; Lévêque-Fort, Sandrine

2013-02-01

75

ULTRA: A Unique Instrument for Time-Resolved Spectroscopy.  

PubMed

We report the development of a high-sensitivity time-resolved infrared and Raman spectrometer with exceptional experimental flexibility based on a 10-kHz synchronized dual-arm femtosecond and picosecond laser system. Ultrafast high-average-power titanium sapphire lasers and optical parametric amplifiers provide wavelength tuning from the ultraviolet (UV) to the mid-infrared region. Customized silicon, indium gallium arsenide, and mercury cadmium telluride linear array detectors are provided to monitor the probe laser intensity in the UV to mid-infrared wavelength range capable of measuring changes in sample absorbance of ?OD ~ 10(-5) in 1 second. The system performance is demonstrated for the time-resolved infrared, two-dimensional (2D) infrared, and femtosecond stimulated Raman spectroscopy techniques with organometallic intermediates, organic excited states, and the dynamics of the tertiary structure of DNA. PMID:21144146

Greetham, Gregory M; Burgos, Pierre; Cao, Qian; Clark, Ian P; Codd, Peter S; Farrow, Richard C; George, Michael W; Kogimtzis, Moschos; Matousek, Pavel; Parker, Anthony W; Pollard, Mark R; Robinson, David A; Xin, Zhi-Jun; Towrie, Michael

2010-12-01

76

Steady State and Time-Resolved Fluorescence Studies of a Hemagglutinin from Moringa oleifera  

Microsoft Academic Search

The saccharide binding and conformational characterization of a hemagglutinin, a low molecular weight protein from the seeds\\u000a of Moringa oleifera was studied using steady state and time resolved fluorescence. The lectin binds sugars LacNAc (K\\u000a a?=?1380 M?1) and fructose (K\\u000a a?=?975 M?1), as determined by the fluorescence spectroscopy. It has a single tryptophan per monomer which is exposed on the surface\\u000a and

Uma V. Katre; C. G. Suresh; M. Islam Khan; Sushama M. Gaikwad

2008-01-01

77

Diffuse optical fluorescence tomography using time-resolved data acquired in transmission  

E-print Network

Diffuse optical fluorescence tomography using time-resolved data acquired in transmission Frederic acquired with a time-resolved system with the goal of reconstructing sources of fluorescence emanating from as the curved nature of the boundary, can have a profound impact on fluorescent images and spectroscopic

Friedlander, Michael P.

78

Multidimensional time-resolved spectroscopy of vibrational coherence in biopolyenes.  

PubMed

Multidimensional femtosecond time-resolved vibrational coherence spectroscopy allows one to investigate the evolution of vibrational coherence in electronic excited states. Methods such as pump-degenerate four-wave mixing and pump-impulsive vibrational spectroscopy combine an initial ultrashort laser pulse with a nonlinear probing sequence to reinduce vibrational coherence exclusively in the excited states. By carefully exploiting specific electronic resonances, one can detect vibrational coherence from 0 cm(-1) to over 2,000 cm(-1) and map its evolution. This review focuses on the observation and mapping of high-frequency vibrational coherence for all-trans biological polyenes such as ?-carotene, lycopene, retinal, and retinal Schiff base. We discuss the role of molecular symmetry in vibrational coherence activity in the S1 electronic state and the interplay of coupling between electronic states and vibrational coherence. PMID:24245903

Buckup, Tiago; Motzkus, Marcus

2014-01-01

79

Thermally activated delayed fluorescence of fluorescein derivative for time-resolved and confocal fluorescence imaging.  

PubMed

Compared with fluorescence imaging utilizing fluorophores whose lifetimes are in the order of nanoseconds, time-resolved fluorescence microscopy has more advantages in monitoring target fluorescence. In this work, compound DCF-MPYM, which is based on a fluorescein derivative, showed long-lived luminescence (22.11 ?s in deaerated ethanol) and was used in time-resolved fluorescence imaging in living cells. Both nanosecond time-resolved transient difference absorption spectra and time-correlated single-photon counting (TCSPC) were employed to explain the long lifetime of the compound, which is rare in pure organic fluorophores without rare earth metals and heavy atoms. A mechanism of thermally activated delayed fluorescence (TADF) that considers the long wavelength fluorescence, large Stokes shift, and long-lived triplet state of DCF-MPYM was proposed. The energy gap (?EST) of DCF-MPYM between the singlet and triplet state was determined to be 28.36 meV by the decay rate of DF as a function of temperature. The ?E(ST) was small enough to allow efficient intersystem crossing (ISC) and reverse ISC, leading to efficient TADF at room temperature. The straightforward synthesis of DCF-MPYM and wide availability of its starting materials contribute to the excellent potential of the compound to replace luminescent lanthanide complexes in future time-resolved imaging technologies. PMID:24936960

Xiong, Xiaoqing; Song, Fengling; Wang, Jingyun; Zhang, Yukang; Xue, Yingying; Sun, Liangliang; Jiang, Na; Gao, Pan; Tian, Lu; Peng, Xiaojun

2014-07-01

80

Noninvasive Multimodal Evaluation of Bioengineered Cartilage Constructs Combining Time-Resolved Fluorescence and Ultrasound Imaging  

PubMed Central

A multimodal diagnostic system that integrates time-resolved fluorescence spectroscopy, fluorescence lifetime imaging microscopy, and ultrasound backscatter microscopy is evaluated here as a potential tool for assessing changes in engineered tissue composition and microstructure nondestructively and noninvasively. The development of techniques capable of monitoring the quality of engineered tissue, determined by extracellular matrix (ECM) content, before implantation would alleviate the need for destructive assays over multiple time points and advance the widespread development and clinical application of engineered tissues. Using a prototype system combining time-resolved fluorescence spectroscopy, FLIM, and UBM, we measured changes in ECM content occurring during chondrogenic differentiation of equine adipose stem cells on 3D biodegradable matrices. The optical and ultrasound results were validated against those acquired via conventional techniques, including collagen II immunohistochemistry, picrosirius red staining, and measurement of construct stiffness. Current results confirm the ability of this multimodal approach to follow the progression of tissue maturation along the chondrogenic lineage by monitoring ECM production (namely, collagen type II) and by detecting resulting changes in mechanical properties of tissue constructs. Although this study was directed toward monitoring chondrogenic tissue maturation, these data demonstrate the feasibility of this approach for multiple applications toward engineering other tissues, including bone and vascular grafts. PMID:21303258

Fite, Brett Z.; Decaris, Martin; Sun, Yinghua; Sun, Yang; Lam, Adrian; Ho, Clark K.L.

2011-01-01

81

Noninvasive multimodal evaluation of bioengineered cartilage constructs combining time-resolved fluorescence and ultrasound imaging.  

PubMed

A multimodal diagnostic system that integrates time-resolved fluorescence spectroscopy, fluorescence lifetime imaging microscopy, and ultrasound backscatter microscopy is evaluated here as a potential tool for assessing changes in engineered tissue composition and microstructure nondestructively and noninvasively. The development of techniques capable of monitoring the quality of engineered tissue, determined by extracellular matrix (ECM) content, before implantation would alleviate the need for destructive assays over multiple time points and advance the widespread development and clinical application of engineered tissues. Using a prototype system combining time-resolved fluorescence spectroscopy, FLIM, and UBM, we measured changes in ECM content occurring during chondrogenic differentiation of equine adipose stem cells on 3D biodegradable matrices. The optical and ultrasound results were validated against those acquired via conventional techniques, including collagen II immunohistochemistry, picrosirius red staining, and measurement of construct stiffness. Current results confirm the ability of this multimodal approach to follow the progression of tissue maturation along the chondrogenic lineage by monitoring ECM production (namely, collagen type II) and by detecting resulting changes in mechanical properties of tissue constructs. Although this study was directed toward monitoring chondrogenic tissue maturation, these data demonstrate the feasibility of this approach for multiple applications toward engineering other tissues, including bone and vascular grafts. PMID:21303258

Fite, Brett Z; Decaris, Martin; Sun, Yinghua; Sun, Yang; Lam, Adrian; Ho, Clark K L; Leach, J Kent; Marcu, Laura

2011-04-01

82

Steady state and time-resolved fluorescence spectroscopic characterization of normal and cancerous urine  

NASA Astrophysics Data System (ADS)

Urine is one of the diagnostically important bio fluids, as it has many metabolites and some of them are native fluorophores. There may be a variation in the distribution and the physiochemical properties of the fluorophores during any metabolic change and pathologic conditions. Native fluorescence spectroscopy has been considered as a promising tool to characterize the fluorophores present in the urine. In this study, we aimed at characterizing the urine of both normal and patients with confirmed cancer using steady state and time-resolved fluorescence spectroscopy at 280 nm and 350 nm excitation. It is observed that the metabolites indoxyl sulphate and neopterin and its derivatives are responsible for altered spectral signatures at 280 nm, and 350 nm excitation. The overall spectral data were subjected to Principal Component Analysis and the resultant components were used as input in the linear discriminant analysis. As a total, 84% and 81.8% of samples were correctly classified at 280 nm and 350 nm respectively.

Rajasekaran, Ramu; Aruna, Prakasa Rao; Balu David, Munusamy; Koteeswaran, Dornadula; Muthuvelu, Kulandaivel; Rai, R.; Ganesan, Singaravelu

2013-03-01

83

Nanosecond-regime correlation time scales for equilibrium protein structural fluctuations of metal-free cytochrome c from picosecond time-resolved fluorescence spectroscopy and the dynamic Stokes shift.  

PubMed

We used picosecond time-resolved fluorescence spectroscopy to characterize the fluorescence Stokes shift (FSS) response function of metal-free (or free-base, fbCytc) cytochrome c under the solution conditions that favor the native states of ferricytochrome c (FeCytc) and Zn(II)-substituted cytochrome c (ZnCytc). The intrinsic porphyrin chromophore serves in these experiments as a fluorescent probe of the structural fluctuations of the surrounding protein and solvent. Demetalation of the porphyrin destabilizes the folded structure of cytochrome c owing to the loss of the axial metal-histidine and metal-methionine bonds. Thus, these experiments examine how the time scales detected in a dynamic solvation experiment in a chromoprotein report changes in the character of motion. The FSS response function in fbCytc in water and pH 7 is well described by a biexponential response over the 100 ps to 50 ns regime with time constants of 1.4 and 9.1 ns; under similar conditions, ZnCytc exhibits a biexponential FSS response with time constants of 250 ps and 1.5 ns [Lampa-Pastirk and Beck, J. Phys. Chem. B 2004, 108, 16288]. These time constants correspond, respectively, to the correlation time scales for motions of the hydrophobic core and the solvent-contact layer of the protein. Both of the time constants observed in fbCytc are further lengthened upon addition of glycerol to the external solvent so that a significant fraction of the protein dynamics is rendered effectively static on the fluorescence time scale. The solvation reorganization energy, the time-integrated Stokes shift of the fluorescence spectrum, is reduced by about a third to 33 cm(-1) in 50% glycerol from 43 cm(-1) in water. These results are interpreted structurally using a model for Brownian diffusive motion with thermally activated barrier crossings on the protein-folding energy landscape. The results suggest that the mean-squared deviations of the structural fluctuations exhibited by fbCytc are nearly a factor of 10 larger than those of ZnCytc. This conclusion is consistent with the suggestion that fbCytc assumes a dynamic, partially unfolded structure with some of the characteristics of a molten globule. PMID:21077593

Tripathy, Jagnyaseni; Beck, Warren F

2010-12-01

84

Electron-transfer acceleration investigated by time resolved infrared spectroscopy.  

PubMed

Ultrafast electron transfer (ET) processes are important primary steps in natural and artificial photosynthesis, as well as in molecular electronic/photonic devices. In biological systems, ET often occurs surprisingly fast over long distances of several tens of angströms. Laser-pulse irradiation is conveniently used to generate strongly oxidizing (or reducing) excited states whose reactions are then studied by time-resolved spectroscopic techniques. While photoluminescence decay and UV-vis absorption supply precise kinetics data, time-resolved infrared absorption (TRIR) and Raman-based spectroscopies have the advantage of providing additional structural information and monitoring vibrational energy flows and dissipation, as well as medium relaxation, that accompany ultrafast ET. We will discuss three cases of photoinduced ET involving the Re(I)(CO)3(N,N) moiety (N,N = polypyridine) that occur much faster than would be expected from ET theories. [Re(4-N-methylpyridinium-pyridine)(CO)3(N,N)](2+) represents a case of excited-state picosecond ET between two different ligands that remains ultrafast even in slow-relaxing solvents, beating the adiabatic limit. This is caused by vibrational/solvational excitation of the precursor state and participation of high-frequency quantum modes in barrier crossing. The case of Re-tryptophan assemblies demonstrates that excited-state Trp ? *Re(II) ET is accelerated from nanoseconds to picoseconds when the Re(I)(CO)3(N,N) chromophore is appended to a protein, close to a tryptophan residue. TRIR in combination with DFT calculations and structural studies reveals an interaction between the N,N ligand and the tryptophan indole. It results in partial electronic delocalization in the precursor excited state and likely contributes to the ultrafast ET rate. Long-lived vibrational/solvational excitation of the protein Re(I)(CO)3(N,N)···Trp moiety, documented by dynamic IR band shifts, could be another accelerating factor. The last discussed process, back-ET in a porphyrin-Re(I)(CO)3(N,N) dyad, demonstrates that formation of a hot product accelerates highly exergonic ET in the Marcus inverted region. Overall, it follows that ET can be accelerated by enhancing the electronic interaction and by vibrational excitation of the reacting system and its medium, stressing the importance of quantum nuclear dynamics in ET reactivity. These effects are experimentally accessible by time-resolved vibrational spectroscopies (IR, Raman) in combination with quantum chemical calculations. It is suggested that structural dynamics play different mechanistic roles in light-triggered ET involving electronically excited donors or acceptors than in ground-state processes. While TRIR spectroscopy is well suitable to elucidate ET processes on a molecular-level, transient 2D-IR techniques combining optical and two IR (or terahertz) laser pulses present future opportunities for investigating, driving, and controlling ET. PMID:25699661

Vl?ek, Antonín; Kvapilová, Hana; Towrie, Michael; Záliš, Stanislav

2015-03-17

85

Time-resolved fluorescence studies of nucleotide flipping by restriction enzymes  

PubMed Central

Restriction enzymes Ecl18kI, PspGI and EcoRII-C, specific for interrupted 5-bp target sequences, flip the central base pair of these sequences into their protein pockets to facilitate sequence recognition and adjust the DNA cleavage pattern. We have used time-resolved fluorescence spectroscopy of 2-aminopurine-labelled DNA in complex with each of these enzymes in solution to explore the nucleotide flipping mechanism and to obtain a detailed picture of the molecular environment of the extrahelical bases. We also report the first study of the 7-bp cutter, PfoI, whose recognition sequence (T/CCNGGA) overlaps with that of the Ecl18kI-type enzymes, and for which the crystal structure is unknown. The time-resolved fluorescence experiments reveal that PfoI also uses base flipping as part of its DNA recognition mechanism and that the extrahelical bases are captured by PfoI in binding pockets whose structures are quite different to those of the structurally characterized enzymes Ecl18kI, PspGI and EcoRII-C. The fluorescence decay parameters of all the enzyme-DNA complexes are interpreted to provide insight into the mechanisms used by these four restriction enzymes to flip and recognize bases and the relationship between nucleotide flipping and DNA cleavage. PMID:19740769

Neely, Robert K.; Tamulaitis, Gintautas; Chen, Kai; Kubala, Marta; Siksnys, Virginijus; Jones, Anita C.

2009-01-01

86

Time-resolved spectroscopy of irradiated n-GaAs  

SciTech Connect

Gallium arsenide films were grown by the metallorganic chemical vapor deposition method and doped n-type with silicon to concentrations of 2 {times} 10{sup 15} and 2 {times} 10{sup 16} cm{sup {minus}3}. The lifetime ({tau}) of the band-to-band recombination process was measured at 77 K using an optical time-resolved spectroscopy technique. The pre-irradiated values ranged from 350 to 550 ps. The samples were irradiated at room temperature with {sup 60}Co gamma rays, fission neutrons, 7 MeV electrons, protons (0.6 to 500 MeV), alpha particles, and lithium and oxygen ions. Degradation constants (K{sub {tau}}) attributed to non-radiative processes generated by radiation-induced defects are reported. K{sub {tau}} is compared to the previously published degradation constants associated with the photoluminescence intensity (K{sub PL}) in the continuous mode, and to the previously published introduction rate (b) of the silicon defect at the arsenic site (Si{sub As}). K{sub {tau}}, K{sub PL} and b(Si{sub As}) are compared to non-ionizing energy loss calculations and to the Rutherford scattering theory of the cross-section.

Parenteau, M.; Carlone, C.; Morris, D. [Univ. de Sherbrooke, Quebec (Canada). Dept. de Physique] [Univ. de Sherbrooke, Quebec (Canada). Dept. de Physique; Khanna, S.M. [Dept. of National Defence, Ottawa, Ontario (Canada). Defence Research Establishment Ottawa] [Dept. of National Defence, Ottawa, Ontario (Canada). Defence Research Establishment Ottawa

1997-12-01

87

Glucose Sensing by Time-Resolved Fluorescence of Sol-Gel Immobilized Glucose Oxidase  

PubMed Central

A monolithic silica gel matrix with entrapped glucose oxidase (GOD) was constructed as a bioactive element in an optical biosensor for glucose determination. Intrinsic fluorescence of free and immobilised GOD was investigated in the visible range in presence of different glucose concentrations by time-resolved spectroscopy with time-correlated single-photon counting detector. A three-exponential model was used for analysing the fluorescence transients. Fractional intensities and mean lifetime were shown to be sensitive to the enzymatic reaction and were used for obtaining calibration curve for glucose concentration determination. The sensing system proposed achieved high resolution (up to 0.17 mM) glucose determination with a detection range from 0.4 mM to 5 mM. PMID:22163807

Esposito, Rosario; Ventura, Bartolomeo Della; De Nicola, Sergio; Altucci, Carlo; Velotta, Raffaele; Mita, Damiano Gustavo; Lepore, Maria

2011-01-01

88

Time-resolved fluorescence of the bacteriophage T4 capsid protein gp23  

Microsoft Academic Search

The time-resolved fluorescence properties of the bacteriophage T4 capsid protein gp23 are investigated. The structural characteristics of this protein are largely unknown and can be probed by recording time-resolved and decay-associated fluorescence spectra and intensity decay curves using a 200 ps-gated intensified CCD-camera. Spectral and decay data are recorded simultaneously, which makes data acquisition fast compared to time-correlated single-photon counting.

Aike Stortelder; Joost B. Buijs; Jaap Bulthuis; Saskia M. van der Vies; Cees Gooijer; Gert van der Zwan

2005-01-01

89

Time resolved laser induced fluorescence measurements: Considerations when using Nd:YAG based system  

NASA Astrophysics Data System (ADS)

Time-resolved laser-induced fluorescence (TR-LIF) and the laser induced breakdown spectroscopy (LIBS) have been shown to be methods which are fast and sensitive to provide information about the constituents in analyzed samples. TR-LIF and LIBS have similar hardware requirements. In this paper, we analyze some characteristics of TR-LIF/LIBS system implemented in our laboratory, considering the fact that the excitation part of the system is based on Nd:YAG laser and Optical Parametric Oscillator (OPO). The laser is more than powerful enough (365 mJ at 1064 nm, variable OPO output >5 mJ) for LIBS, but somehow slow (the length of fundamental laser harmonic output pulse is about 5 ns) for fluorescence measurements in our present area of interest, namely plants and food products. Fortunately, the pulse length of tunable OPO output (320-475 nm) is less then 1 ns, so by means of a correct deconvolution procedure it is possible to measure the fluorescence lifetimes in the range as small as a few nanoseconds. The fluorescence detection part of our system is based on picosecond streak camera. Using the fluorescent dyes (Rhodamine B and Fluorescein) ethanol solutions we verified the analyzing capabilities of our TR-LIF system.

Rabasovic, Maja S.; Sevic, Dragutin; Terzic, Mira; Marinkovic, Bratislav P.

2012-05-01

90

Monitoring tissue metabolism via time-resolved laser fluorescence  

NASA Astrophysics Data System (ADS)

Most assays for drug screening are monitoring the metabolism of cells by detecting the NADH content, which symbolize its metabolic activity, indirectly. Nowadays, the performance of a LASER enables us to monitor the metabolic state of mammalian cells directly and on-line by using time-resolved autofluorescence detection. Therefore, we developed in combination with tissue engineering, an assay for monitoring minor toxic effects of volatile organic compounds (VOC), which are accused of inducing Sick Building Syndrome (SBS). Furthermore, we used the Laserfluoroscope (LF) for pharmacological studies on human bone marrow in vitro with special interest in chemotherapy simulation. In cancer research and therapy, the effect of chemostatica in vitro in the so-called oncobiogram is being tested; up to now without great success. However, it showed among other things that tissue structure plays a vital role. Consequently, we succeeded in simulating a chemotherapy in vitro on human bone marrow. Furthermore, after tumor ektomy we were able to distinguish between tumoric and its surrounding healthy tissue by using the LF. With its sensitive detection of metabolic changes in tissues the LF enables a wide range of applications in biotechnology, e.g. for quality control in artificial organ engineering or biocompatability testing.

Maerz, Holger K.; Buchholz, Rainer; Emmrich, Frank; Fink, Frank; Geddes, Clive L.; Pfeifer, Lutz; Raabe, Ferdinand; Marx, Uwe

1999-05-01

91

Picosecond Time-Resolved Vibrational Circular Dichroism Spectroscopy  

NASA Astrophysics Data System (ADS)

An experimental setup for the recording of time-resolved vibrational circular dichroism (VCD) spectra is presented. Both static and transient VCD signals can be measured with high sensitivity. First results for a transition metal complex show chiral transients that are clearly distinct from the conventional pump-probe signals.

Bonmarin, Mathias; Helbing, Jan

92

Integrated multimodal microscopy, time-resolved fluorescence, and optical-trap rheometry: toward single molecule mechanobiology  

PubMed Central

Cells respond to forces through coordinated biochemical signaling cascades that originate from changes in single-molecule structure and dynamics and proceed to large-scale changes in cellular morphology and protein expression. To enable experiments that determine the molecular basis of mechanotransduction over these large time and length scales, we construct a confocal molecular dynamics microscope (CMDM). This system integrates total-internal-reflection fluorescence (TIRF), epifluorescence, differential interference contrast (DIC), and 3-D deconvolution imaging modalities with time-correlated single-photon counting (TCSPC) instrumentation and an optical trap. Some of the structures hypothesized to be involved in mechanotransduction are the glycocalyx, plasma membrane, actin cytoskeleton, focal adhesions, and cell-cell junctions. Through analysis of fluorescence fluctuations, single-molecule spectroscopic measurements [e.g., fluorescence correlation spectroscopy (FCS) and time-resolved fluorescence] can be correlated with these subcellular structures in adherent endothelial cells subjected to well-defined forces. We describe the construction of our multimodal microscope in detail and the calibrations necessary to define molecular dynamics in cell and model membranes. Finally, we discuss the potential applications of the system and its implications for the field of mechanotransduction. PMID:17343487

Gullapalli, Ramachandra R.; Tabouillot, Tristan; Mathura, Rishi; Dangaria, Jhanvi H.; Butler, Peter J.

2011-01-01

93

Time-resolved fluorescence measurements of actin-phalloidin interactions  

NASA Astrophysics Data System (ADS)

Compounds that interact with the cytoskeleton affect mobility and division, making them useful for treatment of certain types of cancer. Actin binding drugs such as the phallotoxins (small, bicyclic peptides) bind to and stabilize actin polymers (F-actin) without binding to actin monomers (G-actin). It has been shown that the intensity of fluorescently labeled phallotoxins such as fluorescein- phalloidin and rhodamine-phalloidin increases upon bind F- actin. We used LJL BioSystems' new FLAReTM technology to measure excited state lifetime changes of fluorescein- phalloidin and rhodamine-phalloidin upon binding to F- actin.

Helms, Michael K.; French, Todd E.

2000-03-01

94

Feasibility analysis of an epidermal glucose sensor based on time-resolved fluorescence  

E-print Network

-prick capillary blood sampling, which is painful and prevents detection of abnormal glucose levels while sleeping on fluorescence. Currently, diabetic pa- tients measure their blood glucose concentrations by fingerFeasibility analysis of an epidermal glucose sensor based on time-resolved fluorescence Kamal M

Pilon, Laurent

95

Multiplexed Analysis Using Time-Resolved Near-IR Fluorescence for the Detection of Genomic Material  

E-print Network

in a time-correlated single photon counting format to allow acquisition of fluorescence lifetimes onMultiplexed Analysis Using Time-Resolved Near-IR Fluorescence for the Detection of Genomic Material and the need for screening multiple targets simultaneously. We have initiated work on developing time

Davis, Lloyd M.

96

Studies of Minerals, Organic and Biogenic Materials through Time-Resolved Raman Spectroscopy  

NASA Technical Reports Server (NTRS)

A compact remote Raman spectroscopy system was developed at NASA Langley Research center and was previously demonstrated for its ability to identify chemical composition of various rocks and minerals. In this study, the Raman sensor was utilized to perform time-resolved Raman studies of various samples such as minerals and rocks, Azalea leaves and a few fossil samples. The Raman sensor utilizes a pulsed 532 nm Nd:YAG laser as excitation source, a 4-inch telescope to collect the Raman-scattered signal from a sample several meters away, a spectrograph equipped with a holographic grating, and a gated intensified CCD (ICCD) camera system. Time resolved Raman measurements were carried out by varying the gate delay with fixed short gate width of the ICCD camera, allowing measurement of both Raman signals and fluorescence signals. Rocks and mineral samples were characterized including marble, which contain CaCO3. Analysis of the results reveals the short (approx.10-13 s) lifetime of the Raman process, and shows that Raman spectra of some mineral samples contain fluorescence emission due to organic impurities. Also analyzed were a green (pristine) and a yellow (decayed) sample of Gardenia leaves. It was observed that the fluorescence signals from the green and yellow leaf samples showed stronger signals compared to the Raman lines. Moreover, it was also observed that the fluorescence of the green leaf was more intense and had a shorter lifetime than that of the yellow leaf. For the fossil samples, Raman shifted lines could not be observed due the presence of very strong short-lived fluorescence.

Garcia, Christopher S.; Abedin, M. Nurul; Ismail, Syed; Sharma, Shiv K.; Misra, Anupam K.; Nyugen, Trac; Elsayed-Ali, hani

2009-01-01

97

Fluorophore Conjugated Silver Nanoparticles: A Time-resolved Fluorescence Correlation Spectroscopic Study  

PubMed Central

Fluorescence detection is a central component in biological research. In recent years there has been a growing interest in the interactions of fluorophores with metallic surfaces or particles. A single-stranded oligonucleotide was chemically bound to a single 50 nm diameter silver particle and a Cy5-labeled complementary single-stranded oligonucleotide was hybridized with the particle-bound oligonucleotide. The bound Cy5 molecules on the silver particles were spatially separated from the silver surface by the hybridized DNA duplex chains, which were about 8 nm in length, to reduce the competitive quenching. We use fluorescence lifetime correlation spectroscopy (FLCS) with picosecond time-resolved detection to separate the fluorescence correlation spectroscopy (FCS) contributions from fluorophores and metal-conjugated fluorophores. The single Cy5-labeled 50 nm silver particles displayed a factor of 15-fold increase in emission signal and 5-fold decrease in emission lifetimes in solution relative to the Cy5-DNA in the absence of metal. Lifetime measurements support the near-field interaction mechanism between the fluorophore and silver nanoparticle. In this study, FLCS is being applied to a system where the brightness and the fluorescent lifetime of the emitting species are significantly different. Our measurements suggest that FLCS is a powerful method for investigating the metal-fluorophore interaction at the single molecule level and to separate two different species from a mixture solution emitting at the same wavelength. Additionally, the highly bright Cy5-DNA-Ag molecules offer to be excellent probes in high background biological samples. PMID:19609374

Ray, Krishanu; Zhang, Jian; Lakowicz, Joseph R.

2009-01-01

98

Noninvasive assessment of breast cancer risk using time-resolved diffuse optical spectroscopy  

Microsoft Academic Search

Breast density is a recognized strong and independent risk factor for breast cancer. We propose the use of time-resolved transmittance spectroscopy to estimate breast tissue density and potentially provide even more direct information on breast cancer risk. Time-resolved optical mammography at seven wavelengths (635 to 1060 nm) is performed on 49 subjects. Average information on breast tissue of each subject

Paola Taroni; Antonio Pifferi; Giovanna Quarto; Lorenzo Spinelli; Alessandro Torricelli; Francesca Abbate; Anna Villa; Nicola Balestreri; Simona Menna; Enrico Cassano; Rinaldo Cubeddu

2010-01-01

99

Mechanism of the Efficient Tryptophan Fluorescence Quenching in Human ?D-Crystallin Studied by Time-Resolved Fluorescence  

PubMed Central

Human ?D-crystallin (H?D-Crys) is a two-domain, ?-sheet eye lens protein found in the lens nucleus. Its long-term solubility and stability are important to maintain lens transparency throughout life. H?D-Crys has four highly conserved buried tryptophans (Trps), with two in each of the homologous ?-sheet domains. In situ, these Trps will be absorbing ambient UV radiation that reaches the lens. The dispersal of the excited-state energy to avoid covalent damage is likely to be physiologically relevant for the lens crystallins. Trp fluorescence is efficiently quenched in native H?D-Crys. Previous steady-state fluorescence measurements provide strong evidence for energy transfer from Trp42 to Trp68 in the N-terminal domain and from Trp130 to Trp156 in the C-terminal domain [Chen, J., et al. (2006) Biochemistry 45, 11552?11563]. Hybrid quantum mechanical?molecular mechanical (QM-MM) simulations indicated that the fluorescence of Trp68 and Trp156 is quenched by fast electron transfer to the amide backbone. Here we report additional information obtained using time-resolved fluorescence spectroscopy. In the single-Trp-containing proteins (Trp42-only, Trp68-only, Trp130-only, and Trp156-only), the highly quenched Trp68 and Trp156 have very short lifetimes, ? ?0.1 ns, whereas the moderately fluorescent Trp42 and Trp130 have longer lifetimes, ? ?3 ns. In the presence of the energy acceptor (Trp68 or Trp156), the lifetime of the energy donor (Trp42 or Trp130) decreased from ?3 to ?1 ns. The intradomain energy transfer efficiency is 56% in the N-terminal domain and is 71% in the C-terminal domain. The experimental values of energy transfer efficiency are in good agreement with those calculated theoretically. The absence of a time-dependent red shift in the time-resolved emission spectra of Trp130 proves that its local environment is very rigid. Time-resolved fluorescence anisotropy measurements with the single-Trp-containing proteins, Trp42-only and Trp130-only, indicate that the protein rotates as a rigid body and no segmental motion is detected. A combination of energy transfer with electron transfer results in short excited-state lifetimes of all Trps, which, together with the high rigidity of the protein matrix around Trps, could protect H?D-Crys from excited-state reactions causing permanent covalent damage. PMID:18795792

2008-01-01

100

Time-resolved phase-sensitive second harmonic generation spectroscopy.  

PubMed

A methodology based on time-resolved, phase-sensitive second harmonic generation (SHG) for probing the excited state dynamics of species at interfaces is presented. It is based on an interference measurement between the SHG from the sample and a local oscillator generated at a reference together with a lock-in measurement to remove the large constant offset from the interference. The technique is characterized by measuring the phase and excited state dynamics of the dye malachite green at the water/air interface. The key attributes of the technique are that the observed signal is directly proportional to sample concentration, in contrast to the quadratic dependence from non-phase sensitive SHG, and that the real and imaginary parts of the 2nd order non-linear susceptibility can be determined independently. We show that the method is highly sensitive and can provide high quality excited state dynamics in short data acquisition times. PMID:25725724

Nowakowski, Pawe? J; Woods, David A; Bain, Colin D; Verlet, Jan R R

2015-02-28

101

Time-resolved phase-sensitive second harmonic generation spectroscopy  

NASA Astrophysics Data System (ADS)

A methodology based on time-resolved, phase-sensitive second harmonic generation (SHG) for probing the excited state dynamics of species at interfaces is presented. It is based on an interference measurement between the SHG from the sample and a local oscillator generated at a reference together with a lock-in measurement to remove the large constant offset from the interference. The technique is characterized by measuring the phase and excited state dynamics of the dye malachite green at the water/air interface. The key attributes of the technique are that the observed signal is directly proportional to sample concentration, in contrast to the quadratic dependence from non-phase sensitive SHG, and that the real and imaginary parts of the 2nd order non-linear susceptibility can be determined independently. We show that the method is highly sensitive and can provide high quality excited state dynamics in short data acquisition times.

Nowakowski, Pawe? J.; Woods, David A.; Bain, Colin D.; Verlet, Jan R. R.

2015-02-01

102

Proceedings of the Fourteenth International Conference on Time-Resolved Vibrational Spectroscopy (TRVS XIV)  

E-print Network

Abstracts of presentations made at the Fourteenth International Conference on Time-Resolved Vibrational Spectroscopy (TRVS XIV) held May 9-14, 2009 in Meredith, New Hampshire. TRVS is a series of biennial conferences ...

Tokmakoff, Andrei

2011-08-31

103

The application of time-resolved luminescence spectroscopy to a remote uranyl sensor  

SciTech Connect

Time resolved luminescence spectroscopy is an effective method for the determination of a wide range of uranyl concentrations in aqueous samples. We have applied this technique to the development of a remote sensing device using fiber optic cables coupled with a micro flow cell in order to probe for uranyl in aqueous samples. This sensor incorporates a Nafion membrane through which UO{sub 2}{sup 2+} can diffuse in to a reaction/analysis chamber which holds phosphoric acid, a reagent which enhances the uranyl luminescence intensity and lifetime. With this device, anionic and fluorescing organic interferences could be eliminated, allowing for the determination of uranyl over a concentration range of 10{sup 4} to 10{sup {minus}9}M. 17 refs., 5 figs.

Varineau, P.T.; Duesing, R.; Wangen, L.E.

1991-01-01

104

Global and time-resolved monitoring of crop photosynthesis with chlorophyll fluorescence  

E-print Network

February 25, 2014 (received for review October 24, 2013) Photosynthesis is the process by which plantsGlobal and time-resolved monitoring of crop photosynthesis with chlorophyll fluorescence Luis and Bolin Centre for Climate Research, Stockholm University, 10691 Stockholm, Sweden; k Image Processing

Minnesota, University of

105

Time-Resolved Fluorescence Analysis of the Photosystem II Antenna Proteins in Detergent Micelles and Liposomes  

E-print Network

Time-Resolved Fluorescence Analysis of the Photosystem II Antenna Proteins in Detergent Micelles LHCII, CP29, CP26, and CP24 in detergent solution is mostly determined by two lifetime components of 1 in lipid membranes or thylakoids with respect to detergent solution. By increasing the protein density

106

Engineered tryptophan in the adenine-binding pocket of catalytic subunit A of A-ATP synthase demonstrates the importance of aromatic residues in adenine binding, forming a tool for steady-state and time-resolved fluorescence spectroscopy  

PubMed Central

A reporter tryptophan residue was individually introduced by site-directed mutagenesis into the adenine-binding pocket of the catalytic subunit A (F427W and F508W mutants) of the motor protein A1AO ATP synthase from Pyrococcus horikoshii OT3. The crystal structures of the F427W and F508W mutant proteins were determined to 2.5 and 2.6?Å resolution, respectively. The tryptophan substitution caused the fluorescence signal to increase by 28% (F427W) and 33% (F508W), with a shift from 333?nm in the wild-type protein to 339?nm in the mutant proteins. Tryptophan emission spectra showed binding of Mg-ATP to the F427W mutant with a K d of 8.5?µM. In contrast, no significant binding of nucleotide could be observed for the F508W mutant. A closer inspection of the crystal structure of the F427W mutant showed that the adenine-binding pocket had widened by 0.7?Å (to 8.70?Å) in comparison to the wild-type subunit A (8.07?Å) owing to tryptophan substitution, as a result of which it was able to bind ATP. In contrast, the adenine-binding pocket had narrowed in the F508W mutant. The two mutants presented demonstrate that the exact volume of the adenine ribose binding pocket is essential for nucleotide binding and even minor narrowing makes it unfit for nucleotide binding. In addition, structural and fluorescence data confirmed the viability of the fluorescently active mutant F427W, which had ideal tryptophan spectra for future structure-based time-resolved dynamic measurements of the catalytic subunit A of the ATP-synthesizing enzyme A-ATP synthase. PMID:22139149

Tadwal, Vikeramjeet Singh; Manimekalai, Malathy Sony Subramanian; Grüber, Gerhard

2011-01-01

107

Binding of 7-methoxy-4-(aminomethyl)-coumarin to wild-type and W128F mutant cytochrome P450 2D6 studied by time-resolved fluorescence spectroscopy  

PubMed Central

Enzyme structure and dynamics may play a main role in substrate binding and the subsequent steps in the CYP (cytochrome P450) catalytic cycle. In the present study, changes in the structure of human CYP2D6 upon binding of the substrate are studied using steady-state and time-resolved fluorescence methods, focusing not only on the emission of the tryptophan residues, but also on emission of the substrate. As a substrate, MAMC [7-methoxy-4-(aminomethyl)-coumarin] was selected, a compound exhibiting native fluorescence. As well as the wild-type, the W128F (Trp128?Phe) mutant of CYP2D6 was studied. After binding, a variety of energy transfer possibilities exist, and molecular dynamics simulations were performed to calculate distances and relative orientations of donors and acceptors. Energy transfer from Trp128 to haem appeared to be important; its emission was related to the shortest of the three average tryptophan fluorescence lifetimes observed for CYP2D6. MAMC to haem energy transfer was very efficient as well: when bound in the active site, the emission of MAMC was fully quenched. Steady-state anisotropy revealed that besides the MAMC in the active site, another 2.4% of MAMC was bound outside of the active site to wild-type CYP2D6. The tryptophan residues in CYP2D6 appeared to be less accessible for the external quenchers iodide and acrylamide in presence of MAMC, indicating a tightening of the enzyme structure upon substrate binding. However, the changes in the overall enzyme structure were not very large, since the emission characteristics of the enzyme were not very different in the presence of MAMC. PMID:16190863

Stortelder, Aike; Keizers, Peter H. J.; Oostenbrink, Chris; De Graaf, Chris; De Kruijf, Petra; Vermeulen, Nico P. E.; Gooijer, Cees; Commandeur, Jan N. M.; Van Der Zwan, Gert

2005-01-01

108

Time-resolved and steady-state FRET spectroscopy on commercial biocompatible quantum dots  

NASA Astrophysics Data System (ADS)

Semiconductor nanocrystals (quantum dots - QDs) possess unique photophysical properties that make them highly interesting for many biochemical applications. Besides their common use as fluorophores in conventional spectroscopy and microscopy, QDs are well-suited for studying Förster resonance energy transfer (FRET). Size-dependent broadband absorption and narrow emission bands offer several advantages for the use of QDs both as FRET donors and acceptors. QD-based FRET pairs can be efficiently used as biological and chemical sensors for highly sensitive multiplexed detection. In this contribution we present the use of several commercially available QDs (Qdot® Nanocrystals - Invitrogen) as FRET donors in combination with commercial organic dyes as FRET acceptors. In order to investigate the FRET process within our donor-acceptor pairs, we used biotinylated QDs and streptavidin-labeled dyes. The well-known biotinstreptavidin molecular recognition enables effective FRET from QDs to dye molecules and provides defined distances between donor and acceptor. Steady-state and time-resolved fluorescence measurements were performed in order to investigate QD-to-dye FRET. Despite a thick polymer shell around the QDs, our results demonstrate the potential of these QDs as efficient donors both for steady-state and time-resolved FRET applications in nano-biotechnology.

Wegner, D.; Geißler, D.; Stufler, S.; Löhmannsröben, H.-G.; Hildebrandt, N.

2011-03-01

109

Direct Monte Carlo computation of time-resolved fluorescence in heterogeneous turbid media  

PubMed Central

We show that a multiexponential model for time-resolved fluorescence allows the use of an absorption-perturbation Monte Carlo (MC) approach based on stored photon path histories. This enables the rapid fitting of fluorescence yield, lifetimes, and background tissue absorptions in complex heterogeneous media within a few seconds, without the need for temporal convolutions or MC recalculation of photon path lengths. We validate this method using simulations with both a slab and a heterogeneous model of the mouse head. PMID:23164912

Kumar, Anand T. N.

2012-01-01

110

Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments.  

PubMed

The first capillary array scanner for time-resolved fluorescence detection in parallel capillary electrophoresis based on semiconductor technology is described. The system consists essentially of a confocal fluorescence microscope and a x,y-microscope scanning stage. Fluorescence of the labelled probe molecules was excited using a short-pulse diode laser emitting at 640 nm with a repetition rate of 50 MHz. Using a single filter system the fluorescence decays of different labels were detected by an avalanche photodiode in combination with a PC plug-in card for time-correlated single-photon counting (TCSPC). The time-resolved fluorescence signals were analyzed and identified by a maximum likelihood estimator (MLE). The x,y-microscope scanning stage allows for discontinuous, bidirectional scanning of up to 16 capillaries in an array, resulting in longer fluorescence collection times per capillary compared to scanners working in a continuous mode. Synchronization of the alignment and measurement process were developed to allow for data acquisition without overhead. Detection limits in the subzeptomol range for different dye molecules separated in parallel capillaries have been achieved. In addition, we report on parallel time-resolved detection and separation of more than 400 bases of single base extension DNA fragments in capillary array electrophoresis. Using only semiconductor technology the presented technique represents a low-cost alternative for high throughput DNA sequencing in parallel capillaries. PMID:10735310

Neumann, M; Herten, D P; Dietrich, A; Wolfrum, J; Sauer, M

2000-02-25

111

Halide (Cl(super -)) Quenching of Quinine Sulfate Fluorescence: A Time-Resolved Fluorescence Experiment for Physical Chemistry  

ERIC Educational Resources Information Center

The time-resolved fluorescence experiment investigating the halide quenching of fluorescence from quinine sulfate in water is described. The objectives of the experiment include reinforcing student understanding of the kinetics of competing pathways, making connections with microscopic theories of kinetics through comparison of experimental and…

Gutow, Jonathan H.

2005-01-01

112

Fluorescence lifetime heterogeneity in aggregates of LHCII revealed by time-resolved microscopy.  

PubMed Central

Two-photon excitation, time-resolved fluorescence microscopy was used to investigate the fluorescence quenching mechanisms in aggregates of light-harvesting chlorophyll a/b pigment protein complexes of photosystem II from green plants (LHCII). Time-gated microscopy images show the presence of large heterogeneity in fluorescence lifetimes not only for different LHCII aggregates, but also within a single aggregate. Thus, the fluorescence decay traces obtained from macroscopic measurements reflect an average over a large distribution of local fluorescence kinetics. This opens the possibility to resolve spatially different structural/functional units in chloroplasts and other heterogeneous photosynthetic systems in vivo, and gives the opportunity to investigate individually the excited states dynamics of each unit. We show that the lifetime distribution is sensitive to the concentration of quenchers contained in the system. Triplets, which are generated at high pulse repetition rates of excitation (>1 MHz), preferentially quench domains with initially shorter fluorescence lifetimes. This proves our previous prediction from singlet-singlet annihilation investigations (Barzda, V., V. Gulbinas, R. Kananavicius, V. Cervinskas, H. van Amerongen, R. van Grondelle, and L. Valkunas. 2001. Biophys. J. 80:2409-2421) that shorter fluorescence lifetimes originate from larger domains in LHCII aggregates. We found that singlet-singlet annihilation has a strong effect in time-resolved fluorescence microscopy of connective systems and has to be taken into consideration. Despite that, clear differences in fluorescence decays can be detected that can also qualitatively be understood. PMID:11423435

Barzda, V; de Grauw, C J; Vroom, J; Kleima, F J; van Grondelle, R; van Amerongen, H; Gerritsen, H C

2001-01-01

113

Time-resolved fluorescence line-narrowing of Eu3+ in biocompatible eutectic glass-ceramics.  

PubMed

The spectroscopic properties of Eu(3+) in biocompatible glass and glass-ceramic eutectic rods of composition 0.8CaSiO(3)-0.2Ca(3)(PO(4))(2) doped with 0.5 wt% of Eu(2)O(3) are investigated to explore their potential applications as optical probes. The samples were obtained by the laser floating zone technique. Depending on the growth rate, they exhibit three (two crystalline and one amorphous) or two (one crystalline and one amorphous) phases. The crystalline phases correspond to Ca(2)SiO(4) and apatite-like structures. At high growth rates the system presents an amorphous arrangement which gives a glass phase. The results of time-resolved fluorescence line narrowing spectroscopy obtained under excitation within the inhomogeneous broadened (7)F(0)?(5)D(0) absorption band allow to isolate the emission from Eu(3+) ions in the crystalline and amorphous environments and to accurately correlate the spectroscopic properties with the microstructure of these eutectics. PMID:23482227

Sola, D; Balda, R; Al-Saleh, M; Peña, J I; Fernández, J

2013-03-11

114

Time-resolved terahertz transmission spectroscopy of dielectrics  

Microsoft Academic Search

Using the method of time-domain terahertz-transmission spectroscopy we measured the far-infrared complex dielectric function at room temperature in the range of 3 to 80 cm for a variety of samples: Li2Ge7O15 (LGO), (CH3 NHCH2 COOH)3. CaCl2 (TSCC), high-permittivity microwave ceramics, etc. The precision of the measurements and subsequent evaluation of the dielectric function is discussed.

Petr Kužel; Jan Petzelt

2000-01-01

115

Optical characterization of Pseudomonas fluorescens on meat surfaces using time-resolved fluorescence  

NASA Astrophysics Data System (ADS)

A scanning optical system for the detection of bacteria on meat surfaces based on fluorescence lifetime and intensity measurements is described. The system detects autofluorescent light emitted by naturally occurring fluorophores in bacteria. The technique only requires minimal sample preparation and handling, thus the chemical properties of the specimen are preserved. This work presents the preliminary results obtained from a time-resolved fluorescence imaging system for the characterization of a nonpathogenic gram-negative bacteria, Pseudomonas fluorescens. Initial results indicate that the combination of fluorescence lifetime and intensity measurements provides a means for characterizing biological media and for detecting microorganisms on surfaces.

Bouchard, Alain; Frechette, Julie; Vernon, Marcia L.; Cormier, Jean-François; Beaulieu, Rene M.; Vallée, Réal; Mafu, Akier A.

2006-01-01

116

CMOS Time-Resolved, Contact, and Multispectral Fluorescence Imaging for DNA Molecular Diagnostics  

PubMed Central

Instrumental limitations such as bulkiness and high cost prevent the fluorescence technique from becoming ubiquitous for point-of-care deoxyribonucleic acid (DNA) detection and other in-field molecular diagnostics applications. The complimentary metal-oxide-semiconductor (CMOS) technology, as benefited from process scaling, provides several advanced capabilities such as high integration density, high-resolution signal processing, and low power consumption, enabling sensitive, integrated, and low-cost fluorescence analytical platforms. In this paper, CMOS time-resolved, contact, and multispectral imaging are reviewed. Recently reported CMOS fluorescence analysis microsystem prototypes are surveyed to highlight the present state of the art. PMID:25365460

Guo, Nan; Cheung, Ka Wai; Wong, Hiu Tung; Ho, Derek

2014-01-01

117

Frame-Transfer Gating Raman Spectroscopy for Time-Resolved Multiscalar Combustion Diagnostics  

NASA Technical Reports Server (NTRS)

Accurate experimental measurement of spatially and temporally resolved variations in chemical composition (species concentrations) and temperature in turbulent flames is vital for characterizing the complex phenomena occurring in most practical combustion systems. These diagnostic measurements are called multiscalar because they are capable of acquiring multiple scalar quantities simultaneously. Multiscalar diagnostics also play a critical role in the area of computational code validation. In order to improve the design of combustion devices, computational codes for modeling turbulent combustion are often used to speed up and optimize the development process. The experimental validation of these codes is a critical step in accepting their predictions for engine performance in the absence of cost-prohibitive testing. One of the most critical aspects of setting up a time-resolved stimulated Raman scattering (SRS) diagnostic system is the temporal optical gating scheme. A short optical gate is necessary in order for weak SRS signals to be detected with a good signal- to-noise ratio (SNR) in the presence of strong background optical emissions. This time-synchronized optical gating is a classical problem even to other spectroscopic techniques such as laser-induced fluorescence (LIF) or laser-induced breakdown spectroscopy (LIBS). Traditionally, experimenters have had basically two options for gating: (1) an electronic means of gating using an image intensifier before the charge-coupled-device (CCD), or (2) a mechanical optical shutter (a rotary chopper/mechanical shutter combination). A new diagnostic technology has been developed at the NASA Glenn Research Center that utilizes a frame-transfer CCD sensor, in conjunction with a pulsed laser and multiplex optical fiber collection, to realize time-resolved Raman spectroscopy of turbulent flames that is free from optical background noise (interference). The technology permits not only shorter temporal optical gating (down to <1 s, in principle), but also higher optical throughput, thus resulting in a substantial increase in measurement SNR.

Nguyen, Quang-Viet; Fischer, David G.; Kojima, Jun

2011-01-01

118

Comparison of a Time-Resolved Fluorescence Immunoassay and an Enzyme-Linked Immunosorbent Assay for the Analysis of Atrazine  

E-print Network

Comparison of a Time-Resolved Fluorescence Immunoassay and an Enzyme-Linked Immunosorbent Assay of California, Davis, California 95616 Immunoassays for atrazine based on a time-resolved fluorescent label immunoassay (TRFIA) was based on a polyclonal antibody and a europium label, whereas the enzyme

Hammock, Bruce D.

119

Unfolding of Ubiquitin Studied by Picosecond Time-Resolved Fluorescence of the Tyrosine Residue  

PubMed Central

The photophysics of the single tyrosine in bovine ubiquitin (UBQ) was studied by picosecond time-resolved fluorescence spectroscopy, as a function of pH and along thermal and chemical unfolding, with the following results: First, at room temperature (25°C) and below pH 1.5, native UBQ shows single-exponential decays. From pH 2 to 7, triple-exponential decays were observed and the three decay times were attributed to the presence of tyrosine, a tyrosine-carboxylate hydrogen-bonded complex, and excited-state tyrosinate. Second, at pH 1.5, the water-exposed tyrosine of either thermally or chemically unfolded UBQ decays as a sum of two exponentials. The double-exponential decays were interpreted and analyzed in terms of excited-state intramolecular electron transfer from the phenol to the amide moiety, occurring in one of the three rotamers of tyrosine in UBQ. The values of the rate constants indicate the presence of different unfolded states and an increase in the mobility of the tyrosine residue during unfolding. Finally, from the pre-exponential coefficients of the fluorescence decays, the unfolding equilibrium constants (KU) were calculated, as a function of temperature or denaturant concentration. Despite the presence of different unfolded states, both thermal and chemical unfolding data of UBQ could be fitted to a two-state model. The thermodynamic parameters Tm = 54.6°C, ?HTm = 56.5 kcal/mol, and ?Cp = 890 cal/mol//K, were determined from the unfolding equilibrium constants calculated accordingly, and compared to values obtained by differential scanning calorimetry also under the assumption of a two-state transition, Tm = 57.0°C, ?Hm= 51.4 kcal/mol, and ?Cp = 730 cal/mol//K. PMID:15454455

Noronha, Melinda; Lima, João C.; Bastos, Margarida; Santos, Helena; Maçanita, António L.

