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1

Intrinsic photosensitizer fluorescence measured using multi-diameter single-fiber spectroscopy in vivo  

NASA Astrophysics Data System (ADS)

Quantification of fluorescence in vivo is complicated by the influence of tissue optical properties on the collected fluorescence signal. When tissue optical properties in the measurement volume are quantified, one can obtain the intrinsic fluorescence, which equals the product of fluorophore absorption coefficient and quantum yield. We applied this method to in vivo single-fiber fluorescence spectroscopy measurements on mouse tongue, skin, liver, and oral squamous cell carcinoma, where we detected intrinsic fluorescence spectra of the photosensitizers chlorin e6 and Bremachlorin at t=[3,4.5,6,24,48] h incubation time. We observed a tissue-dependent maximum of 35% variation in the total correction factor over the visible wavelength range. Significant differences in spectral shape over time between sensitizers were observed. Although the wavelength position of the fluorescence intensity maximum for ce6 shifted to the red, Bremachlorin showed a blue shift. Furthermore, the Bremachlorin peak appeared to be broader than the ce6 fluorescence peak. Intrinsic fluorescence intensity, which can be related to photosensitizer concentration, was decreasing for all time points but showed significantly more Bremachlorin present compared to ce6 at long incubation times. Results from this study can be used to define an optimal treatment protocol for Bremachlorin-based photodynamic therapy.

van Leeuwen-van Zaane, Floor; Gamm, Ute A.; van Driel, Pieter B. A. A.; Snoeks, Thomas J.; de Bruijn, Henriette S.; van der Ploeg-van den Heuvel, Angelique; Sterenborg, Henricus J. C. M.; Löwik, Clemens W.; Amelink, Arjen; Robinson, Dominic J.

2014-01-01

2

In vivo characterization of myocardial infarction using fluorescence and diffuse reflectance spectroscopy  

NASA Astrophysics Data System (ADS)

We explore the feasibility of using combined fluorescence and diffuse reflectance spectroscopy to characterize a myocardial infarct at different developing stages. An animal study is conducted using rats with surgically induced myocaridal infarction (MI). In vivo fluorescence spectra at 337-nm excitation and diffuse reflectance between 400 and 900 nm are measured from the heart. Spectral acquisition is performed: 1. for normal heart tissue; 2. for the area immediately surrounding the infarct; and 3. for the infarcted tissue itself, one, two, three, and four weeks into MI development. Histological and statistical analyses are used to identify unique pathohistological features and spectral alterations associated with the investigated regions. The main alterations (p<0.05) in diffuse reflectance spectra are identified primarily between 450 and 600 nm. The dominant fluorescence alterations are increases in peak fluorescence intensity at 400 and 460 nm. The extent of these spectral alterations is related to the duration of the infarction. The findings of this study support the concept that optical spectroscopy could be useful as a tool to noninvasively determine the in vivo pathophysiological features of a myocardial infarct and its surrounding tissue, thereby providing real-time feedback to surgeons during various surgical interventions for MI.

Ti, Yalin; Chen, Poching; Lin, Wei-Chiang

2010-05-01

3

Bayesian model selection applied to the analysis of fluorescence correlation spectroscopy data of fluorescent proteins in vitro and in vivo.  

PubMed

Fluorescence correlation spectroscopy (FCS) is a powerful technique to investigate molecular dynamics with single molecule sensitivity. In particular, in the life sciences it has found widespread application using fluorescent proteins as molecularly specific labels. However, FCS data analysis and interpretation using fluorescent proteins remains challenging due to typically low signal-to-noise ratio of FCS data and correlated noise in autocorrelated data sets. As a result, naive fitting procedures that ignore these important issues typically provide similarly good fits for multiple competing models without clear distinction of which model is preferred given the signal-to-noise ratio present in the data. Recently, we introduced a Bayesian model selection procedure to overcome this issue with FCS data analysis. The method accounts for the highly correlated noise that is present in FCS data sets and additionally penalizes model complexity to prevent over interpretation of FCS data. Here, we apply this procedure to evaluate FCS data from fluorescent proteins assayed in vitro and in vivo. Consistent with previous work, we demonstrate that model selection is strongly dependent on the signal-to-noise ratio of the measurement, namely, excitation intensity and measurement time, and is sensitive to saturation artifacts. Under fixed, low intensity excitation conditions, physical transport models can unambiguously be identified. However, at excitation intensities that are considered moderate in many studies, unwanted artifacts are introduced that result in nonphysical models to be preferred. We also determined the appropriate fitting models of a GFP tagged secreted signaling protein, Wnt3, in live zebrafish embryos, which is necessary for the investigation of Wnt3 expression and secretion in development. Bayes model selection therefore provides a robust procedure to determine appropriate transport and photophysical models for fluorescent proteins when appropriate models are provided, to help detect and eliminate experimental artifacts in solution, cells, and in living organisms. PMID:25815704

Sun, Guangyu; Guo, Syuan-Ming; Teh, Cathleen; Korzh, Vladimir; Bathe, Mark; Wohland, Thorsten

2015-04-21

4

Noninvasive fluorescence excitation spectroscopy for the diagnosis of oral neoplasia in vivo  

NASA Astrophysics Data System (ADS)

Fluorescence excitation spectroscopy (FES) is an emerging approach to cancer detection. The goal of this pilot study is to evaluate the diagnostic potential of FES technique for the detection and characterization of normal and cancerous oral lesions in vivo. Fluorescence excitation (FE) spectra from oral mucosa were recorded in the spectral range of 340 to 600 nm at 635 nm emission using a fiberoptic probe spectrofluorometer to obtain spectra from the buccal mucosa of 30 sites of 15 healthy volunteers and 15 sites of 10 cancerous patients. Significant FE spectral differences were observed between normal and well differentiated squamous cell carcinoma (WDSCC) oral lesions. The FE spectra of healthy volunteers consists of a broad emission band around 440 to 470 nm, whereas in WDSCC lesions, a new primary peak was seen at 410 nm with secondary peaks observed at 505, 540, and 580 nm due to the accumulation of porphyrins in oral lesions. The FE spectral bands of the WDSCC lesions resemble the typical absorption spectra of a porphyrin. Three potential ratios (I410/I505, I410/I540, and I410/I580) were calculated from the FE spectra and used as input variables for a stepwise linear discriminant analysis (SLDA) for normal and WDSCC groups. Leave-one-out (LOO) method of cross-validation was performed to check the reliability on spectral data for tissue characterization. The diagnostic sensitivity and specificity were determined for normal and WDSCC lesions from the scatter plot of the discriminant function scores. It was observed that diagnostic algorithm based on discriminant function scores obtained by SLDA-LOO method was able to distinguish WDSCC from normal lesions with a sensitivity of 100% and specificity of 100%. Results of the pilot study demonstrate that the FE spectral changes due to porphyrin have a good diagnostic potential; therefore, porphyrin can be used as a native tumor marker.

Ebenezar, Jeyasingh; Ganesan, Singaravelu; Aruna, Prakasarao; Muralinaidu, Radhakrishnan; Renganathan, Kannan; Saraswathy, Thillai Rajasekaran

2012-09-01

5

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

6

In vivo detection of membrane protein expression using surface plasmon enhanced fluorescence spectroscopy (SPFS)  

Microsoft Academic Search

Surface plasmon enhanced fluorescence spectroscopy (SPFS) was applied for the detection of expression and functional incorporation of integral membrane proteins into plasma membranes of living cells in real time.A vesicular stomatitis virus (VSV) tagged mutant of photoreceptor bovine rhodopsin was generated for high level expression with the semliki forest virus (SFV) system. Adherent baby hamster kidney (BHK-21) cells were cultivated

Simone S. Krupka; Birgit Wiltschi; Ute Reuning; Kerstin Hölscher; Masahiko Hara; Eva-Kathrin Sinner

2006-01-01

7

Stationary spectroscopy of biotissues in vivo: Fluorescent studies of some pathological states  

NASA Astrophysics Data System (ADS)

The stationary spectra of autofluorescence, along with the reflection coefficient at the wavelength of excitation, are measured in vivo for some stomach tissues in the case of different pathological states (dysplasia, superficial gastritis, and cancer) using a nitrogen laser as the source of excitation (?rad=337.1 nm). The fluorescence spectra obtained are decomposed into Gaussian-Lorentzian components. It is found that, in development of dysplasia and tumor processes, at least seven groups of fluorophores can be distinguished that form the entire emission spectrum. The ratio between the fluorescence intensities of flavins and NAD(P)H is determined and the degree of respiratory activity of cells estimated for the states considered. The quantum yields of fluorescence of the biotissues under investigation are estimated.

Giraev, K. M.; Ashurbekov, N. A.; Medzhidov, R. T.

2003-11-01

8

Two-photon excited fluorescence lifetime imaging and spectroscopy of melanins in vitro and in vivo  

PubMed Central

Abstract. Changes in the amounts of cellular eumelanin and pheomelanin have been associated with carcinogenesis. The goal of this work is to develop methods based on two-photon-excited-fluorescence (TPEF) for measuring relative concentrations of these compounds. We acquire TPEF emission spectra (?ex=1000??nm) of melanin in vitro from melanoma cells, hair specimens, and in vivo from healthy volunteers. We find that the pheomelanin emission peaks at approximately 615 to 625 nm and eumelanin exhibits a broad maximum at 640 to 680 nm. Based on these data we define an optical melanin index (OMI) as the ratio of fluorescence intensities at 645 and 615 nm. The measured OMI for the MNT-1 melanoma cell line is 1.6±0.22 while the Mc1R gene knockdown lines MNT-46 and MNT-62 show substantially greater pheomelanin production (OMI=0.5±0.05 and 0.17±0.03, respectively). The measured values are in good agreement with chemistry-based melanin extraction methods. In order to better separate melanin fluorescence from other intrinsic fluorophores, we perform fluorescence lifetime imaging microscopy of in vitro specimens. The relative concentrations of keratin, eumelanin, and pheomelanin components are resolved using a phasor approach for analyzing lifetime data. Our results suggest that a noninvasive TPEF index based on spectra and lifetime could potentially be used for rapid melanin ratio characterization both in vitro and in vivo. PMID:23235925

Krasieva, Tatiana B.; Stringari, Chiara; Liu, Feng; Sun, Chung-Ho; Kong, Yu; Balu, Mihaela; Meyskens, Frank L.; Gratton, Enrico; Tromberg, Bruce J.

2012-01-01

9

In vivo detection of membrane protein expression using surface plasmon enhanced fluorescence spectroscopy (SPFS).  

PubMed

Surface plasmon enhanced fluorescence spectroscopy (SPFS) was applied for the detection of expression and functional incorporation of integral membrane proteins into plasma membranes of living cells in real time. A vesicular stomatitis virus (VSV) tagged mutant of photoreceptor bovine rhodopsin was generated for high level expression with the semliki forest virus (SFV) system. Adherent baby hamster kidney (BHK-21) cells were cultivated on fibronectin-coated gold surfaces and infected with genetically engineered virus driving the expression of rhodopsin. Using premixed fluorescently (Alexa Fluor 647) labeled anti-mouse secondary antibody and monoclonal anti-VSV primary antibody, expression of rhodopsin in BHK-21 cells was monitored by SPFS. Fluorescence enhancement by surface plasmons occurs exclusively in the close vicinity of the gold surface. Thus, only the Alexa Fluor 647 labeled antibodies binding to the VSV-tag at rhodopsin molecules exposed on the cell surface experienced fluorescence enhancement, whereas, unbound antibody molecules in the bulk solution were negligibly excited. With this novel technique, we successfully recorded an increase of fluorescence with proceeding rhodopsin expression. Thus, we were able to observe the incorporation of heterologously expressed rhodopsin in the plasma membrane of living cells in real time using a relatively simple and rapid method. We confirmed our results by comparison with conventional wide field fluorescence microscopy. PMID:16530398

Krupka, Simone S; Wiltschi, Birgit; Reuning, Ute; Hölscher, Kerstin; Hara, Masahiko; Sinner, Eva-Kathrin

2006-08-15

10

In vivo detection of epileptic brain tissue using static fluorescence and diffuse reflectance spectroscopy  

NASA Astrophysics Data System (ADS)

Diffuse reflectance and fluorescence spectroscopy are used to detect histopathological abnormalities of an epileptic brain in a human subject study. Static diffuse reflectance and fluorescence spectra are acquired from normal and epileptic brain areas, defined by electrocorticography (ECoG), from pediatric patients undergoing epilepsy surgery. Biopsy specimens are taken from the investigated sites within an abnormal brain. Spectral analysis reveals significant differences in diffuse reflectance spectra and the ratio of fluorescence and diffuse reflectance spectra from normal and epileptic brain areas defined by ECoG and histology. Using these spectral differences, tissue classification models with accuracy above 80% are developed based on linear discriminant analysis. The differences between the diffuse reflectance spectra from the normal and epileptic brain areas observed in this study are attributed to alterations in the static hemodynamic characteristics of an epileptic brain, suggesting a unique association between the histopathological and the hemodynamic abnormalities in an epileptic brain.

Yadav, Nitin; Bhatia, Sanjiv; Ragheb, John; Mehta, Rupal; Jayakar, Prasanna; Yong, William; Lin, Wei-Chiang

2013-02-01

11

Two-photon excited fluorescence spectroscopy and imaging of melanin in vitro and in vivo  

NASA Astrophysics Data System (ADS)

The ability to detect early melanoma non-invasively would improve clinical outcome and reduce mortality. Recent advances in two-photon excited fluorescence (TPEF) in vivo microscopy offer a powerful tool in early malignant melanoma diagnostics. The goal of this work was to develop a TPEF optical index for measuring relative concentrations of eumelanin and pheomelanin since ex vivo studies show that changes in this ratio have been associated with malignant transformation. We acquired TPEF emission spectra (?ex=1000 nm) of melanin from several specimens, including human hair, malignant melanoma cell lines, and normal melanocytes and keratinocytes in different skin layers (epidermis, papillary dermis) in five healthy volunteers in vivo. We found that the pheomelanin emission peaks at around 620 nm and is blue-shifted from the eumelanin with broad maximum at 640-680nm. We defined "optical melanin index" (OMI) as a ratio of fluorescence signal intensities measured at 645 nm and 615nm. The measured OMI for a melanoma cell line MNT-1 was 1.6+/-0.2. The MNT-46 and MNT-62 lines (Mc1R gene knockdown) showed an anticipated change in melanins production ratio and had OMI of 0.55+/-0.05 and 0.17+/-0.02, respectively, which strongly correlated with HPLC data obtained for these lines. Average OMI measured for basal cells layers (melanocytes and keratinocytes) in normal human skin type I, II-III (not tanned and tanned) in vivo was 0.5, 1.05 and 1.16 respectively. We could not dependably detect the presence of pheomelanin in highly pigmented skin type V-VI. These data suggest that a non-invasive TPEF index could potentially be used for rapid melanin ratio characterization both in vitro and in vivo, including pigmented lesions.

Krasieva, Tatiana B.; Liu, Feng; Sun, Chung-Ho; Kong, Yu; Balu, Mihaela; Meyskens, Frank L.; Tromberg, Bruce J.

2012-03-01

12

Delta-ALA-mediated fluorescence spectroscopy of gastrointestinal tumors: comparison of in vivo and in vitro results  

NASA Astrophysics Data System (ADS)

The limitations of standard endoscopy for detection of dysplastic changes of mucosa are significant challenge and initiate development of new photodiagnostic techniques, additional to diagnostic possibilities of standard endoscopic equipment. One of the most widely examined optical modalities is the laser- or light-induced fluorescence spectroscopy (LIFS), because of its rapid and highly sensitive response to early biochemical and morphological changes in biological tissues. In the recent study delta-aminolevulinic acid/protoporphyrin IX is used as fluorescent marker for dysplasia and tumor detection in esophagus and stomach. The ? -ALA is administered per os six hours before measurements at dose 20mg/kg weight. High-power light-emitting diode at 405 nm is used as an excitation source. Special opto-mechanical device is built to use the light guide of standard video-endoscopic system. Through endoscopic instrumental channel a fiber is applied to return information about fluorescence to microspectrometer. The fluorescence detected from in vivo tumor sites has very complex spectral origins. It consists of autofluorescence, fluorescence from exogenous fluorophores and re-absorption from the chromophores accumulated in the tissue investigated. Mucosa autofluorescence lies at 450-600 nm region. The fluorescence of PpIX is clearly pronounced at the 630-710 nm region. Deep minima in the tumor fluorescence signals are observed in the region 540-575 nm, related to hemoglobin re-absorption. Such high hemoglobin content is an indication of the tumors vascularization and it is clearly pronounced in all dysplastic and tumor sites investigated. After formalin conservation for in vitro samples hemoglobin absorption is strongly reduced that increases mucous fluorescence signal in green-yellow spectral region. Simultaneously the maxima at 635 nm and 720 nm are reduced.