2004-01-01

120

Subcellular localization-dependent changes in EGFP fluorescence lifetime measured by time-resolved flow cytometry  

PubMed Central

Intracellular protein transport and localization to subcellular regions are processes necessary for normal protein function. Fluorescent proteins can be fused to proteins of interest to track movement and determine localization within a cell. Currently, fluorescence microscopy combined with image processing is most often used to study protein movement and subcellular localization. In this contribution we evaluate a high-throughput time-resolved flow cytometry approach to correlate intracellular localization of human LC3 protein with the fluorescence lifetime of enhanced green fluorescent protein (EGFP). Subcellular LC3 localization to autophagosomes is a marker of the cellular process called autophagy. In breast cancer cells expressing native EGFP and EGFP-LC3 fusion proteins, we measured the fluorescence intensity and lifetime of (i) diffuse EGFP (ii) punctate EGFP-LC3 and (iii) diffuse EGFP-?LC3 after amino acid starvation to induce autophagy-dependent LC3 localization. We verify EGFP-LC3 localization with low-throughput confocal microscopy and compare to fluorescence intensity measured by standard flow cytometry. Our results demonstrate that time-resolved flow cytometry can be correlated to subcellular localization of EGFP fusion proteins by measuring changes in fluorescence lifetime. PMID:24010001

Gohar, Ali Vaziri; Cao, Ruofan; Jenkins, Patrick; Li, Wenyan; Houston, Jessica P.; Houston, Kevin D.

2013-01-01

121

Time-resolved fluorescence in lipid bilayers: selected applications and advantages over steady state.  

PubMed

Fluorescence methods are versatile tools for obtaining dynamic and topological information about biomembranes because the molecular interactions taking place in lipid membranes frequently occur on the same timescale as fluorescence emission. The fluorescence intensity decay, in particular, is a powerful reporter of the molecular environment of a fluorophore. The fluorescence lifetime can be sensitive to the local polarity, hydration, viscosity, and/or presence of fluorescence quenchers/energy acceptors within several nanometers of the vicinity of a fluorophore. Illustrative examples of how time-resolved fluorescence measurements can provide more valuable and detailed information about a system than the time-integrated (steady-state) approach will be presented in this review: 1), determination of membrane polarity and mobility using time-dependent spectral shifts; 2), identification of submicroscopic domains by fluorescence lifetime imaging microscopy; 3), elucidation of membrane leakage mechanisms from dye self-quenching assays; and 4), evaluation of nanodomain sizes by time-resolved Förster resonance energy transfer measurements. PMID:25517142

Amaro, Mariana; Šachl, Radek; Jurkiewicz, Piotr; Coutinho, Ana; Prieto, Manuel; Hof, Martin

2014-12-16

122

Time-resolved fluorescence ligand binding for G protein-coupled receptors.  

PubMed

G protein-coupled receptors (GPCRs) and their ligands are traditionally characterized by radioligand-binding experiments. These experiments yield excellent quantitative data, but have low temporal and spatial resolution. In addition, the use of radioligands presents safety concerns. Here we provide a general procedure for an alternative approach with high temporal and spatial resolution, based on Tb(+)-labeled fluorescent receptor ligands and time-resolved fluorescence resonance energy transfer (TR-FRET). This protocol and its design are detailed here for the parathyroid hormone receptor, a class B GPCR, and its fluorescently labeled 34-amino acid peptide ligand, but it can be easily modified for other receptors and their appropriately labeled ligands. We discuss three protocol options that use Tb(+)-labeled fluorescent ligands: a time-resolved fluorescence separation option that works on native receptors but requires separation of bound and unbound ligand; a TR-FRET option using SNAP-tag-labeled receptors for high-throughput screening; and a TR-FRET option that uses fluorescently labeled antibodies directed against an epitope engineered into the Flag-labeled receptors' N terminus. These protocol options can be used as standard procedures with very high signal-to-background ratios in order to characterize ligands and their receptors in living cells and in cell membranes via straightforward plate-reader measurements. PMID:23764938

Emami-Nemini, Alexander; Roux, Thomas; Leblay, Marion; Bourrier, Emmanuel; Lamarque, Laurent; Trinquet, Eric; Lohse, Martin J

2013-01-01

123

Time-Resolved Fluorescence Immunoassay for C-Reactive Protein Using Colloidal Semiconducting Nanoparticles  

PubMed Central

Besides the typical short-lived fluorescence with decay times in the nanosecond range, colloidal II/VI semiconductor nanoparticles dispersed in buffer also possess a long-lived fluorescence component with decay times in the microsecond range. Here, the signal intensity of the long-lived luminescence at microsecond range is shown to increase 1,000-fold for CdTe nanoparticles in PBS buffer. This long-lived fluorescence can be conveniently employed for time-gated fluorescence detection, which allows for improved signal-to-noise ratio and thus the use of low concentrations of nanoparticles. The detection principle is demonstrated with a time-resolved fluorescence immunoassay for the detection of C-reactive protein (CRP) using CdSe-ZnS nanoparticles and green light excitation. PMID:22247668

Härmä, Harri; Toivonen, Juha; Soini, Juhani T.; Hänninen, Pekka; Parak, Wolfgang J.

2011-01-01

124

Time-resolvable fluorescent conjugates for the detection of pathogens in environmental samples containing autofluorescent material  

NASA Astrophysics Data System (ADS)

Water is routinely monitored for environmental pathogens such a Cryptosporidium and Giardia using immunofluorescence microscopy (IFM). Autofluorescence can greatly diminish an operators capacity to resolve labeled pathogens from non-specific background. Naturally fluorescing components (autofluorophores) encountered in biological samples typically have fluorescent lifetimes (?) of less than 100 nanoseconds and their emissions may be excluded through use of time-resolved fluorescence microscopy (TRFM). TRFM relies on the large differences in ? between autofluorescent molecules and long-lived lanthanide chelates. In TRFM, targets labeled with a time-resolvable fluorescent immunoconjugate are excited by an intense (UV) light pulse. A short delay is imposed to permit the decay of autofluorescence before capture of luminescence from the excited chelate using an image intensified CCD camera. In our experience, autofluorescence can be reduced to insignificant levels with a consequent 30-fold increase in target visibility using TRFM techniques. We report conjugation of a novel europium chelate to a monoclonal antibody specific for Giardia lamblia and use of the immunoconjugate for TRFM studies. Initial attempts to conjugate the same chelate to a monoclonal antibody directed against Cryptosporidium parvum led to poorly fluorescent constructs that were prone to denature and precipitate. We successfully conjugated BHHCT to anti-mouse polyvalent immunoglobulin and used this construct to overcome the difficulties in direct labeling of the anti-Cryptosporidium antibody. Both Giardia and Cryptosporidium were labeled using the anti-mouse protocol with a subsequent 20-fold and 6.6-fold suppression of autofluorescence respectively. A rapid protocol for conjugating and purifying the immunoconjugate was found and methods of quantifying the fluorescence to protein ratio determined. Performance of our TRFM was dependent on the quality and brightness of the immunoconjugate and optimization of the conjugation process is necessary to reap the full benefit of time-resolved techniques.

Connally, Russell; Veal, Duncan; Piper, James A.

2003-07-01

125

Molecular diffusivity measurement through an alumina membrane using time-resolved fluorescence imaging  

NASA Astrophysics Data System (ADS)

We present a simple fluorescence imaging method for measuring the time-resolved concentration of a fluorescent molecule diffusing through an anodic alumina membrane with a pore diameter of 20 nm. From the concentration breakthrough curve, the molecular diffusivity of the fluorophore was extracted. The experimentally determined diffusivity was three orders of magnitude lower than reported bulk values. Due to the relative simplicity and ease of use, this method can be applied to provide fundamental information for biomolecular separations applications. One feature of this method is the high sensitivity at intercellular volumes broadening its application to drug delivery and controlled cell growth.

Kennard, Raymond; DeSisto, William J.; Mason, Michael D.

2010-11-01

126

Speciation of uranyl species in nitric acid medium by time-resolved laser-induced fluorescence  

NASA Astrophysics Data System (ADS)

The aim of this work is the development of an on-line analytical procedure for uranyl trace determination in the nuclear fuel reprocessing process using time-resolved laser-induced fluorescence. Because the uranyl fluorescence spectrum is strongly affected by the nitrate concentration, knowledge of composition of the medium is necessary to normalize measurements. This paper reports the assumptions made on the spectral distortion, leading to a spectral deconvolution model. Uranyl complex formation constants are obtained from the spectral deconvolution and validate the method. In this way, spectral distortion allows the determination of the nitrate concentration with good accuracy.

Couston, Laurent; Pouyat, Dominique; Moulin, Christophe; Decambox, Pierre

1995-03-01

127

Towards time resolved core level photoelectron spectroscopy with femtosecond x-ray free-electron lasers  

NASA Astrophysics Data System (ADS)

We have performed core level photoelectron spectroscopy on a W(110) single crystal with femtosecond XUV pulses from the free-electron laser at Hamburg (FLASH). We demonstrate experimentally and through theoretical modelling that for a suitable range of photon fluences per pulse, time-resolved photoemission experiments on solid surfaces are possible. Using FLASH pulses in combination with a synchronized optical laser, we have performed femtosecond time-resolved core-level photoelectron spectroscopy and observed sideband formation on the W 4f lines indicating a cross correlation between femtosecond optical and XUV pulses.

Pietzsch, A.; Föhlisch, A.; Beye, M.; Deppe, M.; Hennies, F.; Nagasono, M.; Suljoti, E.; Wurth, W.; Gahl, C.; Döbrich, K.; Melnikov, A.

2008-03-01

128

Two-Photon Absorption and Time-Resolved Stimulated Emission Depletion Spectroscopy of a New Fluorenyl Derivative  

PubMed Central

The synthesis, comprehensive linear photophysical characterization, two-photon absorption (2PA), steady-state and time-resolved stimulated emission depletion properties of a new fluorene derivative, (E)-1-(2-(di-p-tolylamino)-9,9-diethyl-9H-fluoren-7-yl)-3-(thiophen-2-yl)prop-2-en-1-one (1), are reported. The primary linear spectral properties, including excitation anisotropy, fluorescence lifetimes, and photostability, were investigated in a number of aprotic solvents at room temperature. The degenerate 2PA spectra of 1 were obtained with an open aperture Z-scan and two-photon induced fluorescence methods, using a 1-kHz femtosecond laser system, and maximum 2PA cross-sections of ~400–600 GM were obtained. The nature of the electronic absorption processes in 1 was investigated by DFT-based quantum chemical methods implemented in the Gaussian 09 program. The one- and two-photon stimulated emission spectra of 1 were measured over a broad spectral range using a femtosecond pump probe–based fluorescence quenching technique, while a new methodology for time-resolved fluorescence emission spectroscopy is proposed. An effective application of 1 in fluorescence bioimaging was demonstrated via one- and two-photon fluorescence microscopy images of HCT 116 cells containing the dye encapsulated micelles. PMID:22887914

Bondar, Mykhailo V.; Morales, Alma R.; Yue, Xiling; Luchita, Gheorghe; Przhonska, Olga V.; Kachkovsky, Olexy D.

2012-01-01

129

Time-resolved fluorescence monitoring of cholesterol in peripheral blood mononuclear cells  

NASA Astrophysics Data System (ADS)

Precise evaluation of intracellular cholesterol distribution is crucial for improving diagnostics of diseased states associated with cholesterol alteration. Time-resolved fluorescence techniques are tested for non-invasive investigation of cholesterol in living cells. Fluorescent probe NBD attached to cholesterol was employed to evaluate cholesterol distribution in peripheral blood mononuclear cells (PBMC) isolated from the human blood. Fluorescence Lifetime Imaging Microscopy (FLIM) was successfully applied to simultaneously monitor the spatial distribution and the timeresolved characteristics of the NBD-cholesterol fluorescence in PBMC. Gathered data are the first step in the development of a new perspective non-invasive diagnostic method for evaluation of cholesterol modifications in diseases associated with disorders of lipid metabolism.

Martinakova, Z.; Horilova, J.; Lajdova, I.; Marcek Chorvatova, A.

2014-12-01

130

An instrument for small-animal imaging using time-resolved diffuse and fluorescence optical methods  

NASA Astrophysics Data System (ADS)

We describe time-resolved optical methods that use diffuse near-infrared photons to image the optical properties of tissues and their inner fluorescent probe distribution. The assembled scanner uses picosecond laser diodes at 4 wavelengths, an 8-anode photo-multiplier tube and time-correlated single photon counting. Optical absorption and reduced scattering images as well as fluorescence emission images are computed from temporal profiles of diffuse photons. This method should improve the spatial resolution and the quantification of fluorescence signals. We used the diffusion approximation of the radiation transport equation and the finite element method to solve the forward problem. The inverse problem is solved with an optimization algorithm such as ART or conjugate gradient. The scanner and its performances are presented, together with absorption, scattering and fluorescent images obtained with it.

Montcel, Bruno; Poulet, Patrick

2006-12-01

131

THE JOURNAL OF CHEMICAL PHYSICS 139, 124113 (2013) Time-resolved broadband Raman spectroscopies: A unified  

E-print Network

THE JOURNAL OF CHEMICAL PHYSICS 139, 124113 (2013) Time-resolved broadband Raman spectroscopies in molecules can be studied by an electronically off-resonant Raman process induced by a probe pulse of the technique that involve either spontaneous or stimulated Raman detection and different pulse configurations

Mukamel, Shaul

132

In vivo optical characterization of human prostate tissue using near-infrared time-resolved spectroscopy  

Microsoft Academic Search

The development of photodynamic therapy into a modality for treatment of prostate cancer calls for reliable optical dosimetry. We employ, for the first time, interstitial time-resolved spectroscopy to determine in vivo optical properties of human prostate tissue. Nine patients are included in the study, and measurements are conducted prior to primary brachytherapy treatment of prostate cancer. Intra- subject variability is

Tomas Svensson; Stefan Andersson-Engels; Margre´T. EinarsdóTtíR; Katarina Svanberg

2007-01-01

133

Time-resolved IR laser-assisted XUV photoelectron spectroscopy of metal surfaces  

E-print Network

Time-resolved IR laser-assisted XUV photoelectron spectroscopy of metal surfaces C.-H. Zhang and U by an XUV pulse of length X into the field of a delayed IR laser pulse with carrier period TL allows of the emitted photoelectrons (PEs) oscillates with period TL as a function of the XUV-IR pulse delay, leading

Thumm, Uwe

134

Evidence for a Novel Mechanism of Time-Resolved Flavin Fluorescence Depolarization in Glutathione Reductase  

PubMed Central

Time-resolved flavin fluorescence anisotropy studies on glutathione reductase (GR) have revealed a remarkable new phenomenon: wild-type GR displays a rapid process of fluorescence depolarization, that is absent in mutant enzymes lacking a nearby tyrosine residue that blocks the NADPH-binding cleft. Fluorescence lifetime data, however, have shown a more rigid active-site structure for wild-type GR than for the tyrosine mutants. These results suggest that the rapid depolarization in wild-type GR originates from an interaction with the flavin-shielding tyrosine, and not from restricted reorientational motion of the flavin. A novel mechanism of fluorescence depolarization is proposed that involves a transient charge-transfer complex between the tyrosine and the light-excited flavin, with a concomitant change in the direction of the emission dipole moment of the flavin. This interaction is likely to result from side-chain relaxation of the tyrosine in the minor fraction of enzyme molecules in which this residue is in an unsuitable position for immediate fluorescence quenching at the moment of excitation. Support for this mechanism is provided by binding studies with NADP+ and 2?P-5?ADP-ribose that can intercalate between the flavin and tyrosine and/or block the latter. Fluorescence depolarization analyses as a function of temperature and viscosity confirm the dynamic nature of the process. A comparison with fluorescence depolarization effects in a related flavoenzyme indicates that this mechanism of flavin fluorescence depolarization is more generally applicable. PMID:15454452

van den Berg, Petra A. W.; van Hoek, Arie; Visser, Antonie J. W. G.

2004-01-01

135

Advanced Time-Resolved Fluorescence Microscopy Techniques for the Investigation of Peptide Self-Assembly  

NASA Astrophysics Data System (ADS)

The ubiquitous cross beta sheet peptide motif is implicated in numerous neurodegenerative diseases while at the same time offers remarkable potential for constructing isomorphic high-performance bionanomaterials. Despite an emerging understanding of the complex folding landscape of cross beta structures in determining disease etiology and final structure, we lack knowledge of the critical initial stages of nucleation and growth. In this dissertation, I advance our understanding of these key stages in the cross-beta nucleation and growth pathways using cutting-edge microscopy techniques. In addition, I present a new combined time-resolved fluorescence analysis technique with the potential to advance our current understanding of subtle molecular level interactions that play a pivotal role in peptide self-assembly. Using the central nucleating core of Alzheimer's Amyloid-beta protein, Abeta(16 22), as a model system, utilizing electron, time-resolved, and non-linear microscopy, I capture the initial and transient nucleation stages of peptide assembly into the cross beta motif. In addition, I have characterized the nucleation pathway, from monomer to paracrystalline nanotubes in terms of morphology and fluorescence lifetime, corroborating the predicted desolvation process that occurs prior to cross-beta nucleation. Concurrently, I have identified unique heterogeneous cross beta domains contained within individual nanotube structures, which have potential bionanomaterials applications. Finally, I describe a combined fluorescence theory and analysis technique that dramatically increases the sensitivity of current time-resolved techniques. Together these studies demonstrate the potential for advanced microscopy techniques in the identification and characterization of the cross-beta folding pathway, which will further our understanding of both amyloidogenesis and bionanomaterials.

Anthony, Neil R.

136

Development of a New Time-Resolved Laser-Induced Fluorescence Technique  

NASA Astrophysics Data System (ADS)

We are developing a time-resolved laser-induced fluorescence (LIF) technique to interrogate the ion velocity distribution function (VDF) of EP thruster plumes down to the microsecond time scale. Better measurements of dynamic plasma processes will lead to improvements in simulation and prediction of thruster operation and erosion. We present the development of the new technique and results of initial tests. Signal-to-noise ratio (SNR) is often a challenge for LIF studies, and it is only more challenging for time-resolved measurements since a lock-in amplifier cannot be used with a long time constant. The new system uses laser modulation on the order of MHz, which enables the use of electronic filtering and phase-sensitive detection to improve SNR while preserving time-resolved information. Statistical averaging over many cycles to further improve SNR is done in the frequency domain. This technique can have significant advantages, including (1) larger spatial maps enabled by shorter data acquisition time and (2) the ability to average data without creating a phase reference by modifying the thruster operating condition with a periodic cutoff in discharge current, which can modify the ion velocity distribution.

Durot, Christopher; Gallimore, Alec

2012-10-01

137

A CTRW-based model of time-resolved fluorescence lifetime imaging in a turbid medium  

PubMed Central

We develop an analytic model of time-resolved fluorescent imaging of photons migrating through a semi-infinite turbid medium bounded by an infinite plane in the presence of a single stationary point fluorophore embedded in the medium. In contrast to earlier models of fluorescent imaging in which photon motion is assumed to be some form of continuous diffusion process, the present analysis is based on a continuous-time random walk (CTRW) on a simple cubic lattice, the object being to estimate the position and lifetime of the fluorophore. Such information can provide information related to local variations in pH and temperature with potential medical significance. Aspects of the theory were tested using time-resolved measurements of the fluorescence from small inclusions inside tissue-like phantoms. The experimental results were found to be in good agreement with theoretical predictions provided that the fluorophore was not located too close to the planar boundary, a common problem in many diffusive systems. PMID:21057657

Chernomordik, Victor; Gandjbakhche, Amir H.; Hassan, Moinuddin; Pajevic, Sinisa; Weiss, George H.

2010-01-01

138

Fluorescence time-resolved imaging system embedded in an ultrasound prostate probe  

PubMed Central

Ultrasound imaging (US) of the prostate has a low specificity to distinguish tumors from the surrounding tissues. This limitation leads to systematic biopsies. Fluorescent diffuse optical imaging may represent an innovative approach to guide biopsies to tumors marked with high specificity contrast agents and therefore enable an early detection of prostate cancer. This article describes a time-resolved optical system embedded in a transrectal US probe, as well as the fluorescence reconstruction method and its performance. Optical measurements were performed using a pulsed laser, optical fibers and a time-resolved detection system. A novel fast reconstruction method was derived and used to locate a 45 µL ICG fluorescent inclusion at a concentration of 10 µM, in a liquid prostate phantom. Very high location accuracy (0.15 cm) was achieved after reconstruction, for different positions of the inclusion, in the three directions of space. The repeatability, tested with ten sequential measurements, was of the same order of magnitude. Influence of the input parameters (optical properties and lifetime) is presented. These results confirm the feasibility of using optical imaging for prostate guided biopsies. PMID:21326649

Laidevant, Aurélie; Hervé, Lionel; Debourdeau, Mathieu; Boutet, Jérôme; Grenier, Nicolas; Dinten, Jean-Marc

2011-01-01

139

Time-resolved fluorescence of the bacteriophage T4 capsid protein gp23.  

PubMed

The time-resolved fluorescence properties of the bacteriophage T4 capsid protein gp23 are investigated. The structural characteristics of this protein are largely unknown and can be probed by recording time-resolved and decay-associated fluorescence spectra and intensity decay curves using a 200 ps-gated intensified CCD-camera. Spectral and decay data are recorded simultaneously, which makes data acquisition fast compared to time-correlated single-photon counting. A red-shift of the emission maximum within the first nanosecond of decay is observed, which can be explained by the different decay-associated spectra of fluorescence lifetimes of the protein in combination with dipolar relaxation. In addition, iodide quenching experiments are performed, to study the degree of exposure of the various tryptophan residues. A model for the origin of the observed lifetimes of 0.032 +/- 0.003, 0.39 +/- 0.06, 2.1 +/- 0.1 and 6.8 +/- 0.8 ns is presented: the 32 ps lifetime can be assigned to the emission of a buried tryptophan residue, the 0.4 and 2.1 ns lifetimes to two partly buried residues, and the 6.8 ns lifetime to a single tryptophan outside the bulk of the folded gp23. PMID:15629249

Stortelder, Aike; Buijs, Joost B; Bulthuis, Jaap; van der Vies, Saskia M; Gooijer, Cees; van der Zwan, Gert

2005-01-14

140

High-performance time-resolved fluorescence by direct waveform recording  

PubMed Central

We describe a high-performance time-resolved fluorescence (HPTRF) spectrometer that dramatically increases the rate at which precise and accurate subnanosecond-resolved fluorescence emission waveforms can be acquired in response to pulsed excitation. The key features of this instrument are an intense(1 ?J?pulse), high-repetition rate (10 kHz), and short (1 ns full width at half maximum) laser excitation source and a transient digitizer (0.125 ns per time point) that records a complete and accurate fluorescence decay curve for every laser pulse. For a typical fluorescent sample containing a few nanomoles of dye, a waveform with a signal?noise of about 100 can be acquired in response to a single laser pulse every 0.1 ms, at least 105 times faster than the conventional method of time-correlated single photon counting, with equal accuracy and precision in lifetime determination for lifetimes as short as 100 ps. Using standard single-lifetime samples, the detected signals are extremely reproducible, with waveform precision and linearity to within 1% error for single-pulse experiments. Waveforms acquired in 0.1 s (1000 pulses) with the HPTRF instrument were of sufficient precision to analyze two samples having different lifetimes, resolving minor components with high accuracy with respect to both lifetime and mole fraction. The instrument makes possible a new class of high-throughput time-resolved fluorescence experiments that should be especially powerful for biological applications, including transient kinetics, multidimensional fluorescence, and microplate formats. PMID:21034069

Muretta, Joseph M.; Kyrychenko, Alexander; Ladokhin, Alexey S.; Kast, David J.; Gillispie, Gregory D.; Thomas, David D.

2010-01-01

141

Direct detection of time-resolved Rabi oscillations in a single quantum dot via resonance fluorescence  

NASA Astrophysics Data System (ADS)

Optical Rabi oscillations are coherent population oscillations of a two-level system coupled by an electric dipole transition when driven by a strong nearly resonant optical field. In quantum dot structures, these measurements have typically been performed as a function of the total pulse area ??0(t)dt where the pulse area varies as a function of Rabi frequency. Here, we report direct detection of the time-resolved coherent transient response of the resonance fluorescence to measure the time evolution of the optical Rabi oscillations in a single charged InAs quantum dot. We extract a decoherence rate consistent with the limit from the excited state lifetime.

Schaibley, J. R.; Burgers, A. P.; McCracken, G. A.; Steel, D. G.; Bracker, A. S.; Gammon, D.; Sham, L. J.

2013-03-01

142

Application of Time-Resolved Fluorescence for Direct and Continuous Probing of Release from Polymeric Delivery Vehicles  

PubMed Central

Though accurately evaluating the kinetics of release is critical for validating newly designed therapeutic carriers for in vivo applications, few methods yet exist for release measurement in real time and without the need for any sample preparation. Many of the current approaches (e.g. chromatographic methods, absorption spectroscopy, or NMR spectroscopy) rely on isolation of the released material from the loaded vehicles, which require additional sample purification and can lead to loss of accuracy when probing fast kinetics of release. In this study we describe the use of time-resolved fluorescence for in situ monitoring of small molecule release kinetics from biodegradable polymeric drug delivery systems. This method relies on the observation that fluorescent reporters being released from polymeric drug delivery systems possess distinct excited-state lifetime components, reflecting their different environments in the particle suspensions, i.e., confined in the polymer matrices or free in the aqueous environment. These distinct lifetimes enable real-time quantitative mapping of the relative concentrations of dye in each population to obtain precise and accurate temporal information on the release profile of particular carrier/payload combinations. We found that fluorescence lifetime better distinguishes subtle differences in release profiles, (e.g. differences associated with dye loading) than conventional steady-state fluorescence measurements, which represent the averaged dye behavior over the entire scan. Given the method's applicability to both hydrophobic and hydrophilic cargo, it could be employed to model the release of any drug-carrier combination. PMID:23792808

Viger, Mathieu L.; Sheng, Wangzhong; McFearin, Cathryn L.; Berezin, Mikhail Y.; Almutairi, Adah

2013-01-01

143

Binding and relaxation behavior of Coumarin-153 in lecithin-taurocholate mixed micelles: A time resolved fluorescence spectroscopic study  

NASA Astrophysics Data System (ADS)

The microenvironment of the bile salt-lecithin mixed aggregates has been investigated using steady state and picosecond time resolved fluorescence spectroscopy. The steady state spectra show that the polarity of the bile salt is higher compared to lecithin vesicles or the mixed aggregates. We have observed slow solvent relaxation in bile salt micelles and lecithin vesicles. The solvation time is gradually slowed down due to gradual addition of the bile salt in lecithin vesicles. Addition of bile salt leads to the tighter head group packing in lecithin. Thus, mobility of the water molecules becomes slower and consequently the solvation time is also retarded. We have observed bimodal slow rotational relaxation time in all these systems.

Chakrabarty, Debdeep; Chakraborty, Anjan; Seth, Debabrata; Hazra, Partha; Sarkar, Nilmoni

2005-09-01

144

Time-resolved remote Raman and fluorescence spectrometers for planetary exploration  

NASA Astrophysics Data System (ADS)

At the University of Hawaii, we have developed compact time-resolved (TR) Raman, and fluorescence spectrometers suitable for planetary exploration under NASA's Mars Instrument Development Program. The compact Raman and fluorescence spectrometers consist of custom miniature spectrographs based on volume holographic gratings, and custom miniature intensified CCD cameras. These spectrographs have been interfaced with a regular 50 mm camera lens as well as with a three and a half inch diameter telescope for remotely interrogating minerals, water, water-ice and dry ice. Using a small frequency-doubled Nd:YAG pulsed laser (35 mJ/pulse, 20 Hz) and 50 mm camera lens, TRRaman and LINF spectra of minerals, and bio-minerals can be measured within 30 s under super-critical CO2, and with 3.5-inch telescope these samples can be interrogated to 50 m radial distance during day time and nighttime. The fluorescence spectrograph is capable of measuring TR- laser-induced fluorescence excited with 355 nm laser in the spectral range 400-800 nm spectral range. The TR-fluorescence spectra allow measurement of LINF from rare-earths and transition-metal ions in time domain, and also assist in differentiating between abiogenic minerals from organic and biogenic materials based on the fluorescence lifetime. Biological materials are also identified from their characteristic short-lived (<10 ns) laser-induced fluorescence lifetime. These instruments will play important role in planetary exploration especially in NASA's future Mars Sample Return Mission, and lander and rover missions.

Sharma, Shiv K.; Misra, Anupam K.; Acosta, Tayro E.; Lucey, Paul G.

2012-06-01

145

Protein self-association in crowded protein solutions: A time-resolved fluorescence polarization study  

PubMed Central

The self-association equilibrium of a tracer protein, apomyoglobin (apoMb), in highly concentrated crowded solutions of ribonuclease A (RNase A) and human serum albumin (HSA), has been studied as a model system of protein interactions that occur in crowded macromolecular environments. The rotational diffusion of the tracer protein labeled with two different fluorescent dyes, 8-anilinonaphthalene-1-sulfonate and fluorescein isothiocyanate, was successfully recorded as a function of the two crowder concentrations in the 50–200 mg/mL range, using picosecond-resolved fluorescence anisotropy methods. It was found that apoMb molecules self-associate at high RNase A concentration to yield a flexible dimer. The apparent dimerization constant, which increases with RNase A concentration, could also be estimated from the fractional contribution of monomeric and dimeric species to the total fluorescence anisotropy of the samples. In contrast, an equivalent mass concentration of HSA does not result in tracer dimerization. This different effect of RNase A and HSA is much larger than that predicted from simple models based only on the free volume available to apoMb, indicating that additional, nonspecific interactions between tracer and crowder should come into play. The time-resolved fluorescence polarization methods described here are expected to be of general applicability to the detection and quantification of crowding effects in a variety of macromolecules of biological relevance. PMID:15459331

Zorrilla, Silvia; Rivas, Germán; Acuña, A. Ulises; Lillo, M. Pilar

2004-01-01

146

Fluorescence decay processes following resonant 2p photoexcitation of Ar atoms and clusters studied using a time-resolved fluorescence and photoion coincidence technique  

NASA Astrophysics Data System (ADS)

The novel spectroscopic technique of time-resolved fluorescence-photoion coincidence spectroscopy (TFPICO) has been applied to the investigation of the decay processes of 2p inner-shell excited Ar atoms and clusters. For the Ar atom, only that fluorescence accompanying the production of Ar+ showed a strong dependence on excitation energy. This dependence is discussed in terms of competing Auger decay processes. For Ar clusters, the TFPICO spectra for dimer ions (Ar2)+ revealed long-lifetime fluorescence components which can be attributed to the ‘third excimer continuum’. With this work we demonstrate the usefulness of this technique for investigating the decay processes of inner-shell excited atoms and clusters.

Gejo, T.; Ikegami, T.; Honma, K.; Harries, J. R.; Tamenori, Y.

2013-04-01

147

Time-Resolved Fluorescence Studies Of Ph Effects On The Conformation Of Troponin C  

NASA Astrophysics Data System (ADS)

Using time-resolved nanosecond fluorescence spectroscopy, we investigated the conformational changes of skeletal and cardiac troponin C. A thiol specific fluorescence probe N-(iodoacety1)-N'-(5-sulfo-1-naphthyl)- ethylenediamine (IAEDANS) was attached to cysteine 98 of skeletal troponin C (STnC) and 2-(4'-iodoacetamido-anilino)-naphthalene-6-sulfonic acid (IAANS) was linked to cysteine 35 and 84 of cardiac troponin C (CTnC). With excitation at 340 nm for STnC-IAEDANS and at 335 nm for CTnC-IAANS, apo-STnC and apo-CTnC exhibited biexponential decay kinetics. At 20°C and neutral pH, the following lifetimes were observed: (1) apo-STnC, 9 and 16 ns, and (2) apo-CTnC, 2.3 and 7 ns. The long lived component of the emission in STnC-IAEDANS comprised ~ 61% of observed intensity, however, the corresponding component in CTnC contributed only. A decrease of pH from 7.2 to 5.2 induced an increase of the lifetimes 20% (STnC) and 10% (CTnC). These results suggest that Cys-98 of STnC and Cys-35 and Cys-84 of CTnC became less quenched by their neighboring residues at low pH. Addition of guanidine hydrochloride to STnC resulted in a decrease of ~30% of both lifetimes. The lifetimes increased slightly when the temperature was lowered. Variation of solution viscosity by addition of sucrose did not affect the long component of the lifetimes of STnC. However, the short component did sense a viscosity effect. These results suggest that there was likely a chromophore heterogeneity which may arise from differences in conformation, environment, and or ionization of the excited state of the chromophore. At 20°C and neutral pH, two rotational correlation times were observed: (1) apo-STnC, - 1.2 ns, .2 1 11.3 ns, and (2) apo-CTnC, .~ 0.6 ns, .2 1 13.6 ns. The short rotational correlation times likely reflect rapid motions of the chromophores covalently attached to the side chain of the cysteine residues, and the long correlation times reflect the overall protein motions. The .2 values suggest that both proteins were not highly asymmetric at neutral pH. In the presence of Ca2+, at 20 °C and neutral pH the rotational correlation times were (1) STnC, .1 ~ 2.1 ns, .2 1 13.9 ns, and (2) CTnC, .1 ~ 1.9 ns, .2 1 13.0 ns. A decrease of pH from 7.2 to 5.2 led to an increase of .2 ~ 20% for STnC and a reduction of .2 ~ 7% for CTnC. The two proteins appear to have different hydrodynamic properties in an acidic environment.

Wang, Chien-Kao; Liao, Ronglihi; Cheung, Herbert C.

1988-06-01

148

Time-resolved fluorescence and fluorescence anisotropy of calix[4]arene: Elucidation of the excitation energies of various conformers  

NASA Astrophysics Data System (ADS)

Fluorescence, fluorescence excitation, time-resolved fluorescence spectroscopic, time-dependent fluorescence anisotropy, and time-dependent (TD) DFT studies were carried out to elucidate the excitation energies, fluorescence decay kinetics, and excitation transfer dynamics of calix[4]arene. We found that TD DFT calculations help to analyze the absorption spectra by elucidating the variation of the oscillator strengths with the excitation energies of various conformers involved and to clarify their relative importance. The TD DFT calculations imply that a cone-shaped conformer plays the most important role in the absorption spectrum. Fluorescence in a methanol solvent has a lifetime of 3.11 ± 0.1 ns and the decay kinetics is not found to have particular propensity with respect to solvent polarity. When the molecules in a glass matrix at 77 K were excited by short laser pulses, fluorescence anisotropy was observed. From the time-dependent fluorescence anisotropy, we derived the energy transfer rate constant and fluorescence anisotropy decay time, respectively: ket=1.7×109 s-1, ?h=0.59 ns.

Boo, Bong Hyun; Kim, Hyun Sook; Koh, Sang Gon; Lee, Minyung; No, Kwanghyun

2013-01-01

149

Ultrasensitive detection of microRNA with isothermal amplification and a time-resolved fluorescence sensor.  

PubMed

MicroRNAs (miRNAs) play important roles in a plethora of biological and cellular processes. The levels of miRNAs can be useful biomarkers for cellular events or disease diagnosis, thus the method for sensitive and selective detection of miRNAs is imperative to miRNA discovery, study, and clinical diagnosis. Here we develop a novel method to quantify miRNA expression levels as low as attomolar sensitivity by two-stage exponential amplification reaction (EXPAR) and a time-resolved fluorescence sensor in real samples. The method reveals superior sensitivity with a detection limit of miRNA of 0.1 aM under pure conditions. The method also shows the high selectivity for discriminating differences between miRNA family members, thus providing a promising alternative to standard approaches for quantitative detection of miRNA. PMID:24561522

Wang, Ke; Zhang, Kai; Lv, Zhuwu; Zhu, Xue; Zhu, Ling; Zhou, Fanfan

2014-07-15

150

A CMOS Time-Resolved Fluorescence Lifetime Analysis Micro-System  

PubMed Central

We describe a CMOS-based micro-system for time-resolved fluorescence lifetime analysis. It comprises a 16 × 4 array of single-photon avalanche diodes (SPADs) fabricated in 0.35 ?m high-voltage CMOS technology with in-pixel time-gated photon counting circuitry and a second device incorporating an 8 × 8 AlInGaN blue micro-pixellated light-emitting diode (micro-LED) array bump-bonded to an equivalent array of LED drivers realized in a standard low-voltage 0.35 ?m CMOS technology, capable of producing excitation pulses with a width of 777 ps (FWHM). This system replaces instrumentation based on lasers, photomultiplier tubes, bulk optics and discrete electronics with a PC-based micro-system. Demonstrator lifetime measurements of colloidal quantum dot and Rhodamine samples are presented. PMID:22291564

Rae, Bruce R.; Muir, Keith R.; Gong, Zheng; McKendry, Jonathan; Girkin, John M.; Gu, Erdan; Renshaw, David; Dawson, Martin D.; Henderson, Robert K.

2009-01-01

151

Serological Responses to Experimental Norwalk Virus Infection Measured Using a Quantitative Duplex Time-Resolved Fluorescence Immunoassay ?  

PubMed Central

A quantitative duplex time-resolved fluorescence assay, dissociation-enhanced lanthanide fluorescent immunoassay (DELFIA), was developed to measure Norwalk virus (NV)-specific IgA and IgG antibodies simultaneously. The duplex assay showed superior performance by detecting seroconversion following experimental NV infection at an earlier time point than a reference total immunoglobulin enzyme-linked immunosorbent assay (ELISA). PMID:21593238

Kavanagh, Owen; Estes, Mary K.; Reeck, Amanda; Raju, Ravikiran M.; Opekun, Antone R.; Gilger, Mark A.; Graham, David Y.; Atmar, Robert L.

2011-01-01

152

Detection of membrane packing defects by time-resolved fluorescence depolarization.  

PubMed

Packing defects in lipid bilayer play a significant role in the biological activities of cell membranes. Time-resolved fluorescence depolarization has been used to detect and characterize the onset of packing defects in binary mixtures of dilinoleoylphosphatidylethanolamine/1-palmitoyl-2- oleoylphosphatidylcholine (PE/PC). These PE/PC mixtures exhibit mesoscopic packing defect state (D), as well as one-dimensional lambellar liquid crystalline (L alpha) and two-dimensional inverted hexagonal (HII) ordered phases. Based on previous electron microscopic investigations, this D state is characterized by the presence of interlamellar attachments and precursors of HII phase between the lipid layers. Using a rotational diffusion model for rod-shaped fluorophore in a curved matrix, rotational dynamics parameters, second rank order parameter, localized wobbling diffusion, and curvature-dependent rotational diffusion constants of dipyenylhexatriene (DPH)-labeled PC (DPH-PC) in the host PE/PC matrix were recovered from the measured fluorescence depolarization decays of DPH fluorescence. At approximately 60% PE, abrupt increases in these rotational dynamics parameters were observed, reflecting the onset of packing defects in the host PE/PC matrix. We have demonstrated that rotational dynamics parameters are very sensitive in detecting the onset of curvature-associating packing defects in lipid membranes. In addition, the presence of the D state can be characterized by the enhanced wobbling diffusional motion and order packing of lipid molecules, and by the presence of localized curvatures in the lipid layers. PMID:8842226

Chen, S Y; Cheng, K H

1996-08-01

153

Non-contact characterization of bacteria by time-resolved fluorescence  

NASA Astrophysics Data System (ADS)

Accurate real-time methods for the detection of pathogenic microorganisms in the agri-food industry would represent an improvement over standard methods of analysis. We are currently developing a non-contact, scanning optical system for the detection of bacteria on meat surfaces based on fluorescence lifetime and intensity measurements. The system detects autofluorescent light emitted by the naturally occurring fluorophores in bacteria. Potential expected advantages of this system include accurate and efficient 2D real-time mapping of bacterial contamination of surfaces, and elimination of sample-to-sample cross-contamination. Furthermore, as the technique only requires minimal sample preparation and handling, the chemical properties of the specimen are preserved. This article presents the preliminary results obtained from a time-resolved fluorescence imaging system for the characterization of a non-pathogenic gram-negative bacteria, Pseudomonas fluorescens. Additionally we present a particular application of the system of interest to the agri-food industry, demonstrating its potential as a real-time macroscopic imaging system for mapping bacterial contamination on meat surfaces. Initial results indicate that the combination of fluorescence lifetime and intensity measurements provides a means for characterizing biological media and for detecting microorganisms on surfaces.

Bouchard, Alain; Frechette, Julie; Long, William F.; Vernon, Marcia; Cormier, Jean-Francois; Vallee, Real; Mafu, Akier A.; Lemay, Marie-Josee

2004-07-01

154

Applications of Phasors to In Vitro Time-Resolved Fluorescence Measurements  

PubMed Central

The phasor method of treating fluorescence lifetime data provides a facile and convenient approach to characterize lifetime heterogeneity and to detect the presence of excited state reactions, such as solvent relaxation and Förster Resonance Energy Transfer. The method utilizes a plot of M sin(?) versus M cos(?), where M is the modulation ratio and ? is the phase angle taken from frequency domain fluorometry. A principle advantage of the phasor method is that it provides a model-less approach to time-resolved data, amenable to visual inspection. Although the phasor approach has been recently applied to Fluorescence Lifetime Imaging Microscopy it has not been extensively utilized for cuvette studies. In the present study we explore the applications of the method to in vitro samples. The phasors of binary and ternary mixtures of fluorescent dyes demonstrates the utility of the method for investigating complex mixtures. Data from excited state reactions, such as dipolar relaxation in membrane and protein systems and also energy transfer from the tryptophan residue to the chromophore in EGFP, are also presented. PMID:21078290

Štefl, Martin; James, Nicholas G.; Ross, Justin A.; Jameson, David M.

2010-01-01

155

Noninvasive assessment of breast cancer risk using time-resolved diffuse optical spectroscopy.  

PubMed

Breast density is a recognized strong and independent risk factor for breast cancer. We propose the use of time-resolved transmittance spectroscopy to estimate breast tissue density and potentially provide even more direct information on breast cancer risk. Time-resolved optical mammography at seven wavelengths (635 to 1060 nm) is performed on 49 subjects. Average information on breast tissue of each subject is obtained on oxy- and deoxyhemoglobin, water, lipids, and collagen content, as well as scattering amplitude and power. All parameters, except for blood volume and oxygenation, correlate with mammographic breast density, even if not to the same extent. A synthetic optical index proves to be quite effective in separating different breast density categories. Finally, the estimate of collagen content as a more direct means for the assessment of breast cancer risk is discussed. PMID:21198142

Taroni, Paola; Pifferi, Antonio; Quarto, Giovanna; Spinelli, Lorenzo; Torricelli, Alessandro; Abbate, Francesca; Villa, Anna; Balestreri, Nicola; Menna, Simona; Cassano, Enrico; Cubeddu, Rinaldo

2010-01-01

156

Investigation of RNA Hairpin Loop Folding with Time-Resolved Infrared Spectroscopy  

NASA Astrophysics Data System (ADS)

Ribonucleic acids (RNAs) are a group of functional biopolymers central to the molecular underpinnings of life. To complete the many processes they mediate, RNAs must fold into precise three-dimensional structures. Hairpin loops are the most ubiquitous and basic structural elements present in all folded RNAs, and are the foundation upon which all complex tertiary structures are built. A hairpin loop forms when a single stranded RNA molecule folds back on itself creating a helical stem of paired bases capped by a loop. This work investigates the formation of UNCG hairpin loops with the sequence 5'-GC(UNCG)GC-3' (N = A, U, G, or C) using both equilibrium infrared (IR) and time-resolved IR spectroscopy. Equilibrium IR melting data were used to determine thermodynamic parameters. Melting temperatures ranged from 50 to 60°C, and enthalpies of unfolding were on the order of 100 kJ/mol. In the time-resolved work, temperature jumps of up to 20°C at 2.5°C increments were obtained with transient relaxation kinetics spanning nanoseconds to hundreds of microseconds. The relaxation kinetics for all of the oligomers studied were fit to first or second order exponentials. Multiple vibrational transitions were probed on each oligomer for fully folded and partially denatured structures. In the time-resolved limit, in contrast to equilibrium melting, RNA does not fold according to two-state behavior. These results are some of the first to show that RNA hairpins fold according to a rugged energy landscape, which contradicts their relatively simple nature. In addition, this work has proven that time-resolved IR spectroscopy is a powerful and novel tool for investigating the earliest events of RNA folding, the formation of the hairpin loop.

Stancik, Aaron Lee

157

Correlative Time-Resolved Fluorescence Microscopy To Assess Antibiotic Diffusion-Reaction in Biofilms  

PubMed Central

The failure of antibiotics to inactivate in vivo pathogens organized in biofilms has been shown to trigger chronic infections. In addition to mechanisms involving specific genetic or physiological cell properties, antibiotic sorption and/or reaction with biofilm components may lessen the antibiotic bioavailability and consequently decrease their efficiency. To assess locally and accurately the antibiotic diffusion-reaction, we used for the first time a set of advanced fluorescence microscopic tools (fluorescence recovery after photobleaching, fluorescence correlation spectroscopy, and fluorescence lifetime imaging) that offer a spatiotemporal resolution not available with the commonly used time-lapse confocal imaging method. This set of techniques was used to characterize the dynamics of fluorescently labeled vancomycin in biofilms formed by two Staphylococcus aureus human isolates. We demonstrate that, at therapeutic concentrations of vancomycin, the biofilm matrix was not an obstacle to the diffusion-reaction of the antibiotic that can reach all cells through the biostructure. PMID:22450986

Daddi Oubekka, S.; Briandet, R.; Fontaine-Aupart, M.-P.

2012-01-01

158

Time-resolved spectroscopy using a chopper wheel as a fast shutter  

NASA Astrophysics Data System (ADS)

Widely available, small form-factor, fiber-coupled spectrometers typically have a minimum exposure time measured in milliseconds, and thus cannot be used directly for time-resolved measurements at the microsecond level. Spectroscopy at these faster time scales is typically done with an intensified charge coupled device (CCD) system where the image intensifier acts as a "fast" electronic shutter for the slower CCD array. In this paper, we describe simple modifications to a commercially available chopper wheel system to allow it to be used as a "fast" mechanical shutter for gating a fiber-coupled spectrometer to achieve microsecond-scale time-resolved optical measurements of a periodically pulsed light source. With the chopper wheel synchronized to the pulsing of the light source, the time resolution can be set to a small fraction of the pulse period by using a chopper wheel with narrow slots separated by wide spokes. Different methods of synchronizing the chopper wheel and pulsing of the light sources are explored. The capability of the chopper wheel system is illustrated with time-resolved measurements of pulsed plasmas.

Wang, Shicong; Wendt, Amy E.; Boffard, John B.; Lin, Chun C.

2015-01-01

159

Tryptophan dynamics of the FK506 binding protein: time-resolved fluorescence and simulations.  

PubMed Central

The FK506-binding protein (FKBP12) is important in the immunosuppressant action of FK506 and rapamycin. We have investigated Trp side chain dynamics in FKBP12, with and without a bound immunosuppressant, by measuring the Trp time-resolved fluorescence anisotropy decay r(t). The r(t) for W59 in aqueous uncomplexed FKBP12 at 20 degrees C is well described by a single exponential with a recovered initial anisotropy, r(eff)o, of 0.192 and an overall rotational correlation time for the protein, phi p, of 4.7 ns; r(eff)o = 0.214 and phi p = 4.2 ns for the FKBP12/FK506 complex. Using an expression for the order parameter squared, namely S2 = r(eff)o/rTo, where rTo is the vitrified steady-state excitation anisotropy, we recovered an S2 of 0.75 for W59 fluorescence in uncomplexed FKBP12 and S2 approximately equal to 1 in the FKBP12/FK506 complex. Results obtained for the FKBP12/rapamycin complex are similar to those found for the FKBP12/FK506 complex. Minimum perturbation mapping simulations were performed on the free and complexed forms of FKBP12 and the results were generally in agreement with the experimental data. Images FIGURE 5 FIGURE 6 PMID:8785272

Silva, N D; Prendergast, F G

1996-01-01

160

The use of time-resolved fluorescence in gel-based proteomics for improved biomarker discovery  

NASA Astrophysics Data System (ADS)

This paper describes a new platform for quantitative intact proteomics, entitled Cumulative Time-resolved Emission 2-Dimensional Gel Electrophoresis (CuTEDGE). The CuTEDGE technology utilizes differences in fluorescent lifetimes to subtract the confounding background fluorescence during in-gel detection and quantification of proteins, resulting in a drastic improvement in both sensitivity and dynamic range compared to existing technology. The platform is primarily designed for image acquisition in 2-dimensional gel electrophoresis (2-DE), but is also applicable to 1-dimensional gel electrophoresis (1-DE), and proteins electroblotted to membranes. In a set of proof-of-principle measurements, we have evaluated the performance of the novel technology using the MicroTime 100 instrument (PicoQuant GmbH) in conjunction with the CyDye minimal labeling fluorochromes (GE Healthcare, Uppsala, Sweden) to perform differential gel electrophoresis (DIGE) analyses. The results indicate that the CuTEDGE technology provides an improvement in the dynamic range and sensitivity of detection of 3 orders of magnitude as compared to current state-of-the-art image acquisition instrumentation available for 2-DE (Typhoon 9410, GE Healthcare). Given the potential dynamic range of 7-8 orders of magnitude and sensitivities in the attomol range, the described invention represents a technological leap in detection of low abundance cellular proteins, which is desperately needed in the field of biomarker discovery.

Sandberg, AnnSofi; Buschmann, Volker; Kapusta, Peter; Erdmann, Rainer; Wheelock, Åsa M.

2010-02-01

161

The singlet-oxygen-sensitized delayed fluorescence in mammalian cells: a time-resolved microscopy approach.  

PubMed

The present work provides a proof-of-concept that the singlet oxygen-sensitized delayed fluorescence (SOSDF) can be detected from individual living mammalian cells in a time-resolved microscopy experiment. To this end, 3T3 mouse fibroblasts incubated with 100 ?M TPPS4 or TMPyP were used and the microsecond kinetics of the delayed fluorescence (DF) were recorded. The analysis revealed that SOSDF is the major component of the overall DF signal. The microscopy approach enables precise control of experimental conditions - the DF kinetics are clearly influenced by the presence of the (1)O2 quencher (sodium azide), H2O/D2O exchange, and the oxygen concentration. Analysis of SOSDF kinetics, which was reconstructed as a difference DF kinetics between the unquenched and the NaN3-quenched samples, provides a cellular (1)O2 lifetime of ?? = 1-2 ?s and a TPPS4 triplet lifetime of ?T = 22 ± 5 ?s in agreement with previously published values. The short SOSDF acquisition times, typically in the range of tens of seconds, enable us to study the dynamic cellular processes. It is shown that SOSDF lifetimes increase during PDT-like treatment, which may provide valuable information about changes of the intracellular microenvironment. SOSDF is proposed and evaluated as an alternative tool for (1)O2 detection in biological systems. PMID:25591544

Scholz, Marek; Biehl, Anna-Louisa; D?dic, Roman; Hála, Jan

2015-04-01

162

Nanosecond time-resolved circular polarization of fluorescence: study of NADH bound to horse liver alcohol dehydrogenase.  

PubMed Central

Circularly polarized luminescence (CPL) spectroscopy provides information on the excited-state chirality of a lumiphore analogous but complementary to information regarding the ground-state chirality derived from circular dichroism. The sensitivity of CPL spectra to molecular conformation makes this technique uniquely suited for the study of biomolecular structure, as extensively demonstrated in earlier studies. Unfortunately, the CPL spectra of many biomolecules often contain significantly overlapping contributions from emitting species either because multiple lumiphores are present (e.g., tryptophan residues in a protein) or because multiple conformations of the biomolecule simultaneously exist, each with a unique CPL spectrum. Increased resolution between individual contributions to the CPL may be achieved by time-resolving this signal, thus taking advantage of the fact that, as a rule, each of the emitting species also has a characteristic decay time associated with its electronically excited state. In addition, the time resolution provides information regarding dynamics associated with the different chiral states of the system. The present study describes an instrument for the determination of time-resolved CPL (TR-CPL) with subnanosecond resolution and its application to several chiral systems. The technique was first demonstrated on a model system with a strong time-dependent CPL signal. Subsequently, the circularly polarized component in the fluorescence of reduced nicotinamide adenine dinucleotide (NADH) bound to liver alcohol dehydrogenase was time-resolved. The CPL of NADH in the binary enzyme-coenzyme complex is time-dependent, reflecting structural differences around the reduced nicotinamide possibly due to a dynamic restructuring. In contrast, the CPL of the coenzyme in the ternary complex formed with enzyme and the substrate analog isobutyramide is essentially time-independent, likely reflecting a more rigid binding domain. Since the linear polarization of the fluorescence of the two complexes did not show any local flexibility of the NADH chromophore, the excited-state conformational rearrangement of the binary complex indicates a subtle change in its interactions with group(s) in direct contact with it. PMID:7831331

Schauerte, J A; Schlyer, B D; Steel, D G; Gafni, A

1995-01-01

163

Polarization and time-resolved photoluminescence spectroscopy of excitons in MoSe2 monolayers  

NASA Astrophysics Data System (ADS)

We investigate valley exciton dynamics in MoSe2 monolayers in polarization- and time-resolved photoluminescence (PL) spectroscopy at 4 K. Following circularly polarized laser excitation, we record a low circular polarization degree of the PL of typically ?5%. This is about 10 times lower than the polarization induced under comparable conditions in MoS2 and WSe2 monolayers. The evolution of the exciton polarization as a function of excitation laser energy and power is monitored in PL excitation experiments. Fast PL emission times are recorded for both the neutral exciton of ?3 ps and for the charged exciton (trion) of 12 ps.

Wang, G.; Palleau, E.; Amand, T.; Tongay, S.; Marie, X.; Urbaszek, B.

2015-03-01

164

Time-resolved photoelectron spectroscopy of coupled electron-nuclear motion  

SciTech Connect

We investigate pump-probe electron detachment spectroscopy in a model system which is ideally suited to study coupled electronic and nuclear wave-packet dynamics. Time-resolved photoelectron spectra are calculated within the adiabatic approximation and a discretization of the detachment continuum. These spectra are compared to those which derive from a non-Born-Oppenheimer description and a numerically exact treatment of the detachment process. In this way it is possible to identify the influence of non-adiabatic effects on the spectra in a systematic way and also to test commonly applied approximations.

Falge, Mirjam; Engel, Volker [Institut fuer Physikalische und Theoretische Chemie and Roentgen Research Center for Complex Material Systems, Universitaet Wuerzburg, Am Hubland, 97074 Wuerzburg (Germany); Graefe, Stefanie [Institute for Theoretical Physics, Vienna University of Technology, Wiedner Hauptstr. 8-10, A-1040 Vienna (Austria)

2011-05-14

165

Time-resolved spectroscopy of plasma resonances in highly excited silicon and germanium  

SciTech Connect

The dynamics of the electron-hole plasma in silicon and germanium samples irradiated by 20 ps. 532 nm laser pulses has been investigated in the near infrared by the time-resolved picosecond optical spectroscopy. The experimental reflectivities and transmission are compared with the predictions of the thermal model for degenerate carrier distributions through the Drude formalism. Above a certain fluence, a significant deviation between measured and calculated values indicates a strong increase of the recombination rate as soon as the plasma resonances become comparable with the band gaps. These new plasmon-aided recombination channels are particularly pronounced in germanium. 15 refs., 8 figs.

Huang, C.Y.; Malvezzi, A.M.; Bloembergen, N.; Kurz, H.

1985-01-01

166

A CAMAC system controlled by an IBM AT computer for time-resolved spectroscopy  

SciTech Connect

An IBM AT computer interfaced to a small CAMAC system offers considerable power without the complexity and expense of a large general-purpose system. Our system for time-resolved spectroscopy features menu-driven FORTRAN-based software; high-resolution and high-speed (8K channels, 5-..mu..s fixed dead time) ADCs; segmentable histogram memories (24-bit counts) with large memory space for many histogram segments; independently variable separate histogram dwell times; remote control via a CAMAC serial highway; and ground isolation between the data acquisition equipment and control computer by means of fiber optics.

Lindquist, L.O.; Moss, C.E.

1987-01-01

167

A versatile and reconfigurable setup for all-terahertz time-resolved pump-probe spectroscopy.  

PubMed

A versatile optical setup for all-terahertz (THz) time resolved pump-probe spectroscopy was designed and tested. By utilizing a dual THz pulse generator emitter module, independent and synchronized THz radiation pump and probe pulses were produced, thus eliminating the need for THz beam splitters and the limitations associated with their implementation. The current THz setup allows for precise control of the electric fields splitting ratio between the THz radiation pump and probe pulses, as well as in-phase, out-of-phase, and polarization dependent pump-probe spectroscopy. Since the present THz pump-probe setup does not require specialized THz radiation optical components, such as phase shifters, polarization rotators, or wide bandwidth beam splitters, it can be easily implemented with minimal alterations to a conventional THz time domain spectroscopy system. The present setup is valuable for studying the time dynamics of THz coherent phenomena in solid-state, chemical, and biological systems. PMID:22667602

Elezzabi, A Y; Maraghechi, P

2012-05-01

168

Time-resolved spectroscopy of the Mercury 6 3P1 state  

NASA Technical Reports Server (NTRS)

The time-resolved fluorescence was observed from the Hg 6 3P1 state under the influence of the earth's magnetic field and with applied fields of up to 14 G. Modulation of the fluorescence decay signal was observed as a function of both time and space and can be interpreted in terms of a classical precession of the excited atom about the magnetic field or as quantum beats resulting from interference between coherently populated Zeeman sublevels. This modulation was studied for each of the five resolvable components of the hyperfine structure separately. The fluorescence from the even isotopes was determined to be almost completely modulated while the fluorescence from the odd isotopes was only partially modulated. The frequency of modulation of the fluorescence from the mercury-202 isotope was observed as a function of the applied magnetic field and a value for the Lande factor of 1.46 + or - 0.03 was obtained. This is within experimental error of the accepted value of 1.486. In addition, the frequency of modulation as a function of applied magnetic field was determined for each of the three resolvable components with more than one contributing isotopic hyperfine line. An investigation of the effect of radiation trapping on the degree modulation was also made.

Halstead, J. A.; Reeves, R. R.

1981-01-01

169

Time-resolved fluorescence microscopy to study biologically related applications using sol-gel derived and cellular media  

NASA Astrophysics Data System (ADS)

Fluorescence microscopy provides a non-invasive means for visualising dynamic protein interactions. As well as allowing the calculation of kinetic processes via the use of time-resolved fluorescence, localisation of the protein within cells or model systems can be monitored. These fluorescence lifetime images (FLIM) have become the preferred technique for elucidating protein dynamics due to the fact that the fluorescence lifetime is an absolute measure, in the main independent of fluorophore concentration and intensity fluctuations caused by factors such as photobleaching. In this work we demonstrate the use of a time-resolved fluorescence microscopy, employing a high repetition rate laser excitation source applied to study the influence of a metal surface on fluorescence tagged protein and to elucidate viscosity using the fluorescence lifetime probe DASPMI. These were studied in a cellular environment (yeast) and in a model system based on a sol-gel derived material, in which silver nanostructures were formed in situ using irradiation from a semiconductor laser in CW mode incorporated on a compact time-resolved fluorescence microscope (HORIBA Scientific DeltaDiode and DynaMyc).

Toury, Marion; Chandler, Lin; Allison, Archie; Campbell, David; McLoskey, David; Holmes-Smith, A. Sheila; Hungerford, Graham

2011-03-01

170

Time-Resolved Photoluminescence Spectroscopy and Imaging: New Approaches to the Analysis of Cultural Heritage and Its Degradation  

PubMed Central

Applications of time-resolved photoluminescence spectroscopy (TRPL) and fluorescence lifetime imaging (FLIM) to the analysis of cultural heritage are presented. Examples range from historic wall paintings and stone sculptures to 20th century iconic design objects. A detailed description of the instrumentation developed and employed for analysis in the laboratory or in situ is given. Both instruments rely on a pulsed laser source coupled to a gated detection system, but differ in the type of information they provide. Applications of FLIM to the analysis of model samples and for the in-situ monitoring of works of art range from the analysis of organic materials and pigments in wall paintings, the detection of trace organic substances on stone sculptures, to the mapping of luminescence in late 19th century paintings. TRPL and FLIM are employed as sensors for the detection of the degradation of design objects made in plastic. Applications and avenues for future research are suggested. PMID:24699285

Nevin, Austin; Cesaratto, Anna; Bellei, Sara; D'Andrea, Cosimo; Toniolo, Lucia; Valentini, Gianluca; Comelli, Daniela

2014-01-01

171

Time-resolved photoluminescence spectroscopy and imaging: new approaches to the analysis of cultural heritage and its degradation.  

PubMed

Applications of time-resolved photoluminescence spectroscopy (TRPL) and fluorescence lifetime imaging (FLIM) to the analysis of cultural heritage are presented. Examples range from historic wall paintings and stone sculptures to 20th century iconic design objects. A detailed description of the instrumentation developed and employed for analysis in the laboratory or in situ is given. Both instruments rely on a pulsed laser source coupled to a gated detection system, but differ in the type of information they provide. Applications of FLIM to the analysis of model samples and for the in-situ monitoring of works of art range from the analysis of organic materials and pigments in wall paintings, the detection of trace organic substances on stone sculptures, to the mapping of luminescence in late 19th century paintings. TRPL and FLIM are employed as sensors for the detection of the degradation of design objects made in plastic. Applications and avenues for future research are suggested. PMID:24699285

Nevin, Austin; Cesaratto, Anna; Bellei, Sara; D'Andrea, Cosimo; Toniolo, Lucia; Valentini, Gianluca; Comelli, Daniela

2014-01-01

172

Monitoring cellular stress responses using integrated high-frequency impedance spectroscopy and time-resolved ELISA.  

PubMed

We have developed a lab-on-a-chip system for continuous and non-invasive monitoring of microfluidic cell cultures using integrated high-frequency contactless impedance spectroscopy. Electrically insulated microfabricated interdigitated electrode structures were embedded into four individually addressable microchambers to reliably and reproducibly detect cell-substrate interactions, cell viability and metabolic activity. While silicon nitride passivated sensor substrates provided a homogeneous cell culture surface that minimized cell orientation along interdigitated electrode structures, the application of high-frequency AC fields reduced the impact of the 300 nm thick passivation layer on sensor sensitivity. The additional implementation of multivariate data analysis methods such as partial least square (PLS) for high-frequency impedance spectra provided unambiguous information on intracellular pathway activation, up and down-regulation of protein synthesis as well as global cellular stress responses. A comparative cell analysis using connective tissue fibroblasts showed that high-frequency contactless impedance spectroscopy and time-resolved quantification of IL-6 secretion using ELISA provided similar results following stimulation with circulating pro-inflammatory cytokines IL-1? and TNF?. The combination of microfluidics with contactless impedance sensing and time-resolved quantification of stress factor release will provide biologist with a new tool to (a) establish a variety of uniform cell culture surfaces that feature complex biochemistries, micro- and nanopatterns; and (b) to simultaneously characterize cell responses under physiologically relevant conditions using a complementary non-invasive cell analysis method. PMID:25137192

Charwat, Verena; Joksch, Martin; Sticker, Drago; Purtscher, Michaela; Rothbauer, Mario; Ertl, Peter

2014-10-21

173

Investigation of Prolactin Receptor Activation and Blockade Using Time-Resolved Fluorescence Resonance Energy Transfer  

PubMed Central

The prolactin receptor (PRLR) is emerging as a therapeutic target in oncology. Knowledge-based drug design led to the development of a pure PRLR antagonist (Del1-9-G129R-hPRL) that was recently shown to prevent PRL-induced mouse prostate tumorogenesis. In humans, the first gain-of-function mutation of the PRLR (PRLRI146L) was recently identified in breast tumor patients. At the molecular level, the actual mechanism of action of these two novel players in the PRL system remains elusive. In this study, we addressed whether constitutive PRLR activation (PRLRI146L) or PRLR blockade (antagonist) involved alteration of receptor oligomerization and/or of inter-chain distances compared to unstimulated and PRL-stimulated PRLR. Using a combination of various biochemical and spectroscopic approaches (co-IP, blue native electrophoresis, BRET1), we demonstrated that preformed PRLR homodimers are altered neither by PRL- or I146L-induced receptor triggering, nor by antagonist-mediated blockade. These findings were confirmed using a novel time-resolved fluorescence resonance energy transfer (TR-FRET) technology that allows monitoring distance changes between cell surface tagged receptors. This technology revealed that PRLR blockade or activation did not involve detectable distance changes between extracellular domains of receptor chains within the dimer. This study merges with our previous structural investigations suggesting that the mechanism of PRLR activation solely involves intermolecular contact adaptations leading to subtle intramolecular rearrangements. PMID:22649370

Tallet, Estelle; Fernandez, Isabelle; Zhang, Chi; Salsac, Marion; Gregor, Nathalie; Ayoub, Mohammed Akli; Pin, Jean Philippe; Trinquet, Eric; Goffin, Vincent

2011-01-01

174

Poly(?-amino ester) - DNA complexes: time-resolved fluorescence and cellular transfection studies  

PubMed Central

A large number of different polymers have been developed and studied for application as DNA carriers for non-viral gene delivery, but the DNA binding properties are not understood. This study describes the efficiency of nanoparticle formation by time-resolved fluorescence measurements for poly(?-amino esters), cationic biodegradable polymers with DNA complexation and transfection capability. From the large library of poly(?-amino esters) ten polymers with different transfection efficacies were chosen for this study. The binding constants for nanoparticle formation were determined and compared to polyethylene imines with the same method. Although the DNA binding efficiency of the amine groups are similar for both types of polymers, the overall binding constants are an order of magnitude smaller for poly(?-amino esters) than for 25 kDa polyethylenimines, but yet poly(?-amino esters) show comparable DNA transfection efficacy with polyethyleneimines. Within this series of polymers the transfection efficacy showed increasing trend in association with relative efficiency of nanoparticle formation. PMID:21699928

Vuorimaa, Elina; Ketola, Tiia-Maaria; Green, Jordan J.; Hanzlíková, Martina; Lemmetyinen, Helge; Langer, Robert; Anderson, Daniel G.; Urtti, Arto; Yliperttula, Marjo

2011-01-01

175

Validation and evaluation of a novel time-resolved laser-induced fluorescence technique  

NASA Astrophysics Data System (ADS)

We present a novel technique to measure time-resolved laser-induced fluorescence signals in plasma sources that have a relatively constant Fourier spectrum of oscillations in steady-state operation, but are not periodically pulsed, e.g., Hall thrusters. The technique uses laser modulation of the order of MHz and recovers signal via a combination of band-pass filtering, phase-sensitive detection, and averaging over estimated transfer functions calculated for many different cycles of the oscillation. Periodic discharge current oscillations were imposed on a hollow cathode. Measurements were validated by comparison with independent measurements from a lock-in amplifier and by comparing the results of the transfer function average to an independent analysis technique triggering averaging over many oscillation cycles in the time domain. The performance of the new technique is analyzed and compared to prior techniques, and it is shown that this new technique has a niche in measurements where the analog photomultiplier signal has a nonwhite noise spectral density and cycles of oscillation are not sufficiently repeatable to allow for reliable triggering or a meaningful average waveform in the time domain.

Durot, C. J.; Gallimore, A. D.; Smith, T. B.

2014-01-01

176

Applications of immunomagnetic capture and time-resolved fluorescence detection for Salmonella enteriditis in liquid eggs  

NASA Astrophysics Data System (ADS)

An immuno sandwich method was evaluated for the detection of Salmonella in liquid eggs. Liquid eggs spiked with different out-break strains of Salmonella were mixed with proper enrichment media and incubated at 37 C for 4 to 20 h. After enrichment, immunomagnetic beads (IMB) coated with anti Salmonella antibodies were used to capture the bacteria. Samarium (Sm) labeled anti Salmonella antibodies were then used to form sandwiched complexes with IMB captured bacteria. Sandwiched Salmonella were then treated with Sm-chelator to allow the measurement of the released Sm by time-resolved fluorescence (TRF). The processes ranging from IMB capture to Sm chelation were performed using an automated KingFisher apparatus. With this approach, the presence of ~ 1 CFU of outbreak strains of Salmonella Enteritidis per egg (~50 g of liquid eggs) could be detected after enrichment for 20 h at 37 C. For higher levels of Salmonella Enteritidis contamination, e.g., 10 CFU per 50 g of liquid eggs, the enrichment time could be reduced to 5 h at 37 C. The results demonstrated that a combination of IMB capture and TRF measurement could be a rapid and sensitive method for Salmonella Enteritidis detection in liquid eggs.

Tu, Shu-I.; Gehring, Andrew; Paoli, George

2008-04-01

177

Validation and evaluation of a novel time-resolved laser-induced fluorescence technique.  

PubMed

We present a novel technique to measure time-resolved laser-induced fluorescence signals in plasma sources that have a relatively constant Fourier spectrum of oscillations in steady-state operation, but are not periodically pulsed, e.g., Hall thrusters. The technique uses laser modulation of the order of MHz and recovers signal via a combination of band-pass filtering, phase-sensitive detection, and averaging over estimated transfer functions calculated for many different cycles of the oscillation. Periodic discharge current oscillations were imposed on a hollow cathode. Measurements were validated by comparison with independent measurements from a lock-in amplifier and by comparing the results of the transfer function average to an independent analysis technique triggering averaging over many oscillation cycles in the time domain. The performance of the new technique is analyzed and compared to prior techniques, and it is shown that this new technique has a niche in measurements where the analog photomultiplier signal has a nonwhite noise spectral density and cycles of oscillation are not sufficiently repeatable to allow for reliable triggering or a meaningful average waveform in the time domain. PMID:24517766

Durot, C J; Gallimore, A D; Smith, T B

2014-01-01

178

Time-resolved detection of fluorescent light during inflow of ICG to the brain—a methodological study  

NASA Astrophysics Data System (ADS)

It was reported that time-resolved reflectance measurements carried out during inflow and washout of an optical contrast agent may provide information on the blood supply to the brain cortex of human adults. It was also shown that a measurement of fluorescence excited in the dye circulating in the brain is feasible. Unfortunately, patterns of time-resolved fluorescence signals observed during in vivo measurements are difficult to interpret. The aim of this study was to analyze the influence of several factors on the fluorescence signals measured during in vivo experiments. A laboratory instrument for recording the distributions of arrival of fluorescence photons was constructed and optimized for measurements on humans. Monte Carlo simulations and laboratory measurements on liquid phantoms as well as in vivo measurements on healthy volunteers were carried out. An influence of source-detector separation, position of the source-detector pair on the head, as well as a dose of the injected indocyanine green (ICG) on the fluorescence signals were studied in detail. It was shown that even for a small dose of ICG (0.025 mg kg-1) the time-resolved signals can be successfully detected on the surface of the head. Strong influence of the studied factors on the fluorescence signals was observed. It was also noted that the changes in moments of distributions of arrival times of fluorescence photons depend on the anatomical structure of the tissues located between the source and the detector.

Milej, Daniel; Gerega, Anna; ?o?ek, Norbert; Weigl, Wojciech; Kacprzak, Micha?; Sawosz, Piotr; M?czewska, Joanna; Fronczewska, Katarzyna; Mayzner-Zawadzka, Ewa; Królicki, Leszek; Maniewski, Roman; Liebert, Adam

2012-10-01

179

Picosecond time-resolved absorption and fluorescence dynamics in the artificial bacteriorhodopsin pigment BR6.11.  

PubMed Central

The picosecond molecular dynamics in an artificial bacteriorhodopsin (BR) pigment containing a structurally modified all-trans retinal chromphore with a six-membered ring bridging the C11=C12-C13 positions (BR6.11) are measured by picosecond transient absorption and picosecond time-resolved fluorescence spectroscopy. Time-dependent intensity and spectral changes in absorption in the 570-650-nm region are monitored for delays as long as 5 ns after the 7-ps, 573-nm excitation of BR6.11. Two intermediates, J6.11 and K6.11/1, both with enhanced absorption to the red (> 600 nm) of the BR6.11 spectrum are observed within approximately 50 ps. The J6.11 intermediate decays with a time constant of 12 +/- 3 ps to form K6.11/1. The K6.11/1 intermediate decays with an approximately 100-ps time constant to form a third intermediate, K6.11/2, which is observed through diminished 650-nm absorption (relative to that of K6.11/1). No other transient absorption changes are found during the remainder of the initial 5-ns period of the BR6.11 photoreaction. Fluorescence in the 650-900-nm region is observed from BR6.11, K6.11/1, and K6.11/2, but no emission assignable to J6.11 is found. The BR6.11 fluroescence spectrum has a approximately 725-nm maximum which is blue-shifted by approximately 15 nm relative to that of native BR-570 and is 4.2 +/- 1.5 times larger in intensity (same sample optical density). No differences in the profile of the fluorescence spectra of BR6.11 and the intermediates K6.11/1 and K6.11/2 are observed. Following ground-state depletion of the BR6.11 population, the time-resolved fluroescence intensity monitored at 725 nm increases with two time constants, 12 +/- 3 and approximately 100 ps, both of which correlate well with changes in the picosecond transient absorption data. The resonance Raman spectrum of ground-state BR6.11, measured with low-energy, 560-nm excitation, is significantly different from the spectrum of native BR-570, thus confirming that the picosecond transient absorption and picosecond time resolved fluorescence data are assignable to BR6.11 and its photoreaction alone and not to BR-570 reformed during there constitution process (<5% of the BR6.11 sample could be attributed to native BR-570).The J6.11 and K6.11 absorption and fluorescence data presented here are generally analogous to those measured for native J-625 and K-590, respectively, and therefore, the primary events in the BR6.11 photoreaction can be correlated with those in the native BR photocycle. The BR6.11 photoreaction, however, exhibits important differences including slower formation rates for J and K intermediates as well as the presence of a second K intermediate. These results demonstrate that the restricted motion in the C11=C12-C13 region of retinal found in BR6.11 does not greatly change the overall photoreaction mechanism,but does alter the rates at which processes occur. PMID:8218919

Brack, T. L.; Delaney, J. K.; Atkinson, G. H.; Albeck, A.; Sheves, M.; Ottolenghi, M.

1993-01-01

180

Hyperspectral time-resolved wide-field fluorescence molecular tomography based on structured light and single-pixel detection.  

PubMed

We present a time-resolved fluorescence diffuse optical tomography platform that is based on wide-field structured illumination, single-pixel detection, and hyperspectral acquisition. Two spatial light modulators (digital micro-mirror devices) are employed to generate independently wide-field illumination and detection patterns, coupled with a 16-channel spectrophotometer detection module to capture hyperspectral time-resolved tomographic data sets. The main system characteristics are reported, and we demonstrate the feasibility of acquiring dense 4D tomographic data sets (space, time, spectra) for time domain 3D quantitative multiplexed fluorophore concentration mapping in turbid media. PMID:25680065

Pian, Qi; Yao, Ruoyang; Zhao, Lingling; Intes, Xavier

2015-02-01

181

Highly sensitive detection of human papillomavirus type 16 DNA using time-resolved fluorescence microscopy and long lifetime probes  

NASA Astrophysics Data System (ADS)

We have been interested in the role of Human Papillomavirus (HPV) in cervical cancer and its diagnosis; to that end we have been developing microscopic imaging and fluorescent in situ hybridization (FISH) techniques to genotype and quantitate the amount of HPV present at a single cell level in cervical PAP smears. However, we have found that low levels of HPV DNA are difficult to detect accurately because theoretically obtainable sensitivity is never achieved due to nonspecific autofluorescence, fixative induced fluorescence of cells and tissues, and autofluorescence of the optical components in the microscopic system. In addition, the absorption stains used for PAP smears are intensely autofluorescent. Autofluorescence is a rapidly decaying process with lifetimes in the range of 1-100 nsec, whereas phosphorescence and delayed fluorescence have lifetimes in the range of 1 microsecond(s) ec-10 msec. The ability to discriminate between specific fluorescence and autofluorescence in the time-domain has improved the sensitivity of diagnostic test such that they perform comparably to, or even more sensitive than radioisotopic assays. We have developed a novel time-resolved fluorescence microscope to improve the sensitivity of detection of specific molecules of interest in slide based specimens. This time-resolved fluorescence microscope is based on our recently developed fluorescence lifetime imaging microscopy (FILM) in conjunction with the use of long lifetime fluorescent labels. By using fluorescence in situ hybridization and the long lifetime probe (europium), we have demonstrated the utility of this technique for detection of HPV DNA in cervicovaginal cells. Our results indicate that the use of time-resolved fluorescence microscopy and long lifetime probes increases the sensitivity of detection by removing autofluorescence and will thus lead to improved early diagnosis of cervical cancer. Since the highly sensitive detection of DNA in clinical samples using fluorescence in situ hybridization image is useful for the diagnosis of many other type of diseases, the system we have developed should find numerous applications for the diagnosis of disease states.

Wang, Xue F.; Periasamy, Ammasi; Wodnicki, Pawel; Siadat-Pajouh, M.; Herman, Brian

1995-04-01

182

Monitoring the Folding Kinetics of a ?-Hairpin by Time-Resolved IR Spectroscopy in Silico.  

PubMed

Protein folding is one of the most fundamental problems in modern biochemistry. Time-resolved infrared (IR) spectroscopy in the amide I region is commonly used to monitor folding kinetics. However, associated atomic detail information on the folding mechanism requires simulations. In atomistic simulations structural order parameters are typically used to follow the folding process along the simulated trajectories. However, a rigorous test of the reliability of the mechanisms found in the simulations requires calculation of the time-dependent experimental observable, i.e., in the present case the IR signal in the amide I region. Here, we combine molecular dynamics simulation with a mixed quantum mechanics/molecular mechanics theoretical methodology, the Perturbed Matrix Method, in order to characterize the folding of a ?-hairpin peptide, through modeling the time-dependence of the amide I IR signal. The kinetic and thermodynamic data (folding and unfolding rate constants, and equilibrium folded- and unfolded-state probabilities) obtained from the fit of the calculated signal are in good agreement with the available experimental data [Xu et al. J. Am. Chem. Soc. 2003, 125, 15388-15394]. To the best of our knowledge, this is the first report of the simulation of the time-resolved IR signal of a complex process occurring on a long (microsecond) time scale. PMID:25777154

Daidone, Isabella; Thukral, Lipi; Smith, Jeremy C; Amadei, Andrea

2015-04-01

183

Microcontroller based resonance tracking unit for time resolved continuous wave cavity-ringdown spectroscopy measurements  

NASA Astrophysics Data System (ADS)

We present in this work a new tracking servoloop electronics for continuous wave cavity-ringdown absorption spectroscopy (cw-CRDS) and its application to time resolved cw-CRDS measurements by coupling the system with a pulsed laser photolysis set-up. The tracking unit significantly increases the repetition rate of the CRDS events and thus improves effective time resolution (and/or the signal-to-noise ratio) in kinetics studies with cw-CRDS in given data acquisition time. The tracking servoloop uses novel strategy to track the cavity resonances that result in a fast relocking (few ms) after the loss of tracking due to an external disturbance. The microcontroller based design is highly flexible and thus advanced tracking strategies are easy to implement by the firmware modification without the need to modify the hardware. We believe that the performance of many existing cw-CRDS experiments, not only time-resolved, can be improved with such tracking unit without any additional modification to the experiment.

Votava, Ondrej; Mašát, Milan; Parker, Alexander E.; Jain, Chaithania; Fittschen, Christa

2012-04-01

184

Modelling time-resolved two-dimensional electronic spectroscopy of the primary photoisomerization event in rhodopsin.  

PubMed

Time-resolved two-dimensional (2D) electronic spectra (ES) tracking the evolution of the excited state manifolds of the retinal chromophore have been simulated along the photoisomerization pathway in bovine rhodopsin, using a state-of-the-art hybrid QM/MM approach based on multiconfigurational methods. Simulations of broadband 2D spectra provide a useful picture of the overall detectable 2D signals from the near-infrared (NIR) to the near-ultraviolet (UV). Evolution of the stimulated emission (SE) and excited state absorption (ESA) 2D signals indicates that the S1 ? SN (with N ? 2) ESAs feature a substantial blue-shift only after bond inversion and partial rotation along the cis ? trans isomerization angle, while the SE rapidly red-shifts during the photoinduced skeletal relaxation of the polyene chain. Different combinations of pulse frequencies are proposed in order to follow the evolution of specific ESA signals. These include a two-color 2DVis/NIR setup especially suited for tracking the evolution of the S1 ? S2 transitions that can be used to discriminate between different photochemical mechanisms of retinal photoisomerization as a function of the environment. The reported results are consistent with the available time-resolved pump-probe experimental data, and may be used for the design of more elaborate transient 2D electronic spectroscopy techniques. PMID:24794143

Rivalta, Ivan; Nenov, Artur; Weingart, Oliver; Cerullo, Giulio; Garavelli, Marco; Mukamel, Shaul

2014-07-17

185

Discovery of novel aromatase inhibitors using a homogeneous time-resolved fluorescence assay  

PubMed Central

Aim: Aromatase is an important target for drugs to treat hormone-dependent diseases, including breast cancer. The aim of this study was to develop a homogeneous time-resolved fluorescence (HTRF) aromatase assay suitable for high-throughput screening (HTS). Methods: A 384-well aromatase HTRF assay was established, and used to screen about 7000 compounds from a compound library. Anti-proliferation activity of the hit was evaluated using alamarBlue(R) assay in a hormone-dependent breast cancer cell line T47D. Molecular docking was conducted to elucidate the binding mode of the hit using the Discovery Studio program. Results: The Z? value and signal to background (S/B) ratio were 0.74 and 5.4, respectively. Among the 7000 compounds, 4 hits (XHN22, XHN26, XHN27 and triptoquinone A) were found to inhibit aromatase with IC50 values of 1.60±0.07, 2.76±0.24, 0.81±0.08 and 45.8±11.3 ?mol /L, respectively. The hits XHN22, XHN26 and XHN27 shared the same chemical scaffold of 4-imidazolyl quinoline. Moreover, the most potent hit XHN27 at 10 and 50 ?mol/L inhibited the proliferation of T47D cells by 45.3% and 35.2%, respectively. The docking study revealed that XHN27 docked within the active site of aromatase and might form a hydrogen bond and had a ?-cation interaction with amino acid residues of the protein. Conclusion: XHN27, an imidazolyl quinoline derivative of flavonoid, is a potent aromatase inhibitor with anti-proliferation activity against breast cancer in vitro. The established assay can be used in HTS for discovering novel aromatase inhibitor. PMID:25047514

Ji, Jin-zi; Lao, Ke-jing; Hu, Jie; Pang, Tao; Jiang, Zhen-zhou; Yuan, Hao-liang; Miao, Jing-shan; Chen, Xin; Ning, Shan-shan; Xiang, Hua; Guo, Yu-meng; Yan, Ming; Zhang, Lu-yong

2014-01-01

186

Development of a broadband picosecond infrared spectrometer and its incorporation into an existing ultrafast time-resolved resonance Raman, UV/visible, and fluorescence spectroscopic apparatus.  

PubMed

We have constructed a broadband ultrafast time-resolved infrared (TRIR) spectrometer and incorporated it into our existing time-resolved spectroscopy apparatus, thus creating a single instrument capable of performing the complementary techniques of femto-/picosecond time-resolved resonance Raman (TR3), fluorescence, and UV/visible/infrared transient absorption spectroscopy. The TRIR spectrometer employs broadband (150 fs, approximately 150 cm(-1) FWHM) mid-infrared probe and reference pulses (generated by difference frequency mixing of near-infrared pulses in type I AgGaS2), which are dispersed over two 64-element linear infrared array detectors (HgCdTe). These are coupled via custom-built data acquisition electronics to a personal computer for data processing. This data acquisition system performs signal handling on a shot-by-shot basis at the 1 kHz repetition rate of the pulsed laser system. The combination of real-time signal processing and the ability to normalize each probe and reference pulse has enabled us to achieve a high sensitivity on the order of deltaOD approximately 10(-4) - 10(-5) with 1 min of acquisition time. We present preliminary picosecond TRIR studies using this spectrometer and also demonstrate how a combination of TRIR and TR3 spectroscopy can provide key information for the full elucidation of a photochemical process. PMID:14658632

Towrie, Michael; Grills, David C; Dyer, Joanne; Weinstein, Julia A; Matousek, Pavel; Barton, Robin; Bailey, Philip D; Subramaniam, Naresh; Kwok, Wai M; Ma, Chensheng; Phillips, David; Parker, Anthony W; George, Michael W

2003-04-01

187

Label-Free Toxin Detection by Means of Time-Resolved Electrochemical Impedance Spectroscopy  

PubMed Central

The real-time detection of trace concentrations of biological toxins requires significant improvement of the detection methods from those reported in the literature. To develop a highly sensitive and selective detection device it is necessary to determine the optimal measuring conditions for the electrochemical sensor in three domains: time, frequency and polarization potential. In this work we utilized a time-resolved electrochemical impedance spectroscopy for the detection of trace concentrations of Staphylococcus enterotoxin B (SEB). An anti-SEB antibody has been attached to the nano-porous aluminum surface using 3-aminopropyltriethoxysilane/glutaraldehyde coupling system. This immobilization method allows fabrication of a highly reproducible and stable sensing device. Using developed immobilization procedure and optimized detection regime, it is possible to determine the presence of SEB at the levels as low as 10 pg/mL in 15 minutes. PMID:22315560

Chai, Changhoon; Takhistov, Paul

2010-01-01

188

Time-resolved picosecond spectroscopy of the resonant secondary radiation of F centers in KCl  

NASA Astrophysics Data System (ADS)

The linear polarization (P HL) of hot luminescence (HL) composing of the resonant secondary radiation of the F centers has been measured using a time-resolved picosecond spectroscopy over the whole Stokes wavenumber ? range. The P HL holds constant value of about 40% until the onset of ordinary luminescence (OL), from where it decreases to vanishingly small with decrease of ? This implies that the optically excited F center relaxes down along the 2p-like adiabatic potential energy surface (APES) trough, and transits to the 2s-like APES trough to form the relaxed excited state (RES). The lattice relaxation time and the dynamical transition time are ultra fast estimated to be less than 15 psec.

Akiyama, N.; Nakahara, F.; Ohkura, H.

1995-12-01

189

Time resolved tunable diode laser absoption spectroscopy of dual High Power Impulse Magnetron Sputtering discharges  

NASA Astrophysics Data System (ADS)

Time-resolved measurements have been performed during dual High Power Impulse Magnetron Sputtering (dual-HiPIMS) with two cathodes in a closed magnetic field configuration. The dual-HiPIMS system, operated at a repetition frequency f = 100 Hz and duty cycle of 1 %, was equipped with two different metallic targets (Ti, Cu). The effect of a delay between subsequent pulses on argon excited atom density and temperature was investigated by means of tunable diode laser absorption spectroscopy. It is shown that the peak densities of pulses vary strongly with the delay. We observed an enhancement of metastable density due to pre-ionization effect but more effective than that is the contribution of metal atoms which have smaller ionization energy compare to that of buffer gas atom. Associate with the enhancement of density, the temporal variation of metastable atom temperature in the Cu pulse also transforms from those of low current pulse into the high current one.

Do, Hoang Tung; Stranak, Vitezslav; Hippler, Rainer

2014-08-01

190

Time-resolved broadband cavity-enhanced absorption spectroscopy for chemical kinetics.  

SciTech Connect

Experimental measurements of elementary reaction rate coefficients and product branching ratios are essential to our understanding of many fundamentally important processes in Combustion Chemistry. However, such measurements are often impossible because of a lack of adequate detection techniques. Some of the largest gaps in our knowledge concern some of the most important radical species, because their short lifetimes and low steady-state concentrations make them particularly difficult to detect. To address this challenge, we propose a novel general detection method for gas-phase chemical kinetics: time-resolved broadband cavity-enhanced absorption spectroscopy (TR-BB-CEAS). This all-optical, non-intrusive, multiplexed method enables sensitive direct probing of transient reaction intermediates in a simple, inexpensive, and robust experimental package.

Sheps, Leonid; Chandler, David W.

2013-04-01

191

Characterization of energetic and thermalized sputtered atoms in pulsed plasma using time-resolved tunable diode-laser induced fluorescence  

NASA Astrophysics Data System (ADS)

In this work, a time-resolved tunable diode-laser (DL) induced fluorescence (TR-TDLIF) method calibrated by absorption spectroscopy has been developed in order to determine atom and flux velocity distribution functions (AVDF and FVDF) of the energetic and the thermalized atoms in pulsed plasmas. The experimental set-up includes a low-frequency (˜3 Hz) and high spectral-resolution DL (˜0.005 pm), a fast rise-time pulse generator, and a high power impulse magnetron sputtering (HiPIMS) system. The induced TR-TDLIF signal is recorded every 0.5 ?s with a digital oscilloscope of a second-long trace. The technique is illustrated with determining the AVDF and the FVDF of a metastable state of the sputtered neutral tungsten atoms in the HiPIMS post-discharge. Gaussian functions describing the population of the four W isotopes were used to fit the measured TR-TDLIF signal. These distribution functions provide insight into transition from the energetic to thermalized regimes from the discharge onset. This technique may be extended with appropriate DLs to probe any species with rapidly changing AVDF and FVDF in pulsed and strongly oscillating plasmas.

Desecures, M.; de Poucques, L.; Easwarakhanthan, T.; Bougdira, J.

2014-11-01

192

Time-Resolved Spectroscopy and Near Infrared Imaging for Prostate Cancer Detection: Receptor-targeted and Native Biomarker  

NASA Astrophysics Data System (ADS)

Optical spectroscopy and imaging using near-infrared (NIR) light provides powerful tools for non-invasive detection of cancer in tissue. Optical techniques are capable of quantitative reconstructions maps of tissue absorption and scattering properties, thus can map in vivo the differences in the content of certain marker chromophores and/or fluorophores in normal and cancerous tissues (for example: water, tryptophan, collagen and NADH contents). Potential clinical applications of optical spectroscopy and imaging include functional tumor detection and photothermal therapeutics. Optical spectroscopy and imaging apply contrasts from intrinsic tissue chromophores such as water, collagen and NADH, and extrinsic optical contrast agents such as Indocyanine Green (ICG) to distinguish disease tissue from the normal one. Fluorescence spectroscopy and imaging also gives high sensitivity and specificity for biomedical diagnosis. Recent developments on specific-targeting fluorophores such as small receptor-targeted dye-peptide conjugate contrast agent offer high contrast between normal and cancerous tissues hence provide promising future for early tumour detection. This thesis focus on a study to distinguish the cancerous prostate tissue from the normal prostate tissues with enhancement of specific receptor-targeted prostate cancer contrast agents using optical spectroscopy and imaging techniques. The scattering and absorption coefficients, and anisotropy factor of cancerous and normal prostate tissues were investigated first as the basis for the biomedical diagnostic and optical imaging. Understanding the receptors over-expressed prostate cancer cells and molecular target mechanism of ligand, two small ICG-derivative dye-peptides, namely Cypate-Bombesin Peptide Analogue Conjugate (Cybesin) and Cypate-Octreotate Peptide Conjugate (Cytate), were applied to study their clinical potential for human prostate cancer detection. In this work, the steady-state and time-resolved fluorescence spectroscopy of Cybesin (Cytate) in solution, and in cancerous and normal prostate tissues were studied. It was found that more Cybesin (Cytate) was uptaken in the cancerous prostate tissue than those in the normal tissue. The preferential uptake of Cybesin (Cytate) in cancerous tissue was used to image and distinguish cancerous areas from the normal tissue. To investigate rotational dynamics and fluorescence polarization anisotropy of the contrast agents in prostate tissues, an analytical model was used to extract the rotational times and polarization anisotropies, which were observed for higher values of Cybesin (Cytate)-stained cancerous prostate tissue in comparison with the normal tissue. These reflect changes of microstructures of cancerous and normal tissues and their different binding affinity with contrast agents. The results indicate that the use of optical spectroscopy and imaging combined with receptor-targeted contrast agents is a valuable tool to study microenvironmental changes of tissue, and detect prostate cancer in early stage.

Pu, Yang

193

Multiwavelength time-resolved detection of fluorescence during the inflow of indocyanine green into the adult's brain  

NASA Astrophysics Data System (ADS)

Optical technique based on diffuse reflectance measurement combined with indocyanine green (ICG) bolus tracking is extensively tested as a method for clinical assessment of brain perfusion in adults at the bedside. Methodology of multiwavelength and time-resolved detection of fluorescence light excited in the ICG is presented and advantages of measurements at multiple wavelengths are discussed. Measurements were carried out: 1. on a physical homogeneous phantom to study the concentration dependence of the fluorescence signal, 2. on the phantom to simulate the dynamic inflow of ICG at different depths, and 3. in vivo on surface of the human head. Pattern of inflow and washout of ICG in the head of healthy volunteers after intravenous injection of the dye was observed for the first time with time-resolved instrumentation at multiple emission wavelengths. The multiwavelength detection of fluorescence signal confirms that at longer emission wavelengths, probability of reabsorption of the fluorescence light by the dye itself is reduced. Considering different light penetration depths at different wavelengths, and the pronounced reabsorption at longer wavelengths, the time-resolved multiwavelength technique may be useful in signal decomposition, leading to evaluation of extra- and intracerebral components of the measured signals.

Gerega, Anna; Milej, Daniel; Weigl, Wojciech; Botwicz, Marcin; Zolek, Norbert; Kacprzak, Michal; Wierzejski, Wojciech; Toczylowska, Beata; Mayzner-Zawadzka, Ewa; Maniewski, Roman; Liebert, Adam

2012-08-01

194

BL1.4.2: Time-resolved FTIR spectroscopy at up to 5 nanosecond resolution demonstrated  

E-print Network

BL1.4.2: Time-resolved FTIR spectroscopy at up to 5 nanosecond resolution demonstrated Michael C Transform Infrared Spectroscopy (FTIR) acquires scans on the time scale of one second, so the very fast electronics of the Bruker IFS66v/S FTIR spectrometer on BL1.4.2, we can measure IR spectra with a time

195

Lanthanide labeling of a potent protease activated receptor-2 agonist for time-resolved fluorescence analysis  

PubMed Central

Protease activated receptor-2 (PAR2) is one of four G-protein coupled receptors (GPCRs) that can be activated by exogenous or endogenous proteases, which cleave the extracellular amino-terminus to expose a tethered ligand and subsequent G-protein signaling. Alternatively, PAR2 can be activated by peptide or peptidomimetic ligands derived from the sequence of the natural tethered ligand. Screening of novel ligands that directly bind to PAR2 to agonize or antagonize the receptor has been hindered by the lack of a sensitive, high-throughput, affinity binding assay. In this report we describe the synthesis and use of a modified PAR2 peptidomimetic agonist, 2-furoyl-LIGRLO-(diethylenetriaminepentaacetic acid)-NH2 (2-f-LIGRLO-dtpa), designed for lanthanide-based time resolved fluorescence screening. We first demonstrate that 2-f-LIGRLO-dtpa is a potent and specific PAR2 agonist across a full spectrum of in vitro assays. We then show that 2-f-LIGRLO-dtpa can be utilized in an affinity binding assay to evaluate the ligand-receptor interactions between known high potency peptidomimetic agonists (2-furoyl-LIGRLO-NH2, 2-f-LIGRLO; 2-aminothiazol-4-yl-LIGRL-NH2, 2-at-LIGRL and; 6-aminonicotinyl-LIGRL-NH2, 6-an-LIGRL) and PAR2. A separate N-terminal peptidomimetic modification (3-indoleacetyl-LIGRL-NH2, 3-ia-LIGRL) that does not activate PAR2 signaling was used as a negative control. All three peptidomimetic agonists demonstrated sigmoidal competitive binding curves, with the more potent agonists (2-f-LIGRLO and 2-at-LIGRL) displaying increased competition. In contrast, the control peptide (3-ia-LIGRL) displayed limited competition for PAR2 binding. In summary, we have developed a Europium-containing PAR2 agonist that can be used in a highly sensitive affinity binding assay to screen novel PAR2 ligands in a high-throughput format. This ligand can serve as a critical tool in the screening and development of PAR2 ligands. PMID:22994402

Hoffman, Justin; Flynn, Andrea N.; Tillu, Dipti V.; Zhang, Zhenyu; Patek, Renata; Price, Theodore J.; Vagner, Josef; Boitano, Scott

2012-01-01

196

A Vinblastine Fluorescent Probe for Pregnane X Receptor in a Time-Resolved Fluorescence Resonance Energy Transfer Assay  

PubMed Central

The pregnane X receptor (PXR) regulates the metabolism and excretion of xenobiotics and endobiotics by regulating the expression of drug-metabolizing enzymes and transporters. The unique structure of PXR allows the binding of many drugs and drug leads to it, possibly causing undesired drug-drug interactions. Therefore, it is crucial to evaluate whether lead compounds bind to PXR. Fluorescence-based assays are preferred because of their sensitivity and non-radioactive nature. One fluorescent PXR probe is currently commercially available; however, because its chemical structure is not publicly disclosed, it is not optimal for studying ligand-PXR interactions. Here we report the characterization of BODIPY FL Vinblastine, generated by labeling vinblastine with the fluorophore 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene (BODIPY FL), as a high-affinity ligand for human PXR with a Kd value of 673 nM. We provide evidence that BODIPY FL Vinblastine is a unique chemical entity different from either vinblastine or the fluorophore 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene in its function as a high-affinity human PXR ligand. We describe a BODIPY FL Vinblastine-based human PXR Time-Resolved Fluorescence Resonance Energy Transfer assay, which was used to successfully test a panel of human PXR ligands. The BODIPY FL Vinblastine–based biochemical assay is suitable for high-throughput screening to evaluate whether lead compounds bind to PXR. PMID:24044991

Lin, Wenwei; Chen, Taosheng

2013-01-01

197

Cyclohexene Photo-oxidation over Vanadia Catalyst Analyzed by Time Resolved ATR-FT-IR Spectroscopy  

SciTech Connect

Vanadia was incorporated in the 3-dimensional mesoporous material TUD-1 with a loading of 2percent w/w vanadia. The performance in the selective photo-oxidation of liquid cyclohexene was investigated using ATR-FT-IR spectroscopy. Under continuous illumination at 458 nm a significant amount of product, i.e. cyclohexenone, was identified. This demonstrates for the first time that hydroxylated vanadia centers in mesoporous materials can be activated by visible light to induce oxidation reactions. Using the rapid scan method, a strong perturbation of the vanadyl environment could be observed in the selective oxidation process induced by a 458 nm laser pulse of 480 ms duration. This is proposed to be caused by interaction of the catalytic centre with a cyclohexenyl hydroperoxide intermediate. The restoration of the vanadyl environment could be kinetically correlated to the rate of formation of cyclohexenone, and is explained by molecular rearrangement and dissociation of the peroxide to ketone and water. The ketone diffuses away from the active center and ATR infrared probing zone, resulting in a decreasing ketone signal on the tens of seconds time scale after initiation of the photoreaction. This study demonstrates the high potential of time resolved ATR FT-IR spectroscopy for mechanistic studies of liquid phase reactions by monitoring not only intermediates and products, but by correlating the temporal behavior of these species to molecular changes of the vanadyl catalytic site.

Frei, Heinz; Mul, Guido; Wasylenko, Walter; Hamdy, M. Sameh; Frei, Heinz

2008-06-04

198

Time-resolved tuned diode laser absorption spectroscopy of pulsed plasma  

NASA Astrophysics Data System (ADS)

A novel method for time-resolved tuned diode laser absorption spectroscopy has been developed. In this paper, we describe in detail developed electronic module that controls time-resolution of laser absorption spectroscopy system. The TTL signal triggering plasma pulse is used for generation of two signals: the first one triggers the fine tuning of laser wavelength and second one controls time-defined signal sampling from absorption detector. The described method and electronic system enable us to investigate temporal evolution of sputtered particles in technological low-temperature plasma systems. The pulsed DC planar magnetron sputtering system has been used to verify this method. The 2" in diameter titanium target was sputtered in pure argon atmosphere. The working pressure was held at 2 Pa. All the experiments were carried out for pulse ON time fixed at 100 (is. When changing OFF time the discharge has operated between High Power Impulse Magnetron Sputtering regime and pulsed DC magnetron regime. The effect of duty cycle variation results in decrease of titanium atom density during ON time while length of OFF time elongates. We believe that observed effect is connected with higher degree of ionization of sputtered particles. As previously reported by Bohlmark et al., the measured optical emission spectra in HiPIMS systems were dominated by emission from titanium ions [1].