Vladimirov, B.; Borisova, E.; Avramov, L.

2007-06-01

13

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

14

Ex vivo optical coherence tomography and laser induced fluorescence spectroscopy imaging of murine gastrointestinal tract  

NASA Astrophysics Data System (ADS)

Optical Coherence Tomography (OCT) and Laser Induced Fluorescence Spectroscopy (LIF) have separately been found to have clinical potential in identifying human gastrointestinal (GI) pathologies, yet their diagnostic capability in mouse models of human disease is unknown. We combine the two modalities to survey the GI tract of a variety of mouse strains and sample dysplasias and inflammatory bowel disease (IBD) of the small and large intestine. Segments of duodenum and lower colon 2.5 cm in length and the entire esophagus from 10 mice each of two colon cancer models (ApcMin and AOM treated A/J) and two IBD models (Il-2 and Il-10) and 5 mice each of their respective controls were excised. OCT images and LIF spectra were obtained simultaneously from each tissue sample within 1 hour of extraction. Histology was used to classify tissue regions as normal, Peyer"s patch, dysplasia, adenoma, or IBD. Features in corresponding regions of OCT images were analyzed. Spectra from each of these categories were averaged and compared via the student's t-test. Features in OCT images correlated to histology in both normal and diseased tissue samples. In the diseased samples, OCT was able to identify early stages of mild colitis and dysplasia. In the sample of IBD, the LIF spectra displayed unique peaks at 635nm and 670nm, which were attributed to increased porphyrin production in the proliferating bacteria of the disease. These peaks have the potential to act as a diagnostic for IBD. OCT and LIF appear to be useful and complementary modalities for imaging mouse models.

Hariri, Lida; Tumlinson, Alexandre R.; Wade, Norman; Besselsen, David; Utzinger, Urs; Gerner, Eugene; Barton, Jennifer

2005-04-01

15

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

16

Prostate cancer detection using combined auto-fluorescence and light reflectance spectroscopy: ex vivo study of human prostates  

PubMed Central

This study was conducted to evaluate the capability of detecting prostate cancer (PCa) using auto-fluorescence lifetime spectroscopy (AFLS) and light reflectance spectroscopy (LRS). AFLS used excitation at 447 nm with four emission wavelengths (532, 562, 632, and 684 nm), where their lifetimes and weights were analyzed using a double exponent model. LRS was measured between 500 and 840 nm and analyzed by a quantitative model to determine hemoglobin concentrations and light scattering. Both AFLS and LRS were taken on n = 724 distinct locations from both prostate capsular (nc = 185) and parenchymal (np = 539) tissues, including PCa tissue, benign peripheral zone tissue and benign prostatic hyperplasia (BPH), of fresh ex vivo radical prostatectomy specimens from 37 patients with high volume, intermediate-to-high-grade PCa (Gleason score, GS ?7). AFLS and LRS parameters from parenchymal tissues were analyzed for statistical testing and classification. A feature selection algorithm based on multinomial logistic regression was implemented to identify critical parameters in order to classify high-grade PCa tissue. The regression model was in turn used to classify PCa tissue at the individual aggressive level of GS = 7,8,9. Receiver operating characteristic curves were generated and used to determine classification accuracy for each tissue type. We show that our dual-modal technique resulted in accuracies of 87.9%, 90.1%, and 85.1% for PCa classification at GS = 7, 8, 9 within parenchymal tissues, and up to 91.1%, 91.9%, and 94.3% if capsular tissues were included for detection. Possible biochemical and physiological mechanisms causing signal differences in AFLS and LRS between PCa and benign tissues were also discussed. PMID:24877012

Sharma, Vikrant; Olweny, Ephrem O.; Kapur, Payal; Cadeddu, Jeffrey A.; Roehrborn, Claus G.; Liu, Hanli

2014-01-01

17

Combined fluorescence and reflectance spectroscopy for in vivo quantification of cancer biomarkers in low- and high-grade glioma surgery  

PubMed Central

Biomarkers are indicators of biological processes and hold promise for the diagnosis and treatment of disease. Gliomas represent a heterogeneous group of brain tumors with marked intra- and inter-tumor variability. The extent of surgical resection is a significant factor influencing post-surgical recurrence and prognosis. Here, we used fluorescence and reflectance spectral signatures for in vivo quantification of multiple biomarkers during glioma surgery, with fluorescence contrast provided by exogenously-induced protoporphyrin IX (PpIX) following administration of 5-aminolevulinic acid. We performed light-transport modeling to quantify multiple biomarkers indicative of tumor biological processes, including the local concentration of PpIX and associated photoproducts, total hemoglobin concentration, oxygen saturation, and optical scattering parameters. We developed a diagnostic algorithm for intra-operative tissue delineation that accounts for the combined tumor-specific predictive capabilities of these quantitative biomarkers. Tumor tissue delineation achieved accuracies of up to 94% (specificity = 94%, sensitivity = 94%) across a range of glioma histologies beyond current state-of-the-art optical approaches, including state-of-the-art fluorescence image guidance. This multiple biomarker strategy opens the door to optical methods for surgical guidance that use quantification of well-established neoplastic processes. Future work would seek to validate the predictive power of this proof-of-concept study in a separate larger cohort of patients. PMID:22112112

Valdés, Pablo A.; Kim, Anthony; Leblond, Frederic; Conde, Olga M.; Harris, Brent T.; Paulsen, Keith D.; Wilson, Brian C.; Roberts, David W.

2011-01-01

18

In vivo fluorescence lifetime tomography  

NASA Astrophysics Data System (ADS)

Local molecular and physiological processes can be imaged in vivo through perturbations in the fluorescence lifetime (FLT) of optical imaging agents. In addition to providing functional information, FLT methods can quantify specific molecular events and multiplex diagnostic and prognostic information. We have developed a fluorescence lifetime diffuse optical tomography (DOT) system for in vivo preclinical imaging. Data is captured using a time-resolved intensified charge coupled device (ICCD) system to measure fluorescence excitation and emission in the time domain. Data is then converted to the frequency domain, and we simultaneously reconstruct images of yield and lifetime using an extension to the normalized Born approach. By using differential phase measurements, we demonstrate DOT imaging of short lifetimes (from 350 ps) with high precision (+/-5 ps). Furthermore, this system retains the efficiency, speed, and flexibility of transmission geometry DOT. We demonstrate feasibility of FLT-DOT through a progressive series of experiments. Lifetime range and repeatability are first measured in phantoms. Imaging of subcutaneous implants then verifies the FLT-DOT approach in vivo in the presence of inhomogeneous optical properties. Use in a common research scenario is ultimately demonstrated by imaging accumulation of a targeted near-infrared (NIR) fluorescent-labeled peptide probe (cypate-RGD) in a mouse with a subcutaneous tumor.

Nothdurft, Ralph E.; Patwardhan, Sachin V.; Akers, Walter; Ye, Yunpeng; Achilefu, Samuel; Culver, Joseph P.

2009-03-01

19

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

20

In vivo native fluorescence spectroscopy and nicotinamide adinine dinucleotide/flavin adenine dinucleotide reduction and oxidation states of oral submucous fibrosis for chemopreventive drug monitoring  

NASA Astrophysics Data System (ADS)

Native fluorescence spectroscopy has shown potential to characterize and diagnose oral malignancy. We aim at extending the native fluorescence spectroscopy technique to characterize normal and oral submucous fibrosis (OSF) patients under pre- and post-treated conditions, and verify whether this method could also be considered in the monitoring of therapeutic prognosis noninvasively. In this study, 28 normal subjects and 28 clinically proven cases of OSF in the age group of 20 to 40 years are diagnosed using native fluorescence spectroscopy. The OSF patients are given dexamethasone sodium phosphate and hyaluronidase twice a week for 6 weeks, and the therapeutic response is monitored using fluorescence spectroscopy. The fluorescence emission spectra of normal and OSF cases of both pre- and post-treated conditions are recorded in the wavelength region of 350 to 600 nm at an excitation wavelength of 330 nm. The statistical significance is verified using discriminant analysis. The oxidation-reduction ratio of the tissue is also calculated using the fluorescence emission intensities of flavin adenine dinucleotide and nicotinamide adinine dinucleotide at 530 and 440 nm, respectively, and they are compared with conventional physical clinical examinations. This study suggests that native fluorescence spectroscopy could also be extended to OSF diagnosis and therapeutic prognosis.

Sivabalan, Shanmugam; Vedeswari, C. Ponranjini; Jayachandran, Sadaksharam; Koteeswaran, Dornadula; Pravda, Chidambaranathan; Aruna, Prakasa Rao; Ganesan, Singaravelu

2010-01-01

21

Two-photon excited fluorescence lifetime imaging and spectroscopy of melanins in  

E-print Network

Two-photon excited fluorescence lifetime imaging and spectroscopy of melanins in vitro and in vivo and spectroscopy of melanins in vitro and in vivo Tatiana B. Krasieva,a Chiara Stringari,b Feng Liu,c Chung-Ho Sun ¼ 1000 nm) of melanin in vitro from melanoma cells, hair specimens, and in vivo from healthy volunteers

Chen, Zhongping

22

CHICKEN DISEASE CHARACTERIZATION BY FLUORESCENCE SPECTROSCOPY  

Technology Transfer Automated Retrieval System (TEKTRAN)

Fluorescence spectroscopy was used to characterize chicken carcass spectra. Spectral signatures of three different disease categories of poultry carcasses (airsacculitis, cadaver, and septicemia) were obtained from fluorescence emission measurements in the wavelength range of 360 to 600 nm with 330 ...

23

CHICKEN DISEASE CHARACTERIZATION BY FLUORESCENCE SPECTROSCOPY  

Technology Transfer Automated Retrieval System (TEKTRAN)

Fluorescence spectroscopy was used to characterize chicken carcass diseases. Spectral signatures of three different disease categories of poultry carcasses, airsacculitis, cadaver and septicemia, were obtained from fluorescence emission measurements in the wavelength range of 360 to 600 nm with 330 ...

24

Fluorescent labeling of renal cells in vivo.  

PubMed

In vivo fluorescence imaging, using confocal or multiphoton microscopes, provides a powerful method to analyze kidney function in experimental animals. In this review, the preparation used for physiological studies in rats is described. A variety of fluorescent probes are available to study glomerular permeability, renal blood flow, peritubular capillary permeability, cell ion concentrations, tubule transport properties, and the functional status of renal cells. We have recently used micropuncture techniques and an adenovirus vector to accomplish gene transfer into kidney tubule and endothelial cells; this new methodology will allow the dynamic study of fluorescently-labeled proteins. Two examples of the use of two-photon fluorescence microscopy to study renal pathophysiology, namely polycystic kidney disease and renal ischemia, are presented. Software is available to quantify data collected from in vivo imaging experiments and to construct 3-dimensional images of renal structures. Two-photon or confocal microscopy offers many opportunities for a better understanding of kidney function in health and disease. PMID:16543774

Ashworth, Sharon L; Tanner, George A

2006-01-01

25

Glucose Recognition in Vitro Using Fluorescent Spectroscopy  

SciTech Connect

Diabetes is a disease that affects over 16 million people in the USA at a cost of 100 billion dollars annually. The ability to regulate insulin delivery in people with Type 1 diabetes is imperative as is the need to manage glucose levels in all people with this disease. Our current method for monitoring glucose is a (FDA approved) minimally invasive enzymatic sensor that can measure glucose levels in vivo for three days. We are focused on developing a noninvasive implantable glucose sensor that will be interrogated by an external device. The material must be robust, easy to process, biocompatible and resistant to biofouling. In this Presentation we will discuss the development of a new polymeric matrix that can recognize physiological levels of glucose in vitro using fluorescent spectroscopy.

Noronha, G; Heiss, A M; Reilly, J R; Vachon, Jr, D J; Cary, D R; Zaitseva, N P; Reibold, R A; Lane, S M; Peyser, T A; Satcher, J H

2001-04-25

26

Fluorescence Correlation Spectroscopy of Gold Nanoparticles  

Microsoft Academic Search

Diffusion dynamics of gold nanoparticles (GNPs) was studied by fluorescence correlation spectroscopy (FCS). The fluorescence was studied by exciting the particles by green laser (532 nm), which is far from longitudinal plasmon band of nanorods. Transmission electron microscope (TEM) and UV-Vis-NIR spectrometer were used to characterize the gold nanoparticles. Despite their low quantum yields, GNPs possess the native fluorescence. The excellent

P. Sri Balaji; A. V. R. Murthy; Neha Tiwari; Sulabha Kulkarni

2012-01-01

27

Photon-induced fluorescence spectroscopy (PIFS)  

Microsoft Academic Search

Photon-induced fluorescence spectroscopy (PIFS) using monochromatized synchrotron radiation for exciting the sample selectively and a fluorescence spectrometer with a position-sensitive photon detector for recording its fluorescence spectrum is a very powerful experimental technique especially for the investigation of atomic and molecular photoionization and photodissociation. To reduce the contributions of higher harmonics of the undulator radiation which are also transmitted through

H. Schmoranzer; H. Liebel; F. Vollweiler; R. Müller-Albrecht; A. Ehresmann; K.-H. Schartner; B. Zimmermann

2001-01-01

28

Laser-induced fluorescence spectroscopy in tissue local necrosis detection  

NASA Astrophysics Data System (ADS)

The recent effort leads to reliable imaging techniques which can help to a surgeon during operations. The fluorescence spectroscopy was selected as very useful online in vivo imaging method to organics and biological materials analysis. The presented work scopes to a laser induced fluorescence spectroscopy technique to detect tissue local necrosis in small intestine surgery. In first experiments, we tested tissue auto-fluorescence technique but a signal-to-noise ratio didn't express significant results. Then we applied a contrast dye - IndoCyanine Green (ICG) which absorbs and emits wavelengths in the near IR. We arranged the pilot experimental setup based on highly coherent extended cavity diode laser (ECDL) used for stimulating of some critical areas of the small intestine tissue with injected ICG dye. We demonstrated the distribution of the ICG exciter with the first file of shots of small intestine tissue of a rabbit that was captured by high sensitivity fluorescent cam.

Cip, Ondrej; Buchta, Zdenek; Lesundak, Adam; Randula, Antonin; Mikel, Bretislav; Lazar, Josef; Veverkova, Lenka

2014-03-01

29

Fluorescence-force spectroscopy at the single molecule level  

NASA Astrophysics Data System (ADS)

During the past decade, various powerful single-molecule techniques have evolved and helped to address important questions in life sciences. As the single molecule techniques become mature, there is increasingly pressing need to maximize the information content of the analysis in order to be able to study more complex systems that better approximate in-vivo conditions. Here, we develop a fluorescence-force spectroscopy method to combine single-molecule fluorescence spectroscopy with optical tweezers. Optical tweezers are used to manipulate and observe mechanical properties on the nanometer scale and piconewton force range. However, once the force range is in the low piconewton range or less, the spatial resolution of optical tweezers decreases significantly. In combination with fluorescence spectroscopy, like single molecule Forster (or fluorescence) resonance energy transfer (FRET) whose detectable distance range is approximately 3-10 nm, we are able to observe nanometer fluctuations and internal conformational changes in a low-force regime. The possibility to place fluorescent labels at nearly any desired position and a sophisticated design of the experiment increases the amount of information that can be extracted in contrast to pure mechanical or fluorescence experiments. We demonstrate the applications of this method to various biological systems including: 1) to measure the effect of very low forces on the nanometer scale conformational transitions of the DNA four-way (Holliday) junction; 2) to dissect protein diffusion and dissociation mechanisms on single stranded DNA, 3) to calibrate FRET-based in-vivo force sensors and 4) to study mechanical unfolding of single proteins. The results could not have been obtained with fluorescence or force measurement alone, and clearly demonstrates the power and generality of our approach. Finally, we show that self-quenching of two identical fluorophores can be used to detect small conformational dynamics corresponding to sub-nanometer distance changes of single molecules in a FRET-insensitive short range (< 3 nm), extending the detectable distance range of our fluorescence-force spectroscopy method.