Adámek, P.; Do, H. T.; ?ada, M.; Hubi?ka, Z.; Hippler, R.

2014-05-01

199

Development of a time-resolved fluorometric method for observing hybridization in living cells using fluorescence resonance energy transfer.  

PubMed Central

We previously showed that a specific kind of mRNA (c-fos) was detected in a living cell under a microscope by introducing two fluorescently labeled oligodeoxynucleotides, each labeled with donor or acceptor, into the cytoplasm, making them hybridize to adjacent locations on c-fos mRNA, and taking images of fluorescence resonance energy transfer (FRET) (A. Tsuji, H. Koshimoto, Y. Sato, M. Hirano. Y. Sei-Iida, S. Kondo, and K. Ishibashi, 2000, Biophys. J. 78:3260-3274). On the formed hybrid, the distance between donor and acceptor becomes close and FRET occurs. To observe small numbers of mRNA in living cells using this method, it is required that FRET fluorescence of hybrid must be distinguished from fluorescence of excess amounts of non-hybridizing probes and from cell autofluorescence. To meet these requirements, we developed a time-resolved method using acceptor fluorescence decays. When a combination of a donor having longer fluorescence lifetime and an acceptor having shorter lifetime is used, the measured fluorescence decays of acceptors under FRET becomes slower than the acceptor fluorescence decay with direct excitation. A combination of Bodipy493/503 and Cy5 was selected as donor and acceptor. When the formed hybrid had a configuration where the target RNA has no single-strand part between the two fluorophores, the acceptor fluorescence of hybrid had a sufficiently longer delay to detect fluorescence of hybrid in the presence of excess amounts of non-hybridizing probes. Spatial separation of 10-12 bases between two fluorophores on the hybrid is also required. The decay is also much slower than cell autofluorescence, and smaller numbers of hybrid were detected with less interference of cell autofluorescence in the cytoplasm of living cells under a time-resolved fluorescence microscope with a time-gated function equipped camera. The present method will be useful when observing induced expressions of mRNA in living cells. PMID:11423432

Tsuji, A; Sato, Y; Hirano, M; Suga, T; Koshimoto, H; Taguchi, T; Ohsuka, S

2001-01-01

200

A field programmable gate array-based time-resolved scaler for collinear laser spectroscopy with bunched radioactive potassium beams.  

PubMed

A new data acquisition system including a Field Programmable Gate Array (FPGA) based time-resolved scaler was developed for laser-induced fluorescence and beam bunch coincidence measurements. The FPGA scaler was tested in a collinear laser-spectroscopy experiment on radioactive (37)K at the BEam COoler and LAser spectroscopy (BECOLA) facility at the National Superconducting Cyclotron Laboratory at Michigan State University. A 1.29 ?s bunch width from the buncher and a bunch repetition rate of 2.5 Hz led to a background suppression factor of 3.1 × 10(5) in resonant photon detection measurements. The hyperfine structure of (37)K and its isotope shift relative to the stable (39)K were determined using 5 × 10(4) s(-1) (37)K ions injected into the BECOLA beam line. The obtained hyperfine coupling constants A((2)S(1/2)) = 120.3(1.4) MHz, A((2)P(1/2)) = 15.2(1.1) MHz, and A((2)P(3/2)) = 1.4(8) MHz, and the isotope shift ??(39, 37) = -264(3) MHz are consistent with the previously determined values, where available. PMID:25273722

Rossi, D M; Minamisono, K; Barquest, B R; Bollen, G; Cooper, K; Davis, M; Hammerton, K; Hughes, M; Mantica, P F; Morrissey, D J; Ringle, R; Rodriguez, J A; Ryder, C A; Schwarz, S; Strum, R; Sumithrarachchi, C; Tarazona, D; Zhao, S

2014-09-01

201

A field programmable gate array-based time-resolved scaler for collinear laser spectroscopy with bunched radioactive potassium beams  

SciTech Connect

A new data acquisition system including a Field Programmable Gate Array (FPGA) based time-resolved scaler was developed for laser-induced fluorescence and beam bunch coincidence measurements. The FPGA scaler was tested in a collinear laser-spectroscopy experiment on radioactive {sup 37}K at the BEam COoler and LAser spectroscopy (BECOLA) facility at the National Superconducting Cyclotron Laboratory at Michigan State University. A 1.29 ?s bunch width from the buncher and a bunch repetition rate of 2.5 Hz led to a background suppression factor of 3.1 × 10{sup 5} in resonant photon detection measurements. The hyperfine structure of {sup 37}K and its isotope shift relative to the stable {sup 39}K were determined using 5 × 10{sup 4} s{sup ?1} {sup 37}K ions injected into the BECOLA beam line. The obtained hyperfine coupling constants A({sup 2}S{sub 1/2}) = 120.3(1.4) MHz, A({sup 2}P{sub 1/2}) = 15.2(1.1) MHz, and A({sup 2}P{sub 3/2}) = 1.4(8) MHz, and the isotope shift ??{sup 39,} {sup 37} = ?264(3) MHz are consistent with the previously determined values, where available.

Rossi, D. M., E-mail: rossi@nscl.msu.edu; Davis, M.; Ringle, R.; Rodriguez, J. A.; Ryder, C. A.; Schwarz, S.; Sumithrarachchi, C.; Zhao, S. [National Superconducting Cyclotron Laboratory, Michigan State University, East Lansing, Michigan 48824 (United States); Minamisono, K., E-mail: minamiso@nscl.msu.edu; Barquest, B. R.; Bollen, G.; Hughes, M.; Strum, R.; Tarazona, D. [National Superconducting Cyclotron Laboratory, Michigan State University, East Lansing, Michigan 48824 (United States); Department of Physics and Astronomy, Michigan State University, East Lansing, Michigan 48824 (United States); Cooper, K.; Hammerton, K.; Mantica, P. F.; Morrissey, D. J. [National Superconducting Cyclotron Laboratory, Michigan State University, East Lansing, Michigan 48824 (United States); Department of Chemistry, Michigan State University, East Lansing, Michigan 48824 (United States)

2014-09-15

202

A field programmable gate array-based time-resolved scaler for collinear laser spectroscopy with bunched radioactive potassium beams  

NASA Astrophysics Data System (ADS)

A new data acquisition system including a Field Programmable Gate Array (FPGA) based time-resolved scaler was developed for laser-induced fluorescence and beam bunch coincidence measurements. The FPGA scaler was tested in a collinear laser-spectroscopy experiment on radioactive 37K at the BEam COoler and LAser spectroscopy (BECOLA) facility at the National Superconducting Cyclotron Laboratory at Michigan State University. A 1.29 ?s bunch width from the buncher and a bunch repetition rate of 2.5 Hz led to a background suppression factor of 3.1 × 105 in resonant photon detection measurements. The hyperfine structure of 37K and its isotope shift relative to the stable 39K were determined using 5 × 104 s-1 37K ions injected into the BECOLA beam line. The obtained hyperfine coupling constants A(2S1/2) = 120.3(1.4) MHz, A(2P1/2) = 15.2(1.1) MHz, and A(2P3/2) = 1.4(8) MHz, and the isotope shift ??39, 37 = -264(3) MHz are consistent with the previously determined values, where available.

Rossi, D. M.; Minamisono, K.; Barquest, B. R.; Bollen, G.; Cooper, K.; Davis, M.; Hammerton, K.; Hughes, M.; Mantica, P. F.; Morrissey, D. J.; Ringle, R.; Rodriguez, J. A.; Ryder, C. A.; Schwarz, S.; Strum, R.; Sumithrarachchi, C.; Tarazona, D.; Zhao, S.

2014-09-01

203

Time-resolved pulsed-plasma characterization using broadband terahertz pulses correlated with fluorescence imaging  

E-print Network

Time-resolved pulsed-plasma characterization using broadband terahertz pulses correlated -bandwidth electromagnetic pulses have been used to probe pulsed-discharge argon plasmas at various pressures Lorentz model to the measured time-domain pulse with no plasma and comparing it to the measured pulse

Yoo, S. J. Ben

204

Time-resolved laser-induced fluorescence diagnostics in a HIPIMS discharge  

NASA Astrophysics Data System (ADS)

Laser-induced fluorescence (LIF) combined with resonant optical absorption spectroscopy (ROAS) were utilized for characterization of the Ar-Ti high-power impulse sputtering (HIPIMS) discharge at working pressures of 5 and 20 mTorr. The LIF measurements were performed during the plasma off-time. It was shown that the time-behavior of the measured densities depends strongly on the working pressure in the reactor as well as on the distance from the magnetron target. ROAS measurements indicates that the absolute ground state level density of Ti can reach ? 1011 cm-3 (at 20 mTorr, 10 ?s pulse, and 12 kW of applied power per pulse). Generally, the obtained results indicate highly non-uniform spatial and temporal behavior of the sputtered species such as Ti and Ti+, which correlate with the results previously obtained in the other magnetron discharges.

Britun, Nikolay; Palmucci, Maria; Konstantinidis, Stephanos; Snyders, Rony

2012-09-01

205

The fluorescence resonance energy transfer (FRET) gate: A time-resolved study  

PubMed Central

The two-step energy-transfer process in a self-assembled complex comprising a cationic conjugated polymer (CCP) and a dsDNA is investigated by using pump-dump-emission spectroscopy and time-correlated single-photon counting; energy is transferred from the CCP to an ethidium bromide (EB) molecule intercalated into the dsDNA through a fluorescein molecule linked to one terminus of the DNA. Time-dependent anisotropy measurements indicate that the inefficient direct energy transfer from the CCP to the intercalated EB results from the near orthogonality of their transition moments. These measurements also show that the transition moment of the fluorescein spans a range of angular distributions and lies between that of the CCP and EB. Consequently, the fluorescein acts as a fluorescence resonance energy-transfer gate to relay the excitation energy from the CCP to the EB. PMID:15642946

Xu, Qing-Hua; Wang, Shu; Korystov, Dmitry; Mikhailovsky, Alexander; Bazan, Guillermo C.; Moses, Daniel; Heeger, Alan J.

2005-01-01

206

In vivo time-resolved spectroscopy of the human bronchial early cancer autofluorescence  

NASA Astrophysics Data System (ADS)

Time-resolved measurements of tissue autofluorescence (AF) excited at 405 nm were carried out with an optical-fiber-based spectrometer in the bronchi of 11 patients. The objectives consisted of assessing the lifetime as a new tumor/normal (T/N) tissue contrast parameter and trying to explain the origin of the contrasts observed when using AF-based cancer detection imaging systems. No significant change in the AF lifetimes was found. AF bronchoscopy performed in parallel with an imaging device revealed both intensity and spectral contrasts. Our results suggest that the spectral contrast might be due to an enhanced blood concentration just below the epithelial layers of the lesion. The intensity contrast probably results from the thickening of the epithelium in the lesions. The absence of T/N lifetime contrast indicates that the quenching is not at the origin of the fluorescence intensity and spectral contrasts. These lifetimes (6.9 ns, 2.0 ns, and 0.2 ns) were consistent for all the examined sites. The fact that these lifetimes are the same for different emission domains ranging between 430 and 680 nm indicates that there is probably only one dominant fluorophore involved. The measured lifetimes suggest that this fluorophore is elastin.

Uehlinger, Pascal; Gabrecht, Tanja; Glanzmann, Thomas; Ballini, Jean-Pierre; Radu, Alexandre; Andrejevic, Snezana; Monnier, Philippe; Wagnières, Georges

2009-03-01

207

Detection of fecal contamination on apples with nanosecond-scale time-resolved imaging of laser-induced fluorescence  

NASA Astrophysics Data System (ADS)

Detection of apples contaminated with feces is a public health concern. We found that time-resolved imaging of apples artificially contaminated with feces allowed optimization of timing parameters for detection. Dairy feces were applied to Red Delicious and Golden Delicious apples. Laser-induced fluorescence responses were imaged by use of a gated intensified camera. We developed algorithms to automatically detect contamination iteratively by using one half of the apples and validated them by applying the optimized algorithms to the remaining apples. Results show that consideration of the timing of fluorescence responses to pulsed-laser excitation can enhance detection of feces on apples.

Lefcourt, Alan M.; Kim, Moon S.; Chen, Yud-Ren

2005-03-01

208

[Photodissociation of Acetylene and Acetone using Step-Scan Time-Resolved FTIR Emission Spectroscopy  

NASA Technical Reports Server (NTRS)

The photodissociation of acetylene and acetone was investigated as a function of added quenching gas pressures using step-scan time-resolved FTIR emission spectroscopy. Its main components consist of Bruker IFS88, step-scan Fourier Transform Infrared (FTIR) spectrometer coupled to a flow cell equipped with Welsh collection optics. Vibrationally excited C2H radicals were produced from the photodissociation of acetylene in the unfocused experiments. The infrared (IR) emission from these excited C2H radicals was investigated as a function of added argon pressure. Argon quenching rate constants for all C2H emission bands are of the order of 10(exp -13)cc/molecule.sec. Quenching of these radicals by acetylene is efficient, with a rate constant in the range of 10(exp -11) cc/molecule.sec. The relative intensity of the different C2H emission bands did not change with the increasing argon or acetylene pressure. However, the overall IR emission intensity decreased, for example, by more than 50% when the argon partial pressure was raised from 0.2 to 2 Torr at fixed precursor pressure of 160mTorr. These observations provide evidence for the formation of a metastable C2H2 species, which are collisionally quenched by argon or acetylene. Problems encountered in the course of the experimental work are also described.

McLaren, Ian A.; Wrobel, Jacek D.

1997-01-01

209

Hexamethylcyclopentadiene: time-resolved photoelectron spectroscopy and ab initio multiple spawning simulations.  

PubMed

Progress in our understanding of ultrafast light-induced processes in molecules is best achieved through a close combination of experimental and theoretical approaches. Direct comparison is obtained if theory is able to directly reproduce experimental observables. Here, we present a joint approach comparing time-resolved photoelectron spectroscopy (TRPES) with ab initio multiple spawning (AIMS) simulations on the MS-MR-CASPT2 level of theory. We disentangle the relationship between two phenomena that dominate the immediate molecular response upon light absorption: a spectrally dependent delay of the photoelectron signal and an induction time prior to excited state depopulation in dynamics simulations. As a benchmark molecule, we have chosen hexamethylcyclopentadiene, which shows an unprecedentedly large spectral delay of (310 ± 20) fs in TRPES experiments. For the dynamics simulations, methyl groups were replaced by "hydrogen atoms" having mass 15 and TRPES spectra were calculated. These showed an induction time of (108 ± 10) fs which could directly be assigned to progress along a torsional mode leading to the intersection seam with the molecular ground state. In a stepladder-type approach, the close connection between the two phenomena could be elucidated, allowing for a comparison with other polyenes and supporting the general validity of this finding for their excited state dynamics. Thus, the combination of TRPES and AIMS proves to be a powerful tool for a thorough understanding of ultrafast excited state dynamics in polyenes. PMID:24817114

Wolf, T J A; Kuhlman, T S; Schalk, O; Martínez, T J; Møller, K B; Stolow, A; Unterreiner, A-N

2014-06-21

210

A compact time-resolved system for near infrared spectroscopy based on wavelength space multiplexing  

NASA Astrophysics Data System (ADS)

We designed and developed a compact dual-wavelength and dual-channel time-resolved system for near-infrared spectroscopy studies of muscle and brain. The system employs pulsed diode lasers as sources, compact photomultipliers, and time-correlated single photon counting boards for detection. To exploit the full temporal and dynamic range of the acquisition technique, we implemented an approach based on wavelength space multiplexing: laser pulses at the two wavelengths are alternatively injected into the two channels by means of an optical 2×2 switch. In each detection line (i.e., in each temporal window), the distribution of photon time-of-flights at one wavelength is acquired. The proposed approach increases the signal-to-noise ratio and avoids wavelength cross-talk with respect to the typical approach based on time multiplexing. The instrument was characterized on tissue phantoms to assess its properties in terms of linearity, stability, noise, and reproducibility. Finally, it was successfully tested in preliminary in vivo measurements on muscle during standard cuff occlusion and on the brain during a motor cortex response due to hand movements.

Re, Rebecca; Contini, Davide; Caffini, Matteo; Cubeddu, Rinaldo; Spinelli, Lorenzo; Torricelli, Alessandro

2010-11-01

211

Ultrafast time-resolved spectroscopy of In2O3 nanowires  

NASA Astrophysics Data System (ADS)

Ultrafast carrier dynamics in In2O3 nanowires with an average diameter of ?100±20 nm grown by the vapor-liquid-solid method have been investigated in detail using differential absorption spectroscopy with femtosecond resolution. Measurements revealed that state filling is important for states above the band gap and states just below the band edge, thus demonstrating the critical role that shallow traps play in the relaxation of the photogenerated carriers. Furthermore, time-resolved intensity measurements revealed the importance of Auger recombination in the relaxation of carriers in the In2O3 nanowires and provided the maximum fluence (˜3 ?J/cm2) where this recombination mechanism may be considered negligible. Transient measurements in this low-fluence regime for carriers above the band gap revealed single exponential recovery (˜1.5 ns) associated with recombination of the photogenerated carriers. Similar behavior has been observed for the photogenerated carriers distributed within the shallow traps just below the band edge. Furthermore, measurements at longer probing wavelengths provided an estimate of the nonradiative relaxation of carriers (˜300 ps), which are distributed among the midgap states. Finally, long-lived oscillations in the transient reflection were detected, which corresponds to the presence of longitudinal acoustic phonons in the In2O3 nanowires.

Tsokkou, Demetra; Othonos, Andreas; Zervos, Matthew

2009-10-01

212

Optical analysis of cirrhotic liver by near infrared time resolved spectroscopy  

NASA Astrophysics Data System (ADS)

The severity of liver cirrhosis was related with the optical properties of liver tissue. Various grades of liver cirrhosis were produced in rats by intraperitoneal injection of thioacetamide (TAA) for different periods: 4 weeks, 8 weeks, 12 weeks, and 16 weeks. Optical properties of the liver, absorption, coefficient ((mu) a) and scattering coefficient (microsecond(s) '), were measured by near-infrared time- resolved spectroscopy. Histological examination confirmed cirrhotic changes in the liver, which were more severe in rats with TAA administration for longer periods. The (mu) a increased in 4- and 8-week rats, and then decreased in 12- and 16-week rats. The (mu) a of blood-free liver decreased as liver cirrhosis progressed. The hemoglobin content in the liver calculated from the (mu) a values increased in 4- and 8-week rats and decreased in 12- and 16-week rats. The microsecond(s) ' decreased in the cirrhotic liver, probably reflecting the decrease in the mitochondria content. It was shown that (mu) a and microsecond(s) ' determination is useful to assess the severity of liver cirrhosis.

Nishio, Toshihiro; Kitai, Toshiyuki; Miwa, Mitsuharu; Takahashi, Rei; Yamaoka, Yoshio

1999-10-01

213

Monitoring brain temperature by time-resolved near-infrared spectroscopy: pilot study.  

PubMed

Mild hypothermia (HT(32°C-33°C)) is an effective neuroprotective strategy for a variety of acute brain injuries. However, the wide clinical adaptation of HT(32-33°C) has been hampered by the lack of a reliable noninvasive method for measuring brain temperature, since core measurements have been shown to not always reflect brain temperature. The goal of this work was to develop a noninvasive optical technique for measuring brain temperature that exploits both the temperature dependency of water absorption and the high concentration of water in brain (80%-90%). Specifically, we demonstrate the potential of time-resolved near-infrared spectroscopy (TR-NIRS) to measure temperature in tissue-mimicking phantoms (in vitro) and deep brain tissue (in vivo) during heating and cooling, respectively. For deep brain tissue temperature monitoring, experiments were conducted on newborn piglets wherein hypothermia was induced by gradual whole body cooling. Brain temperature was concomitantly measured by TR-NIRS and a thermocouple probe implanted in the brain. Our proposed TR-NIRS method was able to measure the temperature of tissue-mimicking phantoms and brain tissues with a correlation of 0.82 and 0.66 to temperature measured with a thermometer, respectively. The mean difference between the TR-NIRS and thermometer measurements was 0.15°C ± 1.1°C for the in vitro experiments and 0.5°C ± 1.6°C for the in vivo measurements. PMID:24817622

Bakhsheshi, Mohammad Fazel; Diop, Mamadou; St Lawrence, Keith; Lee, Ting-Yim

2014-05-01

214

Monitoring brain temperature by time-resolved near-infrared spectroscopy: pilot study  

NASA Astrophysics Data System (ADS)

Mild hypothermia (HT) is an effective neuroprotective strategy for a variety of acute brain injuries. However, the wide clinical adaptation of HT has been hampered by the lack of a reliable noninvasive method for measuring brain temperature, since core measurements have been shown to not always reflect brain temperature. The goal of this work was to develop a noninvasive optical technique for measuring brain temperature that exploits both the temperature dependency of water absorption and the high concentration of water in brain (80%-90%). Specifically, we demonstrate the potential of time-resolved near-infrared spectroscopy (TR-NIRS) to measure temperature in tissue-mimicking phantoms (in vitro) and deep brain tissue (in vivo) during heating and cooling, respectively. For deep brain tissue temperature monitoring, experiments were conducted on newborn piglets wherein hypothermia was induced by gradual whole body cooling. Brain temperature was concomitantly measured by TR-NIRS and a thermocouple probe implanted in the brain. Our proposed TR-NIRS method was able to measure the temperature of tissue-mimicking phantoms and brain tissues with a correlation of 0.82 and 0.66 to temperature measured with a thermometer, respectively. The mean difference between the TR-NIRS and thermometer measurements was 0.15°C±1.1°C for the in vitro experiments and 0.5°C±1.6°C for the in vivo measurements.

Bakhsheshi, Mohammad Fazel; Diop, Mamadou; St. Lawrence, Keith; Lee, Ting-Yim

2014-05-01

215

High-resolution Time-resolved Extreme Ultraviolet Spectroscopy on NSTX  

NASA Astrophysics Data System (ADS)

We report on high-resolution, time-resolved spectroscopy in the extreme ultraviolet spectral region (10-200 å) on the NSTX tokamak. This work utilizes two flat-field spectrometers on loan from LLNL's electron beam ion trap facility. XEUS, installed in 2004, has a 2400 line/mm flat-field grating with field of view of ˜50 åthat can be positioned to survey 5 - 135 åwith an instrumental resolution of ˜0.1 åand ?/??˜100 at 10åto ˜1000 at 100 å. LoWEUS, installed in 2008, utilizes a 1200 line/mm grating with field of view of ˜180,s typically positioned to survey 60-280 åwith an instrumental resolution of ˜0.3 åand ?/??˜300 at 100åto ˜600 at 200å. New cameras have achieved a time resolution of 12-13 ms for both instruments. We can now examine time dependence and evolution of both intrinsic and extrinsic impurities on NSTX in the EUV band. Of particular interest is monitoring the entry of molybdenum into the plasma after installation of Mo tiles for the 2011 run. [4pt] Work supported by DOE General Plasma Science program. Part of this work performed under the auspices of DOE by LLNL under contract DE-AC52-07NA27344 and PPPL under contract DE-AC02-09CH11466.

Lepson, J. K.; Beiersdorfer, P.; Clementson, J.; Bitter, M.; Hill, K.; Kaita, R.; Roquemore, L.; Skinner, C. H.; Zimmer, G.

2011-11-01

216

Excited state dynamics of nanocrystalline VO2 with white light continuum time resolved spectroscopy  

NASA Astrophysics Data System (ADS)

In an attempt to use ultrafast pump probe spectroscopy technique with white light continuum to reveal wavelength dependent dynamics of VO2, bandgap needs to be opened. Therefore, nanostructured amorphous and crystalline VO2 thin films were prepared with pulsed DC magnetron reactive sputtering. The mean diameters of grains were measured as 22±0.1 nm and 44±0.1 nm for amorphous and crystalline VO2 thin films, respectively. Temperature dependent resistance measurements show that nanocrystalline VO2 thin film exhibit metal insulator phase transition. The films exhibited dual band gaps (2.3 eV, <0.6 eV for amorphous films and 1.3 eV, 1.8 eV for crystalline film). Increased band gaps made it possible to perform time resolved transmission and reflection experiments with white light continuum at fluences above and below photo induced phase transition threshold. Although transmission chance due to photo induced phase transition of VO2 in the literature usually takes places at infrared region of the spectrum, transmission chance was observed in visible as low as 630 nm in broadband probe spectra. It was observed that measured time scales depend on not only pump fluence but also probe wavelength. Experiments gave the evidence of the long-lived lower energy non-equilibrium state related to the photo induced phase.

Kürüm, Ula?; Yaglioglu, H. Gul; Küçüköz, Betül; Oksuzoglu, R. Mustafa; Y?ld?r?m, Mustafa; Ya?c?, A. Murat; Pekdemir, Sami; Elmali, Ayhan

2014-12-01

217

Time resolved spectroscopy of SGR J1550-5418 bursts detected with Fermi/GBM  

E-print Network

We report on time-resolved spectroscopy of the 63 brightest bursts of SGR J1550-5418, detected with Fermi/Gamma-ray Burst Monitor during its 2008-2009 intense bursting episode. We performed spectral analysis down to 4 ms time-scales, to characterize the spectral evolution of the bursts. Using a Comptonized model, we find that the peak energy, E_peak, anti-correlates with flux, while the low-energy photon index remains constant at -0.8 up to a flux limit F~10^-5 erg s-1 cm-2. Above this flux value the E_peak-flux correlation changes sign, and the index positively correlates with flux reaching 1 at the highest fluxes. Using a two black-body model, we find that the areas and fluxes of the two emitting regions correlate positively. Further, we study here for the first time, the evolution of the temperatures and areas as a function of flux. We find that the area-kT relation follows lines of constant luminosity at the lowest fluxes, R^2 \\propto kT^-4, with a break at higher fluxes ($F>10^-5.5 erg s-1 cm-2). The are...

Younes, G; van der Horst, A J; Baring, M G; Granot, J; Watts, A L; Bhat, P N; Collazzi, A; Gehrels, N; Gorgone, N; Gogus, E; Gruber, D; Grunblatt, S; Huppenkothen, D; Kaneko, Y; von Kienlin, A; van der Klis, M; Lin, L; Mcenery, J; van Putten, T; Wijers, R A M J

2014-01-01

218

Pseudo-bimolecular [2+2] cycloaddition studied by time-resolved photoelectron spectroscopy.  

PubMed

The first study of pseudo-bimolecular cycloaddition reaction dynamics in the gas phase is presented. We used femtosecond time-resolved photoelectron spectroscopy (TRPES) to study the [2+2] photocycloaddition in the model system pseudo-gem-divinyl[2.2]paracyclophane. From X-ray crystal diffraction measurements we found that the ground-state molecule can exist in two conformers; a reactive one in which the vinyl groups are immediately situated for [2+2] cycloaddition and a nonreactive conformer in which they point in opposite directions. From the measured S(1) lifetimes we assigned a clear relation between the conformation and the excited-state reactivity; the reactive conformer has a lifetime of 13?ps, populating the ground state through a conical intersection leading to [2+2] cycloaddition, whereas the nonreactive conformer has a lifetime of 400?ps. Ab initio calculations were performed to locate the relevant conical intersection (CI) and calculate an excited-state [2+2] cycloaddition reaction path. The interpretation of the results is supported by experimental results on the similar but nonreactive pseudo-para-divinyl[2.2]paracyclophane, which has a lifetime of more than 500?ps in the S(1) state. PMID:21365700

Brogaard, Rasmus Y; Boguslavskiy, Andrey E; Schalk, Oliver; Enright, Gary D; Hopf, Henning; Raev, Vitaly A; Jones, Peter G; Thomsen, Ditte L; Sølling, Theis I; Stolow, Albert

2011-03-28

219

Nanosecond Time-Resolved Polarization Spectroscopies: Tools for Probing Protein Reaction Mechanisms  

PubMed Central

Polarization methods, introduced in the 1800’s, offered one of the earliest ways to examine protein structure. Since then, many other structure-sensitive probes have been developed, but circular dichroism (CD) remains a powerful technique because of its versatility and the specificity of protein structural information that can be explored. With improvements in time-resolution, from millisecond to picosecond CD measurements, it has proven to be an important tool for studying the mechanism of folding and function in many biomolecules. For example, nanosecond time-resolved CD (TRCD) studies of the sub-microsecond events of reduced cytochrome c folding have provided direct experimental evidence of kinetic heterogeneity, which is an inherent property of the diffusional nature of early folding dynamics on the energy landscape. In addition, TRCD has been applied to the study of many biochemical processes, such as ligand rebinding in hemoglobin and myoglobin and signaling state formation in photoactive yellow protein and prototropin 1 LOV2. The basic approach to TRCD has also been extended to include a repertoire of nanosecond polarization spectroscopies: optical rotatory dispersion (ORD), magnetic CD and ORD, and linear dichroism. This article will discuss the details of the polarization methods used in this laboratory, as well as the coupling of timeresolved ORD with the temperature-jump trigger so that protein folding can be studied in a larger number of proteins. PMID:20438842

Chen, Eefei; Goldbeck, Robert A.; Kliger, David S.

2010-01-01

220

"Live" Prussian blue fading by time-resolved X-ray absorption spectroscopy  

NASA Astrophysics Data System (ADS)

Prussian blue (PB) is an artists' pigment that has been frequently used in many artworks but poses several problems of conservation because of its fading under light and anoxia treatment. PB fading is due to the reduction of iron(III) into iron(II) and depends a lot on the object investigated. Due to the complexity of the structure, the precise physico-chemical mechanisms behind the redox process remain obscure. In this paper, we present a procedure to investigate light- and anoxia-induced fading of PB-paper samples by means of time resolved X-ray absorption spectroscopy performed at the Fe K-edge. A system composed of a visible light source and a flux-controlled environmental cell allowed light, gas and humidity to be modified in situ. The synchrotron X-ray beam was evidenced to induce a reduction of PB and to play a major role in the kinetics. The analysis of the PB fading kinetics of a sample submitted to various gas and light environments showed that both synchrotron beam and anoxia were influencing PB reduction in a correlated way. In comparison, light was found to play a minor role. Finally, we have demonstrated that the type of paper substrate could influence significantly the kinetics of reduction. Several hypotheses to explain the correlation between PB reduction mechanism and substrate are presented.

Gervais, Claire; Languille, Marie-Angélique; Reguer, Solenn; Gillet, Martine; Vicenzi, Edward P.; Chagnot, Sébastien; Baudelet, François; Bertrand, Loïc

2013-04-01

221

Time-resolved fluorescence for breast cancer detection using an octreotate-indocyanine green derivative dye conjugate  

NASA Astrophysics Data System (ADS)

Time-resolved fluorescence was used to investigate malignant and normal adjacent breast tissues stained with a conjugate of indocyanine green and octreotate. A marked increase in fluorescence lifetime intensity was seen in the breast cancer sample compared to the normal sample. The fluorescent lifetimes were also investigated and showed similar fluorescence decay curves in stained malignant and normal breast tissue. These results confirm that somatostatin receptors occur on human breast carcinomas, suggest that the presence of somatostatin receptors should be investigated as a marker of breast cancer aggressiveness, and suggest that this conjugate might be used to detect the presence of residual breast cancer after surgery, allowing better assessment of tumor margins and reducing the need for second or repeat biopsies in selected patients. These results may also provide clues for designing future treatment options for breast cancer patients.

Sordillo, Laura A.; Das, B. B.; Pu, Yang; Liang, Kexian; Milione, Giovanni; Sordillo, Peter P.; Achilefu, Sam; Alfano, R. R.

2013-03-01

222

Monitoring the State of Cholecystokinin Receptor Oligomerization after Ligand Binding Using Decay of Time-Resolved Fluorescence Anisotropy  

PubMed Central

Oligomeric complexes of G protein–coupled receptors (GPCRs) are now commonly recognized and can provide a mechanism for regulation of signaling systems. Receptor oligomerization has been most extensively studied using coimmunoprecipitation and bioluminescence or fluorescence resonance energy-transfer techniques. Here, we have utilized decay of time-resolved fluorescence anisotropy of yellow fluorescent protein-labeled cholecystokinin receptor constructs to examine the state of oligomerization of this receptor in living cells. The rotational correlation times established that the cholecystokinin receptor is constitutively present in an oligomeric state that is dissociated in response to agonist occupation. In contrast, antagonist occupation failed to modify this signal, leaving the oligomeric structure intact. This dynamic technique complements the other biochemical and steady-state fluorescence techniques to establish the presence of oligomeric receptor complexes in living cells. PMID:19076359

Harikumar, Kaleeckal G.; Miller, Laurence J.

2013-01-01

223

A time-resolved near-infrared fluorescence assay for glucose: opportunities for trans-dermal sensing.  

PubMed

We report a time-resolved near-infrared fluorescence assay for glucose detection that incorporates pulsed diode laser excitation. Reduction in fluorescence resonance energy transfer to a malachite green-Dextran complex from allophycocyanin bound to concanavalin A (ConA) due to displacement of the complex by glucose from ConA provides the basis of the assay. The fluorescence quenching kinetics are analysed and discussed in detail. The change in fluorescence decay kinetics in the presence of glucose is found from dimensionality studies to be brought about by a change in the distribution of malachite green-Dextran acceptors. Glucose concentrations are measured in solution to within +/- 10% over the range 0-30 mM. PMID:10739140

Rolinski, O J; Birch, D J; McCartney, L J; Pickup, J C

2000-01-01

224

Evaluating steady-state and time-resolved fluorescence as a tool to study the behavior of asphaltene in toluene.  

PubMed

A combination of steady-state fluorescence, fluorescence lifetime measurements and the determination of time-resolved emission spectra were employed to characterize asphaltene toluene solutions. Lifetime measurements were shown to be insensitive to the source of asphaltene or the alkane solvent from which asphaltene was precipitated. This insensitivity suggests that either the composition of Athabasca and Cold Lake asphaltene is very similar or that the fluorescence behavior is dominated by the same sub-set of fluorophores for the different samples. These results highlight the limitations in using fluorescence to characterize asphaltene solutions. Different dependencies were observed for the average lifetimes with the asphaltene concentration when measured at two different emission wavelengths (420 nm and 520 nm). This result suggests that different fluorophores underwent diverse interactions with other asphaltene molecules as the asphaltene concentration was raised, suggesting that models for asphaltene aggregation need to include molecular diversity. PMID:24722727

Zhang, Hui Ting; Li, Rui; Yang, Zixin; Yin, Cindy-Xing; Gray, Murray R; Bohne, Cornelia

2014-06-01

225

Time-resolved multi-channel optical system for assessment of brain oxygenation and perfusion by monitoring of diffuse reflectance and fluorescence  

NASA Astrophysics Data System (ADS)

Time-resolved near-infrared spectroscopy is an optical technique which can be applied in tissue oxygenation assessment. In the last decade this method is extensively tested as a potential clinical tool for noninvasive human brain function monitoring and imaging. In the present paper we show construction of an instrument which allows for: (i) estimation of changes in brain tissue oxygenation using two-wavelength spectroscopy approach and (ii) brain perfusion assessment with the use of single-wavelength reflectometry or fluorescence measurements combined with ICG-bolus tracking. A signal processing algorithm based on statistical moments of measured distributions of times of flight of photons is implemented. This data analysis method allows for separation of signals originating from extra- and intracerebral tissue compartments. In this paper we present compact and easily reconfigurable system which can be applied in different types of time-resolved experiments: two-wavelength measurements at 687 and 832 nm, single wavelength reflectance measurements at 760 nm (which is at maximum of ICG absorption spectrum) or fluorescence measurements with excitation at 760 nm. Details of the instrument construction and results of its technical tests are shown. Furthermore, results of in-vivo measurements obtained for various modes of operation of the system are presented.

Milej, D.; Gerega, A.; Kacprzak, M.; Sawosz, P.; Weigl, W.; Maniewski, R.; Liebert, A.

2014-03-01

226

Time-resolved spectroscopy on epitaxial graphene in the infrared spectral range: relaxation dynamics and saturation behavior  

E-print Network

1 Time-resolved spectroscopy on epitaxial graphene in the infrared spectral range: relaxation graphene samples performed in a wide spectral range, namely from the near-infrared (photon energy 1.5 eV) to the terahertz (photon energy 8 meV) spectral range

Boyer, Edmond

227

Probing Reaction Dynamics of Transition-Metal Complexes in Solution via Time-Resolved Soft X-ray Spectroscopy  

SciTech Connect

We report the first time-resolved soft x-ray measurements of solvated transition-metal complexes. L-edge spectroscopy directly probes dynamic changes in ligand-field splitting of 3d orbitals associated with the spin transition, and mediated by changes in ligand-bonding.

Huse, Nils; Kim, Tae Kyu; Khalil, Munira; Jamula, Lindsey; McCusker, James K.; Schoenlein, Robert W.

2010-05-02

228

Probing Folded and Unfolded States of Outer Membrane Protein A using Steady-State and Time-Resolved Tryptophan Fluorescence  

PubMed Central

Steady-state and time-resolved fluorescence measurements on each of five native tryptophan residues in full-length and truncated variants of E. coli outer-membrane protein A (OmpA) have been made in folded and denatured states. Tryptophan singlet excited-state lifetimes are multiexponential and vary among the residues. In addition, substantial increases in excited-state lifetimes accompany OmpA folding, with longer lifetimes in micelles than in phospholipid bilayers. This finding suggests that the Trp environments of OmpA folded in micelles and phospholipid bilayers are different. Measurements of Trp fluorescence decay kinetics with full-length OmpA folded in brominated lipid vesicles reveal that W102 is the most distant fluorophore from the hydrocarbon core, while W7 is the closest. Steady-state and time-resolved polarized fluorescence measurements indicate reduced Trp mobility when OmpA is folded in a micelle, and even lower mobility when the protein is folded in a bilayer. The fluorescence properties of truncated OmpA, in which the soluble periplasmic domain is removed, only modestly differ from those of the full-length form, suggesting similar folded structures for the two forms under these conditions. PMID:16942111

Kim, Judy E.; Arjara, Gitrada; Richards, John H.; Gray, Harry B.; Winkler, Jay R.

2008-01-01

229

Nanosecond rapid freezing of liquid benzene under shock compression studied by time-resolved coherent anti-Stokes Raman spectroscopy  

NASA Astrophysics Data System (ADS)

Nanosecond time-resolved coherent anti-Stokes Raman spectroscopy is used to investigate the shock-induced liquid-solid phase transition and crystallization of liquid benzene. Temporal evolution of the Raman shift of the ring-breathing and C-H stretching modes is investigated. A metastable supercompressed state and a liquid-solid phase transition are observed under shock compression. Time-resolved Raman spectra reveal that the liquid state is initially a metastable state and rapidly transforms to the solid state within 25ns under shock compression at 4.2GPa.

Matsuda, Akitaka; Kondo, Ken-ichi; Nakamura, Kazutaka G.

2006-02-01

230

Liquid-Solid Phase Transition of Benzene under Shock Compression Stuided by Time-Resolved Nonlinear Raman Spectroscopy  

NASA Astrophysics Data System (ADS)

The liquid-solid phase transition of benzene has been studied under laser-shock compression up to 4.2 GPa by using nanosecond time-resolved nonlinear Raman spectroscopy. A shock wave is generated by irradiation of 10-ns pulsed laser beam on the plasma confinement target and its pressure is estimated from the particle velocity, which is measured by a velocity interferometer. The ring-breathing mode shows blue shift under shock compression. Nanosecond time-resolved nonlinear Raman spectra show a rapid phase transition from liquid phase to solid phase under shock compression.

Nakamura, K. G.; Matsuda, A.; Kondo, K.

2006-07-01

231

Global and Time-Resolved Monitoring of Crop Photosynthesis with Chlorophyll Fluorescence  

NASA Technical Reports Server (NTRS)

Photosynthesis is the process by which plants harvest sunlight to produce sugars from carbon dioxide and water. It is the primary source of energy for all life on Earth; hence it is important to understand how this process responds to climate change and human impact. However, model-based estimates of gross primary production (GPP, output from photosynthesis) are highly uncertain, in particular over heavily managed agricultural areas. Recent advances in spectroscopy enable the space-based monitoring of sun-induced chlorophyll fluorescence (SIF) from terrestrial plants. Here we demonstrate that spaceborne SIF retrievals provide a direct measure of the GPP of cropland and grassland ecosystems. Such a strong link with crop photosynthesis is not evident for traditional remotely sensed vegetation indices, nor for more complex carbon cycle models. We use SIF observations to provide a global perspective on agricultural productivity. Our SIF-based crop GPP estimates are 50-75% higher than results from state-of-the-art carbon cycle models over, for example, the US Corn Belt and the Indo-Gangetic Plain, implying that current models severely underestimate the role of management. Our results indicate that SIF data can help us improve our global models for more accurate projections of agricultural productivity and climate impact on crop yields. Extension of our approach to other ecosystems, along with increased observational capabilities for SIF in the near future, holds the prospect of reducing uncertainties in the modeling of the current and future carbon cycle.

Guanter, Luis; Zhang, Yongguang; Jung, Martin; Joiner, Joanna; Voigt, Maximilian; Berry, Joseph A.; Frankenberg, Christian; Huete, Alfredo R.; Zarco-Tejada, Pablo; Lee, Jung-Eun; Moran, M. Susan; Ponce-Campos, Guillermo; Beer, Christian; Camps-Valls, Gustavo; Buchmann, Nina; Gianelle, Damiano; Klumpp, Katja; Cescatti, Alessandro; Baker, John M.; Griffis, Timothy J.

2014-01-01

232

Time-resolved Optical Spectroscopy of the Classical Nova V723 Cas  

NASA Astrophysics Data System (ADS)

We present the results of time-resolved optical spectroscopy of the classical nova and super-soft X-ray source V723 Cas (Nova Cas 1995). The spectra were obtained at the Steward Observatory Bok 2.3-m telescope (range: 4180-5320 Angstroms) on Kitt Peak and at the MDM Observatory Hilter 2.4-m telescope (range: 4000-7500 Angstroms) in 2007, 2008, and 2010. Both sets of spectra were obtained at a spectral resolution of about 2 Angstroms. Exposure times of the individual spectra were about 20 minutes which is short compared to the 16.6-hour binary orbital period, thus minimizing velocity smearing of spectral features. V723 Cas is an unusual system, being an active super-soft X-ray source for more than 15 years since the 1995 outburst, in contrast to the median X-ray turn off time of only 1.4 years. This may be indicative of steady hydrogen burning on the white dwarf due to renewed accretion (Ness et al. 2008, AJ, 135, 1328). Phasing the individual spectra to the 16.6-hour binary photometric orbital period (Goranskij et al. 2007, Ast. Bull. 62, 125), we find that the Balmer emission lines exhibit a double-peaked accretion disk line profile convolved with a variable Gaussian emission line S-wave component. The He II 4686 emission line exhibits phase-dependent line profile variations as well. We have modeled both emission line profiles with a double-peaked accretion disk line component assuming different disk properties and an independent Gaussian component. The results of our line profile modeling compared to the orbital photometric behavior of V723 Cas are presented and we discuss the interpretation of our synoptic observations in the context of the post-outburst evolution of super-soft X-ray sources and this unusual classical nova.

Vonderharr, Thomas; Woodward, C. E.; Wagner, R. M.; Schwarz, G.; Helton, L. A.; Hamilton-Drager, C.; Recine, K. A.

2011-05-01

233

Two-Dimensional Subpicosecond Time-Resolved Fluorescence Anisotropy: Optical Kerr-Gating with a Dynamic Polarization Excitation.  

NASA Astrophysics Data System (ADS)

With an advent of ultrafast lasers, a number of applications are widely adopted to probe photophysical and photochemical properties of a molecule that occurs in an ultrafast (femtosecond to picosecond) time scale. Intramolecular charge transfer (ICT) or proton transfer in photoexcited electron donor-acceptor (EDA) molecules, for instance, has been a topic of very extensive time-resolved studies for several decades. Time-evolution of an anisotropic property can track dipole orientations or conformational changes in their photoexcited molecular systems, which is of extreme importance to examine its structure and excited-state dynamics rather than probing an isotropic "population change".With this respect, we recently developed a subpicosecond time-resolved 2-D fluorescence anisotropy (TRFA) in which method implements a dynamic alternation of laser polarizations to excite a sample using a photoelastic modulator (PEM). In the combination of an ultrafast optical shutter (Kerr-gating) and a spectrograph that is coupled with a CCD, two signal phases so-obtained dynamically, I_{?}( t, ?) and I_{?}( t, ?), provide a 2-D mapped information on both a wide range for spectra and time-resolved kinetics of photoexcited molecules of interest. From the definition of an anisotropy 2-D TRFA, r (t, ?), is given instantly and even more reliably at a single measurement. In this paper we will present benchmark tests of some target samples to establish performance of TRFA.

Fujiwara, Takashige; Romano, Natalie C.; Modarelli, David A.; Lim, Edward C.

2013-06-01

234

Transient Absorption and Time-Resolved Fluorescence Studies of Solvated Ruthenium Di-Bipyridine Pseudo-Halide Complexes  

NASA Astrophysics Data System (ADS)

Time-resolved IR and fluorescence measurements were performed to probe the vibrational and electronic properties, respectively, of ruthenium di-bipyridine pseudo-halide (Ru(Bpy){_2}(X){_2} (where X = CN, N{_3} or NCS)) complexes. Vibrational energy relaxation (VER) times were determined for the complexes dissolved in dimethyl sulfoxide (DMSO) with a trend in VER time of NCS > CN > N{_3}. A similar trend and comparable absolute rates for NCS- and N3- were previously observed by our group and others for simple inorganic anions in solution, suggesting a minimal contribution due to complexation. Measurements of the VER time of the CN complex in various solvents provide VER times in ethanol (42.3 ps) and DMSO (53.3 ps), which shows that protic solvents promote the relaxation. Time-resolved fluorescence measurements indicate a strong ligand dependence, with a factor of five decrease in the excited electronic state decay time from the CN (215 ns) to the NCS (39 ns) complex. A solvent dependence of the CN complex reveals a nearly 3-fold increase in the fluorescence decay time from acetonitrile (70 ns) to DMSO (215 ns).

Compton, R.; Weidinger, D.; Owrutsky, J. C.

2012-06-01

235

A homogeneous europium cryptate-based assay for the diagnosis of mutations by time-resolved fluorescence resonance energy transfer  

PubMed Central

Oligonucleotide ligation assay (OLA) is considered to be a very useful methodology for the detection and characterization of mutations, particularly for clinical purposes. The fluorescence resonance energy transfer between a fluorescent donor and a suitable fluorophore as acceptor has been applied in the past to several scientific fields. This technique is well adapted to nucleic acid analysis such as DNA sequencing, DNA hybridization and polymerase chain reaction. We describe here a homogeneous format based on the use of a rare earth cryptate label as donor: tris-bipyridine-Eu3+. The long-lived fluorescence of this label makes it possible to reach a high sensitivity by using a time-resolved detection mode. A non-radiative energy transfer technology, known as time-resolved amplification of cryptate emission (TRACE®) characterized by a temporal and spectral selectivity has been developed. The TRACE® detection of characterized single nucleotide polymorphism using the OLA for allelic discrimination is proposed. We demonstrate the potentialities of this OLA–TRACE® methodology through the analysis of K-ras oncogene point mutations. PMID:11452039

Lopez-Crapez, E.; Bazin, H.; Andre, E.; Noletti, J.; Grenier, J.; Mathis, G.

2001-01-01

236

Fast-gated intensified charge-coupled device camera to record time-resolved fluorescence spectra of tryptophan.  

PubMed

The possibilities of a 200 ps gated intensified charge-coupled device (CCD) camera to record time-resolved fluorescence were explored using the fluorescing amino acid tryptophan and its derivative Nacetyl-tryptophan amide (NATA) as model compounds. The results were compared to complementary data from time-correlated single-photon counting (TCSPC) experiments. If a spectral resolution of 1-2 nm is desired, the fast-gated intensified CCD (ICCD) camera is the method of choice. For a 10(-5) M tryptophan solution, time-resolved emission spectra and intensity decays (measured over 12 ns at 25 ps resolution) could be obtained in typically 10 minutes, giving the well-known lifetimes of 0.5 and 3 ns. In addition, a longer lifetime of 7 ns was found at the red edge of the spectrum. The very short gate time of the ICCD camera allowed us to observe a shift in the emission maximum of tryptophan even within the first nanosecond of decay of the fluorescence emission. As expected from the tryptophan rotamer model, such a shift is not observed in NATA. Using amplitudes obtained by global analysis, decay-associated spectra of these lifetimes were constructed. PMID:15198823

Stortelder, Aike; Buijs, Joost B; Bulthuis, Jaap; Gooijer, Cees; van der Zwan, Gert

2004-06-01

237

REFINING MEMBRANE PENETRATION BY A COMBINATION OF STEADY-STATE AND TIME-RESOLVED DEPTH-DEPENDENT FLUORESCENCE QUENCHING  

PubMed Central

Accurate determination of the depth of membrane penetration of a fluorescent probe, attached to lipid, protein or other macromolecule of interest, using depth-dependent quenching methodology is complicated by thermal motion in the lipid bilayer. Here we suggest that a combination of steady-state and time-resolved measurements can be used to generate a static quenching profile that reduces the contribution from transverse diffusion occurring during the excited-state lifetime. This procedure results in narrower quenching profiles, compared to those obtained by traditional measurements, and thus improves precision in determination of the underlying depth distribution of the probe. PMID:24141077

Kyrychenko, Alexander; Ladokhin, Alexey S.

2013-01-01

238

Highly Sensitive Time-Resolved Fluoroimmunoassay of Human Immunoglobulin E by Using a New Europium Fluorescent Chelate as a Label  

Microsoft Academic Search

A new europium fluorescent chelate, 4,4?-bis(1?,1?, 1?,2?,2?,3?,3?-heptafluoro-4?,6?-hexanedione-6?-yl)-chlorosulfo-o-terphenyl (BHHCT)–Eu3+, was used as a label for highly sensitive time-resolved fluoroimmunoassay of human IgE. Two assay formats were employed in the analysis. In the first format, an immunoconjugate of rabbit anti-human IgE antibody–human IgE-biotinylated goat anti-human IgE antibody–BHHCT- Eu3+-labeled SA (or BHHCT–Eu3+-labeled BSA–SA; BSA, bovine serum albumin; SA, streptavidin) was used for measurement.

Jingli Yuan; Guilan Wang; Hiroko Kimura; Kazuko Matsumoto

1997-01-01

239

Cyclodextrin supramolecular inclusion-enhanced pyrene excimer switching for time-resolved fluorescence detection of biothiols in serum.  

PubMed

We report here an efficient pyrene excimer signaling-based time-resolved fluorescent sensor for the measurement of biothiols (cysteine (Cys), homocysteine (Hcy), glutathione (GSH)) in human serum based on thymine-Hg(2+)-thymine (T-Hg(2+)-T) coordination chemistry and the inclusion interaction of cyclodextrin. The sensing mechanism of the approach is based on the competitive ligation of Hg(2+) ions by Hcy/Cys/GSH and T-T mismatches in a bis-pyrene-labeled DNA strand with the self-complementary 5' and 3' ends. The introduction of ?-cyclodextrin can provide cooperation for the molecular level space proximity of the two labeled pyrene molecules, moreover the hydrophobic cavity of ?-cyclodextrin can also offer protection for the pyrene dimer's emission from the quenching effect of environmental conditions and enhance the fluorescence intensity of the pyrene excimer. When the biothiols are not presented, the sensing ensemble is in the "off" state due to the long distance between the two labeled pyrene molecules resulted from the formation of a more stable T-Hg(2+)-T structure. While in the presence of biothiols, Hg(2+) interacts very strongly with thiol groups and the T-Hg(2+)-T structure is dehybridized, and then the pyrene excimer will be formed due to the self-complementary 5' and 3' ends of the DNA probe and the cooperation interaction of ?-cyclodextrin to pyrene dimer, thus resulting in switching the sensing ensemble to the "on" state. In the optimum conditions described, the linear concentration range of 1.0-100?M with the limit of detection (LOD) of 0.36?M for GSH was obtained. Moreover, due to the much longer lifetime of the pyrene excimer fluorescence than those of the ubiquitous endogenous fluorescent components, the time-resolved fluorescence technique has been successfully used for application in complicated biological samples. PMID:25590970

Zhang, Qier; Deng, Ting; Li, Jishan; Xu, Weijian; Shen, Guoli; Yu, Ruqin

2015-06-15

240

Time-resolved multicolor two-photon excitation fluorescence microscopy of cells and tissues  

NASA Astrophysics Data System (ADS)

Multilabeling which maps the distribution of different targets is an indispensable technique in many biochemical and biophysical studies. Two-photon excitation fluorescence (TPEF) microscopy of endogenous fluorophores combining with conventional fluorescence labeling techniques such as genetically encoded fluorescent protein (FP) and fluorescent dyes staining could be a powerful tool for imaging living cells. However, the challenge is that the excitation and emission wavelength of these endogenous fluorophores and fluorescent labels are very different. A multi-color ultrafast source is required for the excitation of multiple fluorescence molecules. In this study, we developed a two-photon imaging system with excitations from the pump femtosecond laser and the selected supercontinuum generated from a photonic crystal fiber (PCF). Multiple endogenous fluorophores, fluorescent proteins and fluorescent dyes were excited in their optimal wavelengths simultaneously. A time- and spectral-resolved detection system was used to record the TPEF signals. This detection technique separated the TPEF signals from multiple sources in time and wavelength domains. Cellular organelles such as nucleus, mitochondria, microtubule and endoplasmic reticulum, were clearly revealed in the TPEF images. The simultaneous imaging of multiple fluorophores of cells will greatly aid the study of sub-cellular compartments and protein localization.

Zheng, Wei

2014-11-01

241

Time-Resolved FTIR Difference Spectroscopy for the Study of Photosystem I Particles with Plastoquinone-9 Occupying the A1 Binding Site  

E-print Network

Time-Resolved FTIR Difference Spectroscopy for the Study of Photosystem I Particles binding site. Using time-resolved, step-scan FTIR difference spectroscopy we have produced A1 -/A1 FTIR signatures, we produce an A1 -/A1 FTIR difference spectrum for PS I particles with plastoquinone-9 occupying

Hastings, Gary

242

Ultrafast dynamics in transparent fluids investigated via time-resolved third-order spectroscopy  

NASA Astrophysics Data System (ADS)

The ultrafast dynamics of near critical fluids and other transparent materials are investigated with time-resolved optical heterodyne detected (OHD) third-order spectroscopy. Among the main results of this dissertation are: supercritical fluctuations on a picosecond timescale are revealed by H2 rotational coherence decay; a giant acoustic response due to enhanced near critical compressibility is optically generated and characterized; a new pulse reconstruction algorithm is developed. The decay of J-specific Raman coherence birefringence of H2 rotors in supercritical CO2 (0.8 rhoc) are measured in order to understand the ˜picosecond timescale solvent fluctuations around the critical point. The H2 anisotropic Raman time correlation functions possess long-time exponential tails attributed to motional narrowing. Nonexponential early-time behavior indicates the inhomogeneities in anisotropic local potential interactions. Mixed classical nonadiabatic quantum molecular dynamics simulations are performed. The excellent agreement between experiment and the theoretical treatment supports the use of this methodology. Multiregional studies are conducted for mixtures ranging from gas-like to liquid-like densities. A non-monotonic dependence of the Raman coherence decay times and transition frequencies are observed. A novel third-order polarization due to accumulated acoustic grating is characterized in near critical CHF3 and CO2. The electrostrictively generated acoustic response is pi out of phase with the normal OKE birefringence. Repetition rate dependence identifies the accumulated origin of the phenomenon with an acoustic relaxation timescale of ˜100 ns. The combined effect of efficient coherence coupling excitation and accumulation results in a birefringent signal strength inversely dependent on the sound velocity to the fifth power. In contrast a single-pulse coherence coupling effect routinely observed in transparent media can be shown to originate from nuclear responses. Its impact on frequency selected dichroic Raman spectral density recovery is demonstrated for CCl4. For linearly-chirped pulses, the antisymmetric coherent coupling trace dominates the frequency integrated dichroism. Its absolute sign indicates the sign of the chirp. An assumption-free amplitude-phase reconstruction algorithm combining the coherent coupling dichroism, the intensity autocorrelation and the power spectrum is developed.

Peng, Jian

243

Time resolved fluorescence tomography of turbid media based on lifetime contrast  

PubMed Central

A general linear model for time domain (TD) fluorescence tomography is presented that allows a lifetime-based analysis of the entire temporal fluorescence response from a turbid medium. Simulations are used to show that TD fluorescence tomography is optimally performed using two complementary approaches: A direct TD analysis of a few time points near the peak of the temporal response, which provides superior resolution; and an asymptotic multi-exponential analysis based tomography of the decay portion of the temporal response, which provides accurate localization of yield distributions for various lifetime components present in the imaging medium. These results indicate the potential of TD technology for biomedical imaging with lifetime sensitive targeted probes, and provide useful guidelines for an optimal approach to fluorescence tomography with TD data. PMID:19529654

Kumar, Anand T. N.; Raymond, Scott B.; Boverman, Gregory; Boas, David A.; Bacskai, Brian J.

2009-01-01

244

Active illumination for wide-field time-resolved fluorescence imaging  

NASA Astrophysics Data System (ADS)

The large dynamic range of fluorescence emission collected is one of the major challenges in wide-field fluorescence lifetime imaging. To overcome this challenge, we developed an active illumination strategy to acquire optimal fluorescence signals over the sample imaged even in the presence of large fluorophore concentration distributions. We validated the stability of our approach in a multi-well plate setting with fluorophore concentrations ranging <2 orders of magnitude. We report the ability of our method to retrieve accurately the lifetime over this concentration range based on optimized wide-field data. Our results demonstrate that active wide-field illumination can improve the signal-to-noise ratio and weak-signal sensitivity for enhanced accuracy of fluorescence decay curve fitting and lifetime estimation at high acquisition speed.

Zhao, Lingling; Abe, Ken; Barroso, Margarida; Intes, Xavier

2013-06-01

245

Fluorescence yield and lifetime tomography from time-resolved transmittances of a breast tumor phantom  

NASA Astrophysics Data System (ADS)

In optical tumor detection region, there has been recently a considerable interest in simultaneously reconstructing yield and lifetime distributions of fluorescent imaging agents inside a pathologic tissue, since combined monitoring of these two parameters provides a potential means of in vivo interrogating quantitative and environmental information of specific molecules as well as accessing interactions among them. This paper describes the structure of a multi-channel time-correlated single photon counting (TCSPC) system for early breast tumor detection and how we use it to reconstruct the distribution of fluorescent parameters. By using a normalized Born appropriation algorithm, the proposed examination scheme in a transmission mode is experimentally validated to achieve simultaneous reconstruction of the fluorescent yield and lifetime distributions with reasonable accuracy. The performance of the instrument will be proved by using two targets be of different fluorescent agents embedded in solid phantom for image reconstruction.

Lu, Yiming; Gao, Feng; Zhang, Wei; Wu, Linhui; Zhou, Zhongxing

2012-03-01

246

Steady-state and time-resolved fluorescence studies of stripped Borage oil.  

PubMed

In this study we explored the spectroscopic properties of Borage oil, particularly the use of fluorescence techniques to investigate the presence of conjugated fatty acids (CFAs). This research has important health and dietary applications. The absorption and fluorescence spectra of different CFAs and Borage oil in ethanol were measured. Time-domain fluorescence was employed to establish the life times of the samples. We found that Borage oil contains 1.2x10(-3) mol L(-1) of alpha-eleostearic acid or its isomer (i.e., a conjugated triene), 1.6x10(-4) mol L(-1) of cis-parinaric acid (i.e., a conjugated tetraene) and 1.1x10(-5) mol L(-1) of c-COPA (i.e., a conjugated pentaene). Because of the three-exponential fluorescence intensity decay for Borage oil, other fatty acids with a four conjugated double bond system could not be excluded. PMID:19523559

Smyk, Bogdan; Amarowicz, Ryszard; Szabelski, Mariusz; Gryczynski, Ignacy; Gryczynski, Zygmunt

2009-07-30

247

Characterization of time-resolved fluorescence response measurements for distributed optical-fiber sensing.  

PubMed

A distributed optical-fiber sensing system based on pulsed excitation and time-gated photon counting has been used to locate a fluorescent region along the fiber. The complex Alq3 and the infrared dye IR-125 were examined with 405 and 780 nm excitation, respectively. A model to characterize the response of the distributed fluorescence sensor to a Gaussian input pulse was developed and tested. Analysis of the Alq3 fluorescent response confirmed the validity of the model and enabled the fluorescence lifetime to be determined. The intrinsic lifetime obtained (18.2±0.9 ns) is in good agreement with published data. The decay rate was found to be proportional to concentration, which is indicative of collisional deactivation. The model allows the spatial resolution of a distributed sensing system to be improved for fluorophores with lifetimes that are longer than the resolution of the sensing system. PMID:21102661

Sinchenko, Elena; Gibbs, W E Keith; Davis, Claire E; Stoddart, Paul R

2010-11-20

248

Characterization of the Dynamics of an Essential Helix in the U1A Protein by Time-Resolved Fluorescence Measurements†  

PubMed Central

The RNA recognition motif (RRM), one of the most common RNA-binding domains, recognizes single-stranded RNA. A C-terminal helix that undergoes conformational changes upon binding is often an important contributor to RNA recognition. The N-terminal RRM of the U1A protein contains a C-terminal helix (helix C) that interacts with the RNA-binding surface of a ?-sheet in the free protein (closed conformation), but is directed away from this ?-sheet in the complex with RNA (open conformation). The dynamics of helix C in the free protein have been proposed to contribute to binding affinity and specificity. We report here a direct investigation of the dynamics of helix C in the free U1A protein on the nanosecond time scale using time-resolved fluorescence anisotropy. The results indicate that helix C is dynamic on a 2–3 ns time scale within a 20° range of motion. Steady-state fluorescence experiments and molecular dynamics simulations suggest that the dynamical motion of helix C occurs within the closed conformation. Mutation of a residue on the ?-sheet that contacts helix C in the closed conformation dramatically destabilizes the complex (Phe56Ala) and alters the steady-state fluorescence, but not the time-resolved fluorescence anisotropy, of a Trp in helix C. Mutation of Asp90 in the hinge region between helix C and the remainder of the protein to Ala or Gly subtly alters the dynamics of the U1A protein and destabilizes the complex. Together these results show that helix C maintains a dynamic closed conformation that is stable to these targeted protein modifications and does not equilibrate with the open conformation on the nanosecond time scale. PMID:18293956

Anunciado, Divina; Agumeh, Michael; Kormos, Bethany L.; Beveridge, David L.; Knee, Joseph L.; Baranger, Anne M.

2008-01-01

249

Time-resolved fluorescence microspectroscopy for characterizing crude oils in bulk and hydrocarbon-bearing fluid inclusions.  

PubMed

Time-resolved fluorescence data was collected from a series of 23 bulk crude petroleum oils and six microscopic hydrocarbon-bearing fluid inclusions (HCFI). The data was collected using a diode laser fluorescence lifetime microscope (DLFLM) over the 460-700 nm spectral range using a 405 nm excitation source. The correlation between intensity averaged lifetimes (tau) and chemical and physical parameters was examined with a view to developing a quantitative model for predicting the gross chemical composition of hydrocarbon liquids trapped in HCFI. It was found that tau is nonlinearly correlated with the measured polar and corrected alkane concentrations and that oils can be classified on this basis. However, these correlations all show a large degree of scatter, preventing accurate quantitative prediction of gross chemical composition of the oils. Other parameters such as API gravity and asphaltene, aromatic, and sulfur concentrations do not correlate well with tau measurements. Individual HCFI were analyzed using the DLFLM, and time-resolved fluorescence measurements were compared with tau data from the bulk oils. This enabled the fluid within the inclusions to be classified as either low alkane/high polar or high alkane/low polar. Within the high alkane/low polar group, it was possible to clearly discriminate HCFI from different locales and to see differences in the trapped hydrocarbon fluids from a single geological source. This methodology offers an alternative method for classifying the hydrocarbon content of HCFI and observing small variations in the trapped fluid composition that is less sensitive to fluctuations in the measurement method than fluorescence intensity based methods. PMID:15479528

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

2004-09-01

250

Ultrafast time-resolved coherent degenerate four-wave-mixing spectroscopy for investigating molecular dynamics in different states  

NASA Astrophysics Data System (ADS)

In this paper, ultrafast time-resolved coherent degenerate four-wave-mixing (DFWM) spectroscopy is performed to investigate molecular dynamics in the gaseous phase. Laser pulses lasting for 40 fs are used to create and monitor different vibrational eigenstates of iodine at room temperature (corresponding to a low saturation pressure of about 35 Pa). Using an internal time delay in the DFWM process resonant with the transition between the ground X-state and the excited B-state, the vibrational states of both the electronically excited and the ground states are detected as oscillations in the DFWM transient signal. The dynamics of either the electronically excited or ground state of iodine molecules obtained are consistent with the previous high temperature studies on the femtosecond time-resolved DWFM spectroscopy.

He, Ping; Fan, Rongwei; Chen, Deying; Li, Xiaohui; Xia, Yuanqin; Yu, Xin; Yao, Yong

2011-11-01

251

Molecular Orbital Imaging of the Acetone S2 Excited State Using Time-Resolved (e , 2 e ) Electron Momentum Spectroscopy  

NASA Astrophysics Data System (ADS)

We report a time-resolved (e , 2 e ) experiment on the deuterated acetone molecule in the S2 Rydberg state with a lifetime of 13.5 ps. The acetone S2 state was prepared by a 195 nm pump laser and probed with electron momentum spectroscopy using a 1.2 keV incident electron beam of 1 ps temporal width. In spite of the low data statistics as well as of the limited time resolution (±35 ps ) due to velocity mismatch, the experimental results clearly demonstrate that electron momentum spectroscopy measurements of short-lived transient species are feasible, opening the door to time-resolved orbital imaging in momentum space.

Yamazaki, Masakazu; Oishi, Keiya; Nakazawa, Hiroyuki; Zhu, Chaoyuan; Takahashi, Masahiko

2015-03-01

252

TOPICAL REVIEW: Time-resolved fluorescence imaging in biology and medicine  

NASA Astrophysics Data System (ADS)

Fluorescence lifetime imaging is a rather new and effective tool that can be used to study complex biological samples, either at microscopic or macroscopic levels. The map of the fluorescence lifetime allows one to discriminate amongst different fluorophores and to achieve valuable insights into the behaviour of emitting molecules, leading to information like local pH, oxygen concentration in cells, etc. Moreover, the distribution in space of any fluorescent marker achievable with this technique can be exploited for diagnostic purposes in medicine. After a brief introduction on the motivations for applying fluorescence lifetime imaging in biology and medicine, the basic principles of this technique will be addressed. Then, the two possible implementations of fluorescence lifetime imaging (i.e. the frequency domain and the time domain methods) will be presented. For this purpose, special attention will be devoted to practical aspects of image acquisition and processing, especially for what concerns the time domain method. Then, the analysis of the state-of-the-art systems will include a brief discussion on new concepts that have recently been introduced in this research field. Finally, two interesting applications of fluorescence lifetime imaging will be presented. The former refers to skin tumour detection and has been successfully applied in a preliminary clinical trial, the latter regards DNA chips reading and has been tested only at laboratory level, yet it has produced promising results for its future implementation in commercial systems.

Cubeddu, R.; Comelli, D.; D'Andrea, C.; Taroni, P.; Valentini, G.

2002-05-01

253

ANG-2 for quantitative Na(+) determination in living cells by time-resolved fluorescence microscopy.  

PubMed

Sodium ions (Na(+)) play an important role in a plethora of cellular processes, which are complex and partly still unexplored. For the investigation of these processes and quantification of intracellular Na(+) concentrations ([Na(+)]i), two-photon coupled fluorescence lifetime imaging microscopy (2P-FLIM) was performed in the salivary glands of the cockroach Periplaneta americana. For this, the novel Na(+)-sensitive fluorescent dye Asante NaTRIUM Green-2 (ANG-2) was evaluated, both in vitro and in situ. In this context, absorption coefficients, fluorescence quantum yields and 2P action cross-sections were determined for the first time. ANG-2 was 2P-excitable over a broad spectral range and displayed fluorescence in the visible spectral range. Although the fluorescence decay behaviour of ANG-2 was triexponential in vitro, its analysis indicates a Na(+)-sensitivity appropriate for recordings in living cells. The Na(+)-sensitivity was reduced in situ, but the biexponential fluorescence decay behaviour could be successfully analysed in terms of quantitative [Na(+)]i recordings. Thus, physiological 2P-FLIM measurements revealed a dopamine-induced [Na(+)]i rise in cockroach salivary gland cells, which was dependent on a Na(+)-K(+)-2Cl(-) cotransporter (NKCC) activity. It was concluded that ANG-2 is a promising new sodium indicator applicable for diverse biological systems. PMID:25311309

Roder, Phillip; Hille, Carsten

2014-12-01

254

Time-resolved spectroscopy and microscopic characterization of individual molecular aggregates  

NASA Astrophysics Data System (ADS)

Nanometer size domains play an important role in the photophysics of conjugated organic materials, and it is an open question as to how the properties of such domains depend on their specific structures. To study the properties of individual nano-aggregates, we use a combination of confocal microscopy, ultrafast spectroscopy, and atomic force microscopy. For unsubstituted phenylene-vinylene oligomers, which are used as a model system to understand the properties of the electroluminescent polymer PPV, we find that the morphology of the aggregates strongly influences their picosecond emission properties, with polycrystalline aggregates having enhanced excimer emission relative to crystalline domains of similar size. Temperature-dependent measurements of both the early time fluorescent exciton dynamics and the longer time dark state dynamics will also be presented.

Lim, Sang-Hyun; Bjorklund, Thomas G.; Bardeen, Christopher

2003-03-01

255

Adsorption of Eu(III) on a heterogeneous surface studied by time-resolved laser fluorescence microscopy (TRLFM).  

PubMed

Time-resolved laser fluorescence microscopy (TRLFM) is a useful tool to simultaneously investigate the intensity, location, type, and surrounding chemical environment of a fluorophore. In this study, we demonstrated the applicability of TRLFM for the adsorption of Eu(III) on a natural heterogeneous surface. Different adsorption species of Eu(III) were observed on the Makabe granite surface and its constituents (biotite, plagioclase, potassium feldspar, and quartz). Eu(III) heterogeneously adsorbed on biotite, plagioclase, and quartz and homogeneously on potassium feldspar. The histograms of the fluorescence decay rates of adsorbed Eu(III) indicated efficient quenching of Eu(III) fluorescence probably due to Eu(III)-surface interaction or the formation polynuclear hydoxo Eu(III) species on the surfaces. It was also revealed that single species of Eu(III) was observed on biotite and two species on plagioclase and potassium feldspar. The adsorption of Eu(III) on the granite surface was highly heterogeneous. The TRLFM measurements of different regions of the granite surface turned into the finding of Eu(III) with different fluorescence decay rates. Comparing with the fluorescence decay histograms of the mineral constituents, Eu(III) clearly adsorbed on the feldspar family. It was also found that Eu(III) adsorbed as an outer-sphere complex and on an altered mineral of the granite. PMID:19368166

Ishida, Keisuke; Kimura, Takaumi; Saito, Takumi; Tanaka, Satoru

2009-03-15

256

Ultrafast time-resolved coherent degenerate four-wave-mixing spectroscopy for investigating molecular dynamics in different states  

Microsoft Academic Search

In this paper, ultrafast time-resolved coherent degenerate four-wave-mixing (DFWM) spectroscopy is performed to investigate molecular dynamics in the gaseous phase. Laser pulses lasting for 40fs are used to create and monitor different vibrational eigenstates of iodine at room temperature (corresponding to a low saturation pressure of about 35Pa). Using an internal time delay in the DFWM process resonant with the

Ping He; Rongwei Fan; Deying Chen; Xiaohui Li; Yuanqin Xia; Xin Yu; Yong Yao

2011-01-01

257

Hot photocarrier dynamics in organic solar cells measured by transient absorption and time-resolved terahertz spectroscopy  

NASA Astrophysics Data System (ADS)

We present a study of charge transfer and carrier dynamics in films of zinc phthalocyanine (ZnPc) and buckmisnsterfullerene (C60) by investigated by time-resolved terahertz spectroscopy (TRTS). We compare terahertz photoconductivity dynamics in composite and multi-layered films of C60 and ZnPc. The few picosecond terahertz photoconductivity dynamics arise from autoionization and recombination between C60 molecules and cooling of hot photocarriers following from charge transfer between C60 and ZnPc.

Lane, Paul A.; Cunningham, Paul D.; Melinger, Joseph S.; Heilweil, Edwin J.

2014-10-01

258

Time-resolved fluorescence spectroscopic study of crude petroleum oils: influence of chemical composition.  

PubMed

The fluorescence of crude petroleum oils is sensitive to changes in chemical composition and many different fluorescence methods have been used to characterize crude oils. The use of fluorescence lifetimes to quantitatively characterize oil composition has practical advantages over steady-state measurements, but there have been comparatively few studies in which the lifetime behavior is correlated with gross chemical compositional data. In this study, the fluorescence lifetimes for a series of 23 crude petroleum oils with American Petroleum Institute (API) gravities of between 10 and 50 were measured at several emission wavelengths (450-785 nm) using a 380 nm light emitting diode (LED) excitation source. It was found that the intensity average fluorescence lifetime (tau) at any emission wave-length does not correlate well with either API gravity or aromatic concentration. However, it was found that tau is strongly negatively correlated with both the polar and sulfur concentrations and positively correlated with the corrected alkane concentration. This indicates that the fluorescence behavior of crude petroleum oils is governed primarily by the concentration of quenching species. All the strong lifetime-concentration correlations are nonlinear and show a high degree of scatter, especially for medium to light oils with API gravities of between 25 and 40. The degree of scatter is greatest for oils where the concentrations (wt %) of the polar fraction is approximately 10 +/- 4%, the asphaltene component is approximately 1 +/- 0.5%, and sulfur is 0.5 +/- 0.4%. This large degree of scatter precludes the use of average fluorescence lifetime data obtained with 380 nm excitation for the accurate prediction of the common chemical compositional parameters of crude petroleum oils. PMID:15165340

Ryder, Alan G

2004-05-01

259

Time-resolved protein fluorescence studies of intermediates in the photochemical cycle of bacteriorhodopsin.  

PubMed Central

The photolysis-induced changes in the protein fluorescence intensity (at 320 nm) during the proton-pumping cycle of bacteriorhodopsin were examined by a delayed two-pulse technique in the time range 1 microsecond-20 msec at room temperature. No detectable change in the protein fluorescence intensity was observed on the earliest time scale within the lifetime of the intermediate K590, when retinal apparently undergoes the largest structural changes. The time dependence of the relative changes in fluorescence intensity did, however, display a close correlation with the population of the L550 and M412 intermediates. From a computer numerical fit of the data, with available published kinetic parameters, the protein fluorescence quantum yields of the K590, L550, and M412 intermediates are found to be 1.0, 0.92, and 0.80 of that for native bR570, respectively. The probable mechanisms of the observed fluorescence quenching during the photochemical cycle are qualitatively discussed. PMID:6941246

Fukumoto, J M; Hopewell, W D; Karvaly, B; El-Sayed, M A

1981-01-01

260

High-pressure-low-temperature cryostat designed for use with fourier transform infrared spectrometers and time-resolved infrared spectroscopy.  

PubMed

The design for a new high-pressure-low-temperature infrared (IR) cell for performing experiments using conventional Fourier transform infrared or fast laser-based time-resolved infrared spectroscopy, in a range of solvents, is described. The design builds upon a commercially available compressor and cold end (Polycold PCC(®) and CryoTiger(®)), which enables almost vibration-free operation, ideal for use with sensitive instrumentation. The design of our cell and cryostat allows for the study of systems at temperatures from 77 to 310 K and at pressures up to 250 bar. The CaF2 windows pass light from the mid-IR to the ultraviolet (UV), enabling a number of experiments to be performed, such as Raman, UV-visible absorption spectroscopy, and time-resolved techniques where sample excitation/probing using continuous wave or pulsed lasers is required. We demonstrate the capabilities of this cell by detailing two different applications: (i) the reactivity of a range of Group V-VII organometallic alkane complexes using time-resolved spectroscopy on the millisecond timescale and (ii) the gas-to-liquid phase transition of CO2 at low temperature, which is applicable to measurements associated with transportation issues related to carbon capture and storage. PMID:24666949

Calladine, James A; Love, Ashley; Fields, Peter A; Wilson, Richard G M; George, Michael W

2014-01-01

261

Fluorescence spectroscopy and thermometry for hypersonic flight research  

NASA Technical Reports Server (NTRS)

The present study, which has demonstrated the feasibility of using time-resolved laser-induced fluorescence methods to ascertain boundary layer temperature distributions in real time for hypersonic vehicles in atmospheric flight, employs conventional optics in combination with novel time-resolved spectroscopy. The radiative lifetime and signal intensity involved, both of which are reduced by quenching, are adequate for measurement with state-of-the-art time-resolved fluorescence techniques, except for certain conditions associated with both the highest speeds and the lowest altitudes.

Thompson, B. E.; Fernandez, S. M.; Guignon, E. F.

1990-01-01

262

Europium chelate labels in time-resolved fluorescence immunoassays and DNA hybridization assays  

Microsoft Academic Search

Like many analytical methodologies, immunoassays and nucleic acid hybridization assays rely on the reaction between an analyte of interest and a specific reagent. The analyte concentration is then deduced by measuring either the amount of analyte-reagent complex formed (product) or the amount of residual reagent. The authors describe the application of fluorescent rare-earth chelates to immunoassay and DNA probing.

Eleftherios P. Diamandis; Theodore K. Christopoulos

1990-01-01

263

Global and time-resolved monitoring of crop photosynthesis with chlorophyll fluorescence  

Technology Transfer Automated Retrieval System (TEKTRAN)

Global monitoring of agricultural productivity is critical in a world under a continuous increase of food demand. Here we have used new spaceborne retrievals of chlorophyll fluorescence, an emission quantity intrinsically linked to photosynthesis, to derive spatially explicit photosynthetic uptake r...

264

Europium chelate labels in time-resolved fluorescence immunoassays and DNA hybridization assays  

SciTech Connect

Like many analytical methodologies, immunoassays and nucleic acid hybridization assays rely on the reaction between an analyte of interest and a specific reagent. The analyte concentration is then deduced by measuring either the amount of analyte-reagent complex formed (product) or the amount of residual reagent. The authors describe the application of fluorescent rare-earth chelates to immunoassay and DNA probing.

Diamandis, E.P.; Christopoulos, T.K. (Toronto Western Hospital, Ontario (Canada) Univ. of Toronto, Ontario (Canada))

1990-11-15

265

Quantifying the cerebral metabolic rate of oxygen by combining diffuse correlation spectroscopy and time-resolved near-infrared spectroscopy  

NASA Astrophysics Data System (ADS)

Preterm infants are highly susceptible to ischemic brain injury; consequently, continuous bedside monitoring to detect ischemia before irreversible damage occurs would improve patient outcome. In addition to monitoring cerebral blood flow (CBF), assessing the cerebral metabolic rate of oxygen (CMRO2) would be beneficial considering that metabolic thresholds can be used to evaluate tissue viability. The purpose of this study was to demonstrate that changes in absolute CMRO2 could be measured by combining diffuse correlation spectroscopy (DCS) with time-resolved near-infrared spectroscopy (TR-NIRS). Absolute CBF was determined using bolus-tracking TR-NIRS to calibrate the DCS measurements. Cerebral venous blood oxygenation (SvO2) was determined by multiwavelength TR-NIRS measurements, the accuracy of which was assessed by directly measuring the oxygenation of sagittal sinus blood. In eight newborn piglets, CMRO2 was manipulated by varying the anesthetics and by injecting sodium cyanide. No significant differences were found between the two sets of SvO2 measurements obtained by TR-NIRS or sagittal sinus blood samples and the corresponding CMRO2 measurements. Bland-Altman analysis showed a mean CMRO2 difference of 0.0268±0.8340 mL O2/100 g/min between the two techniques over a range from 0.3 to 4 mL O2/100 g/min.

Verdecchia, Kyle; Diop, Mamadou; Lee, Ting-Yim; St. Lawrence, Keith

2013-02-01

266

A dissociative fluorescence enhancement technique for one-step time-resolved immunoassays  

PubMed Central

The limitation of current dissociative fluorescence enhancement techniques is that the lanthanide chelate structures used as molecular probes are not stable enough in one-step assays with high concentrations of complexones or metal ions in the reaction mixture since these substances interfere with lanthanide chelate conjugated to the detector molecule. Lanthanide chelates of diethylenetriaminepentaacetic acid (DTPA) are extremely stable, and we used EuDTPA derivatives conjugated to antibodies as tracers in one-step immunoassays containing high concentrations of complexones or metal ions. Enhancement solutions based on different ?-diketones were developed and tested for their fluorescence-enhancing capability in immunoassays with EuDTPA-labelled antibodies. Characteristics tested were fluorescence intensity, analytical sensitivity, kinetics of complex formation and signal stability. Formation of fluorescent complexes is fast (5 min) in the presented enhancement solution with EuDTPA probes withstanding strong complexones (ethylenediaminetetra acetate (EDTA) up to 100 mM) or metal ions (up to 200 ?M) in the reaction mixture, the signal is intensive, stable for 4 h and the analytical sensitivity with Eu is 40 fmol/L, Tb 130 fmol/L, Sm 2.1 pmol/L and Dy 8.5 pmol/L. With the improved fluorescence enhancement technique, EDTA and citrate plasma samples as well as samples containing relatively high concentrations of metal ions can be analysed using a one-step immunoassay format also at elevated temperatures. It facilitates four-plexing, is based on one chelate structure for detector molecule labelling and is suitable for immunoassays due to the wide dynamic range and the analytical sensitivity. Figure   PMID:21161513

Mukkala, Veli-Matti; Hakala, Harri H. O.; Mäkinen, Pauliina H.; Suonpää, Mikko U.; Hemmilä, Ilkka A.

2010-01-01

267

Time-resolved fluorescence of 2-aminopurine as a probe of base flipping in M.HhaI–DNA complexes  

PubMed Central

DNA base flipping is an important mechanism in molecular enzymology, but its study is limited by the lack of an accessible and reliable diagnostic technique. A series of crystalline complexes of a DNA methyltransferase, M.HhaI, and its cognate DNA, in which a fluorescent nucleobase analogue, 2-aminopurine (AP), occupies defined positions with respect the target flipped base, have been prepared and their structures determined at higher than 2 ? resolution. From time-resolved fluorescence measurements of these single crystals, we have established that the fluorescence decay function of AP shows a pronounced, characteristic response to base flipping: the loss of the very short (?100 ps) decay component and the large increase in the amplitude of the long (?10 ns) component. When AP is positioned at sites other than the target site, this response is not seen. Most significantly, we have shown that the same clear response is apparent when M.HhaI complexes with DNA in solution, giving an unambiguous signal of base flipping. Analysis of the AP fluorescence decay function reveals conformational heterogeneity in the DNA–enzyme complexes that cannot be discerned from the present X-ray structures. PMID:16340006

Neely, Robert K.; Daujotyte, Dalia; Grazulis, Saulius; Magennis, Steven W.; Dryden, David T. F.; Klimašauskas, Saulius; Jones, Anita C.

2005-01-01

268

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

NASA Astrophysics Data System (ADS)

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.

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

1992-04-01

269

A novel luminescent terbium-3-carboxycoumarin probe for time-resolved fluorescence sensing of pesticides methomyl, aldicarb and prometryne  

NASA Astrophysics Data System (ADS)

The luminescence arising from lanthanide cations offers several advantages over organic fluorescent molecules: sharp, distinctive emission bands allow for easy resolution between multiple lanthanide signals; long emission lifetimes (?s-ms) make them excellent candidates for time-resolved measurements; and high resistance to photo bleaching allow for long or repeated experiments. A time-resolved (gated) luminescence-based method for determination of pesticides methomyl, aldicarb and prometryne in microtiterplate format using the long-lived terbium-3-carboxycoumarin in 1:3 metal:ligand ratio has been developed. The limit of detection is 1.20 × 106, 5.19 × 105 and 2.74 × 106 ng L-1 for methomyl, prometryne and aldicarb, respectively. The quantum yield (QY = 0.08) of Tb(III)-3-carboxycoumarin was determined using 3-(2-benzothiazolyl)-7-diethylamino-coumarin (coumarin 6). Stern-volmer studies at different temperatures indicate that collisional quenching dominates for methomyl, aldicarb and prometryne. Binding constants were determined at 303, 308 and 313 K by using Lineweaver-Burk equation. A thermodynamic analysis showed that the reaction is spontaneous with negative ?G. Effect of some relevant interferents on the detection of pesticides has been investigated.

Azab, Hassan A.; Duerkop, Axel; Saad, E. M.; Awad, F. K.; Abd El Aal, R. M.; Kamel, Rasha M.

2012-11-01

270

A novel luminescent terbium-3-carboxycoumarin probe for time-resolved fluorescence sensing of pesticides methomyl, aldicarb and prometryne.  

PubMed

The luminescence arising from lanthanide cations offers several advantages over organic fluorescent molecules: sharp, distinctive emission bands allow for easy resolution between multiple lanthanide signals; long emission lifetimes (?s-ms) make them excellent candidates for time-resolved measurements; and high resistance to photo bleaching allow for long or repeated experiments. A time-resolved (gated) luminescence-based method for determination of pesticides methomyl, aldicarb and prometryne in microtiterplate format using the long-lived terbium-3-carboxycoumarin in 1:3 metal:ligand ratio has been developed. The limit of detection is 1.20×10(6), 5.19×10(5) and 2.74×10(6)ng L(-1) for methomyl, prometryne and aldicarb, respectively. The quantum yield (QY=0.08) of Tb(III)-3-carboxycoumarin was determined using 3-(2-benzothiazolyl)-7-diethylamino-coumarin (coumarin 6). Stern-volmer studies at different temperatures indicate that collisional quenching dominates for methomyl, aldicarb and prometryne. Binding constants were determined at 303, 308 and 313 K by using Lineweaver-Burk equation. A thermodynamic analysis showed that the reaction is spontaneous with negative ?G. Effect of some relevant interferents on the detection of pesticides has been investigated. PMID:22906968

Azab, Hassan A; Duerkop, Axel; Saad, E M; Awad, F K; Abd El Aal, R M; Kamel, Rasha M

2012-11-01

271

Laguerre-based method for analysis of time-resolved fluorescence data: application to in-vivo characterization and diagnosis of atherosclerotic lesions  

PubMed Central

We report the application of the Laguerre deconvolution technique (LDT) to the analysis of in-vivo time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) data and the diagnosis of atherosclerotic plaques. TR-LIFS measurements were obtained in vivo from normal and atherosclerotic aortas (eight rabbits, 73 areas), and subsequently analyzed using LDT. Spectral and time-resolved features were used to develop four classification algorithms: linear discriminant analysis (LDA), stepwise LDA (SLDA), principal component analysis (PCA), and artificial neural network (ANN). Accurate deconvolution of TR-LIFS in-vivo measurements from normal and atherosclerotic arteries was provided by LDT. The derived Laguerre expansion coefficients reflected changes in the arterial biochemical composition, and provided a means to discriminate lesions rich in macrophages with high sensitivity (>85%) and specificity (>95%). Classification algorithms (SLDA and PCA) using a selected number of features with maximum discriminating power provided the best performance. This study demonstrates the potential of the LDT for in-vivo tissue diagnosis, and specifically for the detection of macrophages infiltration in atherosclerotic lesions, a key marker of plaque vulnerability. PMID:16674179

Jo, Javier A.; Fang, Qiyin; Papaioannou, Thanassis; Baker, J. Dennis; Dorafshar, Amir H.; Reil, Todd; Qiao, Jian-Hua; Fishbein, Michael C.; Freischlag, Julie A.; Marcu, Laura

2007-01-01

272

Communication: Time-resolved fluorescence of highly single crystalline molecular wires of azobenzene  

NASA Astrophysics Data System (ADS)

We report the enhanced fluorescence with the remarkably long lifetime (1.17 ns) in the first excited state (S1) of highly crystalline molecular wires of azobenzene at the excitation wavelength of 467 nm for the first time. This observation suggests that trans-cis photoisomerization through the rotation or inversion mechanism may not be a favorable pathway after excitation to the S1 state in highly single crystalline molecular wires of azobenzene due to the hindered motion within densely packed crystal structure. We also measured the fluorescence lifetime image of a single crystalline molecular wire of azobenzene, indicating that the lifetime was remarkably uniform and that there was only a very minor variation within the crystal.

Jee, Ah-Young; Lee, Yumin; Lee, Minyung; Kim, Myung Hwa

2012-03-01

273

Application of time-resolved fluorescence to the determination of metabolites  

NASA Astrophysics Data System (ADS)

A simple fluorescent methodology for the simultaneous determination of two major metabolites of acetylsalicylic acid - salicylic and gentisic acids - in pharmaceutical preparations and human urine is proposed. Due to the overlapping between the fluorescence spectra of both analytes, the use of the more selective fluorescence decay curves is proposed. Values of dependent instrumental variables affecting the signal-to-noise ratio were fixed with a simplex optimization procedure. A calibration matrix of thirteen standards plus two blank samples was processed using a partial least-squares (PLS) analysis. To assess the goodness of the proposed method, a prediction set of nine synthetic samples was analyzed, obtaining recovery percentages between 95% and 106%. Limits of detection, calculated by means of a new criterion, were 3.49 ?g L-1 and 1.66 ?g L-1 for salicylic and gentisic acids, respectively. The method was also tested in three pharmaceutical preparations containing salicylic acid, obtaining recovery percentages close to 100%. Finally, the simultaneous determination of both analytes in human urine samples was successfully carried out by the PLS-analysis of a matrix of thirteen standards plus five analyte blanks. Although spectra of analytes and urine overlap strongly, no extraction method neither prior separation of the analytes were needed.

Murillo Pulgarín, J. A.; Alañón Molina, A.; Martínez Ferreras, F.

2014-07-01

274

Gene expression analysis with an integrated CMOS microarray by time-resolved fluorescence detection  

PubMed Central

DNA microarrays have proven extraordinarily powerful for differential expression studies across thousands of genes in a single experiment. Microarrays also have the potential for clinical applications, including the detection of infectious and immunological diseases and cancer, if they can be rendered both reliable and cost-effective. Here we report the first practical application of an active microarray based on integrated circuit technology, completely obviating the need for external measurement instrumentation while employing protocols compatible with traditional fluorescence-based surface bioassays. In a gene-expression biodosimetry study, we determine the differential activity of genes from leucocytes in irradiated human blood. Quantum dots are used as fluorescence labels to realize filterless, time-gated fluorescence detection on an active complementary metal-oxide-semiconductor (CMOS) microarray with 100-pM sensitivity. Improvements in surface chemistry should allow sensitivities that approach the microarray hardware limit of less than 10 pM. Techniques for covalent attachment of DNA capture strands to the CMOS active microarrays allow integrated sensors to be placed in immediate proximity to hybridized analyte strands, maximizing photon collection efficiencies. PMID:20392628

Huang, Ta-chien D.; Paul, Sunirmal; Gong, Ping; Levicky, Rastislav; Kymissis, John; Amundson, Sally A.; Shepard, Kenneth L.

2010-01-01

275

Time-resolved imaging system for fluorescence-guided surgery with lifetime imaging capability  

NASA Astrophysics Data System (ADS)

We present a single-photon camera for fluorescence imaging, with a time resolution better than 100ps, capable of providing both intensity and lifetime images. the camera was fabricated in standard CMOS technology. With this FluoCam we show the possibility to study sub-nanosecond fluorescence mechanisms. The FluoCam was used to characterize a near-infrared probe, indocyanine green, conjugated with multimeric cyclic pentapeptide (cRGD). The fluorescent probe-conjugated was used to target and mark tumors with better specificity, in particular aiming at targeting the integrins ?v?3 and ?v?5. As a first step towards clinical studies, preliminary results obtained in-vivo are presented. The first envisioned clinical application would be image-guided surgical oncology to help the surgeon to remove tumor tissue by a better discrimination from normal tissues and also to improve the detection of metastatic lymph nodes. A further application could be the in-vivo determination of the ?v?3 and ?v?5 targets to select patients for therapy with RGD chemotherapy conjugates.

Powolny, F.; Homicsko, K.; Sinisi, R.; Bruschini, Claudio E.; Grigoriev, E.; Homulle, H.; Prior, John O.; Hanahan, D.; Dubikovskaya, E.; Charbon, E.

2014-05-01

276

Liquid-solid phase transition of benzene under shock compression studied by nanosecond time-resolved nonlinear Raman spectroscopy  

NASA Astrophysics Data System (ADS)

Phase transition of benzene has been studied under laser-shock compression up to 4.2 GPa by using nanosecond time-resolved nonlinear Raman spectroscopy. The shock wave is generated by irradiation of 10-ns pulsed laser beam on the plasma confinement target and its pressure is estimated from a particle velocity, which is measured by a velocity interferometer system. Higher frequency shifts in the ring- breathing mode of benzene are observed under shock compression. The shift at pressures below 3.0 GPa agrees well with that of liquid benzene under static compression. A metastable supercooled state and a liquid-solid phase transition are observed at shock pressures above 3.0 GPa. Time-resolved Raman spectra reveal that the liquid state is initially a metastable state and rapidly transforms to the solid state under shock compression at 4.2 GPa. Rapid nucleation and growth occurs within 20 ns.

Nakamura, Kazutaka

2005-07-01

277

Using Time-Resolved Fluorescence to Measure Serum Venom-Specific IgE and IgG  

PubMed Central

We adapted DELFIA™ (dissociation-enhanced lanthanide fluoroimmunoassay), a time resolved fluorescence method, to quantitate whole venom specific and allergenic peptide-specific IgE (sIgE), sIgG1 and sIgG4 in serum from people clinically allergic to Australian native ant venoms, of which the predominant cause of allergy is jack jumper ant venom (JJAV). Intra-assay CV was 6.3% and inter-assay CV was 13.7% for JJAV sIgE. DELFIA and Phadia CAP JJAV sIgE results correlated well and had similar sensitivity and specificity for the detection of JJAV sIgE against intradermal skin testing as the gold standard. DELFIA was easily adapted for detecting sIgE to a panel of other native ant venoms. PMID:21304970

van Eeden, Pauline E.; Wiese, Michael D.; Aulfrey, Susan; Hales, Belinda J.; Stone, Shelley F.; Brown, Simon G. A.

2011-01-01

278

Streptavidin-based macromolecular complex labeled with a europium chelator suitable for time-resolved fluorescence immunoassay applications  

SciTech Connect

The authors found that when they incubated SA-TG-(BCPDA){sub 150} (a conjugate of streptavidin (SA) with thyroglobulin (TG), which is labeled with the europium chelator 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA)), with BCPDA-labeled thyroglobulin (TG-(BCPDA){sub 150}), in the presence of a suitable amount of Eu{sup 3+}, a new macromolecular complex is formed with a molecular weight of about 3 {times} 10{sup 6}. This macromolecular complex is stable and can be used as a streptavidin-based universal detection system for devising time-resolved fluorescence immunoassays. This new reagent yields an 8-26-fold improvement in the detection limits of seven different immunofluorometric assays in comparison to the reagent SA-TG-(BCPDA){sub 150}, which they have recently described.

Morton, R.C. (CyberFluor Inc., Toronto, Ontario (Canada)); Diamandis, E.P. (CyberFluor Inc., Toronto, Ontario (Canada) Univ. of Toronto, Ontario (Canada))

1990-09-01

279

Time resolved laser-induced fluorescence of electrosprayed ions confined in a linear quadrupole trap  

SciTech Connect

We have designed and constructed a linear quadrupole ion trap for the measurement of laser-induced fluorescence (LIF) of mass selected gas-phase ions produced by electrospray ionization. The instrument consists of a simple electrospray source, radiofrequency octopole guide, a dc quadrupole bender, a quadrupole mass filter, the linear quadrupole trap (which is equipped with optics for LIF collection and a channeltron ion detector), and several multielement focusing lenses. With this instrument, the LIF decay lifetime of gas-phase Rhodamine 640 radical cations is determined for the first time.

Friedrich, Jochen; Fu Jinmei; Hendrickson, Christopher L.; Marshall, Alan G.; Wang Yisheng [Ion Cyclotron Resonance Program, National High Magnetic Field Laboratory, Florida State University, 1800 East Paul Dirac Drive, Tallahassee, Florida 32310-4005 (United States); Institute of Atomic and Molecular Sciences, Academia Sinica, P. O. Box 23-166, Taipei, Taiwan (China)

2004-11-01

280

Dynamics of Nuclear Receptor Helix-12 Switch of Transcription Activation by Modeling Time-Resolved Fluorescence Anisotropy Decays  

PubMed Central

Nuclear hormone receptors (NRs) are major targets for pharmaceutical development. Many experiments demonstrate that their C-terminal Helix (H12) is more flexible in the ligand-binding domains (LBDs) without ligand, this increased mobility being correlated with transcription repression and human diseases. Crystal structures have been obtained in which the H12 is extended, suggesting the possibility of large amplitude H12 motions in solution. However, these structures were interpreted as possible crystallographic artifacts, and thus the microscopic nature of H12 movements is not well known. To bridge the gap between experiments and molecular models and provide a definitive picture of H12 motions in solution, extensive molecular dynamics simulations of the peroxisome proliferator-activated receptor-? LBD, in which the H12 was bound to a fluorescent probe, were performed. A direct comparison of the modeled anisotropy decays to time-resolved fluorescence anisotropy experiments was obtained. It is shown that the decay rates are dependent on the interactions of the probe with the surface of the protein, and display little correlation with the flexibility of the H12. Nevertheless, for the probe to interact with the surface of the LBD, the H12 must be folded over the body of the LBD. Therefore, the molecular mobility of the H12 should preserve the globularity of the LBD, so that ligand binding and dissociation occur by diffusion through the surface of a compact receptor. These results advance the comprehension of both ligand-bound and ligand-free receptor structures in solution, and also guide the interpretation of time-resolved anisotropy decays from a molecular perspective, particularly by the use of simulations. PMID:24094408

Batista, Mariana R.B.; Martínez, Leandro

2013-01-01

281

Time-resolved optical writing on a photosensitive and fluorescent polymer film  

NASA Astrophysics Data System (ADS)

Recently a melt-processed blend of 1,4-bis(?-cyano-4-octadecyloxystyryl)-2,5-dimethoxybenzene (C18-RG) dye and polyethylene terephthalate glycol (PETG) has been demonstrated as a promising 3-dimentional optical data storage (ODS) medium 1. ODS in this novel system relies on the laser-induced switching of the aggregation state of the excimerforming fluorescent dye in the inert host polymer. Here we investigate the mechanism and the time scales involved in the writing process. The optical writing was realized by the laser-induced localized excimer to monomer conversion and was characterized by the emergence of the monomer fluorescence. We obtained the dependence of the excimer to monomer conversion on the writing time. Our result indicates that the effective optical writing time is controlled by heating and cooling time of the host polymer and the excimer-to-monomer conversion time. The effective laser writing time, under the specific writing conditions employed in our experiments, is on the order of 10 ms.

Pan, Z.; Akrobetu, R.; Lott, J.; Ryan, C.; Saini, A.; Shan, J.; Mu, R.; Singer, K. D.; Weder, C.; Morgan, S. H.

2011-10-01

282

Ferryl intermediates of catalase captured by time-resolved Weissenberg crystallography and UV-VIS spectroscopy.  

PubMed

Various enzymes use semi-stable ferryl intermediates and free radicals during their catalytic cycle, amongst them haem catalases. Structures for two transient intermediates (compounds I and II) of the NADPH-dependent catalase from Proteus mirabilis (PMC) have been determined by time-resolved X-ray crystallography and single crystal microspectrophotometry. The results show the formation and transformation of the ferryl group in the haem, and the unexpected binding of an anion during this reaction at a site distant from the haem. PMID:8901874

Gouet, P; Jouve, H M; Williams, P A; Andersson, I; Andreoletti, P; Nussaume, L; Hajdu, J

1996-11-01

283

Laser spectroscopy of SrH. Time-resolved measurements of the A 2? state  

NASA Astrophysics Data System (ADS)

In the near-infrared and red wavelength region around 680-760 nm, the spectrum of the A2?- X2? transition of gaseous SrH appears. By applying laser spectroscopic methods, a time-resolved experiment has been performed on the low-lying A 2? state of strontium hdyride. No local influences on the A 2? state were found due to interactions with the low-lying A' 2? state, as in the case of the BaH molecule. The following zero-pressure lifetime was obtained for the SrH molecule: ?( A2? 1/2 (?' = 0)) = 33.8 ± 1.9 ns.

Berg, L.-E.; Ekvall, K.; Hishikawa, A.; Kelly, S.; McGuinness, C.