Zhou, Ruobo

30

Accurate fluorescence quantum yield determination by fluorescence correlation spectroscopy.  

PubMed

Here, we present a comparative method for the accurate determination of fluorescence quantum yields (QYs) by fluorescence correlation spectroscopy. By exploiting the high sensitivity of single-molecule spectroscopy, we obtain the QYs of samples in the microliter range and at (sub)nanomolar concentrations. Additionally, in combination with fluorescence lifetime measurements, our method allows the quantification of both static and collisional quenching constants. Thus, besides being simple and fast, our method opens up the possibility to photophysically characterize labeled biomolecules under application-relevant conditions and with low sample consumption, which is often important in single-molecule studies. PMID:25768035

Kempe, Daryan; Schöne, Antonie; Fitter, Jörg; Gabba, Matteo

2015-04-01

31

Monitoring beer during storage by fluorescence spectroscopy  

Microsoft Academic Search

The present study demonstrates the use of fluorescence spectroscopy and multivariate data analysis for monitoring changes in beer during storage. Total luminescence spectra and synchronous scanning fluorescence spectra were recorded for fresh beer and beer samples stored in clear glass vials for three weeks in darkness at 4°C and at 22°C, and exposed to light at 22°C, respectively. A pronounced

Ewa Sikorska; Tomasz Górecki; Igor V. Khmelinskii; Marek Sikorski; Denis De Keukeleire

2006-01-01

32

Fluorescence correlation spectroscopy near individual gold nanoparticle  

Microsoft Academic Search

Dynamic behavior of fluorescent molecules near an individual gold nanoparticle is investigated experimentally by fluorescence correlation spectroscopy method. The gold particle that acts as an optical nano-antenna presents significant near-field volume reduction. The single molecule diffusion behavior is clearly observed within a reduced near-field volume due to a highly localized field enhancement. The near-field volume and fluorescence enhancement are polarization

Qingyan Wang; Guowei Lu; Lei Hou; Tianyue Zhang; Chunxiong Luo; Hong Yang; Grégory Barbillon; Franck H. Lei; Christophe A. Marquette; Pascal Perriat; Olivier Tillement; Stéphane Roux; Qi Ouyang; Qihuang Gong

2011-01-01

33

Fluorescence spectroscopy of rhodopsins: insights and approaches.  

PubMed

Fluorescence spectroscopy has become an established tool at the interface of biology, chemistry and physics because of its exquisite sensitivity and recent technical advancements. However, rhodopsin proteins present the fluorescence spectroscopist with a unique set of challenges and opportunities due to the presence of the light-sensitive retinal chromophore. This review briefly summarizes some approaches that have successfully met these challenges and the novel insights they have yielded about rhodopsin structure and function. We start with a brief overview of fluorescence fundamentals and experimental methodologies, followed by more specific discussions of technical challenges rhodopsin proteins present to fluorescence studies. Finally, we end by discussing some of the unique insights that have been gained specifically about visual rhodopsin and its interactions with affiliate proteins through the use of fluorescence spectroscopy. This article is part of a Special Issue entitled: Retinal Proteins - You can teach an old dog new tricks. PMID:24183695

Alexiev, Ulrike; Farrens, David L

2014-05-01

34

Predicting in vivo fluorescence lifetime behavior of near-infrared fluorescent contrast agents  

E-print Network

to whole-animal preclinical imaging.1 While steady-state fluorescence imaging intensity versus wavelength reporters for use as FLT probes in vivo, thereby minimizing the use of animals for FLT screening studiesPredicting in vivo fluorescence lifetime behavior of near-infrared fluorescent contrast agents

Larson-Prior, Linda

35

Photodynamics of Red Fluorescent Proteins Studied by Fluorescence Correlation Spectroscopy  

PubMed Central

Red fluorescent proteins are important tools in fluorescence-based life science research. Recently, we have introduced eqFP611, a red fluorescent protein with advantageous properties from the sea anemone Entacmaea quadricolor. Here, we have studied the submillisecond light-driven intramolecular dynamics between bright and dark states of eqFP611 and, for comparison, drFP583 (DsRed) by using fluorescence correlation spectroscopy on protein solutions. A three-state model with one dark and two fluorescent states describes the power-dependence of the flickering dynamics of both proteins at different excitation wavelengths. It involves two light-driven conformational transitions. We have also studied the photodynamics of individual (monomeric) eqFP611 molecules immobilized on surfaces. The flickering rates and dark state fractions of eqFP611 bound to polyethylene glycol-covered glass surfaces were identical to those measured in solution, showing that the bound FPs behaved identically. A second, much slower flickering process was observed on the 10-ms timescale. Deposition of eqFP611 molecules on bare glass surfaces yielded bright fluorescence without any detectable flickering and a >10-fold decreased photobleaching yield. These observations underscore the intimate connection between protein motions and photophysical processes in fluorescent proteins. PMID:14695280

Schenk, Andreas; Ivanchenko, Sergey; Röcker, Carlheinz; Wiedenmann, Jörg; Nienhaus, G. Ulrich

2004-01-01

36

Volumetric tomography of fluorescent proteins through small animals in vivo  

E-print Network

Volumetric tomography of fluorescent proteins through small animals in vivo Giannis Zacharakis of the art and should find wide in vivo imaging applications in drug discovery, immunology, and cancer research. fluorescence whole-body imaging imaging gene expression gene transfer multispectral imaging

Lorenzo, Jorge Ripoll

37

Fluorescence Intensity and Lifetime Distribution Analysis: Toward Higher Accuracy in Fluorescence Fluctuation Spectroscopy  

Microsoft Academic Search

Fluorescence fluctuation methods such as fluorescence correlation spectroscopy and fluorescence intensity distribution analysis (FIDA) have proven to be versatile tools for studying molecular interactions with single molecule sensitivity. Another well-known fluorescence technique is the measurement of the fluorescence lifetime. Here, we introduce a method that combines the benefits of both FIDA and fluorescence lifetime analysis. It is based on fitting

Kaupo Palo; Leif Brand; Christian Eggeling; Stefan Jäger; Peet Kask; Karsten Gall

2002-01-01

38

Ultraviolet, Visible, and Fluorescence Spectroscopy  

NASA Astrophysics Data System (ADS)

Spectroscopy in the ultraviolet-visible (UV-Vis) range is one of the most commonly encountered laboratory techniques in food analysis. Diverse examples, such as the quantification of macrocomponents (total carbohydrate by the phenol-sulfuric acid method), quantification of microcomponents, (thiamin by the thiochrome fluorometric procedure), estimates of rancidity (lipid oxidation status by the thiobarbituric acid test), and surveillance testing (enzyme-linked immunoassays), are presented in this text. In each of these cases, the analytical signal for which the assay is based is either the emission or absorption of radiation in the UV-Vis range. This signal may be inherent in the analyte, such as the absorbance of radiation in the visible range by pigments, or a result of a chemical reaction involving the analyte, such as the colorimetric copper-based Lowry method for the analysis of soluble protein.

Penner, Michael H.

39

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

40

The effects of flow rate on in vivo fluorescence measurements  

E-print Network

THE EFFECTS OF FLOW RATE ON IN VIVO FLUORESCENCE MEASUREMENTS A Thesis by STEPHEN THOMAS SWEET Submitted to the Graduate College of Texas AgiM University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE May... 1988 Major Subject: Oceanography THE EFFECTS OF FLOW RATE ON IN VIVO FLUORESCENCE MEASUREMENTS A Thesis by STEPHEN THOMAS SWEET Approved as to style and content by: D. R. Schink (Chairman) G. A. Fryxe)l (Member) h . . Sweet (Member) G. T...

Sweet, Stephen Thomas

1988-01-01

41

In Vivo Monitoring of Fluorescently Labeled Cancer Cells  

Microsoft Academic Search

Whole animal in vivo imaging has contributed significantly to the detection of disease progression and drug efficacy for the past several years (1). With the introduction of sensitive imaging instruments, applications in fluorescent imaging have expanded to monitoring tumor cell growth and gene expression. The new fluorescent proteins have improved brightness, photostability and better tissue penetration of signals at a

Eva Barton; Angel Ang; Kevin Francis; Jae-Beom Kim

42

Fluorescent Multiblock ?-Conjugated Polymer Nanoparticles for In Vivo Tumor Targeting  

PubMed Central

Highly fluorescent multiblock conjugated polymer nanoparticles with folic acid surface ligands are highly effective for bioimaging and in vivo tumor targeting. The targeted nanoparticles were preferentially localized in tumor cells in vivo, thereby illustrating their potential for diagnostic and therapeutic applications. PMID:23794490

Ahmed, Eilaf; Morton, Stephen W.

2014-01-01

43

In vivo fluorescence lifetime tomography Ralph E. Nothdurft  

E-print Network

and phantoms to whole-body imaging of animals have been limited, likely due to the engineering chal- lenges a fluorescence lifetime diffuse optical tomography DOT system for in vivo preclinical imaging. Data is captured inherent to deep tissue in vivo imaging. This is particu- larly true for near-infrared NIR optical imaging

Larson-Prior, Linda

44

Multicontrast photoacoustic in vivo imaging using near-infrared fluorescent  

E-print Network

images of a mouse as 2D sections of a whole animal, and as localized 3D volumetric images with highMulticontrast photoacoustic in vivo imaging using near-infrared fluorescent proteins Arie Krumholz1 Einstein College of Medicine, Bronx, NY 10461, USA. Non-invasive imaging of biological processes in vivo

Verkhusha, Vladislav V.

45

Diagnosing breast cancer using diffuse reflectance spectroscopy and intrinsic fluorescence spectroscopy  

E-print Network

Using diffuse reflectance spectroscopy and intrinsic fluorescence spectroscopy, we have developed an algorithm that successfully classifies normal breast tissue, fibrocystic change, fibroadenoma, and infiltrating ductal ...

Fitzmaurice, Maryann

46

Plasmon-controlled fluorescence: a new paradigm in fluorescence spectroscopy  

PubMed Central

Fluorescence spectroscopy is widely used in biological research. Until recently, essentially all fluorescence experiments were performed using optical energy which has radiated to the far-field. By far-field we mean at least several wavelengths from the fluorophore, but propagating far-field radiation is usually detected at larger macroscopic distances from the sample. In recent years there has been a growing interest in the interactions of fluorophores with metallic surfaces or particles. Near-field interactions are those occurring within a wavelength distance of an excited fluorophore. The spectral properties of fluorophores can be dramatically altered by near-field interactions with the electron clouds present in metals. These interactions modify the emission in ways not seen in classical fluorescence experiments. In this review we provide an intuitive description of the complex physics of plasmons and near-field interactions. Additionally, we summarize the recent work on metal–fluorophore interactions and suggest how these effects will result in new classes of experimental procedures, novel probes, bioassays and devices. PMID:18810279

Lakowicz, Joseph R.; Ray, Krishanu; Chowdhury, Mustafa; Szmacinski, Henryk; Fu, Yi; Zhang, Jian; Nowaczyk, Kazimierz

2009-01-01

47

In vivo optical spectroscopy for improved detection of pancreatic adenocarcinoma: a feasibility study  

PubMed Central

Pancreatic adenocarcinoma has a five-year survival rate of less than 6%. This low survival rate is attributed to the lack of accurate detection methods, which limits diagnosis to late-stage disease. Here, an in vivo pilot study assesses the feasibility of optical spectroscopy to improve clinical detection of pancreatic adenocarcinoma. During surgery on 6 patients, we collected spectrally-resolved reflectance and fluorescence in vivo. Site-matched in vivo and ex vivo data agreed qualitatively and quantitatively. Quantified differences between adenocarcinoma and normal tissues in vivo were consistent with previous results from a large ex vivo data set. Thus, optical spectroscopy is a promising method for the improved diagnosis of pancreatic cancer in vivo. PMID:24466472

Lloyd, William R.; Wilson, Robert H.; Lee, Seung Yup; Chandra, Malavika; McKenna, Barbara; Simeone, Diane; Scheiman, James; Mycek, Mary-Ann

2013-01-01

48

Fluorescence correlation spectroscopy: principles and applications.  

PubMed

Fluorescence correlation spectroscopy (FCS) is used to study the movements and the interactions of biomolecules at extremely dilute concentrations, yielding results with good spatial and temporal resolutions. Using a number of technical developments, FCS has become a versatile technique that can be used to study a variety of sample types and can be advantageously combined with other methods. Unlike other fluorescence-based techniques, the analysis of FCS data is not based on the average intensity of the fluorescence emission but examines the minute intensity fluctuations caused by spontaneous deviations from the mean at thermal equilibrium. These fluctuations can result from variations in local concentrations owing to molecular mobility or from characteristic intermolecular or intramolecular reactions of fluorescently labeled biomolecules present at low concentrations. Here, we provide a basic introduction to FCS, including its technical development and theoretical basis, experimental setup of an FCS system, adjustment of a setup, data acquisition, and analysis of FCS measurements. Finally, the application of FCS to the study of lipid bilayer membranes and to living cells is discussed. PMID:24987147

Bacia, Kirsten; Haustein, Elke; Schwille, Petra

2014-07-01

49

In vivo imaging with near-infrared fluorescence lifetime contrast  

NASA Astrophysics Data System (ADS)

Fluorescence imaging is a mainstay of biomedical research, allowing detection of molecular events in both fixed and living cells, tissues and whole animals. Such high resolution fluorescence imaging is hampered by unwanted signal from intrinsic background fluorescence and scattered light. The signal to background ratio can be improved by using extrinsic contrast agents and greatly enhanced by multispectral imaging methods. Unfortunately, these methods are insufficient for deep tissue imaging where high contrast and speedy acquisition are necessary. Fluorescence lifetime (FLT) is an inherent characteristic of each fluorescent species that can be independent of intensity and spectral properties. Accordingly, FLT-based detection provides an additional contrast mechanism to optical measurements. This contrast is particularly important in the near-infrared (NIR) due to relative transparency of tissue as well as the broad absorption and emission spectra of dyes that are active in this region. Here we report comparative analysis of signal distribution of several NIR fluorescent polymethine dyes in living mice and their correlations with lifetimes obtained in vitro using solution models. The FLT data obtained from dyes dissolved in serum albumin solution correlated well with FLTs measured in vivo. Thus the albumin solution model could be used as a good predictive model for in vivo FLT behavior of newly developed fluorescent reporters. Subsequent experiments in vivo, including monitoring slow release kinetics and detecting proteinuria, demonstrate the complementary nature of FLT for fluorescence intensity imaging.

Akers, Walter J.; Berezin, Mikhail Y.; Lee, Hyeran; Achilefu, Samuel

2009-02-01

50

Two-Photon Fluorescence Correlation Spectroscopy  

NASA Technical Reports Server (NTRS)

We will describe a two-photon microscope currently under development at the NASA Glenn Research Center. It is composed of a Coherent Mira 900 tunable, pulsed Titanium:Sapphire laser system, an Olympus Fluoview 300 confocal scanning head, and a Leica DM IRE inverted microscope. It will be used in conjunction with a technique known as fluorescence correlation spectroscopy (FCS) to study intracellular protein dynamics. We will briefly explain the advantages of the two-photon system over a conventional confocal microscope, and provide some preliminary experimental results.

Zimmerli, Gregory A.; Fischer, David G.