1996-06-01

284

Solving the structure of reaction intermediates by time-resolved synchrotron x-ray absorption spectroscopy  

E-print Network

by perfect turnover and unique reaction pathways.5 Vibrational spectroscopies,6 magnetic circular dichroism,7 detection, vibrational spectroscopies use a feature identification "fin- gerprinting" approach and thus

Frenkel, Anatoly

285

Cellular Oxygen and Nutrient Sensing in Microgravity Using Time-Resolved Fluorescence Microscopy  

NASA Technical Reports Server (NTRS)

Oxygen and nutrient sensing is fundamental to the understanding of cell growth and metabolism. This requires identification of optical probes and suitable detection technology without complex calibration procedures. Under this project Microcosm developed an experimental technique that allows for simultaneous imaging of intra- and inter-cellular events. The technique consists of frequency-domain Fluorescence Lifetime Imaging Microscopy (FLIM), a set of identified oxygen and pH probes, and methods for fabrication of microsensors. Specifications for electronic and optical components of FLIM instrumentation are provided. Hardware and software were developed for data acquisition and analysis. Principles, procedures, and representative images are demonstrated. Suitable lifetime sensitive oxygen, pH, and glucose probes for intra- and extra-cellular measurements of analyte concentrations have been identified and tested. Lifetime sensing and imaging have been performed using PBS buffer, culture media, and yeast cells as a model systems. Spectral specifications, calibration curves, and probes availability are also provided in the report.

Szmacinski, Henryk

2003-01-01

286

Architecture of polyglutamine-containing fibrils from time-resolved fluorescence decay.  

PubMed

The disease risk and age of onset of Huntington disease (HD) and nine other repeat disorders strongly depend on the expansion of CAG repeats encoding consecutive polyglutamines (polyQ) in the corresponding disease protein. PolyQ length-dependent misfolding and aggregation are the hallmarks of CAG pathologies. Despite intense effort, the overall structure of these aggregates remains poorly understood. Here, we used sensitive time-dependent fluorescent decay measurements to assess the architecture of mature fibrils of huntingtin (Htt) exon 1 implicated in HD pathology. Varying the position of the fluorescent labels in the Htt monomer with expanded 51Q (Htt51Q) and using structural models of putative fibril structures, we generated distance distributions between donors and acceptors covering all possible distances between the monomers or monomer dimensions within the polyQ amyloid fibril. Using Monte Carlo simulations, we systematically scanned all possible monomer conformations that fit the experimentally measured decay times. Monomers with four-stranded 51Q stretches organized into five-layered ?-sheets with alternating N termini of the monomers perpendicular to the fibril axis gave the best fit to our data. Alternatively, the core structure of the polyQ fibrils might also be a zipper layer with antiparallel four-stranded stretches as this structure showed the next best fit. All other remaining arrangements are clearly excluded by the data. Furthermore, the assessed dimensions of the polyQ stretch of each monomer provide structural evidence for the observed polyQ length threshold in HD pathology. Our approach can be used to validate the effect of pharmacological substances that inhibit or alter amyloid growth and structure. PMID:25092288

Röthlein, Christoph; Miettinen, Markus S; Borwankar, Tejas; Bürger, Jörg; Mielke, Thorsten; Kumke, Michael U; Ignatova, Zoya

2014-09-26

287

Full Genotyping of a Highly Polymorphic Human Gene Trait by Time-Resolved Fluorescence Resonance Energy Transfer  

PubMed Central

The ability of detecting the subtle variations occurring, among different individuals, within specific DNA sequences encompassed in highly polymorphic genes discloses new applications in genomics and diagnostics. DQB1 is a gene of the HLA-II DQ locus of the Human Leukocyte Antigens (HLA) system. The polymorphisms of the trait of the DQB1 gene including codons 52–57 modulate the susceptibility to a number of severe pathologies. Moreover, the donor-receiver tissue compatibility in bone marrow transplantations is routinely assessed through crossed genotyping of DQB and DQA. For the above reasons, the development of rapid, reliable and cost-effective typing technologies of DQB1 in general, and more specifically of the codons 52–57, is a relevant although challenging task. Quantitative assessment of the fluorescence resonance energy transfer (FRET) efficiency between chromophores labelling the opposite ends of gene-specific oligonucleotide probes has proven to be a powerful tool to type DNA polymorphisms with single-nucleotide resolution. The FRET efficiency can be most conveniently quantified by applying a time-resolved fluorescence analysis methodology, i.e. time-correlated single-photon counting, which allows working on very diluted template specimens and in the presence of fluorescent contaminants. Here we present a full in-vitro characterization of the fluorescence responses of two probes when hybridized to oligonucleotide mixtures mimicking all the possible genotypes of the codons 52–57 trait of DQB1 (8 homozygous and 28 heterozygous). We show that each genotype can be effectively tagged by the combination of the fluorescence decay constants extrapolated from the data obtained with such probes. PMID:25215592

Totè, Edoardo; Lamperti, Marco; Bondani, Maria; Salerno, Domenico; Cassina, Valeria; Nardo, Luca

2014-01-01

288

TIME-RESOLVED AND BACKWARD-WAVE OSCILLATOR SUBMILLIMETRE SPECTROSCOPY OF SOME  

E-print Network

a time-domain terahertz transmission spectroscopy in the range 3-80 cm-1 . A good agreement of both data-wave oscillator spectroscopy was applied to the transmission measurements of the complex dielectric spectra: the time-domain terahertz spectroscopy (TDTS) and backward wave oscillator (BWO) spectroscopy, and we

Ku?el, Petr

289

Maximum entropy analysis of data simulations and practical aspects of time-resolved fluorescence measurements in the study of molecular interactions  

NASA Astrophysics Data System (ADS)

Time-resolved fluorescence spectroscopy and microscopy is increasingly used to probe molecular interactions and dynamics events in vitro and in vivo. We point out some pitfalls to avoid in the data acquisition procedure using time correlated single photon counting. A good accuracy in fluorescence decay measurements is not only linked to the counts in the peak channel but also to the statistics at the end of the curve. A too narrow time interval between successive excitation leads to an overlap of decays, and that should be taken into account in data analysis. The counting rate in the peak channel of the excitation profile should be close to the one of the fluorescence decay. Many distributions of lifetimes can fit an incomplete or noisy data set to satisfying precisions corresponding to close values of ?2. The maximum entropy principle is appropriate to distinguish among these in a consistent way. It is also shown that encoding a prior knowledge about the system study dramatically improves the quality of the recovered distribution, particularly in case of a set of discrete components. Based on simulated noisy quantify maximum entropy method (MEM) data analysis, we propose a simple strategy for estimating the quality of information and inferences we can draw from experimental results.

Henry, Etienne; Deprez, Eric; Brochon, Jean-Claude

2014-12-01

290

TIME-RESOLVED ULTRAVIOLET SPECTROSCOPY OF THE M-DWARF GJ 876 EXOPLANETARY SYSTEM  

SciTech Connect

Extrasolar planets orbiting M-stars may represent our best chance to discover habitable worlds in the coming decade. The ultraviolet spectrum incident upon both Earth-like and Jovian planets is critically important for proper modeling of their atmospheric heating and chemistry. In order to provide more realistic inputs for atmospheric models of planets orbiting low-mass stars, we present new near- and far-ultraviolet (NUV and FUV) spectroscopy of the M-dwarf exoplanet host GJ 876 (M4V). Using the COS and STIS spectrographs on board the Hubble Space Telescope, we have measured the 1150-3140 A spectrum of GJ 876. We have reconstructed the stellar H I Ly{alpha} emission line profile, and find that the integrated Ly{alpha} flux is roughly equal to the rest of the integrated flux (1150-1210 A + 1220-3140 A) in the entire ultraviolet bandpass (F(Ly{alpha})/F(FUV+NUV) Almost-Equal-To 0.7). This ratio is {approx}2500 Multiplication-Sign greater than the solar value. We describe the ultraviolet line spectrum and report surprisingly strong fluorescent emission from hot H{sub 2} (T(H{sub 2}) > 2000 K). We show the light curve of a chromospheric + transition region flare observed in several far-UV emission lines, with flare/quiescent flux ratios {>=}10. The strong FUV radiation field of an M-star (and specifically Ly{alpha}) is important for determining the abundance of O{sub 2}-and the formation of biomarkers-in the lower atmospheres of Earth-like planets in the habitable zones of low-mass stars.

France, Kevin; Froning, Cynthia S. [Center for Astrophysics and Space Astronomy, University of Colorado, 389 UCB, Boulder, CO 80309 (United States); Linsky, Jeffrey L. [JILA, University of Colorado and NIST, 440 UCB, Boulder, CO 80309 (United States); Tian, Feng [Laboratory for Atmospheric and Space Physics, University of Colorado, Boulder, CO 80309 (United States); Roberge, Aki, E-mail: kevin.france@colorado.edu [Exoplanets and Stellar Astrophysics Laboratory, NASA Goddard Space Flight Center, Greenbelt, MD 20771 (United States)

2012-05-10

291

Time-Resolved Ultraviolet Spectroscopy of The M-Dwarf GJ 876 Exoplanetary System  

NASA Technical Reports Server (NTRS)

Extrasolar planets orbiting M-stars may represent our best chance to discover habitable worlds in the coming decade. The ultraviolet spectrum incident upon both Earth-like and Jovian planets is critically important for proper modeling of their atmospheric heating and chemistry. In order to provide more realistic inputs for atmospheric models of planets orbiting low-mass stars, we present new near- and far-ultraviolet (NUV and FUV) spectroscopy of the M-dwarf exoplanet host GJ 876 (M4V). Using the COS and STIS spectrographs on board the Hubble Space Telescope, we have measured the 1150-3140 A spectrum of GJ 876. We have reconstructed the stellar H1 Ly alpha emission line profile, and find that the integrated Ly alpha flux is roughly equal to the rest of the integrated flux (1150-1210 A + 1220-3140 A) in the entire ultraviolet bandpass (F(Ly alpha)/F(FUV+NUV) equals approximately 0.7). This ratio is approximately 2500x greater than the solar value. We describe the ultraviolet line spectrum and report surprisingly strong fluorescent emission from hot H2 (T(H2) greater than 2000 K). We show the light curve of a chromospheric + transition region flare observed in several far-UV emission lines, with flare/quiescent flux ratios greater than or equal to 10. The strong FUV radiation field of an M-star (and specifically Ly alpha) is important for determining the abundance of O2--and the formation of biomarkers-in the lower atmospheres of Earth-like planets in the habitable zones of low-mass stars.

France, Kevin; Linsky, Jeffrey L.; Tian, Feng; Froning, Cynthia S.; Roberge, Aki

2012-01-01

292

Molecular analysis of mitotic chromosome condensation using a quantitative time-resolved fluorescence microscopy assay  

PubMed Central

Chromosomes condense during mitotic entry to facilitate their segregation. Condensation is typically assayed in fixed preparations, limiting analysis of contributing factors. Here, we describe a quantitative method to monitor condensation kinetics in living cells expressing GFP fused to a core histone. We demonstrate the utility of this method by using it to analyze the molecular requirements for the condensation of holocentric chromosomes during the first division of the Caenorhabditis elegans embryo. In control embryos, the fluorescence intensity distribution for nuclear GFP:histone changes during two distinct time intervals separated by a plateau phase. During the first interval, primary condensation converts diffuse chromatin into discrete linear chromosomes. After the plateau, secondary condensation compacts the curvilinear chromosomes to form shorter bar-shaped structures. We quantitatively compared the consequences on this characteristic profile of depleting the condensin complex, the mitosis-specific histone H3 kinase Aurora B, the centromeric histone CENP-A, and CENP-C, a conserved protein required for kinetochore assembly. Both condensin and CENP-A play critical but distinct roles in primary condensation. In contrast, depletion of CENP-C slows but does not prevent primary condensation. Finally, Aurora B inhibition has no effect on primary condensation, but slightly delays secondary condensation. These results provide insights into the process of condensation, help resolve apparent contradictions from prior studies, and indicate that CENP-A chromatin has an intrinsic role in the condensation of holocentric chromosomes that is independent of its requirement for kinetochore assembly. PMID:17005720

Maddox, Paul S.; Portier, Nathan; Desai, Arshad; Oegema, Karen

2006-01-01

293

Time-resolved measurements of the fluorescence of rhodamine B on semiconductor and glass surfaces  

SciTech Connect

Time-correlated single-photon counting was used to investigate the fluorescence decay of rhodamine B at semiconductor-water and glass-water interfaces. All decays could be fit by the sum of two exponentials with lifetimes tau/sub 1/ (short) and tau/sub 2/ (long). At very low dye concentrations in solution (10/sup -7/ M) tau/sup 1/ = 0.41 ns and tau/sub 2/ = 1.4 ns for semiconductors (tin oxide and indium oxide), and tau/sub 1/ = 0.68 ns and tau/sub 2/ = 2.6 ns for glass. Experiments on dry surfaces prepared with 10/sup -7/ M rhodamine B solutions gave tau/sub 1/ values similar to those obtained for surface-solution samples, but tau/sub 2/ = 3.1 ns with both glass and semiconductor surfaces. Molecules which are capable of excited-state electron transfer to the semiconductor are assigned to glass, and tau/sub 2/ otherwise. Progressively shorter tau/sub 1/ values measured for SnO/sub 2/ with increasing dye concentrations are attributed to the effect of energy-transfer quenching. Treatment of SnO/sub 2/ surfaces in 1 M KCl solutions at pH 13 (reported to reduce the photocurrent yield by two orders of magnitude) had no effect on tau/sub 1/. No changes in tau/sub 1/ which could be attributed to enhanced electron transfer were found with supersensitizers (thiourea and hydroquinone), which are known to increase the photocurrent yield by as much as one order of magnitude. 27 references, 4 figures, 1 table.

Liang, Y.; Goncalves, A.M.P.

1985-07-18

294

Femtosecond time-resolved x-ray photoelectron spectroscopy studies of charge transfer dynamics in novel photovoltaic systems  

NASA Astrophysics Data System (ADS)

Interfacial charge transfer dynamics in dye-sensitized semiconductor films are studied by femtosecond time-resolved x-ray photoelectron spectroscopy. The experiments performed at the Linac Coherent Light Source demonstrate the potential of time-domain inner-shell photoionization studies to monitor charge migration in complex interfacial systems with femtosecond time resolution, chemical sensitivity and atomic specificity. Using this new technique, the transient oxidation state of N3 dye molecules adsorbed to nanocrystalline ZnO is monitored with unprecedented site selectivity, providing an upper bound for the interfacial electron-hole recombination rate.

Gessner, Oliver; Shavorskiy, Andrey; Siefermann, Katrin; Slaughter, Daniel; Sturm, Felix; Weise, Fabian; Strader, Matthew; Cho, Hana; Lin, Ming-Fu; Wright, Travis; Guo, Jinghua; Bluhm, Hendrik; Schoenlein, Robert; Belkacem, Ali; Neumark, Daniel; Leone, Stephen; Cordones, Amy; Vura-Weis, Josh; Gul, Sheraz; Zhang, Jin; Nordlund, Dennis; Ogasawara, Hirohito; Nilsson, Anders; Beye, Martin; Huse, Nils

2012-06-01

295

Probing Reaction Dynamics of Transition-Metal Complexes in Solution via Time-Resolved X-ray Spectroscopy  

SciTech Connect

We report measurements of the photo-induced Fe(II) spin crossover reaction dynamics in solution via time-resolved x-ray absorption spectroscopy. EXAFS measurements reveal that the iron?nitrogen bond lengthens by 0.21+-0.03 Angstrom in the high-spin transient excited state relative to the ground state. XANES measurements at the Fe L-edge show directly the influence of the structural change on the ligand-field splitting of the Fe(II) 3d orbitals associated with the spin transition.

Huse, Nils; Khalil, Munira; Kim, Tae Kyu; Smeigh, Amanda L.; Jamula, Lindsey; McCusker, James K.; Schoenlein, Robert W.

2009-05-24

296

Time-Resolved X-ray Spectroscopy of the Massive Binary delta Ori  

NASA Astrophysics Data System (ADS)

We have obtained 500 ks of Chandra HETG observations of the massive binary delta Ori (O9.5II+unseen companion), one of the fundamental calibrators of the mass-luminosity-radius relation in the upper HR diagram. The program is intended to map the emission line parameters as the secondary moves through the wind of the primary star. Custom extraction techniques have been developed to create 12 time-resolved 40 ks spectra from these observations, each of which is properly calibrated for time and temperature effects. Emission line fluxes for these time slice spectra are presented, as well as phase analysis of the variability of the fluxes. We discuss the interpretation of the resulting data, such as colliding winds and occultation of various temperature regimes of the primary wind by the secondary.

Nichols, Joy S.; Naze, Y.; Corcoran, M. F.; Pollock, A.; Moffat, A. F.; Ignace, R.; Waldron, W. L.; Evans, N. R.

2014-01-01

297

Limitations of Time-Resolved Fluorescence Suggested by Molecular Simulations: Assessing the Dynamics of T cell Receptor Binding Loops  

PubMed Central

Time-resolved fluorescence anisotropy (TRFA) has a rich history in evaluating protein dynamics. Yet as often employed, TRFA assumes that the motional properties of a covalently tethered fluorescent probe accurately portray the motional properties of the protein backbone at the probe attachment site. In an extensive survey using TRFA to study the dynamics of the binding loops of a ?? T cell receptor, we observed multiple discrepancies between the TRFA data and previously published results that led us to question this assumption. We thus simulated several of the experimentally probed systems using a protocol that permitted accurate determination of probe and protein time correlation functions. We found excellent agreement in the decays of the experimental and simulated correlation functions. However, the motional properties of the probe were poorly correlated with those of the backbone of both the labeled and unlabeled protein. Our results warrant caution in the interpretation of TRFA data and suggest further studies to ascertain the extent to which probe dynamics reflect those of the protein backbone. Meanwhile, the agreement between experiment and computation validates the use of molecular dynamics simulations as an accurate tool for exploring the molecular motion of T cell receptors and their binding loops. PMID:23260055

Scott, Daniel R.; Vardeman, Charles F.; Corcelli, Steven A.; Baker, Brian M.

2012-01-01

298

Identifiability analysis of rotational diffusion tensor and electronic transition moments measured in time-resolved fluorescence depolarization experiment  

NASA Astrophysics Data System (ADS)

The subject of this paper is studies of the deterministic identifiability of molecular parameters, such as rotational diffusion tensor components and orientation of electronic transition moments, resulting from the time-resolved fluorescence anisotropy experiment. In the most general case considered, a pair of perpendicularly polarized emissions enables the unique determination of all the rotational diffusion tensor's principal components. The influence of the tensor's symmetry and the associated degeneration of its eigenvalues on the identifiability of the electronic transitions moments is systematically investigated. The analysis reveals that independently of the rotational diffusion tensor's symmetry, the transition moments involved in photoselection and emission processes cannot be uniquely identified without a priori information about their mutual orientation or their orientation with respect to the principal axes of the tensor. Moreover, it is shown that increasing the symmetry of the rotational diffusion tensor deteriorates the degree of the transition moments identifiability. To obtain these results analytically, a novel approach to solve bilinear system of equations for Markov parameters is applied. The effect of the additional information, obtained from fluorescence measurements for different molecular mobilities, to improve the identifiability at various levels of analysis is shown. The effectiveness and reliability of the target analysis method for experimental determination of the molecular parameters is also discussed.

Szubiakowski, Jacek P.

2014-06-01

299

A time-resolved fluorescence immunoassay for the ultrasensitive determination of diethylstilbestrol based on the double-codified gold nanoparticles.  

PubMed

An ultrasensitive and selective method is presented for the determination of diethylstilbestrol (DES) using time-resolved fluorescence immunoassay (TRFIA) based on double-codified gold nanoparticles (DC-AuNPs). In this system, the DC-AuNPs, that are gold nanoparticles (AuNPs) modified with anti-DES antibody and SH-dsDNA-biotin, was regarded as signal amplifier. A competitive immunoreaction was performed on polystyrene microtitration plates, where the DES compete with the immobilized DES-ovalbumin on polystyrene microtitration plates to bind to anti-DES antibodies on DC-AuNPs, and the europium(III)-labeled streptavidin was added to link to the SH-dsDNA-biotin as a tracer. Fluorescence signal was amplified via the AuNPs and the biotin-streptavidin double amplification systems. Under the optimized condition, DES can be quantified by TRFIA. The linear range and the limit of detection of DES were 1.0×10(-6)-10ngmL(-1) and 0.4fgmL(-1), respectively. This method was applied to determine DES in beef sample, with the recoveries ranging from 88% to 105%. PMID:25091151

Wang, Longjun; Zhang, Yuanfu; Liu, Guofu; Zhang, Chunyan; Wang, Shuhao

2014-11-01

300

A new simple cell-based homogeneous time-resolved fluorescence QRET technique for receptor-ligand interaction screening.  

PubMed

In this article, a single-label separation-free fluorescence technique is presented as a potential screening method for cell-based receptor antagonists and agonists.The time-resolved fluorescence technique, quenching resonance energy transfer (QRET), relies on a single-labeled binding partner in combination with a soluble quencher. The quencher efficiently suppresses the luminescence of the unbound labeled ligand, whereas the luminescence of the bound fraction is not affected. This approach allows the development of cell-based screening assays in a simple and cost-effective manner. The authors have applied the technique to the screening of beta(2)-adrenoreceptor (beta(2)AR) antagonists and agonists in intact human embryonic kidney HEK293(i) cells overexpressing human beta(2)-adrenergic receptors. Two antagonists (propranolol, alprenolol) and 2 agonists (metaproterenol, terbutaline) for beta(2)AR were investigated in a displacement assay using europium(III)-labeled pindolol ligand. The assay Z' values ranged from 0.68 to 0.78, the coefficient of variation was less than 10%, and the K(i) values were 19 nM for propranolol and alprenolol and 14 and 5.9 microM for metaproterenol and terbutaline, respectively. The QRET technique with beta(2)AR was also applied to LOPAC compound library screening, yielding nearly error-free recognition of known binders. This simple and cost-effective technique can be readily adapted to laboratory and industrial-scale screening. PMID:19684287

Härmä, Harri; Rozwandowicz-Jansen, Anita; Martikkala, Eija; Frang, Heini; Hemmilä, Ilkka; Sahlberg, Niko; Fey, Vidal; Perälä, Merja; Hänninen, Pekka

2009-09-01

301

Femtosecond broadband time-resolved fluorescence and transient absorption study of the intramolecular charge transfer state of methyl 4-dimethylaminobenzoate.  

PubMed

A combined application of femtosecond broadband time-resolved fluorescence (fs-TRF), fluorescence anisotropy (fs-TRFA) and fs to microsecond (?s) transient absorption (TA) have been used to probe directly the dynamics, nature, formation and decay paths of the singlet intramolecular charge transfer ((1)ICT) state of methyl 4-dimethylaminobenzoate (1a) in acetonitrile. The result reveals explicit evidence for a common electronic origin (the L(a) nature) of the (1)ICT state and its precursor the locally excited ((1)LE) state to account jointly for the dual florescence known to this system. It also shows that the ICT reaction from the (1)LE to (1)ICT state occurs with time constant of ~0.8 ps and the (1)ICT state formed decays with a ~1.9 ns time constant leading mainly to a ??* natured triplet state ((3)T(1)). The (3)T(1) then relaxes with a ~4 ?s lifetime under deoxygenated condition resulting in full recovery of the ground state (S(0)). As a case study, this work contributes novel experimental data for improved understanding of the mechanism of ICT reaction; it also reveals a distinct deactivation pattern for this prototype para-amino substituted aromatic carbonyl compound in acetonitrile. PMID:21847483

Chan, Chris Tsz-Leung; Cheng, Chopen Chan-Wut; Ho, Keith Yat-Fung; Kwok, Wai-Ming

2011-09-28

302

Time-resolved fluorescence resonance energy transfer kinase assays using physiological protein substrates: Applications of terbium–fluorescein and terbium–green fluorescent protein fluorescence resonance energy transfer pairs  

Microsoft Academic Search

Fluorescence-based kinase assays using peptide substrates are an established format for high-throughput screening and profiling of kinases. Among fluorescence-based formats, time-resolved fluorescence resonance energy transfer (TR-FRET) using a lanthanide donor species has advantages over other fluorescent formats in being resistant to many types of optical interference such as autofluorescent compounds, scattered light from precipitated compounds, or colored compounds that absorb

Steven M. Riddle; Kevin L. Vedvik; George T. Hanson; Kurt W. Vogel

2006-01-01

303

Time-resolved spectroscopy of the excited singlet states of tirapazamine and desoxytirapazamine.  

PubMed

Laser flash photolysis (LFP, 400 nm excitation) of the anti-cancer drug tirapazamine (TPZ) in acetonitrile produces the singlet excited-state S1 with lambda(max) = 544 nm. The lifetime of this state is 130 ps, in good agreement with the reported fluorescence lifetime. The excited state is reduced to the corresponding radical anion by KSCN or KI. The spectrum of the radical anion is in good agreement with previously reported pulse radiolysis studies and time-dependent density functional theory (TD-DFT) calculations. LFP of desoxytirapazamine (dTPZ) also produces the first excited singlet state, S1. The fluorescence quantum yield and lifetime (5.4 ns) of the dTPZ singlet excited state are both much greater than the corresponding values of TPZ. This is explained by DFT calculations that predict that cyclization of TPZ to form an oxaziridine is thermodynamically facile but that cyclization of dTPZ to form an oxadiaziridine is not. Thus, the S1 state of TPZ has a short lifetime and low fluorescence quantum yield due to ready cyclization whereas the cyclization of the S1 state of dTPZ is unimportant and does not limit either the fluorescence quantum yield or the fluorescence lifetime. This conclusion is confirmed by studies of dTPZ', an isomer of dTPZ containing the C=N-O moiety which has a low quantum yield and short fluorescence lifetime similar to that of TPZ. PMID:16833470

Shi, Xiaofeng; Poole, James S; Emenike, Ijeoma; Burdzinski, Gotard; Platz, Matthew S

2005-03-01

304

Time-Resolved Spectroscopy Measurements of Self Emission From Shocked Liquid Deuterium  

NASA Astrophysics Data System (ADS)

We are studying the deuterium equation of state using shock waves created by the impact of a flyer plate launched by the 20 MA current generated by the Z accelerator. The flyer strikes a cryogenic sample consisting of an Al pusher, a D2 cell, and sapphire window. The shock pressure in the D2 reaches 0.2-0.7 Mbar with a 10-30 nsec duration. The D2 cell consists of 300 and 600 micron thick steps with a 6 mm total diameter. The shock velocity is measured with a suite of VISAR and active shock breakout diagnostics. The self emission from the D2 regions is measured with four time-resolved absolutely-calibrated fiber-coupled streaked spectrographs. Three spectrographs operate in the 4000-7000 Åvisible regime and one operates in the 2500-4000 Åultraviolet. The steadiness of the shock as it traverses the D2 cell is confirmed by the constant self emission. The temperature in the shocked D2 is inferred from the absolute intensity and spectrum. Interesting variations in the emission are also observed as the shock reverberates between the Al pusher and the sapphire window. Sandia is a multiprogram laboratory operated by Sandia Corporation, a Lockheed Martin Company, for the U.S. Dept. of Energy under contract DE-AC04-94AL85000

Bailey, J. E.; Knudson, M. D.; Asay, J. R.; Bernard, M. A.; Carlson, A. L.; Dunham, G.; Hall, C. A.; Hanson, D. L.; Hickman, R. J.; Johnston, R. R.; Lake, P. W.

2001-06-01

305

Diffusion optical spectroscopy of cancerous and normal prostate tissues in time-resolved and frequency domain  

NASA Astrophysics Data System (ADS)

It is well-known that light transport can be well described using Maxwell's electromagnetic theory. In biological tissue, the scattering particles cause the interaction of scattered waves from neighboring particles. Since such interaction cannot be ignored, multiple scattering occurs. The theoretical solution of multiple scattering is complicated. A suitable description is that the wavelike behavior of light is ignored and the transport of an individual photon is considered to be absorbed or scattered. This is known as the Radiative Transfer Equation (RTE) theory. Analytical solutions to the RTE that explicitly describes photon migration can be obtained by introducing some proper approximations. One of the most popular models used in the field of tissue optics is the Diffusion Approximation (DA). In this study, we report on the results of our initial study of optical properties of ex vivo normal and cancerous prostate tissues and how tissue parameters affect the near infrared light transporting in the two types of tissues. The time-resolved transport of light is simulated as an impulse isotropic point source of energy within a homogeneous unbounded medium with different absorption and scattering properties of cancerous and normal prostate tissues. Light source is also modulated sinusoidally to yield a varied fluence rate in frequency domain at a distant observation point within the cancerous and normal prostate tissues. Due to difference of the absorption and scattering coefficients between cancerous and normal tissues, the expansion of light pulse, intensity, phase are found to be different.

Zhou, Kenneth J.; Pu, Yang; Chen, Jun

2014-03-01

306

Ascertaining free histidine from mixtures with histidine-containing proteins using time-resolved photoluminescence spectroscopy.  

PubMed

The use of photoluminescent probes for differentiating free amino acids from biomolecules containing the same amino acids is challenging. Photoluminescent probes generally present similar emission spectra when in the presence of either free-amino acids or protein containing those same amino acids. Probes based on cyclometalated iridium(III) complexes Ir(L)2(sol)2 (where L is 2-phenylpyridine, 2-(2,4-difluorophenyl)pyridine, or benzo[h]quinolone, and sol is a solvent molecule) present long-lived emission when bound to histidine. This emission is tuned by the microenvironment around the complex and therefore its lifetime is different for free histidine (487 ns) than from histidine-containing proteins such as bovine serum albumin (average lifetime > 700 ns). As a proof-of-concept we demonstrate that free histidine can be discerned from a mixture with histidine-containing proteins by using time-resolved photoluminescence decays. In the presence of multiple sources of histidine, iridium(III) probes display a multiexponential decay, which can be fitted by nonlinear least-squares methods to separate the different components. Because the pre-exponential factor of the 487 ns lifetime is proportional to the concentration of free histidine, we can use it to assess the amount of free histidine in solution even in the presence of proteins such as bovine serum albumin. We also show that iridium(III) probes displaying different photoluminescence maxima can be produced by modifying the ancillary ligands of the metal complex. PMID:25313943

Huang, Kewei; Jiang, Chengmin; Martí, Angel A

2014-11-13

307

Rotational and Translational Dynamics of Rhodamine 6G in a Pyrrolidinium Ionic Liquid: A Combined Time-Resolved Fluorescence Anisotropy Decay and NMR Study  

SciTech Connect

NMR spectroscopy and time-resolved fluorescence anisotropy decay (TRFAD) are two of the most commonly used methods to study solute-solvent interactions. However, only a few studies have been reported to date using a combined NMR and TRFAD approach to systematically investigate the overall picture of diffusional and rotational dynamics of both the solute and solvent. In this paper, we combined NMR and TRFAD to probe fluorescent rhodamine dyes in a pyrrolidinium-based room temperature ionic liquid (RTIL), an emergent environmentally-friendly solvent type used in several energy-related applications. A specific interaction of the R6G cation and [Tf2N]- anion was identified, resulting in near-stick boundary condition rotation of R6G in this RTIL. The diffusional rates of the R6G solute and [C4mpyr][Tf2N] solvent derived from 1H NMR suggest the rates are proportional to their corresponding hydrodynamic radii. The 1H and 13C NMR studies of self-rotational dynamics of [C4mpyr][Tf2N] showed that the self-rotational correlation time of [C4mpyr]+ is 47 2 ps at 300 K. At the same temperature, we find that the correlation time for N-CH3 rotation in [C4mpyr]+ is 77 2 ps, comparable to overall molecular reorientation. This slow motion is attributed to properties of the cation structure.

Guo, Jianchang [ORNL; Han, Kee Sung [ORNL; Mahurin, Shannon Mark [ORNL; Baker, Gary A [ORNL; Hillesheim, Patrick C [ORNL; Dai, Sheng [ORNL; Hagaman, Edward {Ed} W [ORNL; Shaw, Robert W [ORNL

2012-01-01

308

Molecular dynamics in low vibrational states investigated by fs time-resolved coherent anti-Stokes Raman spectroscopy technique  

NASA Astrophysics Data System (ADS)

The molecular dynamics process is investigated in this paper using a broadband fs time-resolved coherent anti-Stokes Raman spectroscopy (CARS) technique. By varying the timing of laser pulses, low vibrational states are started and studied on both the electronically excited B( 3?0 u+) state and ground X( 1?0 g+) state of iodine in the gas phase at room temperature. According to change the pump wavelength or Stokes pulse as well as the wavelength of the detection window for the CARS signal, dynamics on different potential-energy surfaces can be accessed and detected by the CARS spectroscopy. Results show that the period of the oscillation is decreased for the excited B( 3?0 u+) state as the wavelength of the pump pulses is increased, while it is increased for the ground X( 1?0 g+) state with the increase of the Stokes wavelength.

He, Ping; Li, Sining; Fan, Rongwei; Xia, Yuanqin; Yu, Xin; Yao, Yong; Chen, Deying

2011-09-01

309

Fluorescence Correlation Spectroscopy  

NSDL National Science Digital Library

This paper, which was previously published as part of an online biophysics textbook, provides detailed information about concepts related to fluorescence correlation spectroscopy. Sections of the document include writing on experimental realization, theoretical concepts, and applications of this technology.

Haustein, Elke

310

Developments in time-resolved ultrafast imaging and spectroscopy at terahertz frequencies  

E-print Network

Prior to the advent of high energy pulsed femtosecond lasers, the field of terahertz (THz) spectroscopy was stagnated by the lack of both high power THz sources and sensitive THz detectors. Over the past few years, it has ...

Teo, Stephanie M

2014-01-01

311

A Time-Resolved Fluorescence–Resonance Energy Transfer Assay for Identifying Inhibitors of Hepatitis C Virus Core Dimerization  

PubMed Central

Abstract Binding of hepatitis C virus (HCV) RNA to core, the capsid protein, results in the formation of the nucleocapsid, the first step in the assembly of the viral particle. A novel assay was developed to discover small molecule inhibitors of core dimerization. This assay is based on time-resolved fluorescence resonance energy transfer (TR-FRET) between anti-tag antibodies labeled with either europium cryptate (Eu) or allophycocyanin (XL-665). The N-terminal 106-residue portion of core protein (core106) was tagged with either glutathione-S-transferase (GST) or a Flag peptide. Tag-free core106 was selected as the reference inhibitor. The assay was used to screen the library of pharmacologically active compounds (LOPAC) consisting of 1,280 compounds and a 2,240-compound library from the Center for Chemical Methodology and Library Development at Boston University (CMLD-BU). Ten of the 28 hits from the primary TR-FRET run were confirmed in a secondary amplified luminescent proximity homogeneous assay (ALPHA screen). One hit was further characterized by dose–response analysis yielding an IC50 of 9.3 ?M. This 513 Da compound was shown to inhibit HCV production in cultured hepatoma cells. PMID:20035614

Kota, Smitha; Scampavia, Louis; Spicer, Timothy; Beeler, Aaron B.; Takahashi, Virginia; Snyder, John K.; Porco, John A.; Hodder, Peter

2010-01-01

312

Nanosecond dynamics of calmodulin and ribosome-bound nascent chains studied by time-resolved fluorescence anisotropy.  

PubMed

We report a time-resolved fluorescence anisotropy study of ribosome-bound nascent chains (RNCs) of calmodulin (CaM), a prototypical member of the EF-hand family of calcium-sensing proteins. As shown in numerous studies, in vitro protein refolding can differ substantially from biosynthetic protein folding, which takes place cotranslationally and depends on the rate of polypeptide chain elongation. A challenge in this respect is to characterize the adopted conformations of nascent chains before their release from the ribosome. CaM RNCs (full-length, half-length, and first EF-hand only) were synthesized in vitro. All constructs contained a tetracysteine motif site-specifically incorporated in the first N-terminal helix; this motif is known to react with FlAsH, a biarsenic fluorescein derivative. As the dye is rotationally locked to this helix, we characterized the structural properties and folding states of polypeptide chains tethered to ribosomes and compared these with released chains. Importantly, we observed decelerated tumbling motions of ribosome-tethered and partially folded nascent chains, compared to released chains. This indicates a pronounced interaction between nascent chains and the ribosome surface, and might reflect chaperone activity of the ribosome. PMID:24644265

Lamprou, Paraskevas; Kempe, Daryan; Katranidis, Alexandros; Büldt, Georg; Fitter, Jörg

2014-05-01

313

Time-resolved spectroscopy and photometry of the dwarf nova FS Aurigae in quiescence  

E-print Network

We present results of non-simultaneous time-resolved photometric and spectroscopic observations of the little-studied dwarf nova FS Aur in quiescence. The spectrum of FS Aur shows strong and broad emission lines of hydrogen and HeI, and of weaker HeII 4686 and CIII/NIII blend, similar to other quiescent dwarf novae. All emission lines are single-peaked, however their form varies with orbital phase. Absorption lines from a late-type secondary are not detected. From the radial velocity measurements of the hydrogen lines H$_\\beta$ and H$_\\gamma$ we determined a most probable orbital period P=0.059+-0.002. This period agrees well with the 0.0595+-0.0001 estimate by Thorstensen et al. (1996). On the other hand, the period of photometric modulations is longer than the spectroscopic period and can be estimated as 3 hours. Longer time coverage during a single night is needed to resolve this problem. Using the semi-amplitude of the radial velocities, obtained from measurements of hydrogen and helium lines, and some empirical and theoretical relations we limited the basic parameters of the system: a mass ratio q>=0.22, a primary mass M_1=0.34 - 0.46 M_sun, a secondary mass M_2<= 0.1M_sun, and an inclination angle i=51^{\\circ} - 65^{\\circ}. Doppler tomography has shown at least two bright regions in the accretion disk of FS Aur. The first, brighter spot is located at phase about 0.6. The second spot is located opposite the first one and occupies an extensive area at phases about 0.85 - 1.15.

V. V. Neustroev

2001-10-26

314

TIME-RESOLVED SPECTROSCOPY OF THE POLAR EU CANCRI IN THE OPEN CLUSTER MESSIER 67  

SciTech Connect

We present time-resolved spectroscopic and polarimetric observations of the AM Her system EU Cnc. EU Cnc is located near the core of the old open cluster Messier 67; new proper motion measurements indicate that EU Cnc is indeed a member of the star cluster, and this system therefore is useful to constrain the formation and evolution of magnetic cataclysmic variables. The spectra exhibit two-component emission features with independent radial velocity variations as well as time-variable cyclotron emission indicating a magnetic field strength of 41 MG. The period of the radial velocity and cyclotron hump variations are consistent with the previously known photometric period, and the spectroscopic flux variations are consistent in amplitude with previous photometric amplitude measurements. The secondary star is also detected in the spectrum. We also present polarimetric imaging measurements of EU Cnc that show a clear detection of polarization, and the degree of polarization drops below our detection threshold at phases when the cyclotron emission features are fading or not evident. The combined data are all consistent with the interpretation that EU Cnc is a low-state polar in the cluster Messier 67. The mass function of the system gives an estimate of the accretor mass of M{sub WD} {>=} 0.68 M{sub Sun} with M{sub WD} Almost-Equal-To 0.83 M{sub Sun} for an average inclination. We are thus able to place a lower limit on the progenitor mass of the accreting white dwarf of {>=}1.43 M{sub Sun }.

Williams, Kurtis A. [Department of Physics and Astronomy, Texas A and M University-Commerce, P.O. Box 3011, Commerce, TX 75429 (United States); Howell, Steve B. [NASA Ames Research Center, P.O. Box 1, M/S 244-30, Moffett Field, CA 94035 (United States); Liebert, James; Smith, Paul S. [Steward Observatory, University of Arizona, Tucson, AZ (United States); Bellini, Andrea [Space Telescope Science Institute, 3700 San Martin Drive, Baltimore, MD 21218 (United States); Rubin, Kate H. R. [Max-Planck-Institut fuer Astronomie, Koenigstuhl 17, D-69117 Heidelberg (Germany); Bolte, Michael, E-mail: kurtis.williams@tamuc.edu, E-mail: steve.b.howell@nasa.gov, E-mail: jamesliebert@gmail.com, E-mail: psmith@as.arizona.edu, E-mail: bellini@stsci.edu, E-mail: rubin@mpia.de, E-mail: bolte@ucolick.org [UCO/Lick Observatory, University of California, 1156 High St., Santa Cruz, CA 95064 (United States)

2013-05-15

315

Time-resolved Spectroscopy of the Polar EU Cancri in the Open Cluster Messier 67  

NASA Astrophysics Data System (ADS)

We present time-resolved spectroscopic and polarimetric observations of the AM Her system EU Cnc. EU Cnc is located near the core of the old open cluster Messier 67; new proper motion measurements indicate that EU Cnc is indeed a member of the star cluster, and this system therefore is useful to constrain the formation and evolution of magnetic cataclysmic variables. The spectra exhibit two-component emission features with independent radial velocity variations as well as time-variable cyclotron emission indicating a magnetic field strength of 41 MG. The period of the radial velocity and cyclotron hump variations are consistent with the previously known photometric period, and the spectroscopic flux variations are consistent in amplitude with previous photometric amplitude measurements. The secondary star is also detected in the spectrum. We also present polarimetric imaging measurements of EU Cnc that show a clear detection of polarization, and the degree of polarization drops below our detection threshold at phases when the cyclotron emission features are fading or not evident. The combined data are all consistent with the interpretation that EU Cnc is a low-state polar in the cluster Messier 67. The mass function of the system gives an estimate of the accretor mass of M WD >= 0.68 M ? with M WD ? 0.83 M ? for an average inclination. We are thus able to place a lower limit on the progenitor mass of the accreting white dwarf of >=1.43 M ?. Some of the data presented herein were obtained at the W. M. Keck Observatory, which is operated as a scientific partnership among the California Institute of Technology, the University of California, and the National Aeronautics and Space Administration. The Observatory was made possible by the generous financial support of the W. M. Keck Foundation.

Williams, Kurtis A.; Howell, Steve B.; Liebert, James; Smith, Paul S.; Bellini, Andrea; Rubin, Kate H. R.; Bolte, Michael

2013-05-01

316

Time Resolved Photoelectron Spectroscopy of Thioflavin T Photoisomerization: A Simulation Study  

E-print Network

environment. The fluorescence quantum yield increases substantially upon binding to amyloid fibers configuration (300 fs). 1. INTRODUCTION Amyloid fibrils are insoluble filaments formed by proteins with ordered,2 The accumulation of amyloid fibrils in organism tissues can lead to a number of diseases, such as Alzheimer

Mukamel, Shaul

317

DNA binding induces dissociation of the multimeric form of HIV-1 integrase: A time-resolved fluorescence anisotropy study  

PubMed Central

Self-assembly of HIV-1 integrase (IN) in solution has been studied previously by time-resolved fluorescence, using tryptophan anisotropy decay. This approach provides information on the size of macromolecules via the determination of rotational correlation times (?). We have shown that, at submicromolar concentration, IN is characterized by a long rotational correlation time (?20°C = 90–100 ns) corresponding to a high-order oligomeric form, likely a tetramer. In the present work, we investigated the self-assembly properties of the DNA-bound IN by using three independent fluorophores. Under enzymatic assay conditions (10?7 M IN, 2 × 10?8 M DNA), using either fluorescein-labeled or fluorescent guanosine analog-containing oligonucleotides that mimic a viral end long terminal repeat sequence, we found that the DNA–IN complex was characterized by shorter ?20°C values of 15.5–19.5 and 23–27 ns, calculated from experiments performed at 25°C and 37°C, respectively. These results were confirmed by monitoring the Trp anisotropy decay as a function of the DNA substrate concentration: the ? of IN shifted from 90–100 ns to lower values (<30 ns) upon increasing the DNA concentration. Again, the normalized ?20°C values were significantly higher when monitored at 37°C as compared with 25°C. These results indicate that upon binding the viral DNA end, the multimeric enzyme undergoes a dissociation, most likely into a homogenous monomeric form at 25°C and into a monomer–dimer equilibrium at 37°C. PMID:11504911

Deprez, Eric; Tauc, Patrick; Leh, Hervé; Mouscadet, Jean-François; Auclair, Christian; Hawkins, Mary E.; Brochon, Jean-Claude

2001-01-01

318

Identification of Pregnane X Receptor Ligands Using Time-Resolved Fluorescence Resonance Energy Transfer and Quantitative High-Throughput Screening  

PubMed Central

Abstract The human pregnane X nuclear receptor (PXR) is a xenobiotic-regulated receptor that is activated by a range of diverse chemicals, including antibiotics, antifungals, glucocorticoids, and herbal extracts. PXR has been characterized as an important receptor in the metabolism of xenobiotics due to induction of cytochrome P450 isozymes and activation by a large number of prescribed medications. Developing methodologies that can efficiently detect PXR ligands will be clinically beneficial to avoid potential drug–drug interactions. To facilitate the identification of PXR ligands, a time-resolved fluorescence resonance energy transfer (TR-FRET) assay was miniaturized to a 1,536-well microtiter plate format to employ quantitative high-throughput screening (qHTS). The optimized 1,536-well TR-FRET assay showed Z?-factors of ?0.5. Seven- to 15-point concentration–response curves (CRCs) were generated for 8,280 compounds using both terbium and fluorescein emission data, resulting in the generation of 241,664 data points. The qHTS method allowed us to retrospectively examine single concentration screening datasets to assess the sensitivity and selectivity of the PXR assay at different compound screening concentrations. Furthermore, nonspecific assay artifacts such as concentration-based quenching of the terbium signal and compound fluorescence were identified through the examination of CRCs for specific emission channels. The CRC information was also used to define chemotypes associated with PXR ligands. This study demonstrates the feasibility of profiling thousands of compounds against PXR using the TR-FRET assay in a high-throughput format. PMID:19505231

Shukla, Sunita J.; Nguyen, Dac-Trung; MacArthur, Ryan; Simeonov, Anton; Frazee, William J.; Hallis, Tina M.; Marks, Bryan D.; Singh, Upinder; Eliason, Hildegard C.; Printen, John; Austin, Christopher P.; Inglese, James

2009-01-01

319

SIMULTANEOUS DETECTION OF ESCHERICHIA COLI 0157:H7 AND SALMONELLA TYPHIMURIUM IN FOODS USING IMMUNOMAGNETIC CAPTURE AND LANTHANIDE TIME-RESOLVED FLUORESCENCE  

Technology Transfer Automated Retrieval System (TEKTRAN)

A procedure, based on immunomagnetic capture and time-resolved fluorescence, was developed to detect Escherichia coli O157:H7 and Salmonella Typhimurium in ground meats and fresh sprouts. After a brief enrichment period, streptavidin coated magnetic beads conjugated with biotin-labeled specific ant...

320

SIMULTANEOUS DETECTION OF ESCHERICHIA COLI 0157:H7 AND SALMONELLA TYPHIMURIUM IN FOODS USING IMMUNOMAGNETIC CAPTURE AND LANTHANIDE TIME-RESOLVED FLUORESCENCE  

Technology Transfer Automated Retrieval System (TEKTRAN)

A time-resolved fluorescence procedure was developed to detect Escherichia coli O157:H7 and Salmonella typhimurium in ground meats. After a 4.5 hour enrichment period, streptavidin coated magnetic beads conjugated with biotin-labeled specific anti-bacteria antibodies were used to capture targeted ...

321

Electrolyte-Concentration and Ion-Size Dependence of Excited-State Intramolecular Charge-Transfer Reaction in (Alkylamino)benzonitriles: Time-Resolved Fluorescence  

E-print Network

-Transfer Reaction in (Alkylamino)benzonitriles: Time-Resolved Fluorescence Emission Studies Tuhin Pradhan and Ranjit & Technology, S. N. Bose National Centre for Basic Sciences, JD Block, Sector III, Salt Lake, Kolkata 700 098)benzonitrile (P6C) in ethyl acetate in presence of LiClO4 indicate that the average reaction time for LE f CT

Biswas, Ranjit

322

Time-Resolved Fluorescence and Fourier Transform Infrared Spectroscopic Investigations of Lateral Packing Defects and Superlattice Domains in Compositionally Uniform Cholesterol/Phosphatidylcholine Bilayers  

PubMed Central

Time-resolved fluorescence and Fourier transform infrared spectroscopies were used to investigate the lateral organization of lipids in compositionally uniform and fully equilibrated 1-palmitoyl-2-oleoyl-phosphatidylcholine/cholesterol (POPC/CHOL) liposomes prepared by a recently devised low-temperature trapping method. Independent fluorescence decay lifetime and rotational dynamics parameters of diphenylhexatriene (DPH) chain-labeled phosphatidylcholine (DPH-PC) in these liposomes were recovered from the time-resolved fluorescence measurements as a function of cholesterol molar fraction (XCHOL) at 23°C. The results indicate significantly greater lifetime heterogeneity, shorter average lifetime, rotational correlation time, and lower order parameter of the DPH moiety at XCHOL ? 0.40 and 0.50 as compared to the adjacent cholesterol concentrations. Less prominent changes were also detected at, for example, XCHOL ? 0.20 and 0.33. These XCHOL's coincide with the “critical” XCHOL's predicted by the previously proposed superlattice (SL) model, thus indicating that POPC and cholesterol molecules tend to form SL domains where the components tend to be regularly distributed. The data also support another prediction of the SL model, namely that lateral packing defects coexist with the ordered SL domains. It appears that unfavorable interaction of the DPH-moiety of DPH-PC with cholesterol results in a preferential partition of DPH-PC to the defect regions. Fourier transform infrared analysis of the native lipid O=P=O, C=O, and C-H vibrational bands of POPC/CHOL liposomes in the absence of DPH-PC revealed an increase in the conformational order of the acyl chains and a decrease in the conformational order (or increased hydration) of the interfacial and headgroup regions at or close to the predicted critical XCHOL's. This provides additional but probe-independent evidence for SL domain formations in the POPC/CHOL bilayers. We propose that the defect regions surrounding the putative SL domains could play an important role in modulating the activity of various membrane-associated enzymes, e.g., those regulating the lipid compositions of cell membranes. PMID:12770884

Cannon, Brian; Heath, Garrett; Huang, Juyang; Somerharju, Pentti; Virtanen, Jorma A.; Cheng, Kwan Hon

2003-01-01

323

Excited state non-adiabatic dynamics of pyrrole: A time-resolved photoelectron spectroscopy and quantum dynamics study  

NASA Astrophysics Data System (ADS)

The dynamics of pyrrole excited at wavelengths in the range 242-217 nm are studied using a combination of time-resolved photoelectron spectroscopy and wavepacket propagations performed using the multi-configurational time-dependent Hartree method. Excitation close to the origin of pyrrole's electronic spectrum, at 242 and 236 nm, is found to result in an ultrafast decay of the system from the ionization window on a single timescale of less than 20 fs. This behaviour is explained fully by assuming the system to be excited to the A2(???) state, in accord with previous experimental and theoretical studies. Excitation at shorter wavelengths has previously been assumed to result predominantly in population of the bright A1(???) and B2(???) states. We here present time-resolved photoelectron spectra at a pump wavelength of 217 nm alongside detailed quantum dynamics calculations that, together with a recent reinterpretation of pyrrole's electronic spectrum [S. P. Neville and G. A. Worth, J. Chem. Phys. 140, 034317 (2014)], suggest that population of the B1(???) state (hitherto assumed to be optically dark) may occur directly when pyrrole is excited at energies in the near UV part of its electronic spectrum. The B1(???) state is found to decay on a timescale of less than 20 fs by both N-H dissociation and internal conversion to the A2(???) state.

Wu, Guorong; Neville, Simon P.; Schalk, Oliver; Sekikawa, Taro; Ashfold, Michael N. R.; Worth, Graham A.; Stolow, Albert

2015-02-01

324

Time resolved spectroscopy and gain studies of Fullerenes C60 and C70  

NASA Astrophysics Data System (ADS)

The fluorescence decay time of Fullerenes C60 and C70 in pure form as well as in mixture with Coumarin C440 and Quinizarine dyes are studied. Results indicate that the decay of pure fullerenes is constant throughout the solute concentration and it is also independent of excitation wavelength, whereas in the case of mixture with dyes different behavior is noticed. We have also calculated the Stern-Volmer quenching constant and optical gain of both the fullerenes from which it is found that the optical gain is positive for Fullerene C70 only in a very narrow range of concentration.

Qaiser, Darakhshan; Khan, Mohd. Shahid; Singh, R. D.; Khan, Zahid H.

2013-09-01

325

Time-resolved absorption difference spectroscopy of the LH1 antenna of Rhodopseudomonas viridis  

Microsoft Academic Search

The LH-1 antenna of the purple bacterium RhodopseudomonasViridis has been investigated using femtosecond pump-probe spectroscopy. We find that qualitatively the relaxation processes are comparable to those in Rhodobacter sphaeroidesand Rhodospirillum rubrum. Both the spectral relaxation and depolarization take place in approximately 150 fs, which suggests that the excitations in the LH-1 ring are delocalized over two to three pigments. Strong

R. Monshouwer; A. Baltuska; Rienk van Grondelle

1998-01-01

326

Steady-state and time-resolved fluorescence studies on wild type and mutant chromatium vinosum high potential iron proteins: holo- and apo-forms.  

PubMed Central

Detailed circular dichroism (CD), steady-state and time-resolved tryptophan fluorescence studies on the holo- and apo- forms of high potential iron protein (HiPIP) from Chromatium vinosum and its mutant protein have been carried out to investigate conformational properties of the protein. CD studies showed that the protein does not have any significant secondary structure elements in the holo- or apo- HiPIP, indicating that the metal cluster does not have any effect on formation of secondary structure in the protein. Steady-state fluorescence quenching studies however, suggested that removal of the iron-sulfur ([Fe(4)S(4)](3+)) cluster from the protein leads to an increase in the solvent accessibility of tryptophans, indicating change in the tertiary structure of the protein. CD studies on the holo- and apo- HiPIP also showed that removal of the metal prosthetic group drastically affects the tertiary structure of the protein. Time-resolved fluorescence decay of the wild type protein was fitted to a four-exponentials model and that of the W80N mutant was fitted to a three-exponentials model. The time-resolved fluorescence decay was also analyzed by maximum entropy method (MEM). The results of the MEM analysis agreed with those obtained from discrete exponentials model analysis. Studies on the wild type and mutants helped to assign the fast picosecond lifetime component to the W80 residue, which exhibits fast fluorescence energy transfer to the [Fe(4)S(4)](3+) cluster of the protein. Decay-associated fluorescence spectra of each tryptophan residues were calculated from the time-resolved fluorescence results at different emission wavelengths. The results suggested that W80 is in the hydrophobic core of the protein, but W60 and W76 are partially or completely exposed to the solvent. PMID:11566801

Sau, A K; Chen, C A; Cowan, J A; Mazumdar, S; Mitra, S

2001-01-01

327

Time-resolved Fluorescence Resonance Energy Transfer (TR-FRET) to Analyze the Disruption of EGFR/HER2 Dimers  

PubMed Central

In oncology, simultaneous inhibition of epidermal growth factor receptor (EGFR) and HER2 by monoclonal antibodies (mAbs) is an efficient therapeutic strategy but the underlying mechanisms are not fully understood. Here, we describe a time-resolved fluorescence resonance energy transfer (TR-FRET) method to quantify EGFR/HER2 heterodimers on cell surface to shed some light on the mechanism of such therapies. First, we tested this antibody-based TR-FRET assay in NIH/3T3 cell lines that express EGFR and/or HER2 and in various tumor cell lines. Then, we used the antibody-based TR-FRET assay to evaluate in vitro the effect of different targeted therapies on EGFR/HER2 heterodimers in the ovarian carcinoma cell line SKOV-3. A simultaneous incubation with Cetuximab (anti-EGFR) and Trastuzumab (anti-HER2) disturbed EGFR/HER2 heterodimers resulting in a 72% reduction. Cetuximab, Trastuzumab or Pertuzumab (anti-HER2) alone induced a 48, 44, or 24% reduction, respectively. In contrast, the tyrosine kinase inhibitors Erlotinib and Lapatinib had very little effect on EGFR/HER2 dimers concentration. In vivo, the combination of Cetuximab and Trastuzumab showed a better therapeutic effect (median survival and percentage of tumor-free mice) than the single mAbs. These results suggest a correlation between the extent of the mAb-induced EGFR/HER2 heterodimer reduction and the efficacy of such mAbs in targeted therapies. In conclusion, quantifying EGFR/HER2 heterodimers using our antibody-based TR-FRET assay may represent a useful method to predict the efficacy and explain the mechanisms of action of therapeutic mAbs, in addition to other commonly used techniques that focus on antibody-dependent cellular cytotoxicity, phosphorylation, and cell proliferation. PMID:21282108

Gaborit, Nadège; Larbouret, Christel; Vallaghe, Julie; Peyrusson, Frédéric; Bascoul-Mollevi, Caroline; Crapez, Evelyne; Azria, David; Chardès, Thierry; Poul, Marie-Alix; Mathis, Gérard; Bazin, Hervé; Pèlegrin, André

2011-01-01

328

Heat stress induces in leaves an increase of the minimum level of chlorophyll fluorescence, Fo: A time-resolved analysis.  

PubMed

A time-resolved study of the effects of heat stress (23 to 50°C) on Fo level of chlorophyll fluorescence of leaves having different antenna content has been performed in order to elucidate the causes of heat induced increase of Fo in vivo. The multi-exponential deconvolution of the decays after a picosecond flash at Fo have shown that the best fit in both wild-type and the mutant chlorina F2 of barley leaves is obtained with three components in the temperature range utilized (100, 400 and 1200 ps at 23°C). In intermittent light greened pea leaves, a fourth long lifetime component (4 ns at 23°C) is needed. The comparison of the three types of leaves at 23°C shows that the content of the LHCII b complex does not affect the lifetimes of the two main components (100 and 400 ps) and affects their preexponential factors. This result suggests that in the PS II unit the exciton transfer from LHC IIb to the rest of the antenna is irreversible. The effects of heat stress on individual lifetime components, Ti, included several changes. Utilizing for PS II unit an extended 'Reversible Radical Pair' model, having three compartments, to interpret the variations of Ti and Ai induced by temperature increases, it can be inferred that heat determines: (i) an irreversible disconnection of a monor antenna complex which is not the LHC IIb complex, this effect is induced by temperatures higher than 40°C; (ii) a decrease of the quantum efficiency of Photosystem II photochemistry which is due to several effects: a decrease of the rate of charge separation, an increase of P(+)I(-) recombination rate constant and a decrease of the stabilization of charges. These effects on Photosystem II photochemistry start to occur above 30°C and are partially reversible. PMID:24271298

Briantais, J M; Dacosta, J; Goulas, Y; Ducruet, J M; Moya, I

1996-05-01

329

Time-resolved spectroscopy measurements of hydrogen-alpha, -beta, and -gamma emissions  

SciTech Connect

Hydrogen emission spectroscopy results are reported following laser-induced optical breakdown with infrared Nd:YAG laser radiation focused into a pulsed methane flow. Measurements of Stark-broadened atomic hydrogen-alpha, -beta, and -gamma lines show electron number densities of 0.3 to 4x10{sup 17} cm{sup -3} for time delays of 2.1 to 0.4 {mu}s after laser-induced optical breakdown. In methane flow, recombination molecular spectra of the {delta}{nu}=+2 progression of the C2 Swan system are discernable in the H{beta} and H{gamma} plasma emissions within the first few microseconds. The recorded atomic spectra indicate the occurrence of hydrogen self-absorption for pulsed CH4 flow pressures of 2.7x10{sup 5} Pa (25 psig) and 6.5x10{sup 5} Pa (80 psig)

Parigger, Christian G.; Dackman, Matthew; Hornkohl, James O

2008-11-01

330

Photosynthetic Dioxygen Formation Monitored by Time-Resolved X-Ray Spectroscopy  

SciTech Connect

Photosynthetic water oxidation provides the dioxygen of the atmosphere. Its partial reactions proceed at a Mn4Ca complex bound to photosystem II of plants and cyanobacteria. Understanding the mechanism of this biological oxidation of water molecules to O2 is one of the major challenges in life sciences. We have developed and employed X-ray absorption Spectroscopy (XAS) techniques facilitating measurements on metalloenzymes at room temperature. By these techniques, we were able to resolve structural changes at the Mn ions, to follow oxidation-state changes in the microseconds time domain, and to detect a novel and likely crucial intermediate in the oxygen-evolving step of the catalytic cycle of the Mn complex. Based on the obtained results, we replace the classic S-state model of the catalytic cycle by a more elaborated reaction scheme which solves apparent inconsistencies of earlier models, explains a large body of experimental results, and provides a fresh twist in photosynthesis research.

Haumann, Michael; Dau, Holger [Freie Universitaet Berlin, Inst. f. Experimentalphysik, Arnimallee 14, D-14195 Berlin (Germany)

2007-02-02

331

Time-resolved FTIR spectroscopy for monitoring protein dynamics exemplified by functional studies of Ras protein bound to a lipid bilayer  

E-print Network

online 22 August 2011 Keywords: Infrared Time-resolved Difference spectroscopy Rapid scan Step scan of difference spectroscopy. We dis- cuss triggering techniques and both the rapid scan and the step scan technology followed by methods for data evaluation and band assignment. This chapter ends with a discussion

Gerwert, Klaus

332

Time resolved spectroscopy of GRB 021004 reveals a clumpy extended wind  

E-print Network

High resolution spectroscopy of GRB 021004 revealed a wealth of absorption lines from several intermediate ionization species. The velocity structure of the absorber is complex and material with velocity up to >3000 km/s is observed. Since only the blueshifted component is observed, the absorber is very likely to be material closely surrounding the gamma-ray burst. We use a time-dependent photoionization code to track the abundance of the ions over time. Thanks to the presence of absorption from intermediate ionization states at long times, we can estimate the location and mass of the components of the absorber. We interpret those constraints within the hypernova scenario showing that the mass loss rate of the progenitor must have been ~10^{-4} solar masses per year, suggestive of a very massive star. In addition, the wind termination shock must lie at a distance of at least 100 pc, implying a low density environment. The velocity structure of the absorber also requires clumping of the wind at those large distances.

D. Lazzati; R. Perna; J. Flasher; V. Dwarkadas; F. Fiore

2006-08-21

333

Development of a High Harmonic Beamline for Time-Resolved XUV Spectroscopy  

NASA Astrophysics Data System (ADS)

In order to better understand bond breaking and other photochemical processes it is critical to determine the valence electron dynamics occurring during such phenomena. Extreme ultraviolet (XUV) light induces transitions between narrowly confined core electronic states and valence states. Thus ultrafast XUV absorption provides a route to determine electron distributions during chemical change. We present the design of our new femtoseconds XUV absorption spectrometer. The XUV pulses are generated in a rare gas cell in a high harmonic generation (HHG) process. Strong laser field HHG yields a promising probe source in the 10-100 eV spectral range, making it an ideal tool for XUV absorption spectroscopy of molecules containing 3d transition metals with M2,3 edges between 40-70 eV. The femtosecond duration pulses intrinsically produced by HHG allow for the necessary temporal resolution. We plan to study organometallic molecules such as the transition metal carbonyls which undergo ligand dissociation under the influence of ultraviolet light. After UV excitation a radiationless non-Born-Oppenheimer processes occur before dissociation. The understanding of these non-Born-Oppenheimer dynamics is important to the general field of photocatalysis. This work is supported by the Office of Science Early Career Research Program.

Sistrunk, Emily; Grilj, Jakob; Guehr, Markus

2012-06-01

334

Structural dynamics of membrane proteins - time-resolved and surface-enhanced IR spectroscopy  

NASA Astrophysics Data System (ADS)

Membrane proteins are the target of more than 50% of all drugs and are encoded by about 30% of the human genome. Electrophysiological techniques, like patch-clamp, unravelled many functional aspects of membrane proteins but suffer from structural sensitivity. We have developed Surface Enhanced Infrared Difference Absorption Spectroscopy (SEIDAS) to probe potential-induced structural changes of a protein on the level of a monolayer. A novel concept is introduced to incorporate membrane proteins into solid supported lipid bilayers in an orientated manner via the affinity of the His-tag to the Ni-NTA terminated gold surface. General applicability of the methodological approach is shown by tethering photosystem II to the gold surface. In conjunction with hydrogenase, the basis is set towards a biomimetic system for hydrogen production. Recently, we succeeded to record IR difference spectra of a monolayer of sensory rhodopsin II under voltage-clamp conditions. This approach opens an avenue towards mechanistic studies of voltage-gated ion channels with unprecedented structural and temporal sensitivity. Initial vibrational studies on the novel light-gated channelrhodopsin-2 (ChR2) will be presented. ChR2 represents a versatile tool in the new field of optogenetics where physiological reactions are controlled by light.

Heberle, Joachim

2013-03-01

335

Time-resolved visible and extreme ultraviolet spectroscopy of laser-produced tin plasma  

NASA Astrophysics Data System (ADS)

Previous experimental studies of laser-matter interactions have often been conducted without sufficient accuracy or attention to critical laser parameters. Moreover, much of the work published in the open literature lacks the essential theoretical underpinnings necessary to explain observations and provide predictive capability for future experiments. In this study, we use nanosecond-resolved spectroscopic techniques to investigate fundamental physics in laser-produced tin plasma, and overcome these shortcomings by implementing several metrological innovations to ensure the accuracy of experimental data. Furthermore, we present a side-by-side comparison of experimental results with computational modeling to advance our understanding of the many nonlinear, interrelated processes that occur within transient tin plasma. This dissertation is divided into three primary sections. In the first section, we study the physics governing the generation and early-time evolution of tin plasma in the low-irradiance regime: IL ˜ 4 x 1011 - 1 x 1012W/cm2 . A two-channel XUV photodiode spectrometer has been developed to measure tin plasma temperature, as well as diagnose radiation transport processes during the laser irradiation phase. During laser heating, the radiation spectrum from semi-infinite tin targets was found to approach the blackbody limit in the 10--80 nm spectral range. Through one-dimensional numerical modeling, this is shown to be due to the penetration of a radiative diffusion wave beyond the critical depth. Analysis of the time-dependent tin emission spectrum has shown that nearly 30% of the incident laser energy is converted to energetic photons in the spectral range of 15 < hv < 120 eV. The equilibrium radiation temperature, characteristic of the optically thick ablation front, has shown reasonable agreement with numerical predictions despite the model's limited dimensionality. The second part of this work examines the late-time hydrodynamics associated with the radiative plasma phase studied in the preceding section. Nanosecond-gated optical emission spectroscopy is employed to diagnose electron temperature, electron density, and propagation velocity of the ablation plume. In contrast to the large change in radiation temperature observed for a factor of three increase in laser intensity, it is found that the post-pulse plume hydrodynamics is not significantly affected for the same variation in irradiation conditions. At late times, the ion kinetic energy is found to exceed electron thermal energy by more than 100 times, which serves as a lower bound on the ratio to the ion thermal counterpart. The expanding laser-produced tin plasma is well described by a cylindrical hydrodynamic transport model; a comparison between time-integrated experimental and numerical plasma energy density has shown convergence to within a factor of two. At distances > 3 mm from the target, it was found that the heavy ion tin plasma transitions from Boltzmann to coronal equilibrium, rendering LTE assumptions in the spectral deconvolution procedure invalid. In the final section of this study, we investigate the radiative properties of tin ablation plasma as the laser irradiance is varied by more than an order of magnitude. The effect of increased focused laser energy is manifested in a weak scaling of radiation temperature, and a significant broadening of the emission lifetime at the highest laser intensities. It is found that the resulting radiation conversion efficiency is not a strong function of laser intensity within the parameter regime of this work. It is shown that agreement between experimental and simulated plasma conditions becomes progressively worse in the high-irradiance regime as the ionization and radiative transfer models play increasingly dominant roles in the plasma energetics.

O'Shay, Joseph Fred

336

Probing the hydrogen-bond network of water via time-resolved soft X-ray spectroscopy.  

PubMed

We report time-resolved studies of hydrogen bonding in liquid H(2)O, in response to direct excitation of the O-H stretch mode at 3 mum, probed via soft X-ray absorption spectroscopy at the oxygen K-edge. This approach employs a newly developed nanofluidic cell for transient soft X-ray spectroscopy in the liquid phase. Distinct changes in the near-edge spectral region (XANES) are observed, and are indicative of a transient temperature rise of 10 K following transient laser excitation and rapid thermalization of vibrational energy. The rapid heating occurs at constant volume and the associated increase in internal pressure, estimated to be 8 MPa, is manifested by distinct spectral changes that differ from those induced by temperature alone. We conclude that the near-edge spectral shape of the oxygen K-edge is a sensitive probe of internal pressure, opening new possibilities for testing the validity of water models and providing new insight into the nature of hydrogen bonding in water. PMID:19440624

Huse, Nils; Wen, Haidan; Nordlund, Dennis; Szilagyi, Erzsi; Daranciang, Dan; Miller, Timothy A; Nilsson, Anders; Schoenlein, Robert W; Lindenberg, Aaron M

2009-05-28

337

Characterization of post mortem arterial tissue using time-resolved photoacoustic spectroscopy at 436, 461 and 532 nm.  

PubMed

Time-resolved photoacoustic spectroscopy has been used to characterize post mortem arterial tissue for the purpose of discriminating between normal and atheromatous areas of tissue. Ultrasonic thermoelastic waves were generated in post mortem human aorta by the absorption of nanosecond laser pulses at 436, 461 and 532 nm produced by a frequency doubled Q-switched Nd:YAG laser in conjunction with a gas filled Raman cell. A PVDF membrane hydrophone was used to detect the thermoelastic waves. At 436 nm, differences in the photoacoustic signatures of normal tissue and atherorma were found to be highly variable. At 461 nm, there was a clear and reproducible difference between the photacoustic response of atheroma and normal tissue as a result of increased optical attenuation in atheroma. At 532 nm, the generation of subsurface thermoelastic waves provided a means of determining the structure and thickness of the tissue sample. It is suggested that pulsed photoacoustic spectroscopy at 461 and 532 nm may find application in characterizing arterial tissue in situ by providing information about both the composition and thickness of the vessel wall. PMID:9015817

Beard, P C; Mills, T N

1997-01-01

338

Probing the hydrogen-bond network of water via time-resolved soft x-ray spectroscopy  

SciTech Connect

We report time-resolved studies of hydrogen bonding in liquid H2O, in response to direct excitation of the O-H stretch mode at 3 mu m, probed via soft x-ray absorption spectroscopy at the oxygen K-edge. This approach employs a newly developed nanofluidic cell for transient soft x-ray spectroscopy in liquid phase. Distinct changes in the near-edge spectral region (XANES) are observed, and are indicative of a transient temperature rise of 10K following transient laser excitation and rapid thermalization of vibrational energy. The rapid heating occurs at constant volume and the associated increase in internal pressure, estimated to be 8MPa, is manifest by distinct spectral changes that differ from those induced by temperature alone. We conclude that the near-edge spectral shape of the oxygen K-edge is a sensitive probe of internal pressure, opening new possibilities for testing the validity of water models and providing new insight into the nature of hydrogen bonding in water.

Huse, Nils; Wen, Haidan; Nordlund, Dennis; Szilagyi, Erzsi; Daranciang, Dan; Miller, Timothy A.; Nilsson, Anders; Schoenlein, Robert W.; Lindenberg, Aaron M.

2009-04-24

339

Fluorescence Spectroscopy of Neoplastic and Non-Neoplastic Tissues  

PubMed Central

Abstract Fast and non-invasive, diagnostic techniques based on fluorescence spectroscopy have the potential to link the biochemical and morphologic properties of tissues to individual patient care. One of the most widely explored applications of fluorescence spectroscopy is the detection of endoscopically invisible, early neoplastic growth in epithelial tissue sites. Currently, there are no effective diagnostic techniques for these early tissue transformations. If fluorescence spectroscopy can be applied successfully as a diagnostic technique in this clinical context, it may increase the potential for curative treatment, and thus, reduce complications and health care costs. Steady-state, fluorescence measurements from small tissue regions as well as relatively large tissue fields have been performed. To a much lesser extent, time-resolved, fluorescence measurements have also been explored for tissue characterization. Furthermore, sources of both intrinsic (endogenous fluorophores) and extrinsic fluorescence (exogenous fluorophores) have been considered. The goal of the current report is to provide a comprehensive review on steady-state and time-resolved, fluorescence measurements of neoplastic and non-neoplastic, biologic systems of varying degrees of complexity. First, the principles and methodology of fluorescence spectroscopy are discussed. Next, the endogenous fluorescence properties of cells, frozen tissue sections and excised and intact bulk tissues are presented; fluorescence measurements from both animal and human tissue models are discussed. This is concluded with future perspectives. PMID:10933071

Ramanujam, Nirmala

2000-01-01

340

Synthesis and Characterization of Time-resolved Fluorescence Probes for Evaluation of Competitive Binding to Melanocortin Receptors  

PubMed Central

Probes for use in time-resolved fluorescence competitive binding assays at melanocortin receptors based on the parental ligands MSH(4), MSH(7), and NDP-?-MSH were prepared by solid phase synthesis methods, purified, and characterized. The saturation binding of these probes was studied using HEK-293 cells engineered to overexpress the human melanocortin 4 receptor (hMC4R) as well as the human cholecystokinin 2 receptor (hCCK2R). The ratios of non-specific binding to total binding approached unity at high concentrations for each probe. At low probe concentrations, receptor-mediated binding and uptake was discernable, and so probe concentrations were kept as low as possible in determining Kd values. The Eu-DTPA-PEGO-MSH(4) probe exhibited low specific binding relative to non-specific binding, even at low nanomolar concentrations, and was deemed unsuitable for use in competition binding assays. The Eu-DTPA-PEGO probes based on MSH(7) and NDP-?-MSH exhibited Kd values of 27±3.9 nM and 4.2±0.48 nM, respectively, for binding with hMC4R. These probes were employed in competitive binding assays to characterize the interactions of hMC4R with monovalent and divalent MSH(4), MSH(7), and NDP-?-MSH constructs derived from squalene. Results from assays with both probes reflected only statistical enhancements, suggesting improper ligand spacing on the squalene scaffold for the divalent constructs. The Ki values from competitive binding assays that employed the MSH(7)-based probe were generally lower than the Ki values obtained when the probe based on NDP-?-MSH was employed, which is consistent with the greater potency of the latter probe. The probe based on MSH(7) was also competed with monovalent, divalent, and trivalent MSH(4) constructs that previously demonstrated multivalent binding in competitive binding assays against a variant of the probe based on NDP-?-MSH. Results from these assays confirm multivalent binding, but suggest a more modest increase in avidity for these MSH(4) constructs than was previously reported. PMID:23890524

Alleti, Ramesh; Vagner, Josef; Dehigaspitiya, Dilani Chathurika; Moberg, Valerie E.; Elshan, N. G. R. D.; Tafreshi, Narges K.; Brabez, Nabila; Weber, Craig S.; Lynch, Ronald M.; Hruby, Victor J.; Gillies, Robert J.; Morse, David L.; Mash, Eugene A.

2013-01-01

341

Two-dimensional subpicosecond time-resolved fluorescence anisotropy: Optical Kerr-gating with the excitation of alternating polarizations of light  

NASA Astrophysics Data System (ADS)

We have developed a subpicosecond time-resolved fluorescence anisotropy (TRFA) that newly implements a photoelastic modulator to alternate the polarizations of an excitation laser light. The setup facilitates virtually simultaneous detection of the parallel I? and the perpendicular I? emission from a photoexcited molecule of interest by means of an ultra-short (optical Kerr-gating) shutter and a spectrograph coupled with a charge-coupled device. From a set of I?(?,t) and I?(?,t) that comprise 2-D (two-dimensional) information on both a full range of spectra and subpicosecond time-resolved fluorescence decays, the 2-D TRFA, R(?,t), is directly given with better accuracy. To claim the merit of the technique we carried out a test for 2-D TRFA of Coumarin 153 in methanol.

Fujiwara, Takashige; Romano, Natalie C.; Lim, Edward C.

2014-03-01

342

Effect of sphere to rod transition on the probe microenvironment in sodium dodecyl sulphate micelles: A time resolved fluorescence anisotropy study  

Microsoft Academic Search

The effect of different hydrotropic salts on the microenvironment at the anionic head group region of sodium dodecyl sulphate (SDS) micelle has been studied through time-resolved fluorescence anisotropy measurements of a solubilized probe, coumarin-153 (C153). The organic cations of the hydrotropic salts used in this study, i.e. aniline hydrochloride (AHC) and o-, m- and p-toluidine hydrochlorides (OTHC, MTHC and PTHC,

Teena Goel; Manoj Kumbhakar; Tulsi Mukherjee; Haridas Pal

2010-01-01

343

Monitoring changes of cellular metabolism and microviscosity in vitro based on time-resolved endogenous fluorescence and its anisotropy decay dynamics  

NASA Astrophysics Data System (ADS)

Reduced nicotinamide adenine dinucleotide (NADH) is a well-known metabolic coenzyme and endogenous fluorophore. In this study, we develop a system that simultaneously measures time- and wavelength-resolved fluorescence to extract free and protein-bound NADH signals from total cellular fluorescence. We analyze temporal characteristics of NADH fluorescence in a mixture of NADH and lactate dehydrogenase (LDH) as well as in living cell samples. The results show that in both the NADH/LDH mixture and cell samples, a fraction of free NADH and protein-bound components can be identified. The extracted free and bound NADH signals are confirmed by time-resolved measurement of anisotropy decay of NADH fluorescence, based on the fact that free NADH is a small fluorescent molecule with much shorter rotational diffusion time than bound NADH. The ratio of free NADH signal to bound NADH signal is very different between normal and cancer cervical epithelial cells. In addition, the ratio changes significantly when the cell samples are treated with a mitochondrial inhibitor or uncoupler, demonstrating that the method is sensitive to monitor cellular metabolic activity. Finally, we demonstrate that the microviscosity for relatively small molecules such as NADH in cells could be extracted from wavelength- and time-resolved NADH fluorescence of living cell samples.

Zheng, Wei; Li, Dong; Qu, Jianan Y.

2010-05-01

344

Continuous monitoring of absolute cerebral blood flow by combining diffuse correlation spectroscopy and time-resolved near-infrared technology  

NASA Astrophysics Data System (ADS)

Continuous bedside monitoring of cerebral blood flow (CBF) in patients recovering from brain injury could improve the detection of impaired substrate delivery, which can exacerbate injury and worsen outcome. Diffuse correlation spectroscopy (DCS) provides the ability to monitor perfusion changes continuously, but it is difficult to quantify absolute blood flow - leading to uncertainties as to whether or not CBF has fallen to ischemic levels. To continuously measure CBF, we propose to calibrate DCS data using a single time-point, time-resolved near-infrared (TR-NIR) technique for measuring absolute CBF. Experiments were conducted on newborn piglets in which CBF was increased by raising the arterial tension of CO2 (40-62 mmHg) and decreased by carotid occlusion. For validation, values of CBF measured by TR-NIR were converted into blood flow changes and compared to CBF changes measured by DCS. A strong correlation between perfusion changes from the two techniques was revealed (slope = 0.98 and R2 = 0.96), suggesting that a single time-point CBF measurement by TR-NIR can be used to convert continuous DCS data into units of CBF (ml/100g/min).

Diop, Mamadou; Lee, Ting-Yim; St. Lawrence, Keith

2011-02-01

345

Picosecond Dynamics of G-Protein Coupled Receptor Activation in Rhodopsin from Time-Resolved UV Resonance Raman Spectroscopy  

PubMed Central

The protein response to retinal chromophore isomerization in the visual pigment rhodopsin is studied using picosecond time-resolved UV resonance Raman spectroscopy. High signal-to-noise Raman spectra are obtained using a 1 kHz Ti:Sapphire laser apparatus that provides <3 ps visible (466 nm) pump and UV (233 nm) probe pulses. When there is no time delay between the pump and probe events, tryptophan modes W18, W16, and W3 exhibit decreased Raman scattering intensity. At longer pump-probe time delays of +5 and +20 ps, both tryptophan (W18, W16, W3, and W1) and tyrosine (Y1 + 2xY16a, Y7a, Y8a) peak intensities drop by up to 3%. These intensity changes are attributed to decreased hydrophobicity in the microenvironment near at least one tryptophan and one tyrosine residue that likely arise from weakened interaction with the ?-ionone ring of the chromophore following cis-to-trans isomerization. Examination of the crystal structure suggests that W265 and Y268 are responsible for these signals. These UV Raman spectral changes are nearly identical to those observed for the rhodopsinto-Meta I transition, implying that impulsively driven protein motion by the isomerizing chromophore during the 200 fs primary transition drives key structural changes that lead to protein activation. PMID:12731857

Kim, Judy E.; Pan, Duohai; Mathies, Richard A.

2005-01-01

346

Communication: Ultrafast time-resolved ion photofragmentation spectroscopy of photoionization-induced proton transfer in phenol-ammonia complex  

NASA Astrophysics Data System (ADS)

Photoionization-induced proton transfer (PT) in phenol-ammonia (PhOH-NH3) complex has been studied using ultrafast time-resolved ion photofragmentation spectroscopy. Neutral PhOH-NH3 complexes prepared in a free jet are photoionized by femtosecond [1+1] resonance-enhanced multiphoton ionization via the S1 state, and the subsequent dynamics occurring in the cations is probed by delayed pulses that result in ion fragmentation. The observed temporal evolutions of the photofragmentation spectra are consistent with an intracomplex PT reaction. The experiments revealed that PT in [PhOH-NH3]+ cation proceeds in two distinct steps: an initial impulsive wave-packet motion in ˜70 fs followed by a slower relaxation of about 1 ps that stabilizes the system into the final PT configuration. These results indicate that for a barrierless PT system, even though the initial PT motions are impulsive and ultrafast, the reaction may take a much longer time scale to complete.

Shen, Ching-Chi; Tsai, Tsung-Ting; Ho-Wei, Jr.; Chen, Yi-Wei; Cheng, Po-Yuan

2014-11-01

347

Photoexcited State Properties of H2-Porphyrin/C60-Based Rotaxanes as Studied by Time-Resolved EPR Spectroscopy  

PubMed Central

Light-driven intramolecular electron transfer (ET) and energy transfer (EnT) processes in two rotaxanes, the first containing two free base porphyrins and C60 fullerene moieties incorporated around a Cu(I)bisphenanthroline core ((H2P)2-Cu(I)(phen)2-C60) and a second rotaxane lacking the fullerene moiety ((H2P)2-Cu(I)(phen)2) were studied by X-band (9.5 GHz) time-resolved electron paramagnetic resonance (TREPR) spectroscopy. The experiments were performed in a frozen toluene and ethanol, and different phases of the nematic liquid crystal (E-7). It is demonstrated that the ET and EnT processes in the (H2P)2-Cu(I)(phen)2-C60 rotaxane in different media result in formation of the same charge separated state, namely (H2P)2•+-Cu(I)(phen)2•?-C60, while photoexcitation of the (H2P)2-Cu(I)(phen)2 rotaxane does not induce noticeable transfer processes in these matrices. The results are discussed in terms of the high conformational mobility of the rotaxanes, which enables changes in the molecular topography and resultant modification of the rates and routes of photoinduced processes occurring in these systems. The parameters of the transfer processes are compared with those obtained in our previous study of (ZnP)2-Cu(I)(phen)2-C60 and (ZnP)2-Cu(I)(phen) rotaxanes under the same experimental conditions. PMID:21528881

Jakob, Manuela; Berg, Alexander; Levanon, Haim; Schuster, David I.; Megiatto, Jackson D.

2011-01-01

348

Time-resolved and photoluminescence spectroscopy of ?-Al?O? nanowires for promising fast optical sensor applications.  

PubMed

Herein, we have demonstrated the high yield facile growth of Al2O3 nanowires of uniform morphology with different polymorph phases (e.g. ?, ? and ?) via a hydrothermal method with varying calcination temperatures. The synthesized ?-Al2O3 nanowires were well characterized by XRD, FTIR, SEM/EDAX, AFM and HRTEM techniques. Microstructural analysis confirmed that the dimensions of the individual ?-Al2O3 nanowires are approximately in the ranges 5-20 nm in width and 40-150 nm in length, and the aspect ratio is up to 20. AFM results evidenced the uniform distribution of the nanowires with controlled morphology. Furthermore, UV-vis spectroscopic data reveal that the estimated optical band gap of the ?-Al2O3 nanowires was ~5.16 eV. The photoluminescence spectrum exhibits blue emission upon excitation at a wavelength of 252 nm. Time-resolved spectroscopy demonstrates that these nanowires illustrate a decay time of ~2.23 nanoseconds. The obtained photoluminescence results with a decay time of nanoseconds suggest that the ?-Al2O3 phase could be an exceptional choice for next generation fast optical sensors. PMID:25300301

Gangwar, Jitendra; Gupta, Bipin Kumar; Kumar, Pawan; Tripathi, Surya Kant; Srivastava, Avanish Kumar

2014-12-01

349

Time-Resolved Frequency Comb Spectroscopy of Transient Free Radicals in the Mid-Infrared Spectral Region  

NASA Astrophysics Data System (ADS)

The chemical kinetics of transient free radicals, such as HOCO and Criegee intermediates, play important roles in combustion and atmospheric processes. Establishing accurate kinetics models for these complex systems require knowledge of the reaction rates and lifetimes of all molecules along a particular reaction pathway. However, standard spectroscopic techniques lack a combination of sensitivity, frequency resolution, and adequate temporal resolution to survey these reactions on the ?s timescale. To answer this challenge, we have developed time-resolved frequency comb spectroscopy (TRFCS). This novel technique allows for the detection of transient intermediates with high time-resolution and sensitivity while also permitting the direct determination of rotational state distributions of all relevant molecules. We demonstrate this technique in the mid-infrared spectral region, at 3.7 ?m, by studying the photolysis of deuterated acrylic acid. We simultaneously observe the time-dependent concentrations of photoproducts trans-DOCO, HOD, and D_2O, identified through their unique rovibrational structure, with 5 × 1010 molecules cm-3 sensitivity, and with a time resolution of 25 ?s. We aim to apply this technique to detect directly the formation of the DOCO intermediate in the OD + CO chemical reaction at atmospherically relevant pressures, in order to validate statistical rate models of this reaction.

Bjork, Bryce J.; Fleisher, Adam J.; Changala, Bryan; Bui, Thinh Quoc; Cossel, Kevin; Okumura, Mitchio; Ye, Jun

2014-06-01

350

Time-resolved energy-momentum spectroscopy of electric and magnetic dipole transitions in Cr3+:MgO.  

PubMed

Due to the recent interest in magnetic light-matter interactions, the magnetic dipole (MD) transitions in lanthanide ions have been studied for potential applications in nano-optics. Similar to lanthanide ions, transition-metal ions also exhibit strong MD emission at room temperature, but their prominent MD zero-phonon lines are often accompanied by significant electric dipole (ED) sideband emission. Here, we extend energy-momentum spectroscopy to time-resolved measurements, and use this technique to quantify the ED and MD contributions to light emission from trivalent chromium doped magnesium oxide (Cr(3+):MgO). This allows us to differentiate the MD (2)E ? (4)A2 zero-phonon line from phonon-assisted (2)E ? (4)A2 and (4)T2 ? (4)A2 ED sidebands. We also demonstrate how the relative intensities of the sharp MD zero-phonon line and the broad ED sidebands can be used as a qualitative measure of the MD and ED local density of optical states. PMID:23879390

Karaveli, Sinan; Wang, Shutong; Xiao, Gang; Zia, Rashid

2013-08-27

351

Fast CCD camera for x-ray photon correlation spectroscopy and time-resolved x-ray scattering and imaging  

SciTech Connect

A new, fast x-ray detector system is presented for high-throughput, high-sensitivity, time-resolved, x-ray scattering and imaging experiments, most especially x-ray photon correlation spectroscopy (XPCS). After a review of the architectures of different CCD chips and a critical examination of their suitability for use in a fast x-ray detector, the new detector hardware is described. In brief, its principal component is an inexpensive, commercial camera - the SMD1M60 - originally designed for optical applications, and modified for use as a direct-illumination x-ray detector. The remainder of the system consists of two Coreco Imaging PC-DIG frame grabber boards, located inside a Dell Power-edge 6400 server. Each frame grabber sits on its own PCI bus and handles data from 2 of the CCD's 4 taps. The SMD1M60 is based on a fast, frame-transfer, 4-tap CCD chip, read out at12-bit resolution at frame rates of up to 62 Hz for full frame readout and up to 500 Hz for one-sixteenth frame readout. Experiments to characterize the camera's suitability for XPCS and small-angle x-ray scattering (SAXS) are presented. These experiments show that single photon events are readily identified, and localized to within a pixel index or so. This is a sufficiently fine spatial resolution to maintain the speckle contrast at an acceptable value for XPCS measurements. The detective quantum efficiency of the SMD1M60 is 49% for directly-detected 6.3 keV x rays. The effects of data acquisition strategies that permit near-real-time data compression are also determined and discussed. Overall, the SMD1M60 detector system represents a major improvement in the technology for time-resolved x-ray experiments, that require an area detector with time-resolutions in few-milliseconds-to-few-seconds range, and it should have wide applications, extending beyond XPCS.

Falus, P.; Borthwick, M.A.; Mochrie, S.G.J. [Department of Physics, Yale University, New Haven, Connecticut 06520 and Department of Physics, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 (United States); Department of Physics, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 (United States); Departments of Physics and Applied Physics, Yale University, New Haven, Connecticut 06520 (United States)

2004-11-01

352

Micro-systems for time-resolved fluorescence analysis using CMOS single-photon avalanche diodes and micro-LEDs   

E-print Network

Fluorescence based analysis is a fundamental research technique used in the life sciences. However, conventional fluorescence intensity measurements are prone to misinterpretation due to illumination and fluorophore ...

Rae, Bruce R.

2009-01-01

353

Femtosecond time-resolved transient absorption spectroscopy of CH3NH3PbI3 perovskite films: evidence for passivation effect of PbI2.  

PubMed

CH3NH3PbI3 perovskite layered films deposited on substrates with and without a titania support structure have been prepared and studied using time-resolved femtosecond transient absorption (fs-TA) spectroscopy in the visible light range (450-800 nm). The electron injection dynamics from the photoexcited perovskite layers to the neighboring film structures could be directly monitored via the transient bleaching dynamics of the perovskite at ?750 nm and thus systematically studied as a function of the layer-by-layer architecture. In addition, for the first time we could spectrally distinguish transient bleaching at ?750 nm from laser-induced fluorescence that occurs red-shifted at ?780 nm. We show that an additional bleach feature at ?510 nm appears when PbI2 is present in the perovskite film. The amplitudes of the PbI2 and perovskite TA peaks were compared to estimate relative amounts of PbI2 in the samples. Kinetic analysis reveals that perovskite films with less PbI2 show faster relaxation rates than those containing more PbI2. These fast dynamics are attributed to charge carrier trapping at perovskite grain boundaries, and the slower dynamics in samples containing PbI2 are due to a passivation effect, in line with other recently reported work. PMID:25145978

Wang, Lili; McCleese, Christopher; Kovalsky, Anton; Zhao, Yixin; Burda, Clemens

2014-09-01

354

Time-resolved optical emission spectroscopy on three-dimensionally integrated micro solution plasma in He/H2O mixture  

NASA Astrophysics Data System (ADS)

We have performed time-averaged and time-resolved optical emission spectroscopy (OES) on three-dimensionally integrated micro solution plasma (3D IMSP) in He/H2O mixture. The results of time-resolved OES on 3D IMSP in He/H2O mixture have shown that duration of the optical emission of OH(A2?+ ? X2? around 309 nm) is shorter than that in Ar/H2O mixture. Possible causes of this difference are discussed by means of difference in the energy of metastable states of Ar and He.

Himeno, Y.; Ogura, Y.; Shirafuji, T.

2014-06-01

355

Time resolved infrared spectroscopy: kinetic studies of weakly binding ligands in an iron-iron hydrogenase model compound.  

PubMed

Solution photochemistry of (?-pdt)[Fe(CO)(3)](2) (pdt = ?(2)-S(CH(2))(3)S), a precursor model of the 2-Fe subsite of the H-cluster of the hydrogenase enzyme, has been studied using time-resolved infrared spectroscopy. Following the loss of CO, solvation of the Fe center by the weakly binding ligands cyclohexene, 3-hexyne, THF, and 2,3-dihydrofuran (DHF) occurred. Subsequent ligand substitution of these weakly bound ligands by pyridine or cyclooctene to afford a more stable complex was found to take place via a dissociative mechanism on a seconds time scale with activation parameters consistent with such a pathway. That is, the ?S(‡) values were positive and the ?H(‡) parameters closely agreed with bond dissociation enthalpies (BDEs) obtained from DFT calculations. For example, for cyclohexene replacement by pyridine, experimental ?H(‡) and ?S(‡) values were determined to be 19.7 ± 0.6 kcal/mol (versus a theoretical prediction of 19.8 kcal/mol) and 15 ± 2 eu, respectively. The ambidentate ligand 2,3-DHF was shown to initially bind to the iron center via its oxygen atom followed by an intramolecular rearrangement to the more stable ?(2)-olefin bound species. DFT calculations revealed a transition state structure with the iron atom almost equidistant from the oxygen and one edge of the olefinic bond. The computed ?H(‡) of 10.7 kcal/mol for this isomerization process was found to be in excellent agreement with the experimental value of 11.2 ± 0.3 kcal/mol. PMID:22680284

Muhammad, Sohail; Moncho, Salvador; Brothers, Edward N; Darensbourg, Marcetta Y; Darensbourg, Donald J; Bengali, Ashfaq A

2012-07-01

356

Microsecond Time-Resolved Absorption Spectroscopy Used to Study CO Compounds of Cytochrome bd from Escherichia coli  

PubMed Central

Cytochrome bd is a tri-heme (b558, b595, d) respiratory oxygen reductase that is found in many bacteria including pathogenic species. It couples the electron transfer from quinol to O2 with generation of an electrochemical proton gradient. We examined photolysis and subsequent recombination of CO with isolated cytochrome bd from Escherichia coli in one-electron reduced (MV) and fully reduced (R) states by microsecond time-resolved absorption spectroscopy at 532-nm excitation. Both Soret and visible band regions were examined. CO photodissociation from MV enzyme possibly causes fast (?<1.5 µs) electron transfer from heme d to heme b595 in a small fraction of the protein, not reported earlier. Then the electron migrates to heme b558 (??16 µs). It returns from the b-hemes to heme d with ??180 µs. Unlike cytochrome bd in the R state, in MV enzyme the apparent contribution of absorbance changes associated with CO dissociation from heme d is small, if any. Photodissociation of CO from heme d in MV enzyme is suggested to be accompanied by the binding of an internal ligand (L) at the opposite side of the heme. CO recombines with heme d (??16 µs) yielding a transient hexacoordinate state (CO-Fe2+-L). Then the ligand slowly (??30 ms) dissociates from heme d. Recombination of CO with a reduced heme b in a fraction of the MV sample may also contribute to the 30-ms phase. In R enzyme, CO recombines to heme d (??20 µs), some heme b558 (??0.2–3 ms), and finally migrates from heme d to heme b595 (??24 ms) in ?5% of the enzyme population. Data are consistent with the recent nanosecond study of Rappaport et al. conducted on the membranes at 640-nm excitation but limited to the Soret band. The additional phases were revealed due to differences in excitation and other experimental conditions. PMID:24755641

Siletsky, Sergey A.; Zaspa, Andrey A.; Poole, Robert K.; Borisov, Vitaliy B.

2014-01-01

357

Photochemistryand Photobiology, 1996,64(3): 552-563 Comparative Time-Resolved Photosystem II Chlorophyll a Fluorescence  

E-print Network

increased the fraction- al intensity of a short lifetime distribution at the expense of a longer lifetime had different effects on the relationship between the fluorescence lifetimes and intensity. The ob- phyll a (CNa)fluorescence yield. In this paper we present a comparative fluorescence lifetime analysis

Govindjee

358

Hexamerization of the bacteriophage T4 capsid protein gp23 and its W13V mutant studied by time-resolved tryptophan fluorescence.  

PubMed

The bacteriophage T4 capsid protein gp23 was studied using time-resolved and steady-state fluorescence of the intrinsic protein fluorophore tryptophan. In-vitro gp23 consists mostly of monomers at low temperature but forms hexamers at room temperature. To extend our knowledge of the structure and hexamerization characteristics of gp23, the temperature-dependent fluorescence properties of a tryptophan mutant (W13V) were compared to those of wild-type gp23. The W13V mutation is located in the N-terminal part of the protein, which is cleaved off after prohead formation in the live bacteriophage. Results show that W13 plays a role in the hexamerization process but is not needed to stabilize the hexamer once it is formed. Furthermore, besides the monomer-to-hexamer temperature transition (15-23 degrees C and 12-43 degrees C for wild-type and W13V gp23, respectively), we were able to observe denaturation of the N-terminus in hexameric wild-type gp23 around 40 degrees C. In addition, with the aid of a recently published homology model of gp23, the lifetimes obtained from time-resolved fluorescence measurements could tentatively be assigned to specific tryptophan residues. PMID:17149929

Stortelder, Aike; Hendriks, Johnny; Buijs, Joost B; Bulthuis, Jaap; Gooijer, Cees; van der Vies, Saskia M; van der Zwan, Gert

2006-12-14

359

Steady-state and time-resolved fluorescence studies indicate an unusual conformation of 2-aminopurine within ATAT and TATA duplex DNA sequences  

PubMed Central

2-Aminopurine (2-AP), a fluorescent analog of adenine, has been widely used as a probe for local DNA conformation, since excitation and emission characteristics and fluoresence lifetimes of 2-AP vary in a sequence-dependent manner within DNA. Using steady-state and time-resolved fluorescence techniques, we report that 2-AP appears to be unusually stacked in the internal positions of ATAT and TATA in duplex DNA. The excitation wavelength maxima for 2-AP within these contexts were red shifted, indicating reduced solvent exposure for the fluorophore. Furthermore, in these contexts, 2-AP fluorescence was resistant to acrylamide-dependent collisional quenching, suggesting that the fluorophore is protected by its stacked position within the duplex. This conclusion was further reinforced by the presence of a secondary peak at 275 nm in the fluorescence excitation spectra that is indicative of efficient excitation energy transfer from nearby non-fluorescent DNA bases. Fluorescence anisotropy decay and internal angular ‘wobbling’ motion measurements of 2-AP within these alternating AT contexts were also consistent with the fluorophore being highly constrained and immobile within the base stack. When these fluorescence characteristics are compared with those of 2-AP within other duplex DNA sequence contexts, they are unique. PMID:12711677

Rai, Priyamvada; Cole, Timothy David; Thompson, Elizabeth; Millar, David P.; Linn, Stuart

2003-01-01

360

Organization and dynamics of tryptophan residues in tetrameric and monomeric soybean agglutinin: studies by steady-state and time-resolved fluorescence, phosphorescence and chemical modification.  

PubMed

We have investigated the organization and dynamics of tryptophan residues in tetrameric, monomeric and unfolded states of soybean agglutinin (SBA) by selective chemical modification, steady-state and time-resolved fluorescence, and phosphorescence. Oxidation with N-bromosuccinimide (NBS) modifies two tryptophans (Trp 60 and Trp 132) in tetramer, four (Trp 8, Trp 203 and previous two) in monomer, and all six (Trp 8, Trp 60, Trp 132, Trp 154, Trp 203 and Trp 226) in unfolded state. Utilizing wavelength-selective fluorescence approach, we have observed a red-edge excitation shift (REES) of 10 and 5 nm for tetramer and monomer, respectively. A more pronounced REES (21 nm) is observed after NBS oxidation. These results are supported by fluorescence anisotropy experiments. Acrylamide quenching shows the Stern-Volmer constant (K(SV)) for tetramer, monomer and unfolded SBA being 2.2, 5.0 and 14.6 M(-1), respectively. Time-resolved fluorescence studies exhibit biexponential decay with the mean lifetime increasing along tetramer (1.0 ns) to monomer (1.9 ns) to unfolded (3.6 ns). Phosphorescence studies at 77 K give more structured spectra, with two (0,0) bands at 408.6 (weak) and 413.2 nm for tetramer. However, a single (0,0) band appears at 411.8 and 407.2 nm for monomer and unfolded SBA, respectively. The exposure of hydrophobic surface in SBA monomer has been examined by 8-anilino-1-naphthalenesulfonate (ANS) binding, which shows approximately 20-fold increase in ANS fluorescence compared to that for tetramer. The mean lifetime of ANS also shows a large increase (12.0 ns) upon binding to monomer. These results may provide important insight into the role of tryptophans in the folding and association of SBA, and oligomeric proteins in general. PMID:19383525

Molla, Anisur R; Maity, Shyam S; Ghosh, Sanjib; Mandal, Dipak K

2009-07-01

361

Local structures in ionic liquids probed and characterized by microscopic thermal diffusion monitored with picosecond time-resolved Raman spectroscopy.  