2002-01-01

51

Broadband Proton Decoupling for In Vivo Brain Spectroscopy in Humans  

E-print Network

Broadband Proton Decoupling for In Vivo Brain Spectroscopy in Humans Peter B. Barker,1,2* Xavier, PBAR, is described for broadband heteronuclear decoupling in vivo in humans at 1.5T. The se- quence Wiley-Liss, Inc. Key words: decoupling; broadband; spectroscopy; in vivo; car- bon-13; phosphorus-31

Ouwerkerk, Ronald

52

Handheld multispectral fluorescence lifetime imaging system for in vivo applications  

PubMed Central

There is an increasing interest in the application of fluorescence lifetime imaging (FLIM) for medical diagnosis. Central to the clinical translation of FLIM technology is the development of compact and high-speed clinically compatible systems. We present a handheld probe design consisting of a small maneuverable box fitted with a rigid endoscope, capable of continuous lifetime imaging at multiple emission bands simultaneously. The system was characterized using standard fluorescent dyes. The performance was then further demonstrated by imaging a hamster cheek pouch in vivo, and oral mucosa tissue both ex vivo and in vivo, all using safe and permissible exposure levels. Such a design can greatly facilitate the evaluation of FLIM for oral cancer imaging in vivo. PMID:24688824

Cheng, Shuna; Cuenca, Rodrigo M.; Liu, Boang; Malik, Bilal H.; Jabbour, Joey M.; Maitland, Kristen C.; Wright, John; Cheng, Yi-Shing Lisa; Jo, Javier A.

2014-01-01

53

Kindling fluorescent proteins for precise in vivo photolabeling.  

PubMed

Photobleaching of green fluorescent protein (GFP) is a widely used approach for tracking the movement of subcellular structures and intracellular proteins. Although photobleaching is a powerful technique, it does not allow direct tracking of an object's movement and velocity within a living cell. Direct tracking becomes possible only with the introduction of a photoactivated fluorescent marker. A number of previous studies have reported optically induced changes in the emission spectra of fluorescent proteins. However, the ideal photoactivated fluorescent marker should be a nonfluorescent tag capable of "switching on" (i.e., becoming fluorescent) in response to irradiation by light of a particular wavelength, intensity, and duration. In this report, we generated a mutant of Anemonia sulcata chromoprotein asCP. The mutant protein is capable of unique irreversible photoconversion from the nonfluorescent to a stable bright-red fluorescent form ("kindling"). This "kindling fluorescent protein" (KFP1) can be used for precise in vivo photolabeling to track the movements of cells, organelles, and proteins. We used KFP1 for in vivo cell labeling in mRNA microinjection assays to monitor Xenopus laevis embryo development and to track mitochondrial movement in mammalian cells. PMID:12524551

Chudakov, Dmitriy M; Belousov, Vsevolod V; Zaraisky, Andrey G; Novoselov, Vladimir V; Staroverov, Dmitry B; Zorov, Dmitry B; Lukyanov, Sergey; Lukyanov, Konstantin A

2003-02-01

54

Fluorescence Spectroscopy Investigations of Cutaneous Tissues  

NASA Astrophysics Data System (ADS)

Fluorescence Spectroscopy of the human skin is very prominent for early diagnosis and differentiation of cutaneous diseases. Selection of proper excitation sources and sensitive detectors gives wide range of possibilities related to real-time determination of existing pathological conditions. A problem with using laser as an excitation source is the high expenses associated with the operation of these types of installations. This is why we test capability of a cheaper excitation sources - ultraviolet and blue light-emitting diodes. Initially, we investigate the spectral response of normal skin from different anatomic areas, as well as from different phototypes volunteers. Our first results obtained demonstrated promising possibility to implement an inexpensive system for detection of cutaneous lesions with wide clinical applications. The results achieved will be introduced in development of diagnostic algorithms for improvement of diagnostic sensitivity of benign and malignant tumor lesions determination.

Borisova, E.; Bliznakova, I.; Momchilov, N.; Troyanova, P.; Avramov, L.

2007-04-01

55

Laser Induced Fluorescence Spectroscopy of Soft Tissues of the Oral Cavity  

NASA Astrophysics Data System (ADS)

The present study deals with the in vivo measurement of auto-fluorescence from different anatomical sites of oral cavities of healthy volunteers, using a homebuilt Laser Induced Fluorescence (LIF) Spectroscopy setup. Excitation wave length of 325 nm from a He-Cd laser was used as the source. From the 7 anatomical sites (say buccal mucosa, tongue, palate etc) of each oral cavity of 113 subjects, 1266 fluorescence spectra were recorded. The spectra were analysed using Principal Component Analysis (PCA) to see the correlation between different sites.

Patil, Ajeetkumar; Unnikrishnan, V. K.; Bernard, Rodney; Pai, Keerthilatha M.; Ongole, Ravikiran; Kartha, V. B.; Chidangil, Santhosh

2011-07-01

56

Combined Raman spectroscopy and autofluoresence imaging method for in vivo skin tumor diagnosis  

NASA Astrophysics Data System (ADS)

The fluorescence and Raman spectroscopy (RS) combined method of in vivo detection of malignant human skin cancer was demonstrated. The fluorescence analysis was used for detection of abnormalities during fast scanning of large tissue areas. In suspected cases of malignancy the Raman spectrum analysis of biological tissue was performed to determine the type of neoplasm. A special RS phase method was proposed for in vivo identification of skin tumor. Quadratic Discriminant Analysis was used for tumor type classification on phase planes. It was shown that the application of phase method provides a diagnosis of malignant melanoma with a sensitivity of 89% and a specificity of 87%.

Zakharov, V. P.; Bratchenko, I. A.; Myakinin, O. O.; Artemyev, D. N.; Khristoforova, Y. A.; Kozlov, S. V.; Moryatov, A. A.

2014-09-01

57

Clinical applications of in vivo fluorescence confocal laser scanning microscopy  

NASA Astrophysics Data System (ADS)

Living skin for basic and clinical research can be evaluated by Confocal Laser Scanning Microscope (CLSM) non-invasively. CLSM imaging system can achieve skin image its native state either "in vivo" or "fresh biopsy (ex vivo)" without fixation, sectioning and staining that is necessary for routine histology. This study examines the potential fluorescent CLSM with a various exogenous fluorescent contrast agent, to provide with more resolution images in skin. In addition, in vivo fluorescent CLSM researchers will be extended a range of potential clinical application. The prototype of our CLSM system has been developed by Prof. Gweon's group. The operating parameters are composed of some units, such as illuminated wavelength 488 nm, argon illumination power up to 20mW on the skin, objective lens, 0.9NA oil immersion, axial resolution 1.0?m, field of view 200?m x 100?m (lateral resolution , 0.3?m). In human volunteer, fluorescein sodium was administrated topically and intradermally. Animal studies were done in GFP transgenic mouse, IRC mouse and pig skin. For imaging of animal skin, fluorescein sodium, acridine orange, and curcumine were used for fluorescein contrast agent. We also used the GFP transgenic mouse for fluorescein CLSM imaging. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. Curcumin is a yellow food dye that has similar fluorescent properties to fluorescein sodium. Acridin Orange can be highlight nuclei in viable keratinocyte. In vivo CLSM of transgenic GFP mouse enable on in vivo, high resolution view of GFP expressing skin tissue. GFP signals are brightest in corneocyte, kertinocyte, hair and eccrine gland. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. In papillary dermis, fluorescein distribution is more homogeneous. Curcumin is a yellow food dye that has similar fluorescent properties to fluorescein sodium. In vivo CLSM of transgenic GFP mouse enable on in vivo, high resolution view of GFP expressing skin tissue. GFP signals are brightest in corneocyte, kertinocyte, skin appendage and blood vessels. In conclusion, this study demonstrates the usefulness of CLSM as technique for imaging skin in vivo. In addition, CLSM is non-invasive, the same tissue site may be imaged over a period of time to monitor the various change such as wound healing, severity of skin diseases and effect of therapeutic management.

Oh, Chilhwan; Park, Sangyong; Kim, Junhyung; Ha, Seunghan; Park, Gyuman; Lee, Gunwoo; Lee, Onseok; Chun, Byungseon; Gweon, Daegab

2008-02-01

58

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

59

Fluorescence spectroscopic analysis (FSA) detects quantitative changes in atherosclerotic plaque collagen and elastin content In Vivo  

NASA Astrophysics Data System (ADS)

In order to assess the capacity for in vivo fluorescence spectroscopi8c analysis of arterial collagen and elastin, fluorescence emission intensity was recorded form rabbit aorta after angioplasty and stent implant, and correlated with extracted elastin and collagen content. FEI from saline treated rabbits after stent implant was higher between 485 and 500 nm than after anti-inflammatory treatment. FEI was significantly decreased after implantation of shorter stents at 476-500 nm. Multiple regression analysis demonstrated an excellent correlation between FEI and elastin and HPLC- measured collagen content at 486-500 nm and 476-480 nm respectively. Conclusions: FEI recorded in vivo form arterial intimal surface, can be successfully used for quantitative assessment of compositional changes in connective tissue. Stent implant can induce changes in intimal arterial structure at discrete sites distant from the stent implant site.

Christov, Alexander M.; Dai, Erbin; Liu, Liying; Guan, Haiyan; Bernards, Mark A.; Cavers, Paul B.; Susko, David; Lucas, Alexandra

2002-05-01

60

In vivo Raman spectroscopy of cervix cancers  

NASA Astrophysics Data System (ADS)

Cervix-cancer is the third most common female cancer worldwide. It is the leading cancer among Indian females with more than million new diagnosed cases and 50% mortality, annually. The high mortality rates can be attributed to late diagnosis. Efficacy of Raman spectroscopy in classification of normal and pathological conditions in cervix cancers on diverse populations has already been demonstrated. Our earlier ex vivo studies have shown the feasibility of classifying normal and cancer cervix tissues as well as responders/non-responders to Concurrent chemoradiotherapy (CCRT). The present study was carried out to explore feasibility of in vivo Raman spectroscopic methods in classifying normal and cancerous conditions in Indian population. A total of 182 normal and 132 tumor in vivo Raman spectra, from 63 subjects, were recorded using a fiberoptic probe coupled HE-785 spectrometer, under clinical supervision. Spectra were acquired for 5 s and averaged over 3 times at 80 mW laser power. Spectra of normal conditions suggest strong collagenous features and abundance of non-collagenous proteins and DNA in case of tumors. Preprocessed spectra were subjected to Principal Component-Linear Discrimination Analysis (PCLDA) followed by leave-one-out-cross-validation. Classification efficiency of ~96.7% and 100% for normal and cancerous conditions respectively, were observed. Findings of the study corroborates earlier studies and suggest applicability of Raman spectroscopic methods in combination with appropriate multivariate tool for objective, noninvasive and rapid diagnosis of cervical cancers in Indian population. In view of encouraging results, extensive validation studies will be undertaken to confirm the findings.

Rubina, S.; Sathe, Priyanka; Dora, Tapas Kumar; Chopra, Supriya; Maheshwari, Amita; Krishna, C. Murali

2014-03-01

61

Structured illumination fluorescence correlation spectroscopy for velocimetry in Zebrafish embryos  

NASA Astrophysics Data System (ADS)

The vascular system of Zebrafish embryos is studied by means of Fluorescence Correlation and Image Correlation Spectroscopy. The long term project addresses biologically relevant issues concerning vasculogenesis and cardiogenesis and in particular mechanical interaction between blood flow and endothelial cells. To this purpose we use Zebrafish as a model system since the transparency of its embryos facilitates morphological observation of internal organs in-vivo. The correlation analysis provides quantitative characterization of fluxes in blood vessels in vivo. We have pursued and compared two complementary routes. In a first one we developed a two-spots two-photon setup in which the spots are spaced at adjustable micron-size distances (1-40 ?m) along a vessel and the endogenous (autofluorescence) or exogenous (dsRed transgenic erythrocytes) signal is captured with an EM-CCD and cross-correlated. In this way we are able to follow the morphology of the Zebrafish embryo, simultaneously measure the heart pulsation, the velocity of red cells and of small plasma proteins. These data are compared to those obtained by image correlations on Zebrafish vessels. The two methods allows to characterize the motion of plasma fluids and erythrocytes in healthy Zebrafish embryos to be compared in the future to pathogenic ones.

Pozzi, Paolo; Rossetti, Leone; Sironi, Laura; Freddi, Stefano; D'Alfonso, Laura; Caccia, Michele; Bouzin, Margaux; Collini, Maddalena; Chirico, Giuseppe

2013-02-01

62

Near-infrared fluorescent proteins for multicolor in vivo imaging  

PubMed Central

Near-infrared fluorescent proteins are in high demand for in vivo imaging. We developed four spectrally distinct fluorescent proteins, iRFP670, iRFP682, iRFP702, and iRFP720, from bacterial phytochromes. iRFPs exhibit high brightness in mammalian cells and tissues and are suitable for long-term studies. iRFP670 and iRFP720 enable two-color imaging in living cells and mice using standard approaches. Five iRFPs including previously engineered iRFP713 allow multicolor imaging in living mice with spectral unmixing. PMID:23770755

Shcherbakova, Daria M.; Verkhusha, Vladislav V.

2013-01-01

63

Native fluorescence spectroscopy of thymus and fat tissues  

NASA Astrophysics Data System (ADS)

Fluorescence spectroscopy of the human thymus gland and surrounding mediastinal fat were measured to evaluate this approach in distinguishing between thymus and fat tissues during therapeutic surgery for myasthenia gravis disease.

Tang, Gui C.; Oz, Mehmet C.; Reid, V.; Steinglass, K.; Ginsberg, Mark D.; Jacobowitz, Larry; Alfano, Robert R.

1993-08-01

64

Quantitative Determination of DNA-Ligand Binding Using Fluorescence Spectroscopy  

ERIC Educational Resources Information Center

The effective use of fluorescence spectroscopy for determining the binding of the intercalcating agent crhidium bromide to DNA is being described. The analysis used simple measurement techniques and hence can be easily adopted by the students for a better understanding.

Healy, Eamonn F.

2007-01-01

65

Effect of probe pressure on cervical fluorescence spectroscopy measurements  

E-print Network

Fluorescence spectroscopy is a promising technology for detection of epithelial precancers and cancers. While age and menopausal status influence measurements in the cervix, other variables do not significantly affect the ...

Nath, Audrey

66

Fluorescent-Spectroscopic Research of in Vivo Tissues Pathological Conditions  

NASA Astrophysics Data System (ADS)

The steady-state spectra of autofluorescence and the reflection coefficient on the excitation wavelength of some stomach tissues in vivo with various pathological conditions (surface gastritis, displasia, cancer) are measured under excitation by the nitrogen laser irradiation (?ex=337.1 nm). The contour expansion of obtained fluorescence spectra into contributions of components is conducted by the Gaussian-Lorentzian curves method. It is shown that at least 7 groups of fluorophores forming a total luminescence spectrum can be distinguished during the development of displasia and tumor processes. The correlation of intensities of flavins and NAD(P)·H fluorescence is determined and the degree of respiratory activity of cells for the functional condition considered is estimated. The evaluations of the fluorescence quantum yield of the tissue's researched are given.

Giraev, K. M.; Ashurbekov, N. A.; Medzhidov, R. T.

67

Ex Vivo Fluorescence Molecular Tomography of the Spine  

PubMed Central

We investigated the potential of fluorescence molecular tomography to image ex vivo samples collected from a large animal model, in this case, a dog spine. Wide-field time-gated fluorescence tomography was employed to assess the impact of multiview acquisition, data type, and intrinsic optical properties on the localization and quantification accuracy in imaging a fluorescent inclusion in the intervertebral disk. As expected, the TG data sets, when combining early and late gates, provide significantly better performances than the CW data sets in terms of localization and quantification. Moreover, the use of multiview imaging protocols led to more accurate localization. Additionally, the incorporation of the heterogeneous nature of the tissue in the model to compute the Jacobians led to improved imaging performances. This preliminary imaging study provides a proof of concept of the feasibility of quantitatively imaging complex ex vivo samples nondestructively and with short acquisition times. This work is the first step towards employing optical molecular imaging of the spine to detect and characterize disc degeneration based on targeted fluorescent probes. PMID:23197973

Pimpalkhare, Monish; Chen, Jin; Venugopal, Vivek; Intes, Xavier

2012-01-01

68

Dual-Color Fluorescence Cross-Correlation Spectroscopy on a  

E-print Network

fluorescence correlation spectroscopy (SPIM-FCS) is a new method for imaging FCS in 3D samples, providing and theory," Biopolymers 13, 1­27 (1974). 2. D. Magde, E. L. Elson, and W. W. Webb, "Fluorescence correlation for BioImaging Sciences, National University of Singapore, 14 science Drive 4, Singapore 117557 3

Garbe, Christoph S.