PubMed

Vibrational cooling rate of the first excited singlet (S(1)) state of trans-stilbene and bulk thermal diffusivity are measured for seven room temperature ionic liquids, C(2)mimTf(2)N, C(4)mimTf(2)N, C(4)mimPF(6), C(5)mimTf(2)N, C(6)mimTf(2)N, C(8)mimTf(2)N, and bmpyTf(2)N. Vibrational cooling rate measured with picosecond time-resolved Raman spectroscopy reflects solute-solvent and solvent-solvent energy transfer in a microscopic solvent environment. Thermal diffusivity measured with the transient grating method indicates macroscopic heat conduction capability. Vibrational cooling rate of S(1) trans-stilbene is known to have a good correlation with bulk thermal diffusivity in ordinary molecular liquids. In the seven ionic liquids studied, however, vibrational cooling rate shows no correlation with thermal diffusivity; the observed rates are similar (0.082 to 0.12 ps(-1) in the seven ionic liquids and 0.08 to 0.14 ps(-1) in molecular liquids) despite large differences in thermal diffusivity (5.4-7.5 × 10(-8) m(2) s(-1) in ionic liquids and 8.0-10 × 10(-8) m(2) s(-1) in molecular liquids). This finding is consistent with our working hypothesis that there are local structures characteristically formed in ionic liquids. Vibrational cooling rate is determined by energy transfer among solvent ions in a local structure, while macroscopic thermal diffusion is controlled by heat transfer over boundaries of local structures. By using "local" thermal diffusivity, we are able to simulate the vibrational cooling kinetics observed in ionic liquids with a model assuming thermal diffusion in continuous media. The lower limit of the size of local structure is estimated with vibrational cooling process observed with and without the excess energy. A quantitative discussion with a numerical simulation shows that the diameter of local structure is larger than 10 nm. If we combine this lower limit, 10 nm, with the upper limit, 100 nm, which is estimated from the transparency (no light scattering) of ionic liquids, an order of magnitude estimate of local structure is obtained as 10 nm < L < 100 nm, where L is the length or the diameter of the domain of local structure. PMID:22423845

Yoshida, Kyousuke; Iwata, Koichi; Nishiyama, Yoshio; Kimura, Yoshifumi; Hamaguchi, Hiro-o

2012-03-14

362

Relaxation dynamics of Au25L18 nanoclusters studied by femtosecond time-resolved near infrared transient absorption spectroscopy  

NASA Astrophysics Data System (ADS)

The relaxation dynamics of electronically excited [Au25(SR)18]q, where q = 0 or -1 and SR = S(CH2)2Ph, were studied using femtosecond time-resolved transient absorption spectroscopy. Nanoclusters excited by 400 nm light were probed using temporally delayed broad-bandwidth continuum probe pulses. Continuum pulses were generated in both the visible and near infrared (NIR) spectral regions, providing access to a wide range of transient spectral features. The use of NIR probe pulses allowed the relaxation dynamics of the excited states located near the HOMO-LUMO energy gap to be monitored in the probe step via the sp <-- LUMO and sp <-- LUMO+1 transitions. These NIR measurements yielded excited state absorption (ESA) data that were much less congested than the typical visible transient spectrum. For the neutral nanocluster, the time-domain data were composed of three components: (1) a few-picosecond decay, (2) a slower decay taking a few hundred picoseconds and (3) a non-decaying plateau function. Component 1 reflected energy relaxation to semi-ring ligand states; component 2 was attributed to relaxation via a manifold of states located near the HOMO-LUMO energy gap. Component 3 arose from slow radiative recombination. The dynamics of the anion depended upon the identity of the excited state from which the particle was relaxing. The LUMO+1 state of the anion exhibited relaxation dynamics that were similar to those observed for the neutral nanocluster. By comparison, the time-domain data observed for the LUMO state contained only two components: (1) a 3.3 +/- 0.2 ps decay and (2) a 5 +/- 1 ns decay. The amplitude coefficients of each component were also analyzed. Taken together, the amplitude coefficients and lifetimes were indicative of an activation barrier located approximately 100 meV above the HOMO-LUMO energy gap, which mediated a previously unobserved excited state decay process for [Au25(SR)18]0. These data suggested that NIR ESA measurements will be instrumental in describing the relaxation processes of quantum-confined nanoclusters.

Green, Thomas D.; Knappenberger, Kenneth L.

2012-06-01

363

Steady-state and time-resolved fluorescence study of some dyes in polymer microspheres showing morphology dependent resonances  

NASA Astrophysics Data System (ADS)

Fluorescence emission spectra of N,N'-bis(2,5-di-tert-butylphenyl)-3,4:9,10- Perylenebis(dicarboximide) (DBPI), rhodamine 6G (R6G), and cresyl violet (CV) in spherical polymer beads of less than 20 ?m diameter show sharp ripple structures. The observed peak positions and the intervals of the structures are consistent with the calculations of the morphology dependent resonances (MDR). Observed intensities of the MDR in the fluorescence emission spectra are found to show excitation energy dependence. The fluorescence spectra have also been measured as a function of the refractive indexes of the medium and the bead. These MDR in the beads up to 4 ?m diameter do not appear to affect the fluorescence decay of the dyes, since the fluorescence lifetime remains constant irrespective of the size of the bead and the refractive index of a surrounding medium. Simulations based on the Lorentz-Mie theory for the microspheres of different refractive indexes have been used to quantify the observed effect on the basis of the available data on the homogeneous widths of the dye molecules. A fluorescence study of microcrystals of DBPI is also presented here from the point of view of comparison with fluorescence decay of dye impregnated beads. The microcrystals exhibit a size effect in the fluorescence decay which has been attributed mainly to the self-absorption effect.

Bisht, Prem B.; Fukuda, Kazuhiro; Hirayama, Satoshi

1996-11-01

364

Separation of indocyanine green boluses in the human brain and scalp based on time-resolved in-vivo fluorescence measurements  

NASA Astrophysics Data System (ADS)

Non-invasive detection of fluorescence from the optical tracer indocyanine green is feasible in the adult human brain when employing a time-domain technique with picosecond resolution. A fluorescence-based assessment may offer higher signal-to-noise ratio when compared to bolus tracking relying on changes in time-resolved diffuse reflectance. The essential challenge is to discriminate the fluorescence originating from the brain from contamination by extracerebral fluorescence and hence to reconstruct the bolus kinetics; however, a method to reliably perform the necessary separation is missing. We present a novel approach for the decomposition of the fluorescence contributions from the two tissue compartments. The corresponding sensitivity functions pertaining to the brain and to the extracerebral compartment are directly derived from the in-vivo measurement. This is achieved by assuming that during the initial and the late phase of bolus transit the fluorescence signal originates largely from one of the compartments. Solving the system of linear equations allows one to approximate time courses of a bolus for each compartment. We applied this method to repetitive measurements on two healthy subjects with an overall 34 boluses. A reconstruction of the bolus kinetics was possible in 62% of all cases.

Jelzow, Alexander; Wabnitz, Heidrun; Obrig, Hellmuth; Macdonald, Rainer; Steinbrink, Jens

2012-05-01

365

Solvent-induced changes on the polarity of the triplet excited state of 2-chlorothioxanthone: From time-resolved absorption and resonance Raman spectroscopies  

NASA Astrophysics Data System (ADS)

Solvent polarity has been known to influence the triplet state structure and reactivity. Here, we present our experimental and theoretical study on the effect of solvent on the lowest triplet excited state structure of 2-chlorothioxanthone (CTX). Time-resolved absorption (TA) spectroscopy has been employed to understand the triplet state electronic structure; whereas solvent-induced structural changes have been studied using time-resolved resonance Raman (TR3) spectroscopy. Both the DFT and TD-DFT calculations have been performed in the solution phase employing self-consistent reaction field implicit solvation model to support the experimental data. It has been observed that CO stretching frequencies of the excited triplet state are sensitive to the solvent polarity and increase with the increase in the solvent polarity. Both TA and TR3 studies reveal that specific solvent effect (H-bonding) is more pronounced in comparison to the nonspecific solvent effect.

Pandey, Rishikesh; Umapathy, Siva

2014-01-01

366

Determination of aggregates as charge trapping and recombination centers in poly[2-methoxy-5-(2'-ethylhexyloxy)-1,4-phenylene vinylene] by time-resolved electroluminescence spectroscopy  

Microsoft Academic Search

A presence of aggregates can give critical influences on photophysical properties of conjugated polymer. For poly[2-methoxy-5-(2'-ethylhexyloxy)-1,4-phenylene vinylene], the authors found that the aggregates can serve as charge traps and recombination centers by using time-resolved electroluminescence spectroscopy, and that the emissions from isolated chains and aggregates show different dynamics. The authors also found that the highest occupied molecular orbital (HOMO) and

Hao-En Tseng; Ching-Yang Liu; Show-An Chen

2006-01-01

367

Time-resolved detection enables standard two-photon fluorescence microscopy for in vivo label-free imaging of microvasculature in tissue.  

PubMed

We conducted a systematic study on two-photon excited fluorescence (TPEF) of hemoglobin using the near transform-limited and Gaussian-shaped femtosecond pulse sources. We found that the two-photon action cross section of hemoglobin drops over 2 orders of magnitude in the wavelength range from 550 to 800 nm, while the spectral and temporal characteristics of hemoglobin TPEF are insensitive to the change of excitation wavelength. In particular, our new findings showed that the hemoglobin fluorescence could be excited with sufficient efficiency using a conventional Ti:sapphire laser tuned at the wavelength close to 700 nm. With the employment of a time-resolved detection method, we demonstrated that the TPEF signals of hemoglobin excited by a Ti:sapphire laser could be clearly differentiated from other nonlinear signals presented within the living biological tissues, indicating that a standard TPEF microscope can become a routine tool for in vivo label-free microangiography imaging. PMID:21765493

Li, Dong; Zheng, Wei; Zhang, Wei; Teh, Seng Khoon; Zeng, Yan; Luo, Yi; Qu, Jianan Y

2011-07-15

368

Inhibition of NADH oxidation by chloramphenicol in the freely moving rat measured by picosecond time-resolved emission spectroscopy.  

PubMed

Owing to the lack of methods capable to monitor the energetic processes taking place within small brain regions (i.e. nucleus raphe dorsalis, nRD), the neurotoxicity of various categories of substances, including antibiotics and psycho-active drugs, still remains difficult to evaluate. Using an in vivo picosecond optical spectroscopy imaging method, we report that chloramphenicol (CAP), besides its well-known ability to inhibit the mitochondria protein synthesis, also influences the NADH/NAD+ redox processes of the respiratory chain. At a 200-mg/kg dose, CAP indeed produces a marked increase in the fluorescent signal of the nRD which, according to clear evidence, is likely to be related to the NADH concentration. This effect also implies an efficient inhibition of complex I of the respiratory chain by CAP. It refers to the mechanism through which the adverse effects of the antibiotic may take place. It could explain why paradoxical sleep, a state needing aerobic energy to occur, is suppressed after CAP administration. The present approach constitutes the first attempt to determine by fluorescence methods the effects of substances on deep brain structures of the freely moving animal. It points out that in vivo ultrafast optical methods are innovative and adequate tools for combined neurochemical and behavioural approaches. PMID:12562508

Mottin, Stéphane; Laporte, Pierre; Cespuglio, Raymond

2003-02-01

369

Time-Resolved Down-Conversion of 2-Aminopurine in a DNA Hairpin: Fluorescence Anisotropy and Solvent Effects  

NASA Astrophysics Data System (ADS)

Femtosecond fluorescence anisotropy decay measured by type II difference frequency generation provides new insight into the local structural dynamics of ?P(-)PBS fragments of the HIV- 1 DNA primary binding sequence, labeled with 2-aminopurine.

Tourón Touceda, Patricia; Gelot, Thomas; Crégut, Olivier; Léonard, Jérémie; Haacke, Stefan

2013-03-01

370

Simultaneously reconstructing fluorescent yield and lifetime from time-resolved transmittances of a small-animal-sized phantom  

Microsoft Academic Search

A full three-dimensional, featured-data algorithm for time-domain diffuse fluorescence tomography is presented, which inverts the Laplace-transformed time-domain coupled diffusion equations and employs a pair of real-domain transform-factors to effectively separate the fluorescent yield and lifetime parameters. By use of a multi-channel time-correlation single photon counting system and a normalized Born formulation for the inversion, the proposed scheme is experimentally validated

Feng Gao; Limin Zhang; Patrick Poulet; Jiao Li; Huijuan Zhao; Yukio Yamada

2009-01-01

371

Selective Nonpeptidic Fluorescent Ligands for Oxytocin Receptor: Design, Synthesis, and Application to Time-Resolved FRET Binding Assay.  

PubMed

The design and the synthesis of the first high-affinity fluorescent ligands for oxytocin receptor (OTR) are described. These compounds enabled the development of a TR-FRET based assay for OTR, readily amenable to high throughput screening. The validation of the assay was achieved by competition experiments with both peptide and nonpeptide OTR ligands as competitors. These probes represent the first selective fluorescent ligands for the oxytocin G protein-coupled receptor. PMID:25642985

Karpenko, Iuliia A; Margathe, Jean-François; Rodriguez, Thiéric; Pflimlin, Elsa; Dupuis, Elodie; Hibert, Marcel; Durroux, Thierry; Bonnet, Dominique

2015-03-12

372

Picosecond time-resolved fluorescence of ribonuclease T1. A pH and substrate analogue binding study.  

PubMed Central

The tryptophyl fluorescence of ribonuclease T1 decays monoexponentially at pH 5.5, tau = 4.04 ns but on increasing pH, a second short-lived component of 1.5 ns appears with a midpoint between pH 6.5 and 7.0. Both components have the same fluorescence spectrum. Acrylamide quenches both fluorescence components, and the short-lived component is quenched fivefold faster than the predominant long component. Binding of the substrate analogue 2'-guanylic acid at pH 5.5 quenches the fluorescence by 20% and introduces a second decay component, tau = 1.16 ns. Acrylamide quenches both tryptophyl decay components, with similar quenching rates. The fluorescence anisotropy decay of ribonuclease T1 was consistent with a molecule the size of ribonuclease T1 surrounded by a single layer of water at pH 7.4, even though the anisotropy decay at pH 5.5 deviated from Stokes-Einstein behavior. The fluorescence data were interpreted with a model where the tryptophyl residue exists in two conformations, remaining in a hydrophobic pocket. The acrylamide quenching is interpreted with electron transfer theory and suggests that one conformer has the nearest atom approximately 3 A from the protein surface, and the other, approximately 2 A. PMID:3038204

Chen, L X; Longworth, J W; Fleming, G R

1987-01-01

373

A high-throughput time-resolved mini-silicon photomultiplier with embedded fluorescence lifetime estimation in 0.13 ?m CMOS.  

PubMed

We describe a miniaturized, high-throughput, time-resolved fluorescence lifetime sensor implemented in a 0.13 m CMOS process, combining single photon detection, multiple channel timing and embedded pre-processing of fluorescence lifetime estimations on a single device. Detection is achieved using an array of single photon avalanche diodes (SPADs) arranged in a digital silicon photomultiplier (SiPM) architecture with 400 ps output pulses and a 10% fill-factor. An array of time-to-digital converters (TDCs) with ?50 ps resolution records up to 8 photon events during each excitation period. Data from the TDC array is then processed using a centre-of-mass method (CMM) pre-calculation to produce fluorescence lifetime estimations in real-time. The sensor is believed to be the first reported implementation of embedded fluorescence lifetime estimation. The system is demonstrated in a practical laboratory environment with measurements of a variety of fluorescent dyes with different single exponential lifetimes, successfully showing the sensor's ability to overcome the classic pile-up limitation of time-correlated single photon counting (TCSPC) by over an order of magnitude. PMID:23853257

Tyndall, David; Rae, Bruce R; Li, David Day-Uei; Arlt, Jochen; Johnston, Abigail; Richardson, Justin A; Henderson, Robert K

2012-12-01

374

Early Amyloidogenic Oligomerization Studied through Fluorescence Lifetime Correlation Spectroscopy  

PubMed Central

Amyloidogenic protein aggregation is a persistent biomedical problem. Despite active research in disease-related aggregation, the need for multidisciplinary approaches to the problem is evident. Recent advances in single-molecule fluorescence spectroscopy are valuable for examining heterogenic biomolecular systems. In this work, we have explored the initial stages of amyloidogenic aggregation by employing fluorescence lifetime correlation spectroscopy (FLCS), an advanced modification of conventional fluorescence correlation spectroscopy (FCS) that utilizes time-resolved information. FLCS provides size distributions and kinetics for the oligomer growth of the SH3 domain of ?-spectrin, whose N47A mutant forms amyloid fibrils at pH 3.2 and 37 °C in the presence of salt. The combination of FCS with additional fluorescence lifetime information provides an exciting approach to focus on the initial aggregation stages, allowing a better understanding of the fibrillization process, by providing multidimensional information, valuable in combination with other conventional methodologies. PMID:22949804

Paredes, Jose M.; Casares, Salvador; Ruedas-Rama, Maria J.; Fernandez, Elena; Castello, Fabio; Varela, Lorena; Orte, Angel

2012-01-01

375

Non-invasive measurement of blood glucose level by time-resolved transmission spectroscopy: A feasibility study  

NASA Astrophysics Data System (ADS)

An optical spectroscopic method is investigated theoretically for in vivo measurement of blood glucose concentration. This method is based on dynamic dual wavelength (610 nm and 810 nm) time-resolved measurements under a condition of artificial blood flow kinetics in a human finger. The influence of glucose concentration on absorption and reduced scattering coefficients of the whole blood is simulated using the T-matrix method. The scattering centers, RBC aggregation, under the artificial — kinetics condition are modeled as spheroid. The modified parametric slopes were derived from the Laplace transformed data of the time-resolved transmittance. The results show that an appropriate selection of the Laplace parameter can lead to enhanced sensitivity for glucose measurement.

Sun, Meixiu; Chen, Nanguang

2012-03-01

376

Time-resolved photoluminescence spectroscopy of Ce:Gd3Al2Ga3O12 crystals  

NASA Astrophysics Data System (ADS)

Time-resolved photoluminescence (PL) spectra of Ce-doped Gd3Al2Ga3O12 (Ce:GAGG) crystal have been measured at various temperatures under Ce3+ 4f ? 5d excitation. All time-resolved PL spectra show only a Ce3+ 5d ? 4f band around 2.3 eV. Decay curves of this band consist of a fast and slow components. The temperature dependence of integrated intensities of the fast and slow components are analyzed by considering nonradiative processes starting from the ground level of an electron trap. From the results of analysis, we discuss relaxation processes of 5d electrons created under Ce3+ 4f ? 5d excitation.

Satoh, Azusa; Kitaura, Mamoru; Kamada, Kei; Ohnishi, Akimasa; Sasaki, Minoru; Hara, Kazuhiko

2014-01-01

377

Picosecond time resolved photoluminescence spectroscopy of a tetracene film on highly oriented pyrolytic graphite: Dynamical relaxation, trap emission, and superradiance  

NASA Astrophysics Data System (ADS)

A detailed time resolved investigation of the photoluminescence of a thin tetracene film deposited on highly oriented pyrolytic graphite is presented. In agreement with Lim et al. [Phys. Rev. Lett. 92, 107402 (2004)], we find strong evidence for superradiance: an increase of the relative intensity of the pure electronic transition with respect to the vibronic sideband and a concomitant decrease of the radiative lifetime from 10to1.83ns upon cooling from 300to4K. For lower temperatures, a redshift (˜200cm-1) of the free exciton is observed. Previously, this shift was attributed to a structural phase transition. Our time resolved spectra reveal that the spectral shift is related to a dynamical relaxation process which occurs within the first 50ps.

Voigt, M.; Langner, A.; Schouwink, P.; Lupton, J. M.; Mahrt, R. F.; Sokolowski, M.

2007-09-01

378

Bayesian approach to inverse problem in the case of time-resolved polarized fluorescence investigation of microscopically ordered systems  

NASA Astrophysics Data System (ADS)

In the case of fluorescence investigation of systems, which are isotropic in the macro scale but anisotropic in the micro scale, a Bayesian approach to an inverse problem allows finding distributions of model parameters. Next, this approach provides capacity to ascertain, whether the aligning potential alters during an electronic excitation of the fluorescence probe. The usage of the synthetic data set allows to specify an extent of a priori information necessary to a description of the data. As a numerical basement for Bayesian calculations the Differential Evolution Markov Chain method is employed.

Buczkowski, Marcin; Budzi?ski, Maciej P.; Fisz, Jacek J.

2013-06-01

379

Liquid film characterization in horizontal, annular, two-phase, gas-liquid flow using time-resolved laser-induced fluorescence  

NASA Astrophysics Data System (ADS)

A non-intrusive optical technique was developed to provide time-resolved longitudinal and cross-sectional images of the liquid film in horizontal annular pipe flow of air and water, revealing the interfacial wave behavior. Quantitative information on the liquid film dynamics was extracted from the time-resolved images. The planar laser-induced fluorescence technique was utilized to allow for optical separation of the light emitted by the film from that scattered by the air-water interface. The visualization test section was fabricated from a tube presenting nearly the same refractive index as water, which allowed the visualization of the liquid film at regions very close to the pipe wall. Longitudinal images of the liquid film were captured using a high-frame-rate digital video camera synchronized with a high-repetition-rate laser. An image processing algorithm was developed to automatically detect the position of the air-water interface in each image frame. The thickness of the liquid film was measured at two axial stations in each processed image frame, providing time history records of the film thickness at two different positions. Wave frequency information was obtained by analyzing the time-dependent signals of film thickness for each of the two axial positions recorded. Wave velocities were measured by cross-correlating the amplitude signals from the two axial positions. For the film cross-section observations, two high-speed digital video cameras were used in a stereoscopic arrangement. Comparisons with results from different techniques available in literature indicate that the technique developed presents equivalent accuracy in measuring the liquid film properties. Time-resolved images of longitudinal and cross-section views of the film were recorded, which constitute valuable information provided by the technique implemented.

Farias, P. S. C.; Martins, F. J. W. A.; Sampaio, L. E. B.; Serfaty, R.; Azevedo, L. F. A.

2012-03-01

380

Density relaxation and particle motion characteristics in a non-ionic deep eutectic solvent (acetamide + urea): Time-resolved fluorescence measurements and all-atom molecular dynamics simulations  

NASA Astrophysics Data System (ADS)

Temperature dependent relaxation dynamics, particle motion characteristics, and heterogeneity aspects of deep eutectic solvents (DESs) made of acetamide (CH3CONH2) and urea (NH2CONH2) have been investigated by employing time-resolved fluorescence measurements and all-atom molecular dynamics simulations. Three different compositions (f) for the mixture [fCH3CONH2 + (1 - f)NH2CONH2] have been studied in a temperature range of 328-353 K which is ˜120-145 K above the measured glass transition temperatures (˜207 K) of these DESs but much lower than the individual melting temperature of either of the constituents. Steady state fluorescence emission measurements using probe solutes with sharply different lifetimes do not indicate any dependence on excitation wavelength in these metastable molten systems. Time-resolved fluorescence anisotropy measurements reveal near-hydrodynamic coupling between medium viscosity and rotation of a dissolved dipolar solute. Stokes shift dynamics have been found to be too fast to be detected by the time-resolution (˜70 ps) employed, suggesting extremely rapid medium polarization relaxation. All-atom simulations reveal Gaussian distribution for particle displacements and van Hove correlations, and significant overlap between non-Gaussian (?2) and new non-Gaussian (?) heterogeneity parameters. In addition, no stretched exponential relaxations have been detected in the simulated wavenumber dependent acetamide dynamic structure factors. All these results are in sharp contrast to earlier observations for ionic deep eutectics with acetamide [Guchhait et al., J. Chem. Phys. 140, 104514 (2014)] and suggest a fundamental difference in interaction and dynamics between ionic and non-ionic deep eutectic solvent systems.

Das, Anuradha; Das, Suman; Biswas, Ranjit

2015-01-01

381

Density relaxation and particle motion characteristics in a non-ionic deep eutectic solvent (acetamide + urea): time-resolved fluorescence measurements and all-atom molecular dynamics simulations.  

PubMed

Temperature dependent relaxation dynamics, particle motion characteristics, and heterogeneity aspects of deep eutectic solvents (DESs) made of acetamide (CH3CONH2) and urea (NH2CONH2) have been investigated by employing time-resolved fluorescence measurements and all-atom molecular dynamics simulations. Three different compositions (f) for the mixture [fCH3CONH2 + (1 - f)NH2CONH2] have been studied in a temperature range of 328-353 K which is ?120-145 K above the measured glass transition temperatures (?207 K) of these DESs but much lower than the individual melting temperature of either of the constituents. Steady state fluorescence emission measurements using probe solutes with sharply different lifetimes do not indicate any dependence on excitation wavelength in these metastable molten systems. Time-resolved fluorescence anisotropy measurements reveal near-hydrodynamic coupling between medium viscosity and rotation of a dissolved dipolar solute. Stokes shift dynamics have been found to be too fast to be detected by the time-resolution (?70 ps) employed, suggesting extremely rapid medium polarization relaxation. All-atom simulations reveal Gaussian distribution for particle displacements and van Hove correlations, and significant overlap between non-Gaussian (?2) and new non-Gaussian (?) heterogeneity parameters. In addition, no stretched exponential relaxations have been detected in the simulated wavenumber dependent acetamide dynamic structure factors. All these results are in sharp contrast to earlier observations for ionic deep eutectics with acetamide [Guchhait et al., J. Chem. Phys. 140, 104514 (2014)] and suggest a fundamental difference in interaction and dynamics between ionic and non-ionic deep eutectic solvent systems. PMID:25612718

Das, Anuradha; Das, Suman; Biswas, Ranjit

2015-01-21

382

Employing time-resolved terahertz spectroscopy to analyze carrier dynamics in thin-film Cu{sub 2}ZnSn(S,Se){sub 4} absorber layers  

SciTech Connect

We report the application of time-resolved terahertz spectroscopy (TRTS) to measure photoexcited carrier lifetimes and mobility, and to determine recombination mechanisms in Cu{sub 2}ZnSn(S,Se){sub 4} (CZTSSe) thin films fabricated from nanocrystal inks. Ultrafast time resolution permits tracking the evolution of carrier density to determine recombination rates and mechanisms. The carrier generation profile was manipulated by varying the photoexcitation wavelength and fluence to distinguish between surface, Shockley-Read-Hall (SRH), radiative, and Auger recombination mechanisms and determine rate constants. Surface and SRH recombination are the dominant mechanisms for the air/CZTSSe/SiO{sub 2}/Si film stack. Diffusion to, and then recombination at, the air-CZTSSe interface occurred on the order of 100 picoseconds, while SRH recombination lifetimes were 1–2 nanoseconds. TRTS measurements can provide information that is complementary to conventional time-resolved photoluminescence measurements and can direct the design of efficient thin film photovoltaics.

Guglietta, Glenn W.; Baxter, Jason B. [Department of Chemical and Biological Engineering, Drexel University, Philadelphia, Pennsylvania 19104 (United States); Choudhury, Kaushik Roy; Caspar, Jonathan V. [DuPont Central Research and Development, Experimental Station, Wilmington, Delaware 19880 (United States)

2014-06-23

383

Homodimerization of Amyloid Precursor Protein at the Plasma Membrane: A homoFRET Study by Time-Resolved Fluorescence Anisotropy Imaging  

PubMed Central

Classical FRET (Förster Resonance Energy Transfer) using two fluorescent labels (one for the donor and another one for the acceptor) is not efficient for studying the homodimerization of a protein as only half of the homodimers formed can be identified by this technique. We thus resorted to homoFRET detected by time-resolved Fluorescence Anisotropy IMaging (tr-FAIM). To specifically image the plasma membrane of living cells, an original combination of tr-FAIM and Total Internal Reflection Fluorescence Lifetime Imaging Microscope (TIRFLIM) was implemented. The correcting factor accounting for the depolarization due to the high numerical aperture (NA) objective, mandatory for TIRF microscopy, was quantified on fluorescein solutions and on HEK293 cells expressing enhanced Green Fluorescence Protein (eGFP). Homodimerization of Amyloid Precursor Protein (APP), a key mechanism in the etiology of Alzheimer’s disease, was measured on this original set-up. We showed, both in epifluorescence and under TIRF excitation, different energy transfer rates associated with the homodimerization of wild type APP-eGFP or of a mutated APP-eGFP, which forms constitutive dimers. This original set-up thus offers promising prospects for future studies of protein homodimerization in living cells in control and pathological conditions. PMID:22973448

Devauges, Viviane; Marquer, Catherine; Lécart, Sandrine; Cossec, Jack-Christophe; Potier, Marie-Claude; Fort, Emmanuel; Suhling, Klaus; Lévêque-Fort, Sandrine

2012-01-01

384

Homogeneous time-resolved fluorescence-based assay to screen for ligands targeting the growth hormone secretagogue receptor type 1a.  

PubMed

The growth hormone secretagogue receptor type 1a (GHS-R1a) belongs to class A G-protein-coupled receptors (GPCR). This receptor mediates pleiotropic effects of ghrelin and represents a promising target for dysfunctions of growth hormone secretion and energy homeostasis including obesity. Identification of new compounds which bind GHS-R1a is traditionally achieved using radioactive binding assays. Here we propose a new fluorescence-based assay, called Tag-lite binding assay, based on a fluorescence resonance energy transfer (FRET) process between a terbium cryptate covalently attached to a SNAP-tag fused GHS-R1a (SNAP-GHS-R1a) and a high-affinity red fluorescent ghrelin ligand. The long fluorescence lifetime of the terbium cryptate allows a time-resolved detection of the FRET signal. The assay was made compatible with high-throughput screening by using prelabeled cells in suspension under a 384-well plate format. K(i) values for a panel of 14 compounds displaying agonist, antagonist, or inverse agonist properties were determined using both the radioactive and the Tag-lite binding assays performed on the same batches of GHS-R1a-expressing cells. Compound potencies obtained in the two assays were nicely correlated. This study is the first description of a sensitive and reliable nonradioactive binding assay for GHS-R1a in a format amenable to high-throughput screening. PMID:20937574

Leyris, Jean-Philippe; Roux, Thomas; Trinquet, Eric; Verdié, Pascal; Fehrentz, Jean-Alain; Oueslati, Nadia; Douzon, Stéphanie; Bourrier, Emmanuel; Lamarque, Laurent; Gagne, Didier; Galleyrand, Jean-Claude; M'kadmi, Céline; Martinez, Jean; Mary, Sophie; Banères, Jean-Louis; Marie, Jacky

2011-01-15

385

Lateral Distribution of the Transmembrane Domain of Influenza Virus Hemagglutinin Revealed by Time-resolved Fluorescence Imaging*  

PubMed Central

Influenza virus hemagglutinin (HA) has been suggested to be enriched in liquid-ordered lipid domains named rafts, which represent an important step in virus assembly. We employed Förster resonance energy transfer (FRET) via fluorescence lifetime imaging microscopy to study the interaction of the cytoplasmic and transmembrane domain (TMD) of HA with agly co sylphos pha tidyl ino si tol (GPI)-anchored peptide, an established marker for rafts in the exoplasmic leaflet of living mammalian plasma membranes. Cyan fluorescent protein (CFP) was fused to GPI, whereas the HA sequence was tagged with yellow fluorescent protein (YFP) on its exoplasmic site (TMD-HA-YFP), avoiding any interference of fluorescent proteins with the proposed role of the cytoplasmic domain in lateral organization of HA. Constructs were expressed in Chinese hamster ovary cells (CHO-K1) for which cholesterol-sensitive lipid nanodomains and their dimension in the plasma membrane have been described (Sharma, P., Varma, R., Sarasij, R. C., Ira, Gousset, K., Krishnamoorthy, G., Rao, M., and Mayor, S. (2004) Cell 116, 577–589). Upon transfection in CHO-K1 cells, TMD-HA-YFP is partially expressed as a dimer. Only dimers are targeted to the plasma membrane. Clustering of TMD-HA-YFP with GPI-CFP was observed and shown to be reduced upon cholesterol depletion, a treatment known to disrupt rafts. No indication for association of TMD-HA-YFP with GPI-CFP was found when palmitoylation, an important determinant of raft targeting, was suppressed. Clustering of TMD-HA-YFP and GPI-CFP was also observed in purified plasma membrane suspensions by homoFRET. We concluded that the pal mit oy lated TMD-HA alone is sufficient to recruit HA to cholesterol-sensitive nanodomains. The corresponding construct of the spike protein E2 of Semliki Forest virus did not partition preferentially in such domains. PMID:19349276

Scolari, Silvia; Engel, Stephanie; Krebs, Nils; Plazzo, Anna Pia; De Almeida, Rodrigo F. M.; Prieto, Manuel; Veit, Michael; Herrmann, Andreas

2009-01-01

386

Direct on-strip analysis of size- and time-resolved aerosol impactor samples using laser induced fluorescence spectra excited at 263 and 351 nm.  

PubMed

We report a novel atmospheric aerosol characterization technique, in which dual wavelength UV laser induced fluorescence (LIF) spectrometry marries an eight-stage rotating drum impactor (RDI), namely UV-LIF-RDI, to achieve size- and time-resolved analysis of aerosol particles on-strip. The UV-LIF-RDI technique measured LIF spectra via direct laser beam illumination onto the particles that were impacted on a RDI strip with a spatial resolution of 1.2mm, equivalent to an averaged time resolution in the aerosol sampling of 3.6 h. Excited by a 263 nm or 351 nm laser, more than 2000 LIF spectra within a 3-week aerosol collection time period were obtained from the eight individual RDI strips that collected particles in eight different sizes ranging from 0.09 to 10 ?m in Djibouti. Based on the known fluorescence database from atmospheric aerosols in the US, the LIF spectra obtained from the Djibouti aerosol samples were found to be dominated by fluorescence clusters 2, 5, and 8 (peaked at 330, 370, and 475 nm) when excited at 263 nm and by fluorescence clusters 1, 2, 5, and 6 (peaked at 390 and 460 nm) when excited at 351 nm. Size- and time-dependent variations of the fluorescence spectra revealed some size and time evolution behavior of organic and biological aerosols from the atmosphere in Djibouti. Moreover, this analytical technique could locate the possible sources and chemical compositions contributing to these fluorescence clusters. Advantages, limitations, and future developments of this new aerosol analysis technique are also discussed. PMID:24745745

Wang, Chuji; Pan, Yong-Le; James, Deryck; Wetmore, Alan E; Redding, Brandon

2014-04-11

387

STEADY-STATE AND TIME-RESOLVED FLUORESCENCE QUENCHING WITH TRANSITION METAL IONS AS SHORT-DISTANCE PROBES FOR PROTEIN CONFORMATION  

PubMed Central

A series of model dye-labeled histidine-containing peptides was used to investigate the nature of the quenching mechanism with Cu2+ and Ni2+. The strong reduction in steady-state fluorescence was found to be unaccompanied by any noticeable changes in lifetime kinetics. This static nature of quenching is not consistent with the dynamic FRET phenomenon, which was assumed to dominate the quenching mechanism, and is likely caused by shorter range orbital coupling. Our results indicate that the FRET-like 6th power of distance dependence of quenching cannot be automatically assumed for transition metal ions, and time-resolved measurements should be used to distinguish various quenching mechanisms. PMID:20707982

Posokhov, Yevgen O.; Kyrychenko, Alexander; Ladokhin, Alexey S.

2011-01-01

388

Continuous wave and time resolved spectroscopy of InAsN/GaAsN based quantum dots  

NASA Astrophysics Data System (ADS)

We present a study of the optical properties of quantum dots based on a new family of semiconductors: III-V dilute nitrides such as (In,Ga)(N,As). Continuous wave and time resolved photoluminescence (PL) experiments allowed us to evaluate the impact of N incorporation during the growth of InAs/GaAs quantum dots. Previous work [V. Sallet et al., to be submitted to J. Cryst. Growth (2005); O. Schumann et al., J. Appl. Phys. 96, 2832 (2004)] showed that increasing the flux of N atoms into the growth chamber modifies drastically the size of the dots which leads to a bimodal growth. Two populations of dots with different sizes appear. The quantum dot PL line broadens and a second PL line appears at higher energy. Time resolved PL allows us to identify the nature of this second PL line: second population of quantum dots. A second decay time is observed which we interpret as being the consequence of the perturbation of the electronic states of the quantum dots.

Taliercio, T.; Valvin, P.; Intartaglia, R.; Sallet, V.; Harmand, J. C.; Guillet, T.; Lefebvre, P.; Bretagnon, T.; Gil, B.

2005-11-01

389

Effect of Ca(2+) on the Steady-State and Time-Resolved Emission Properties of the Genetically Encoded Fluorescent Sensor CatchER.  

PubMed

We previously designed a calcium sensor CatchER (a GFP-based Calcium sensor for detecting high concentrations in the high calcium concentration environment such as ER) with a capability for monitoring calcium ion responses in various types of cells. Calcium binding to CatchER induces the ratiometric changes in the absorption spectra, as well as an increase in fluorescence emission at 510 nm upon excitation at both 395 and 488 nm. Here, we have applied the combination of the steady-state and time-resolved optical methods and Hydrogen/Deuterium isotope exchange to understand the origin of such calcium-induced optical property changes of CatchER. We first demonstrated that calcium binding results in a 44% mean fluorescence lifetime increase of the indirectly excited anionic chromophore. Thus, CatchER is the first protein-based calcium indicator with the single fluorescent moiety to show the direct correlation between the lifetime and calcium binding. Calcium exhibits a strong inhibition on the excited-state proton transfer nonadiabatic geminate recombination in protic (vs deuteric) medium. Analysis of CatchER crystal structures and the MD simulations reveal the proton transfer mechanism in which the disrupted proton migration path in CatchER is rescued by calcium binding. Our finding provides important insights for a strategy to design calcium sensors and suggests that CatchER could be a useful probe for FLIM imaging of calcium in situ. PMID:24836743

Zhuo, You; Solntsev, Kyril M; Reddish, Florence; Tang, Shen; Yang, Jenny J

2015-02-12

390

The use of time-resolved fluorescence imaging in the study of protein kinase C localisation in cells  

PubMed Central

Background Two-photon-excitation fluorescence lifetime imaging (2P-FLIM) was used to investigate the association of protein kinase C alpha (PKC?) with caveolin in CHO cells. PKC? is found widely in the cytoplasm and nucleus in most cells. Upon activation, as a result of increased intracellular Ca2+ and production of DAG, through G-protein coupled-phospholipase C signalling, PKC translocates to a variety of regions in the cell where it phosphorylates and interacts with many signalling pathways. Due to its wide distribution, discerning a particular interaction from others within the cell is extremely difficult Results Fluorescence energy transfer (FRET), between GFP-PKC? and DsRed-caveolin, was used to investigate the interaction between caveolin and PKC, an aspect of signalling that is poorly understood. Using 2P-FLIM measurements, the lifetime of GFP was found to decrease (quench) in certain regions of the cell from ~2.2 ns to ~1.5 ns when the GFP and DsRed were sufficiently close for FRET to occur. This only occurred when intracellular Ca2+ increased or in the presence of phorbol ester, and was an indication of PKC and caveolin co-localisation under these conditions. In the case of phorbol ester stimulated PKC translocation, as commonly used to model PKC activation, three PKC areas could be delineated. These included PKC? that was not associated with caveolin in the nucleus and cytoplasm, PKC? associated with caveolin in the cytoplasm/perinuclear regions and probably in endosomes, and PKC in the peripheral regions of the cell, possibly indirectly interacting with caveolin. Conclusion Based on the extent of lifetime quenching observed, the results are consistent with a direct interaction between PKC? and caveolin in the endosomes, and possibly an indirect interaction in the peripheral regions of the cell. The results show that 2P-FLIM-FRET imaging offers an approach that can provide information not only confirming the occurrence of specific protein-protein interactions but where they occur within the cell. PMID:15854225

Stubbs, Christopher D; Botchway, Stanley W; Slater, Simon J; Parker, Anthony W

2005-01-01

391

Probing Electronic and Vibrational Dynamics in Molecules by Time-Resolved Photoelectron, Auger-Electron, and X-ray Photon Scattering Spectroscopy  

PubMed Central

We present a unified description for time-resolved electron and photon scattering spectroscopies from molecules prepared in nonstationary states. Signals are expressed in terms of superoperator Green’s functions and a systematic procedure for treating various degrees of freedom consistently at different levels of theory is developed. The standard Fermi Golden Rule expressions for photelectron spectra, which are limited to broad, slowly-varying signals, are obtained as a limiting case of our more general theory that applies to broader parameter regimes. PMID:25730500

Bennett, Kochise; Kowalewski, Markus; Mukamel, Shaul

2015-01-01

392

Direct observation of back energy transfer in blue phosphorescent materials for organic light emitting diodes by time-resolved optical waveguide spectroscopy  

PubMed Central

We demonstrate a high-sensitive transient absorption technique for detection of excited states in an organic thin film by time-resolved optical waveguide spectroscopy. By using a laser beam as a probe light, we detect small change in the transient absorbance which is equivalent to 10?7 absorbance unit in a conventional method. This technique was applied to organic thin films of blue phosphorescent materials for organic light emitting diodes. We directly observed the back energy transfer from emitting guest molecules to conductive host molecules. PMID:23526833

Hirayama, H.; Sugawara, Y.; Miyashita, Y.; Mitsuishi, M.; Miyashita, T.

2013-01-01

393

Time-Resolved IR-Absorption Spectroscopy of Hot-Electron Dynamics in Satellite and Upper Conduction Bands in GaP  

NASA Technical Reports Server (NTRS)

The relaxation dynamics of hot electrons in the X6 and X7 satellite and upper conduction bands in GaP was directly measured by femtosecond UV-pump-IR-probe absorption spectroscopy. From a fit to the induced IR-absorption spectra the dominant scattering mechanism giving rise to the absorption at early delay times was determined to be intervalley scattering of electrons out of the X7 upper conduction-band valley. For long delay times the dominant scattering mechanism is electron-hole scattering. Electron transport dynamics of the upper conduction band of GaP has been time resolved.

Cavicchia, M. A.; Alfano, R. R.

1995-01-01

394

Tracking Local Conformational Changes of Ribonuclease A Using Picosecond Time-Resolved Fluorescence of the Six Tyrosine Residues  

PubMed Central

The six tyrosine residues of ribonuclease A (RNase A) are used as individual intrinsic probes for tracking local conformational changes during unfolding. The fluorescence decays of RNase A are well described by sums of three exponentials with decay times (?1 = 1.7 ns, ?2 = 180 ps, and ?3 = 30 ps) and preexponential coefficients (A1 = 1, A2 = 1, and A3 = 4) at pH 7, 25°C. The decay times are controlled by photo-induced electron transfer from individual tyrosine residues to the nearest disulphide (–SS–), bridge, which is distance (R) dependent. We assign ?1 to Tyr-76 (R = 12.8 Å), ?2 to Tyr-115 (R = 6.9 Å), and ?3 to Tyr-25, Tyr-73, Tyr-92, and Tyr-97 (all four at R = 5.5 ± 0.3 Å) at 23°C. On the basis of this assignment, the results show that, upon thermal or chemical unfolding only Tyr-25, Tyr-92, and Tyr-76 undergo significant displacement from their nearest –SS– bridge. Despite reporting on different regions of the protein, the concordance between the transition temperatures, Tm, obtained from Tyr-76 (Tm = 59.2°C) and Tyr-25 and Tyr-92 (Tm = 58.2°C) suggests a single unfolding event in this temperature range that affects all these regions similarly. PMID:17384067

Noronha, Melinda; Lima, João C.; Paci, Emanuele; Santos, Helena; Maçanita, António L.

2007-01-01

395

Investigation of verbal and visual working memory by multi-channel time-resolved functional near-infrared spectroscopy  

NASA Astrophysics Data System (ADS)

Working memory (WM) is fundamental for a number of cognitive processes, such as comprehension, reasoning and learning. WM allows the short-term maintenance and manipulation of the information selected by attentional processes. The goal of this study was to examine by time-resolved fNIRS neural correlates of the verbal and visual WM during forward and backward digit span (DF and DB, respectively) tasks, and symbol span (SS) task. A neural dissociation was hypothesised between the maintenance and manipulation processes. In particular, a dorsolateral/ventrolateral prefrontal cortex (DLPFC/VLPFC) recruitment was expected during the DB task, whilst a lateralised involvement of Brodmann Area (BA) 10 was expected during the execution of the DF task. Thirteen subjects were monitored by a multi-channel, dual-wavelength (690 and 829 nm) time-resolved fNIRS system during 3 minutes long DF and DB tasks and 4 minutes long SS task. The participants' mean memory span was calculated for each task: DF: 6.46+/-1.05 digits; DB: 5.62+/-1.26 digits; SS: 4.69+/-1.32 symbols. No correlation was found between the span level and the heart rate data (measured by pulse oximeter). As expected, DB elicited a broad activated area, in the bilateral VLPFC and the right DLPFC, whereas a more localised activation was observed over the right hemisphere during either DF (BA 10) or SS (BA 10 and 44). The robust involvement of the DLPFC during DB, compared to DF, is compatible with previous findings and with the key role of the central executive subserving in manipulating processes.

Contini, D.; Caffini, M.; Re, R.; Zucchelli, L.; Spinelli, L.; Basso Moro, S.; Bisconti, S.; Ferrari, M.; Quaresima, V.; Cutini, S.; Torricelli, A.

2013-03-01

396

Time-resolved fluorescence investigation of the human immunodeficiency virus type 1 nucleocapsid protein: influence of the binding of nucleic acids.  

PubMed Central

Depending on the HIV-1 isolate, MN or BH10, the nucleocapsid protein, NCp7, corresponds to a 55- or 71-amino acid length product, respectively. The MN NCp7 contains a single Trp residue at position 37 in the distal zinc finger motif, and the BH10 NCp7 contains an additional Trp, at position 61 in the C-terminal chain. The time-resolved intensity decay parameters of the zinc-saturated BH10 NCp7 were determined and compared to those of single-Trp-containing derivatives. The fluorescence decay of BH10 NCp7 could be clearly represented as a linear combination (with respect to both lifetimes and fractional intensities) of the individual emitting Trp residues. This suggested the absence of interactions between the two Trp residues, a feature that was confirmed by molecular modeling and fluorescence energy transfer studies. In the presence of tRNAPhe, taken as a RNA model, the same conclusions hold true despite the large fluorescence decrease induced by the binding of tRNAPhe. Indeed, the fluorescence of Trp37 appears almost fully quenched, in keeping with a stacking of this residue with the bases of tRNAPhe. Despite the multiple binding sites in tRNAPhe, the large prevalence of ultrashort lifetimes, associated with the stacking of Trp37, suggests that this stacking constitutes a major feature in the binding process of NCp7 to nucleic acids. In contrast, Trp61 only stacked to a small extent with tRNAPhe. The behavior of this residue in the tRNAPhe-NCp7 complexes appeared to be rather heterogeneous, suggesting that it does not constitute a major determinant in the binding process. Finally, our data suggested that the binding of NCp7 proteins from the two HIV-1 strains to nonspecific nucleic acid sequences was largely similar. PMID:10049336

Bombarda, E; Ababou, A; Vuilleumier, C; Gérard, D; Roques, B P; Piémont, E; Mély, Y

1999-01-01

397

Time-resolved fluorescence investigation of the human immunodeficiency virus type 1 nucleocapsid protein: influence of the binding of nucleic acids.  

PubMed

Depending on the HIV-1 isolate, MN or BH10, the nucleocapsid protein, NCp7, corresponds to a 55- or 71-amino acid length product, respectively. The MN NCp7 contains a single Trp residue at position 37 in the distal zinc finger motif, and the BH10 NCp7 contains an additional Trp, at position 61 in the C-terminal chain. The time-resolved intensity decay parameters of the zinc-saturated BH10 NCp7 were determined and compared to those of single-Trp-containing derivatives. The fluorescence decay of BH10 NCp7 could be clearly represented as a linear combination (with respect to both lifetimes and fractional intensities) of the individual emitting Trp residues. This suggested the absence of interactions between the two Trp residues, a feature that was confirmed by molecular modeling and fluorescence energy transfer studies. In the presence of tRNAPhe, taken as a RNA model, the same conclusions hold true despite the large fluorescence decrease induced by the binding of tRNAPhe. Indeed, the fluorescence of Trp37 appears almost fully quenched, in keeping with a stacking of this residue with the bases of tRNAPhe. Despite the multiple binding sites in tRNAPhe, the large prevalence of ultrashort lifetimes, associated with the stacking of Trp37, suggests that this stacking constitutes a major feature in the binding process of NCp7 to nucleic acids. In contrast, Trp61 only stacked to a small extent with tRNAPhe. The behavior of this residue in the tRNAPhe-NCp7 complexes appeared to be rather heterogeneous, suggesting that it does not constitute a major determinant in the binding process. Finally, our data suggested that the binding of NCp7 proteins from the two HIV-1 strains to nonspecific nucleic acid sequences was largely similar. PMID:10049336

Bombarda, E; Ababou, A; Vuilleumier, C; Gérard, D; Roques, B P; Piémont, E; Mély, Y

1999-03-01

398

Time-resolved spectrofluorometer for clinical tissue characterization during endoscopy  

NASA Astrophysics Data System (ADS)

Time-resolved fluorescence spectroscopy has the potential to provide more information for the detection of early cancer than continuous wave spectroscopy. A new optical fiber-based spectrofluorometer for time-resolved fluorescence spectroscopy of biological tissue during clinical endoscopy is presented. The apparatus is based on a nitrogen laser pumping a dye laser as excitation source and a streak camera coupled with a spectrograph as time-resolved spectrometer. The excitation and fluorescence light is carried by an optical fiber to the tissue under investigation and back to the detector, respectively. This optical fiber can be inserted into the biopsy channel of a conventional endoscope. Hence, the apparatus can be used to perform in situ tissue characterization during endoscopy. The instrument enables the measurement of the decays of entire fluorescence spectra within 15 s with a dynamic range of the spectro-temporal images of up to three orders of magnitude. Luminescence lifetimes from the sub ns up to the ms range can be measured. Spectral and temporal resolution, sensitivity, and dynamic range of the instrumentation were determined. The accuracy of the apparatus was checked by the measurement of the fluorescence lifetimes of various fluorophores with known lifetimes. For the first time, two-dimensional time-resolved spectra with sub-ns temporal resolution of tissue fluorescence of the human bladder, the bronchi, and the esophagus taken during endoscopy are presented as a demonstration of performance of the instrumentation. The excitation wavelengths were 337 nm in the case of the bladder and the esophagus and 480 nm in the case of the bronchi. Lifetime contrasts between normal and neoplastic tissue were found in all three organs. The spectral analysis of the fluorescence decays showed that the fluorescence between 370 and 490 nm, excited at 337 nm, consisted in several overlapping spectra. In the case of the esophagus, the contrast between normal and tumoral tissue was inverse in two different spectral bands proving the importance of the choice of the appropriate spectral range for time-resolved autofluorescence measurements for an optimal contrast. The in vivo fluorescence decay of the photosensitizers 5-aminolevulinic acid hexylester hydrochloride-induced protoporphyrin IX was measured in the human bladder and found to be mono-exponential with a lifetime of 15.9 (±1.2) ns. An in vivo fluorescence lifetime of 8.5 (±0.8) ns was found in the case of the photosensitizer 5, 10, 15, 20-tetra(m-hydroxyphenyl)chlorin (mTHPC) in the esophagus.

Glanzmann, Thomas; Ballini, Jean-Pierre; van den Bergh, Hubert; Wagnières, Georges

1999-10-01