69

Continuous Fluorescence Microphotolysis and Correlation Spectroscopy Using 4Pi Microscopy  

Microsoft Academic Search

Continuous fluorescence microphotolysis (CFM) and fluorescence correlation spectroscopy (FCS) permit measurement of molecular mobility and association reactions in single living cells. CFM and FCS complement each other ideally and can be realized using identical equipment. So far, the spatial resolution of CFM and FCS was restricted by the resolution of the light microscope to the micrometer scale. However, cellular functions

Anton Arkhipov; Jana Hüve; Martin Kahms; Reiner Peters; Klaus Schulten

2007-01-01

70

In vivo lipidomics using single-cell Raman spectroscopy  

PubMed Central

We describe a method for direct, quantitative, in vivo lipid profiling of oil-producing microalgae using single-cell laser-trapping Raman spectroscopy. This approach is demonstrated in the quantitative determination of the degree of unsaturation and transition temperatures of constituent lipids within microalgae. These properties are important markers for determining engine compatibility and performance metrics of algal biodiesel. We show that these factors can be directly measured from a single living microalgal cell held in place with an optical trap while simultaneously collecting Raman data. Cellular response to different growth conditions is monitored in real time. Our approach circumvents the need for lipid extraction and analysis that is both slow and invasive. Furthermore, this technique yields real-time chemical information in a label-free manner, thus eliminating the limitations of impermeability, toxicity, and specificity of the fluorescent probes common in currently used protocols. Although the single-cell Raman spectroscopy demonstrated here is focused on the study of the microalgal lipids with biofuel applications, the analytical capability and quantitation algorithms demonstrated are applicable to many different organisms and should prove useful for a diverse range of applications in lipidomics. PMID:21310969

Wu, Huawen; Volponi, Joanne V.; Oliver, Ann E.; Parikh, Atul N.; Simmons, Blake A.; Singh, Seema

2011-01-01

71

Noncontact point spectroscopy guided by two-channel fluorescence imaging in a hamster cheek pouch model  

NASA Astrophysics Data System (ADS)

A system for in vivo, fluorescence image-guided, non-contact point fluorescence spectroscopy is presented. A 442 nm HeCd laser is used as the fluorescence excitation source. An intensified CCD serves as the detector for both imaging and spectroscopy, on which two regions of 300 X 300 pixels were used for green (500 +/- 18 nm) and red (630 +/- 18 nm) imaging channels, and a strip of 600 X 120 pixels are used for emission spectroscopy (450 - 750 nm). At a working distance of 40 mm, the system has a spatial resolution of 0.16 mm and a spectral resolution of 5 nm. System performance is demonstrated in a carcinogenesis model in hamsters, where tumors were induced by painting DMBA in the cheek pouch. Autofluorescence and Photofrin-induced fluorescence measurements were performed every 2 weeks during the 18 weeks of tumor induction. Punch biopsies on selected animals were taken for histological staging. The results show that autofluorescence fluorescence can distinguish dysplasia from normal mucosal tissue model, utilizing the peak red intensity (or the red-to-green intensity ratio). Photofrin-induced fluorescence was superior to autofluorescence for differentiating high grade dysplasia from invasive cancer.

Yang, Victor X.; Yeow, Jenny; Lilge, Lothar D.; Kost, James; Mang, Thomas S.; Wilson, Brian C.

1999-07-01

72

Quantum dots fluorescence quantum yield measured by Thermal Lens Spectroscopy.  

PubMed

An essential parameter to evaluate the light emission properties of fluorophores is the fluorescence quantum yield, which quantify the conversion efficiency of absorbed photons to emitted photons. We detail here an alternative nonfluorescent method to determine the absolute fluorescence quantum yield of quantum dots (QDs). The method is based in the so-called Thermal Lens Spectroscopy (TLS) technique, which consists on the evaluation of refractive index gradient thermally induced in the fluorescent material by the absorption of light. Aqueous dispersion carboxyl-coated cadmium telluride (CdTe) QDs samples were used to demonstrate the Thermal Lens Spectroscopy technical procedure. PMID:25103802

Estupiñán-López, Carlos; Dominguez, Christian Tolentino; Cabral Filho, Paulo E; Fontes, Adriana; de Araujo, Renato E

2014-01-01

73

MRI-coupled spectrally-resolved fluorescence tomography for in vivo imaging  

NASA Astrophysics Data System (ADS)

A unique fluorescence imaging system incorporates multi-channel spectrometer-based optical detection directly into clinical MRI for simultaneous MR and spectrally-resolved fluorescence tomography acquisition in small animal and human breast-sized volumes. A custom designed MRI rodent coil adapted to accommodate optical fibers in a circular geometry for contact mode acquisition provides small animal imaging capabilities, and human breast-sized volumes are imaged using a clinical breast coil modified with an optical fiber patient array. Spectroscopy fibers couple light emitted from the tissue surface to sixteen highly sensitive CCD-based spectrometers operating in parallel. Tissue structural information obtained from standard and contrast enhanced T1-weighted images is used to spatially constrain the diffuse fluorescence tomography reconstruction algorithm, improving fluorescence imaging capabilities qualitatively and quantitatively. Simultaneous acquisition precludes the use of complex co-registration processes. Calibration procedures for the optical acquisition system are reviewed and the imaging limits of the system are investigated in homogeneous and heterogeneous gelatin phantoms containing Indocyanine Green (ICG). Prior knowledge of fluorescence emission spectra is used to de-couple fluorescence emission from residual excitation laser cross-talk. Preliminary in vivo data suggests improved fluorescence imaging in mouse brain tumors using MR-derived spatial priors. U-251 human gliomas were implanted intracranially into nude mice and combined contrast enhanced MRI/fluorescence tomography acquisition was completed at 24 hour intervals over the course of 72 hours after administration of an EGFR targeted NIR fluorophore. Reconstructed images demonstrate an inability to recover reasonable images of fluorescence activity without the use of MRI spatial priors.

Davis, Scott C.; Gibbs-Strauss, Summer L.; Tuttle, Stephen B.; Jiang, Shudong; Springett, Roger; Dehghani, Hamid; Pogue, Brian W.; Paulsen, Keith D.

2008-02-01

74

Validation of temperature-modulated fluorescence tomography in vivo  

NASA Astrophysics Data System (ADS)

To overcome the strong scattering in biological tissue that has long afflicted fluorescence tomography, we have developed a novel technique, "temperature-modulated fluorescence tomography" (TM-FT) to combine the sensitivity of fluorescence imaging with focused ultrasound resolution. TM-FT relies on two key elements: temperature sensitive ICG loaded pluronic nanocapsules we termed ThermoDots and high intensity focused ultrasound (HIFU). TM-FT localizes the position of the fluorescent ThermoDots by irradiating and scanning a HIFU beam across the tissue while conventional fluorescence tomography measurements are acquired. The HIFU beam produces a local hot spot, in which the temperature suddenly increases changing the quantum efficiency of the ThermoDots. The small size of the focal spot (~1 mm) up to a depth of 6 cm, allows imaging the distribution of these temperature sensitive agents with not only high spatial resolution but also high quantitative accuracy in deep tissue using a proper image reconstruction algorithm. Previously we have demonstrated this technique with a phantom study with ThermoDots sensitive in the 20-25°C range. We recently optimized the ThermoDots for physiological temperatures. In this work, we will demonstrate a new HIFU scanning method which is optimized for in vivo studies. The performance of the system is tested using a phantom that resembles a small animal bearing a small tumor targeted by ThermoDots.

Kwong, Tiffany C.; Nouizi, Farouk; Lin, Yuting; Rajyaguru, Rushi; Nguyen, Trinh; Alptekin, Lara; Sampathkumaran, Uma; Zhu, Yue; Ahmed, Shaaz; Gulsen, Gultekin

2014-02-01

75

In vivo Diagnosis of Cervical Intraepithelial Neoplasia Using 337-nm- Excited Laser-Induced Fluorescence  

NASA Astrophysics Data System (ADS)

Laser-induced fluorescence at 337-nm excitation was used in vivo to differentiate neoplastic [cervical intraepithelial neoplasia (CIN)], nonneoplastic abnormal (inflammation and human papilloma viral infection), and normal cervical tissues. A colposcope (low-magnification microscope used to view the cervix with reflected light) was used to identify 66 normal and 49 abnormal (5 inflammation, 21 human papilloma virus infection, and 23 CIN) sites on the cervix in 28 patients. These sites were then interrogated spectroscopically. A two-stage algorithm was developed to diagnose CIN. The first stage differentiated histologically abnormal tissues from colposcopically normal tissues with a sensitivity, specificity, and positive predictive value of 92%, 90%, and 88%, respectively. The second stage differentiated preneoplastic and neoplastic tissues from nonneoplastic abnormal tissues with a sensitivity, specificity, and positive predictive value of 87%, 73%, and 74%, respectively. Spectroscopic differences were consistent with a decrease in the absolute contribution of collagen fluorescence, an increase in the absolute contribution of oxyhemoglobin attenuation, and an increase in the relative contribution of reduced nicotinamide dinucleotide phosphate [NAD(P)H] fluorescence as tissue progresses from normal to abnormal in the same patient. These results suggest that in vivo fluorescence spectroscopy of the cervix can be used to diagnose CIN at colposcopy.

Ramanujam, N.; Mitchell, M. F.; Mahadevan, A.; Warren, S.; Thomsen, S.; Silva, E.; Richards-Kortum, R.

1994-10-01

76

Spectral unmixing of multi-color tissue specific in vivo fluorescence in mice  

NASA Astrophysics Data System (ADS)

Fluorescence Molecular Tomography (FMT) has emerged as a powerful tool for monitoring biological functions in vivo in small animals. It provides the means to determine volumetric images of fluorescent protein concentration by applying the principles of diffuse optical tomography. Using different probes tagged to different proteins or cells, different biological functions and pathways can be simultaneously imaged in the same subject. In this work we present a spectral unmixing algorithm capable of separating signal from different probes when combined with the tomographic imaging modality. We show results of two-color imaging when the algorithm is applied to separate fluorescence activity originating from phantoms containing two different fluorophores, namely CFSE and SNARF, with well separated emission spectra, as well as Dsred- and GFP-fused cells in F5-b10 transgenic mice in vivo. The same algorithm can furthermore be applied to tissue-specific spectroscopy data. Spectral analysis of a variety of organs from control, DsRed and GFP F5/B10 transgenic mice showed that fluorophore detection by optical systems is highly tissue-dependent. Spectral data collected from different organs can provide useful insight into experimental parameter optimisation (choice of filters, fluorophores, excitation wavelengths) and spectral unmixing can be applied to measure the tissue-dependency, thereby taking into account localized fluorophore efficiency. Summed up, tissue spectral unmixing can be used as criteria in choosing the most appropriate tissue targets as well as fluorescent markers for specific applications.

Zacharakis, Giannis; Favicchio, Rosy; Garofalakis, Anikitos; Psycharakis, Stylianos; Mamalaki, Clio; Ripoll, Jorge

2007-07-01

77

Spectroscopy detection of green and red fluorescent proteins in genetically modified plants using a fiber optics system  

NASA Astrophysics Data System (ADS)

In this paper, fiber optic spectroscopy is developed to detect and quantify recombinant green (EGFP) and red (DsRED) fluorescent proteins in vitro and in vivo. The bacterial expression vectors carrying the coding regions of EGFP and DsRED were introduced into Escherichia coli host cells and fluorescent proteins were produced following induction with IPTG. Soluble EGFP and DsRED proteins were isolated from lysed bacterial cells and serially diluted for quantitative analysis by fiber optic spectroscopy. Fluorescence at the appropriate emission wavelengths could be detected up to 64X dilution for EGFP and 40X dilution for DsRED. To determine the capability of spectroscopy detection in vivo, transgenic potato hairy roots expressing EGFP and DsRED were regenerated. This was achieved by cloning the EGFP and DsRED genes into the plant binary vector, pTMV35S, to create the recombinant vectors pGLOWGreen and pGLOWRed. These latter binary vectors were introduced into Agrobacterium rhizogenes strain A4T. Infection of potato cells with transformed agrobacteria was used to insert the fluorescent protein genes into the potato genome. Genetically modified potato cells were then regenerated into hairy roots. A panel of transformed hairy roots expressing varying levels of fluorescent proteins was selected by fluorescence microscopy. We are now assessing the capability of spectroscopic detection system for in vivo quantification of green and red fluorescence levels in transformed roots.

Liew, Oi Wah; Asundi, Anand K.; Chen, Jun-Wei; Chew, Yiwen; Yu, Shangjuan; Yeo, Gare H.

2001-05-01

78

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

79

Near-infrared multichannel Raman spectroscopy toward real-timein vivo cancer diagnosis  

Microsoft Academic Search

Using a newly developed InP\\/InGaAsP multichannel detector, we developed a near-infrared Raman spectroscopic system that can measure human tissues efficiently without interference from fluorescence. This enabled us to measure an in vivo Raman spectrum of live human tissue (skin) in 1 min using fiber probe optics. By applying this system to human lung tissues, we found that Raman spectroscopy makes

Shoji Kaminaka; Toshiaki Ito; Hiroya Yamazaki; Ehiichi Kohda; Hiro-O. Hamaguchi

2002-01-01

80

Fluorescence imaging in vivo: recent advances Jianghong Rao, Anca Dragulescu-Andrasi and Hequan Yao  

E-print Network

Fluorescence imaging in vivo: recent advances Jianghong Rao, Anca Dragulescu-Andrasi and Hequan Yao In vivo fluorescence imaging uses a sensitive camera to detect fluorescence emission from fluorophores in whole-body living small animals. To overcome the photon attenuation in living tissue, fluorophores

Rao, Jianghong

81

Optical spectroscopy for differentiation of liver tissue under distinct stages of fibrosis: an ex vivo study  

NASA Astrophysics Data System (ADS)

Liver fibrosis is the decisive step towards the development of cirrhosis; its early detection affects crucially the diagnosis of liver disease, its prognosis and therapeutic decision making. Nowadays, several techniques are employed to this task. However, they have the limitation in estimating different stages of the pathology. In this paper we present a preliminary study to evaluate if optical spectroscopy can be employed as an auxiliary tool of diagnosis of biopsies of human liver tissue to differentiate the fibrosis stages. Ex vivo fluorescence and diffuse reflectance spectra were acquired from biopsies using a portable fiber-optic system. Empirical discrimination algorithms based on fluorescence intensity ratio at 500 nm and 680 nm as well as diffuse reflectance intensity at 650 nm were developed. Sensitivity and specificity of around 80% and 85% were respectively achieved. The obtained results show that combined use of fluorescence and diffuse reflectance spectroscopy could represent a novel and useful tool in the early evaluation of liver fibrosis.

Fabila, D. A.; Hernández, L. F.; de la Rosa, J.; Stolik, S.; Arroyo-Camarena, U. D.; López-Vancell, M. D.; Escobedo, G.

2013-11-01

82

Profiling the near field of nanoshells using surface enhanced Raman spectroscopy and fluorescence spectroscopy  

Microsoft Academic Search

Plasmon resonances in metal nanoparticles control the far field and near field optical properties of these metallic structures. The enhanced electromagnetic near field is strongest at the surface of the nanoparticles and rapidly decays away from the surface. This enhanced near field is exploited in surface enhanced spectroscopies including Surface Enhanced Raman Spectroscopy (SERS) and Metal Enhanced Fluorescence Spectroscopy (MEFS).

Surbhi Lal

2006-01-01

83

Fluorescence lifetime imaging microscopy of nanodiamonds in vivo  

NASA Astrophysics Data System (ADS)

The negatively charged nitrogen-vacancy (NV-) center in bulk diamond is a photostable fluorophore with a radiative lifetime of 11.6 ns at room temperature. The lifetime substantially increases to ~20 ns for diamond nanoparticles (size ~ 100 nm) suspended in water due to the change in refractive index of the surrounding medium of the NV- centers. This fluorescence decay time is much longer than that (typically 1 - 4 ns) of endogenous and exogenous fluorophores commonly used in biological imaging, making it possible to detect NV--containing nanodiamonds in vivo at the single particle level by fluorescence lifetime imaging microscopy (FLIM). We demonstrate the feasibility of this approach using Caenorhabditis elegans (C. elegans) as a model organism.

Kuo, Yung; Hsu, Tsung-Yuan; Wu, Yi-Chun; Hsu, Jui-Hung; Chang, Huan-Cheng

2013-03-01

84

Deep tissue fluorescence imaging and in vivo biological applications  

NASA Astrophysics Data System (ADS)

We describe a novel technical approach with enhanced fluorescence detection capabilities in two-photon microscopy that achieves deep tissue imaging, while maintaining micron resolution. Compared to conventional two-photon microscopy, greater imaging depth is achieved by more efficient harvesting of fluorescence photons propagating in multiple-scattering media. The system maintains the conventional two-photon microscopy scheme for excitation. However, for fluorescence collection the detection system harvests fluorescence photons directly from a wide area of the turbid sample. The detection scheme relies on a wide area detector, minimal optical components and an emission path bathed in a refractive-index-matching fluid that minimizes emission photon losses. This detection scheme proved to be very efficient, allowing us to obtain high resolution images at depths up to 3 mm. This technique was applied to in vivo imaging of the murine small intestine (SI) and colon. The challenge is to image normal and diseased tissue in the whole live animal, while maintaining high resolution imaging at millimeter depth. In Lgr5-GFP mice, we have been successful in imaging Lgr5-eGFP positive stem cells, present in SI and colon crypt bases.

Crosignani, Viera; Dvornikov, Alexander; Aguilar, Jose S.; Stringari, Chiara; Edwards, Robert; Mantulin, William W.; Gratton, Enrico

2012-11-01

85

In Vivo Blood Characterization from Bioimpedance Spectroscopy of Blood  

E-print Network

In Vivo Blood Characterization from Bioimpedance Spectroscopy of Blood Pooling Tao Dai Department@sce.carleton.ca Abstract Characterization of blood impedance properties is important to estimate clinical diagnos- tic in the measurement field, rather than the blood individually. This paper describes a novel in vivo measurement

Adler, Andy

86

Fulvic acid oxidation state detection using fluorescence spectroscopy.  

PubMed

Humic substances are a heterogeneous class of moderate molecular weight, yellow-colored biomolecules present in all soils, sediments, and natural waters. Although humic substances are generally resistant to microbial degradation under anaerobic conditions, some microorganisms in soils and sediments can use quinone moieties in humic substances as electron acceptors. Laboratory experiments have shown that humic substances can act as electron shuttles in the microbial reduction of ferric iron. Field studies of electron shuttling processes have been constrained by the lack of methods to characterize the oxidation state of quinone moieties in humic substances at natural concentrations. All humic substances have fluorescent properties, and fluorescence spectroscopy can indicate differences in precursor organic source of humic substances. Here we show that the quinone moieties responsible for electron transfer reactions contribute significantly to the fluorescence of humic substances. Further we use fluorescence spectroscopy to elucidate the oxidation state of quinone moieties in humic substances at natural concentrations found in sediment interstitial waters. PMID:12141500

Klapper, Lisa; McKnight, Diane M; Fulton, J Robin; Blunt-Harris, Elizabeth L; Nevin, Kelly P; Lovley, Derek R; Hatcher, Patrick G

2002-07-15

87

Fluorescence correlation spectroscopy and its potential for intracellular applications  

Microsoft Academic Search

Fluorescence correlation spectroscopy (FCS) is a time-averaging fluctuation analysis of small molecular ensembles, combining\\u000a maximum sensitivity with high statistical confidence. Among a multitude of physical parameters that are, in principle, accessible\\u000a by FCS, it most conveniently allows to determine local concentrations, mobility coefficients, and characteristic rate constants\\u000a of fast-reversible and slow-irreversible reactions of fluorescently labeled biomolecules at very low (nanomolar)

Petra Schwille

2001-01-01

88

Wide-field in vivo background free imaging by selective magnetic modulation of nanodiamond fluorescence  

PubMed Central

The sensitivity and resolution of fluorescence-based imaging in vivo is often limited by autofluorescence and other background noise. To overcome these limitations, we have developed a wide-field background-free imaging technique based on magnetic modulation of fluorescent nanodiamond emission. Fluorescent nanodiamonds are bright, photo-stable, biocompatible nanoparticles that are promising probes for a wide range of in vitro and in vivo imaging applications. Our readily applied background-free imaging technique improves the signal-to-background ratio for in vivo imaging up to 100-fold. This technique has the potential to significantly improve and extend fluorescent nanodiamond imaging capabilities on diverse fluorescence imaging platforms. PMID:24761300

Sarkar, Susanta K.; Bumb, Ambika; Wu, Xufeng; Sochacki, Kem A.; Kellman, Peter; Brechbiel, Martin W.; Neuman, Keir C.

2014-01-01

89

Pancreatic tissue assessment using fluorescence and reflectance spectroscopy  

NASA Astrophysics Data System (ADS)

The ability of multi-modal optical spectroscopy to detect signals from pancreatic tissue was demonstrated by studying human pancreatic cancer xenografts in mice and freshly excised human pancreatic tumor tissue. Measured optical spectra and fluorescence decays were correlated with tissue morphological and biochemical properties. The measured spectral features and decay times correlated well with expected pathological differences in normal, pancreatitis and adenocarcinoma tissue states. The observed differences between the fluorescence and reflectance properties of normal, pancreatitis and adenocarcinoma tissue indicate a possible application of multi-modal optical spectroscopy to differentiating between the three tissue classifications.

Chandra, Malavika; Heidt, David; Simeone, Diane; McKenna, Barbara; Scheiman, James; Mycek, Mary-Ann

2007-07-01

90

Eclipsing thermal lens spectroscopy for fluorescence quantum yield measurement.  

PubMed

A modified spatial filtering method that improves the sensitivity of single-beam and mode-mismatched thermal lens spectroscopy (TLS) for fluorescence quantum yield measurement is presented. The method is based on the detection of the external part of a laser beam transmitted by the fluorescent sample (eclipsing detection mode). The experimental results show that the signal/noise (S/N) ratio of the absolute quantum yield of Rh6G can be enhanced up to ~1400% using the eclipsing detection mode on the TLS experimental setup. The method was evaluated by measuring the fluorescence quantum yield of varying concentration of ethanolic solutions of Rhodamine 6G. PMID:23938731

Estupiñán-López, C; Tolentino Dominguez, C; de Araujo, R E

2013-07-29

91

Intramolecular Fluorescence Correlation Spectroscopy in a Feedback Tracking Microscope  

PubMed Central

Abstract We derive the statistics of the signals generated by shape fluctuations of large molecules studied by feedback tracking microscopy. We account for the influence of intramolecular dynamics on the response of the tracking system and derive a general expression for the fluorescence autocorrelation function that applies when those dynamics are linear. We show that in comparison to traditional fluorescence correlation spectroscopy, tracking provides enhanced sensitivity to translational diffusion, molecular size, heterogeneity, and long-timescale decays. We demonstrate our approach using a three-dimensional tracking microscope to study genomic ?-phage DNA molecules with various fluorescence label configurations. PMID:20655860

McHale, Kevin; Mabuchi, Hideo

2010-01-01

92

Oil Classification with Fluorescence Spectroscopy Engineering Physics  

E-print Network

#12;2 For my parents and my niece Fatima A. Ferra #12;3 ABSTRACT This thesis is a part of an EU oil and bilge oil: the channels relationships method; artificial neural networks (ANNs); and principal and the input fluorescence signature of the oil play a very important role in the efficiency

Oldenburg, Carl von Ossietzky Universität

93

Fluorescence Imaging In Vivo up to 1700 nm  

E-print Network

Compared to visible and near-infrared regions below ~ 900 nm, imaging in the second near-infrared window beyond 1000 nm (NIR-II, 1000-1700 nm) is promising for deep-tissue high-resolution optical imaging in vivo owing to reduced scattering of photons traversing through tissues. Here, we succeeded fluorescence imaging in vivo in the long 1500-1700 nm (NIR-IIb) region using a novel, chemical separation enriched large-diameter semiconducting single-walled carbon nanotube material. Imaging in the 1500-1700 nm window resolved 3-4 um wide capillary blood vessels at ~ 3 millimeters depth through the intact body and brain of mice with the ability of blood-flow speed mapping in individual capillary vessels. Further, non-invasive single fluorophore imaging inside the tumor of a live mouse was achieved in the 1500-1700 nm window. NIR-IIb imaging can be generalized to a wide range of fluorophores emitting up to 1700 nm for a new paradigm of high performance in vivo optical imaging.

Diao, Shuo; Hong, Guosong; Antaris, Alexander L; Chang, Junlei; Wu, Justin Z; Zhang, Bo; Kuo, Calvin J; Dai, Hongjie

2015-01-01

94

Fluorescent Lifetime Spectroscopy in Random Media  

NASA Astrophysics Data System (ADS)

Recently, an abundance of near-infrared phosphorescent and fluorescent probes have been developed whose lifetime is sensitive to changes in the local environment. The lifetime of these probes can be readily determined in a dilute, non-scattering media using conventional time- and frequency-domain techniques. From the lifetime, the concentration of metabolites can be found using the Stern-Volmer relationship. However, in highly scattering media such as tissues and particulate process streams, measurement of lifetime is complicated by the time delay associated with light scatter. In this dissertation, frequency-domain measurements of photon migration are developed for measuring fluorescent lifetimes independent of absorption and scattering properties of tissues and other random media. The measurement consists of launching, onto the surface of the medium, excitation light whose intensity is sinusoidally modulated at megahertz frequencies. The fluorescent light generated within the medium is intensity modulated at the same frequency, but phase-shifted and amplitude demodulated relative to the incident excitation source. In addition, the excitation light is also phase-shifted and amplitude demodulated relative to the incident excitation source. From Green's function analysis, finite element computations, and experimental measurements of fluorescent phase-shift and amplitude demodulation, we show it is possible to determine fluorophore lifetime of common laser dyes in a tissue mimicking phantom. Furthermore, the finite element computations of excitation and fluorescent light fluence show that when the fluorophore is uniformly distributed within a medium, signals re-emitted at the surface do not originate from significant depths if its lifetime is greater than the photon migration time associated with scatter. Consequently, this research points to the development of short-lived fluorescent compounds for biodiagnostics using properly referenced frequency-domain measurements. Applications for lifetime-based sensing in tissues and other random media (such as monitoring changes in glucose, pH, pCO _2, pO_2, and other metabolites) depend upon the continued development of the referencing procedures developed herein, as well as engineering interactions with synthetic chemists for the future development of optical probes.

Hutchinson, Christina Laura

95

"FluSpec": A Simulated Experiment in Fluorescence Spectroscopy  

ERIC Educational Resources Information Center

The "FluSpec" educational software package is a fully contained tutorial on the technique of fluorescence spectroscopy as well as a simulator on which experiments can be performed. The procedure for each of the experiments is also contained within the package along with example analyses of results that are obtained using the software.

Bigger, Stephen W.; Bigger, Andrew S.; Ghiggino, Kenneth P.

2014-01-01

96

Flame front tracking by laser induced fluorescence spectroscopy and advanced  

E-print Network

information from high speed Planar Laser Induced Fluores- cence (PLIF) data obtained from turbulent flamesFlame front tracking by laser induced fluorescence spectroscopy and advanced image analysis Rafeef. The application of non-linear anisotropic diffusion filtering and of Active Contour Mod- els (Snakes) is described

Hamarneh, Ghassan

97

Sensitive fluorescence spectroscopy of jet cooled 15NO 2  

Microsoft Academic Search

A spectroscopic setup designed for high resolution spectroscopy of jet cooled molecules is described. The primary features of the new setup are an exceedingly low gas consumption and a high detection efficiency, which are achieved by optimizing the detection geometry of the time gated fluorescence in combination with a special piezo valve. This makes the setup particularly suitable for studying

E. A. Volkers; A. Vredenborg; H. V. J. Linnartz; J. Bulthuis; S. Stolte; R. Jost

2004-01-01

98

Classification of edible oils using synchronous scanning fluorescence spectroscopy  

Microsoft Academic Search

Total luminescence and synchronous scanning fluorescence spectroscopy techniques were tested as regards their ability to characterize and differentiate edible oils, including soybean, sunflower, rapeseed, peanut, olive, grapeseed, linseed and corn oils. Total luminescence spectra of all oils studied as n-hexane solutions exhibit an intense peak, which appears at 290 nm in excitation and 320 nm in emission, attributed to tocopherols.

Ewa Sikorska; Tomasz Górecki; Igor V Khmelinskii; Marek Sikorski; Jacek Kozio?

2005-01-01

99

CHARACTERIZATION OF EDIBLE OILS USING SYNCHRONOUS SCANNING FLUORESCENCE SPECTROSCOPY  

Microsoft Academic Search

Synchronous scanning fluorescent spectroscopy is explored as a tool for edible oil quality control and identification. Characteristic bands existing in the respective spectra and significant changes of these bands resulting from thermal oxidation and photo-oxidation of the oils confirm the potential of the method for the edible oil analysis. The method consists in monitoring and quantification of minor luminescent constituents,

Ewa Sikorska; Anna Romaniuk; Igor. Khmelinskii; Marek Sikorski

100

Fibre optic fluorescence spectroscopy for monitoring fish freshness  

NASA Astrophysics Data System (ADS)

In this study, a portable Y-type fibreoptic fluorescence spectroscopy measurement system was used to evaluate the freshness of eight cobias (Rachycentron canadum). The results showed that the ratio of fluorescent intensity, which F480 nm/Fexci+50 nm was belong with the range of collagen type I and type V characteristic spectra, was positive correlated to the frozen time by hours. It was a strong approach to be a potential index for differentiating the fish freshness during delivery process. Besides, the different pattern results of dorsum and abdomen were shown in this study. In further, fibreoptic fluorescence spectroscopy could be a way not only to measure and quantify the freshness of different fish body but also to verify the level of taste.

Wu, Chi-Wu; Hsiao, Tzu-Chien; Chu, Shou-Chia; Hu, Hung-Hsi; Chen, Jyh-Cheng

2012-01-01

101

In vivo fluorescence imaging of lysosomes: a potential technique to follow dye accumulation in the context of PDT?  

NASA Astrophysics Data System (ADS)

Lysosomes and intracellular acidic compartments seem to play an important role in the context of PDT. Some photosensitizers are localized in the lysosomes of tumor-associated macrophages. Liposomes, which are lysosomotropic drug carriers, are used to deliver photosensitizers in tumors. Liposomes are taken up by the liver cells after intravenous injection. Intracellular pathway and liposomes localization in the different liver cells require sacrifice of the animals, cell separation, and observation by electronic microscopy. Little is known about liposomes kinetic uptake by the acidic intracellular compartments in vivo. We propose in this study a new method to follow liposomes uptake in the liver in vivo using a fluorescent pH-sensitive probe. We have already demonstrated the ability of fluorescence spectroscopy and imaging using a pH-dependent probe to monitor pH in living tissues. As pH of lysosome is very low, the kinetic of liposome uptake in this intracellular acidic compartment is followed by monitoring the pH of the whole liver in vivo and ex vivo. Liposomes-encapsulated carboxyfluorescein are prepared by the sonication procedure. Carboxyfluorescein is used at high concentration (100 mM) in order to quench its fluorescence. Liposomes are injected to Wistar rats into the peinil vein. After laparotomy, fluorescence spectra and images are recorded during two hours. Results show a rapid fluorescence increase followed by a slow phase of fluorescence decrease. pH decreases from physiological value to 6.0. After sacrifice and flush with cold saline solution, pH of liver ex vivo is found to be 5.0 - 5.5. These data show a rapid clearance of released dye and an uptake of liposomes by the liver cells and, as liposomes penetrate in the acidic compartment, dye is released from liposomes and is delivered in lysosomes leading to the decrease of pH.

Devoisselle, Jean-Marie; Mordon, Serge R.; Soulie-Begu, Sylvie

1995-03-01

102

Histologic differences between orthotopic xenograft pancreas models affect Verteporfin uptake measured by fluorescence microscopy and spectroscopy  

NASA Astrophysics Data System (ADS)

Photodynamic therapy (PDT) that uses the second generation photosensitizer, verteporfin (VP), is a developing therapy for pancreatic cancer. The optimal timing of light delivery related to VP uptake and distribution in pancreatic tumors will be important information to obtain to improve treatment for this intractable disease. In this work we examined uptake and distribution of VP in two orthotopic pancreatic tumors with different histological structure. ASPC-1 (fast-growing) and Panc-1 (slower growing) tumors were implanted in SCID mice and studied when tumors were approximately 100mm3. In a pilot study, these tumors had been shown to differ in uptake of VP using lightinduced fluorescence spectroscopy (LIFS) in vivo and fluorescence imaging ex vivo and that work is extended here. In vivo fluorescence mean readings of tumor and liver increased rapidly up to 15 minutes after photosensitizer injection for both tumor types, and then continued to increase up to 60 minutes post injection to a higher level in ASPC-1 than in Panc-1. There was variability among animals with the same tumor type, in both liver and tumor uptake and no selectivity of tumor over liver. In this work we further examined VP uptake at multiple time points in relation to microvascular density and perfusion, using DiOC7 (to mark blood vessels) and VP fluorescence in the same tissue slices. Analysis of DiOC7 fluorescence indicates that AsPC-1 and Panc-1 have different vascular densities but AsPC-1 vasculature is more perfusive. Analysis of colocalized DiOC7 and VP fluorescence showed ASPC-1 with higher accumulation of VP 3 hrs after injection and more VP at a distance from blood vessels compared to Panc-1. This work shows the need for techniques to analyze photosensitizer distribution in order to optimize photodynamic therapy as an effective treatment for pancreatic tumors.

O'Hara, Julia A.; Samkoe, Kimberley S.; Chen, Alina; Isabelle, Martin; Hoopes, P. J.; Hasan, Tayyaba; Pogue, Brian W.

2012-02-01

103

Extracting decay curves of the correlated fluorescence photons measured in fluorescence correlation spectroscopy  

NASA Astrophysics Data System (ADS)

We have developed a new method to extract the decay curves of the correlated fluorescence photons from the data of fluorescence correlation spectroscopy using time-correlated single photon counting. In this method, a two-dimensional correlation map of photon pairs is generated at an absolute delay time with reference to the excitation-emission delay of each photon. Using a dye-labeled DNA as an example, we have demonstrated that the decay curve of the correlated fluorescence photons is separated from the uncorrelated background signals simply by subtracting a two-dimensional correlation map at sufficiently long delay time without additional prior information.

Ishii, Kunihiko; Tahara, Tahei

2012-01-01

104

In vivo fluorescent imaging of the mouse retina using adaptive optics  

E-print Network

In vivo fluorescent imaging of the mouse retina using adaptive optics David P. Biss, Daniel Sumorok- tive optics. We demonstrate an adaptive optics system for in vivo imaging of fluorescent structures.1­3 In human retinal imaging and more recently in primates, adaptive optics (AO) has provided near

105

Sucrose Monoester Micelles Size Determined by Fluorescence Correlation Spectroscopy (FCS)  

PubMed Central

One of the several uses of sucrose detergents, as well as other micelle forming detergents, is the solubilization of different membrane proteins. Accurate knowledge of the micelle properties, including size and shape, are needed to optimize the surfactant conditions for protein purification and membrane characterization. We synthesized sucrose esters having different numbers of methylene subunits on the substituent to correlate the number of methylene groups with the size of the corresponding micelles. We used Fluorescence Correlation Spectroscopy (FCS) and two photon excitation to determine the translational D of the micelles and calculate their corresponding hydrodynamic radius, Rh. As a fluorescent probe we used LAURDAN (6-dodecanoyl-2-dimethylaminonaphthalene), a dye highly fluorescent when integrated in the micelle and non-fluorescent in aqueous media. We found a linear correlation between the size of the tail and the hydrodynamic radius of the micelle for the series of detergents measured. PMID:22216230

Sanchez, Susana A.; Gratton, Enrico; Zanocco, Antonio L.; Lemp, Else; Gunther, German

2011-01-01

106

Detection of residual concentration of imidazoline inhibitors in oilfield production water based on fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Fluorescence spectroscopy is applied to detect the residual concentration of imidazoline inhibitors in this study. Imidazoline inhibitors emit a weak fluorescence spectrum at 445 nm excitation wavelength, the addition of eosin Y can enhance the fluorescence intensity obviously and the excitation wavelength is located at 521 nm. The fluorescence intensity has a good linear relationship with the inhibitor concentration. The common ions in oilfield production water do not affect imidazoline detection using eosin Y as fluorescence agent based on fluorescence spectroscopy.

Chen, Zhenyu; Guo, Xianxian; Qiu, Yubing; Guo, Xingpeng

2012-09-01

107

Quantification of osmotic water transport in vivo using fluorescent albumin.  

PubMed

Osmotic water transport across the peritoneal membrane is applied during peritoneal dialysis to remove the excess water accumulated in patients with end-stage renal disease. The discovery of aquaporin water channels and the generation of transgenic animals have stressed the need for novel and accurate methods to unravel molecular mechanisms of water permeability in vivo. Here, we describe the use of fluorescently labeled albumin as a reliable indicator of osmotic water transport across the peritoneal membrane in a well-established mouse model of peritoneal dialysis. After detailed evaluation of intraperitoneal tracer mass kinetics, the technique was validated against direct volumetry, considered as the gold standard. The pH-insensitive dye Alexa Fluor 555-albumin was applied to quantify osmotic water transport across the mouse peritoneal membrane resulting from modulating dialysate osmolality and genetic silencing of the water channel aquaporin-1 (AQP1). Quantification of osmotic water transport using Alexa Fluor 555-albumin closely correlated with direct volumetry and with estimations based on radioiodinated ((125)I) serum albumin (RISA). The low intraperitoneal pressure probably accounts for the negligible disappearance of the tracer from the peritoneal cavity in this model. Taken together, these data demonstrate the appropriateness of pH-insensitive Alexa Fluor 555-albumin as a practical and reliable intraperitoneal volume tracer to quantify osmotic water transport in vivo. PMID:25100279

Morelle, Johann; Sow, Amadou; Vertommen, Didier; Jamar, François; Rippe, Bengt; Devuyst, Olivier

2014-10-15

108

Long-Term Retention of Fluorescent Quantum Dots In Vivo  

NASA Astrophysics Data System (ADS)

Quantum dots that emit in the near-infrared can be used in vivo to follow circulation, to target the reticuloendothelial system, and to map lymphatic drainage from normal tissues and tumors. We have explored the role of surface charge and passivation by polyethylene glycol in determining circulating lifetimes and sites of deposition. Use of long polyethylene glycol polymers increases circulating lifetime. Changing surface charge can partially direct quantum dots to the liver and spleen, or the lymph nodes. Quantum dots are cleared in the order liver > spleen > bone marrow > lymph nodes. Quantum dots retained by lymph nodes maintained fluorescence for two years, suggesting either that the coating is extremely stable or that some endosomes preserve quantum dot function. We also explored migration from tumors to sentinel lymph nodes using tumor models in mice; surface charge and size make little difference to transport from tumors. Antibody and Fab-conjugates of polymer-coated quantum dots failed to target tumors in vivo, probably because of size.

Ballou, Byron; Ernst, Lauren A.; Andreko, Susan; Eructiez, Marcel P.; Lagerholm, B. Christoffer; Waggoner, Alan S.

109

Multiphoton microscopy, fluorescence lifetime imaging and optical spectroscopy for the diagnosis of neoplasia  

NASA Astrophysics Data System (ADS)

Cancer morbidity and mortality is greatly reduced when the disease is diagnosed and treated early in its development. Tissue biopsies are the gold standard for cancer diagnosis, and an accurate diagnosis requires a biopsy from the malignant portion of an organ. Light, guided through a fiber optic probe, could be used to inspect regions of interest and provide real-time feedback to determine the optimal tissue site for biopsy. This approach could increase the diagnostic accuracy of current biopsy procedures. The studies in this thesis have characterized changes in tissue optical signals with carcinogenesis, increasing our understanding of the sensitivity of optical techniques for cancer detection. All in vivo studies were conducted on the dimethylbenz[alpha]anthracene treated hamster cheek pouch model of epithelial carcinogenesis. Multiphoton microscopy studies in the near infrared wavelength region quantified changes in tissue morphology and fluorescence with carcinogenesis in vivo. Statistically significant morphological changes with precancer included increased epithelial thickness, loss of stratification in the epithelium, and increased nuclear diameter. Fluorescence changes included a statistically significant decrease in the epithelial fluorescence intensity per voxel at 780 nm excitation, a decrease in the fluorescence lifetime of protein-bound nicotinamide adenine dinucleotide (NADH, an electron donor in oxidative phosphorylation), and an increase in the fluorescence lifetime of protein-bound flavin adenine dinucleotide (FAD, an electron acceptor in oxidative phosphorylation) with precancer. The redox ratio (fluorescence intensity of FAD/NADH, a measure of the cellular oxidation-reduction state) did not significantly change with precancer. Cell culture experiments (MCF10A cells) indicated that the decrease in protein-bound NADH with precancer could be due to increased levels of glycolysis. Point measurements of diffuse reflectance and fluorescence spectra in the ultraviolet to visible wavelength range indicated that the most diagnostic optical signals originate from sub-surface tissue layers. Optical properties extracted from these spectroscopy measurements showed a significant decrease in the hemoglobin saturation, absorption coefficient, reduced scattering coefficient and fluorescence intensity (at 400 nm excitation) in neoplastic compared to normal tissues. The results from these studies indicate that multiphoton microscopy and optical spectroscopy can non-invasively provide information on tissue structure and function in vivo that is related to tissue pathology.

Skala, Melissa Caroline

2007-12-01

110

Phytoplankton in Lake Tanganyika – vertical and horizontal distribution of in vivo fluorescence  

Microsoft Academic Search

Determinations of chlorophyll a and in vivo fluorescence of photosynthetic pigments were used to study vertical and horizontal distribution of phytoplankton in Lake Tanganyika (East Africa). Blue excited fluorescence (IVFb) was an approximate predictor of chlorophyll a at different depths and locations. Green excited fluorescence (IVFg), which reflects phycoerythrin in cyanobacteria, explained chlorophyll a variation equally well, and in combination

K. Salonen; J. Sarvala; M. Järvinen; V. Langenberg; M. Nuottajärvi; K. Vuorio; D. B. R. Chitamwebwa

1999-01-01

111

Fluorescence Lifetime Correlation Spectroscopy (FLCS): Concepts, Applications and Outlook  

PubMed Central

Fluorescence Lifetime Correlation Spectroscopy (FLCS) is a variant of fluorescence correlation spectroscopy (FCS), which uses differences in fluorescence intensity decays to separate contributions of different fluorophore populations to FCS signal. Besides which, FLCS is a powerful tool to improve quality of FCS data by removing noise and distortion caused by scattered excitation light, detector thermal noise and detector after pulsing. We are providing an overview of, to our knowledge, all published applications of FLCS. Although these are not numerous so far, they illustrate possibilities for the technique and the research topics in which FLCS has the potential to become widespread. Furthermore, we are addressing some questions which may be asked by a beginner user of FLCS. The last part of the text reviews other techniques closely related to FLCS. The generalization of the idea of FLCS paves the way for further promising application of the principle of statistical filtering of signals. Specifically, the idea of fluorescence spectral correlation spectroscopy is here outlined. PMID:23202928

Kapusta, Peter; Machá?, Radek; Benda, Aleš; Hof, Martin

2012-01-01

112

Dynamic tissue analysis using time- and wavelength-resolved fluorescence spectroscopy for atherosclerosis diagnosis  

PubMed Central

Simultaneous time- and wavelength-resolved fluorescence spectroscopy (STWRFS) was developed and tested for the dynamic characterization of atherosclerotic tissue ex vivo and arterial vessels in vivo. Autofluorescence, induced by a 337 nm, 700 ps pulsed laser, was split to three wavelength sub-bands using dichroic filters, with each sub-band coupled into a different length of optical fiber for temporal separation. STWRFS allows for fast recording/analysis (few microseconds) of time-resolved fluorescence emission in these sub-bands and rapid scanning. Distinct compositions of excised human atherosclerotic aorta were clearly discriminated over scanning lengths of several centimeters based on fluorescence lifetime and the intensity ratio between 390 and 452 nm. Operation of STWRFS blood flow was further validated in pig femoral arteries in vivo using a single-fiber probe integrated with an ultrasound imaging catheter. Current results demonstrate the potential of STWRFS as a tool for real-time optical characterization of arterial tissue composition and for atherosclerosis research and diagnosis. PMID:21369214

Sun, Yinghua; Sun, Yang; Stephens, Douglas; Xie, Hongtao; Phipps, Jennifer; Saroufeem, Ramez; Southard, Jeffrey; Elson, Daniel S.; Marcu, Laura

2011-01-01

113

Fluorescence Tomography and Magnetic Resonance Imaging of Myocardial Macrophage Infiltration in Infarcted Myocardium In Vivo  

Microsoft Academic Search

Background—Fluorescence imaging of the heart is currently limited to invasive ex vivo or in vitro applications. We hypothesized that the adaptation of advanced transillumination and tomographic techniques would allow noninvasive fluorescence images of the heart to be acquired in vivo and be coregistered with in vivo cardiac magnetic resonance images. Methods and Results—The uptake of the magnetofluorescent nanoparticle CLIO-Cy5.5 by

David E. Sosnovik; Matthias Nahrendorf; Nikolaos Deliolanis; Mikhail Novikov; Elena Aikawa; Lee Josephson; Anthony Rosenzweig; Ralph Weissleder; Vasilis Ntziachristos

2010-01-01

114

Two-dimensional fluorescence lifetime correlation spectroscopy. 1. Principle.  

PubMed

Fluorescence correlation spectroscopy (FCS) is a unique tool for investigating microsecond molecular dynamics of complex molecules in equilibrium. However, application of FCS in the study of molecular dynamics has been limited, owing to the complexity in the extraction of physically meaningful information. In this work, we develop a new method that combines FCS and time-correlated single photon counting (TCPSC) to extract unambiguous information about equilibrium dynamics of complex molecular systems. In this method, which we name two-dimensional fluorescence lifetime correlation spectroscopy (2D FLCS), we analyze the correlation of the fluorescence photon pairs, referring to the fluorescence lifetime. We first obtain the correlations of the photon pairs with respect to the excitation-emission delay times in the form of a two-dimensional (2D) map. Then, the 2D map is converted to the correlations between different species that have distinct fluorescence lifetimes using inverse Laplace transformation. This 2D FLCS is capable of visualizing the equilibration dynamics of complex molecules with microsecond time resolution at the single-molecule level. We performed a kinetic Monte Carlo simulation of a TCPSC-FCS experiment as a proof-of-principle example. The result clearly shows the validity of the proposed method and its high potential in analyzing the photon data of dynamic systems. PMID:23977832

Ishii, Kunihiko; Tahara, Tahei

2013-10-01

115

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

116

Femtosecond broadband fluorescence upconversion spectroscopy: Improved setup and photometric correction  

SciTech Connect

A setup for fluorescence upconversion spectroscopy (FLUPS) is described which has 80 fs temporal response (fwhm) for emission in the spectral range 425-750 nm. Broadband phase matching is achieved with tilted gate pulses at 1340 nm. Background from harmonics of the gate pulse is removed and sensitivity increased compared to previous designs. Photometric calibration of the upconversion process is performed with a set of fluorescent dyes. For Coumarin 153 in methanol the peak position, bandwidth, and asymmetry depending on delay time are reported.

Zhang, X.-X. [Photonics Center, College of Physical Science, Nankai University, Tianjin (China); Department of Chemistry, Humboldt Universitaet zu Berlin (Germany); Wuerth, C.; Resch-Genger, U. [Federal Institute for Materials Research and Testing, Berlin (Germany); Zhao, L. [Photonics Center, College of Physical Science, Nankai University, Tianjin (China); Ernsting, N. P.; Sajadi, M. [Department of Chemistry, Humboldt Universitaet zu Berlin (Germany)

2011-06-15

117

Diffusivity of asphaltene molecules by fluorescence correlation spectroscopy.  

PubMed

Using fluorescence correlation spectroscopy (FCS) we measure the translational diffusion coefficient of asphaltene molecules in toluene at extremely low concentrations (0.03-3.0 mg/L): where aggregation does not occur. We find that the translational diffusion coefficient of asphaltene molecules in toluene is about 0.35 x 10(-5) cm(2)/s at room temperature. This diffusion coefficient corresponds to a hydrodynamic radius of approximately 1 nm. These data confirm previously estimated size from rotational diffusion studied using fluorescence depolarization. The implication of this concurrence is that asphaltene molecular structures are monomeric, not polymeric. PMID:16805495

Andrews, A Ballard; Guerra, Rodrigo E; Mullins, Oliver C; Sen, Pabitra N

2006-07-01

118

Fluorescence correlation spectroscopy: Diagnostics for sparse?molecules  

PubMed Central

The robust glow of molecular fluorescence renders even sparse molecules detectable and susceptible to analysis for concentration, mobility, chemistry, and photophysics. Correlation spectroscopy, a statistical-physics-based tool, gleans quantitative information from the spontaneously fluctuating fluorescence signals obtained from small molecular ensembles. This analytical power is available for studying molecules present at minuscule concentrations in liquid solutions (less than one nanomolar), or even on the surfaces of living cells at less than one macromolecule per square micrometer. Indeed, routines are becoming common to detect, locate, and examine individual molecules under favorable conditions. PMID:9342306

Maiti, Sudipta; Haupts, Ulrich; Webb, Watt W.

1997-01-01

119

Surface-plasmon field-enhanced fluorescence spectroscopy  

Microsoft Academic Search

We describe the combination of surface plasmon- and fluorescence spectroscopy for sensor applications. The resonant excitation of PSP modes at a metal\\/buffer-interface in a flow cell results in optical field intensities largely enhanced compared to the incoming laser light: a factor of 16, calculated for a Au\\/water interface by Fresnel formulas was experimentally confirmed. This field enhancement can be used

Thorsten Liebermann; Wolfgang Knoll

2000-01-01

120

Three-dimensional in vivo fluorescence diffuse optical tomography of breast cancer in humans.  

PubMed

We present three-dimensional (3D) in vivo images of human breast cancer based on fluorescence diffuse optical tomography (FDOT). To our knowledge, this work represents the first reported 3D fluorescence tomography of human breast cancer in vivo. In our protocol, the fluorophore Indocyanine Green (ICG) is injected intravenously. Fluorescence excitation and detection are accomplished in the soft-compression, parallel-plane, transmission geometry using laser sources at 786 nm and spectrally filtered CCD detection. Phantom and in vivo studies confirm the signals are due to ICG fluorescence, rather than tissue autofluorescence and excitation light leakage. Fluorescence images of breast tumors were in good agreement with those of MRI, and with DOT based on endogenous contrast. Tumorto- normal tissue contrast based on ICG fluorescence was two-to-four-fold higher than contrast based on hemoglobin and scattering parameters. In total the measurements demonstrate that FDOT of breast cancer is feasible and promising. PMID:19546980

Corlu, Alper; Choe, Regine; Durduran, Turgut; Rosen, Mark A; Schweiger, Martin; Arridge, Simon R; Schnall, Mitchell D; Yodh, Arjun G

2007-05-28

121

In vivo fluorescence lifetime tomography of a FRET probe expressed in mouse.  

PubMed

Förster resonance energy transfer (FRET) is a powerful biological tool for reading out cell signaling processes. In vivo use of FRET is challenging because of the scattering properties of bulk tissue. By combining diffuse fluorescence tomography with fluorescence lifetime imaging (FLIM), implemented using wide-field time-gated detection of fluorescence excited by ultrashort laser pulses in a tomographic imaging system and applying inverse scattering algorithms, we can reconstruct the three dimensional spatial localization of fluorescence quantum efficiency and lifetime. We demonstrate in vivo spatial mapping of FRET between genetically expressed fluorescent proteins in live mice read out using FLIM. Following transfection by electroporation, mouse hind leg muscles were imaged in vivo and the emission of free donor (eGFP) in the presence of free acceptor (mCherry) could be clearly distinguished from the fluorescence of the donor when directly linked to the acceptor in a tandem (eGFP-mCherry) FRET construct. PMID:21750768

McGinty, James; Stuckey, Daniel W; Soloviev, Vadim Y; Laine, Romain; Wylezinska-Arridge, Marzena; Wells, Dominic J; Arridge, Simon R; French, Paul M W; Hajnal, Joseph V; Sardini, Alessandro

2011-07-01

122

In vivo fluorescence lifetime tomography of a FRET probe expressed in mouse  

PubMed Central

Förster resonance energy transfer (FRET) is a powerful biological tool for reading out cell signaling processes. In vivo use of FRET is challenging because of the scattering properties of bulk tissue. By combining diffuse fluorescence tomography with fluorescence lifetime imaging (FLIM), implemented using wide-field time-gated detection of fluorescence excited by ultrashort laser pulses in a tomographic imaging system and applying inverse scattering algorithms, we can reconstruct the three dimensional spatial localization of fluorescence quantum efficiency and lifetime. We demonstrate in vivo spatial mapping of FRET between genetically expressed fluorescent proteins in live mice read out using FLIM. Following transfection by electroporation, mouse hind leg muscles were imaged in vivo and the emission of free donor (eGFP) in the presence of free acceptor (mCherry) could be clearly distinguished from the fluorescence of the donor when directly linked to the acceptor in a tandem (eGFP-mCherry) FRET construct. PMID:21750768

McGinty, James; Stuckey, Daniel W.; Soloviev, Vadim Y.; Laine, Romain; Wylezinska-Arridge, Marzena; Wells, Dominic J.; Arridge, Simon R.; French, Paul M. W.; Hajnal, Joseph V.; Sardini, Alessandro

2011-01-01

123

The spectroscopic basis of Fluorescence Triple Correlation Spectroscopy  

PubMed Central

We have developed Fluorescence Triple Correlation Spectroscopy (F3CS) as an extension of the widely-used fluorescence microscopy technique Fluorescence Correlation Spectroscopy. F3CS correlates three signals at once and provides additional capabilities for the study of systems with complex stoichiometry, kinetic processes and irreversible reactions. A general theory of F3CS was developed to describe the interplay of molecular dynamics and microscope optics, leading to an analytical function to predict experimental triple correlations of molecules that freely diffuse through the tight focus of the microscope. Experimental correlations were calculated from raw fluorescence data using triple correlation integrals that extend multiple-tau correlation theory to delay times in two dimensions. The quality of experimental data was improved by tuning specific spectroscopic parameters and employing multiple independent detectors to minimize optoelectronic artifacts. Experiments with the reversible system of freely-diffusing 16S rRNA revealed that triple correlation functions contain symmetries predicted from time-reversal arguments. Irreversible systems are shown to break these symmetries and correlation strategies were developed to detect time-reversal asymmetries in a comprehensive way with respect to two delay times, each spanning many orders of magnitude in time. The correlation strategies, experimental approaches and theory developed here enable studies of the composition and dynamics of complex systems using F3CS. PMID:22229664

Ridgeway, William K.; Millar, David P.; Williamson, James R.

2012-01-01

124

Fluorescence spectroscopy using indocyanine green for lymph node mapping  

NASA Astrophysics Data System (ADS)

The principles of cancer treatment has for years been radical resection of the primary tumor. In the oncologic surgeries where the affected cancer site is close to the lymphatic system, it is as important to detect the draining lymph nodes for metastasis (lymph node mapping). As a replacement for conventional radioactive labeling, indocyanine green (ICG) has shown successful results in lymph node mapping; however, most of the ICG fluorescence detection techniques developed are based on camera imaging. In this work, fluorescence spectroscopy using a fiber-optical probe was evaluated on a tissue-like ICG phantom with ICG concentrations of 6-64 ?M and on breast tissue from five patients. Fiber-optical based spectroscopy was able to detect ICG fluorescence at low intensities; therefore, it is expected to increase the detection threshold of the conventional imaging systems when used intraoperatively. The probe allows spectral characterization of the fluorescence and navigation in the tissue as opposed to camera imaging which is limited to the view on the surface of the tissue.

Haj-Hosseini, Neda; Behm, Pascal; Shabo, Ivan; Wârdell, Karin

2014-02-01

125

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

126

Fluorescence recovery spectroscopy as a probe of slow rotational motions.  

PubMed Central

Pump-and-probe techniques can be used to follow the slow rotational motions of fluorescent labels bound to macromolecules in solution. A strong pulse of polarized light initially anisotropically depletes the ground-state population. A continuous low-intensity beam of variable polarization then probes the anisotropic ground-state distribution. Using an additional emission polarizer, the generated fluorescence can be recorded as it rises towards its prepump value. A general theory of fluorescence recovery spectroscopy (FRS) is presented that allows for irreversible depletion processes like photobleaching as well as slowly reversible processes like triplet formation. In either case, rotational motions modulate recovery through cosine-squared laws for dipolar absorption and emission processes. Certain pump, probe, and emission polarization directions eliminate the directional dependence of either dipole and simplify the resulting expressions. Two anisotropy functions can then be constructed to independently monitor the rotations of either dipole. These functions are identical in form to the anisotropy used in fluorescence depolarization measurements and all rotational models developed there apply here with minor modifications. Several setups are discussed that achieve the necessary polarization alignments. These include right-angle detection equipment that is commonly available in laboratories using fluorescence methods. Images FIGURE 1 FIGURE 2 FIGURE 3 PMID:6518257

Wegener, W A

1984-01-01

127

Synchronous fluorescence spectroscopy for analysis of wine and wine distillates  

NASA Astrophysics Data System (ADS)

Wine and brandies are multicomponent systems and conventional fluorescence techniques, relying on recording of single emission or excitation spectra, are often insufficient. In such cases synchronous fluorescence spectra can be used for revealing the potential of the fluorescence techniques. The technique is based on simultaneously scanning of the excitation and emission wavelength with constant difference (??) maintained between them. In this study the measurements were made using FluoroLog3 spectrofluorimeter (HORIBA Jobin Yvon, France) and collected for excitation and emission in the wavelength region 220 - 700 nm using wavelength interval ?? from 10 to 100 nm in 10 nm steps. This research includes the results obtained for brandy and red wine samples. Fluorescence analysis takes advantage in the presence of natural fluorophores in wines and brandies, such as gallic, vanillic, p-coumaric, syringic, ferulic acid, umbelliferone, scopoletin and etc. Applying of synchronous fluorescence spectroscopy for analysis of these types of alcohols allows us to estimate the quality of wines and also to detect adulteration of brandies like adding of a caramel to wine distillates for imitating the quality of the original product aged in oak casks.

Andreeva, Ya.; Borisova, E.; Genova, Ts.; Zhelyazkova, Al.; Avramov, L.

2015-01-01

128

Stability of siRNA polyplexes from poly(ethylenimine) and poly(ethylenimine)-g-poly(ethylene glycol) under in vivo conditions: Effects on pharmacokinetics and biodistribution measured by Fluorescence Fluctuation Spectroscopy and Single Photon Emission Computed Tomography (SPECT) imaging  

Microsoft Academic Search

In search of optimizing siRNA delivery systems for systemic application, one critical parameter remains their stability in blood circulation. In this study, we have traced pharmacokinetics and biodistribution of each component of siRNA polyplexes formed with polyethylenimine 25 kDa (PEI) or PEGylated PEIs by in vivo real-time gamma camera recording, SPECT imaging, and scintillation counting of blood samples and dissected organs.

Olivia M. Merkel; Damiano Librizzi; Andreas Pfestroff; Tino Schurrat; Kevin Buyens; Niek N. Sanders; Stefaan C. De Smedt; Martin Béhé; Thomas Kissel

2009-01-01

129

The study of blue LED to induce fluorescence spectroscopy and fluorescence imaging for oral carcinoma detection  

NASA Astrophysics Data System (ADS)

Fluorescence spectroscopy and fluorescence imaging diagnosis of malignant lesions provides us with a new method to diagnose diseases in precancerous stage. Early diagnosis of disease has significant importance in cancer treatment, because most cancers can be cured well in precancerous, especially when the diffusion of cancer is limited in a restricted region. In this study, Golden hamster models were applied to 5% 9, 10 dimethyl-1, 2-benzanthracene (DMBA) to induce hamster buccal cheek pouch carcinoma three times a week. Rose Bengal, which has been used in clinican for years and avoids visible side-effect to human was chosen as photosensitizer. 405 nm blue LED was used to induce the fluorescence of photosensitizer. After topical application of photosensitizer, characteristic red emission fluorescence peak was observed around 600nm. Similar, normal oral cavity has special luminescence around 480nm. Fluorescence spectroscopy technology is based on analysing emission peaks of photosensitizer in the areas of oral carcinoma, moreover, red-to-green (IR/IG) intensity ratio is also applied as a diagnostic algorithm. A CCD which is connected with a computer is used to take pictures at carcinoma areas through different filters. Fluorescence images from normal hamster buccal cheek pouch are compared with those from carcinogen-induced models of carcinoma, and morphological differences between normal and lesion tissue can be distinguished. The pictures are analyzed by Matlab and shown on the screen of computer. This paper demonstrates that Rose Bengal could be used as photosensitizer to detect oral carcinoma, and blue LED as excitation source could not only have a good effect to diagnose oral carcinoma, but also decrease cost greatly.

Zheng, Longjiang; Hu, Yuanting

2009-07-01

130

In vivo imaging with near-infrared fluorescence lifetime contrast  

Microsoft Academic Search

Fluorescence imaging is a mainstay of biomedical research, allowing detection of molecular events in both fixed and living cells, tissues and whole animals. Such high resolution fluorescence imaging is hampered by unwanted signal from intrinsic background fluorescence and scattered light. The signal to background ratio can be improved by using extrinsic contrast agents and greatly enhanced by multispectral imaging methods.

Walter J. Akers; Mikhail Y. Berezin; Hyeran Lee; Samuel Achilefu

2009-01-01

131

Dynamic and unique nucleolar microenvironment revealed by fluorescence correlation spectroscopy.  

PubMed

Organization and functions of the nucleolus is maintained by mobilities and interactions of nucleolar factors. Because the nucleolus is a densely packed structure, molecular crowding effects determined by the molecular concentrations and mobilities in the nucleolus should also be important for regulating nucleolar organization and functions. However, such molecular property of nucleolar organization is not fully understood. To understand the biophysical property of nucleolar organization, the diffusional behaviors of inert green fluorescent protein (GFP) oligomers with or without nuclear localization signals (NLSs) were analyzed under various conditions by fluorescence correlation spectroscopy. Our result demonstrates that the mobility of GFPs inside the nucleolus and the nucleoplasm can be represented by single free diffusion under normal conditions, even though the mobility in the nucleolus is considerably slower than that in the chromatin region. Moreover, the free diffusion of GFPs is found to be significantly size- and NLS-dependent only in the nucleolus. Interestingly, the mobility in the nucleolus is highly sensitive to ATP depletion, as well as actinomycin D (ActD) treatment. In contrast, the ultra-structure of the nucleolus was not significantly changed by ATP depletion but was changed by ActD treatment. These results suggest that the nucleolus behaves similarly to an open aqueous-phase medium with an increased molecular crowding effect that depends on both energy and transcription.-Park, H., Han, S.-S., Sako, Y., Pack, C.-G. Dynamic and unique nucleolar microenvironment revealed by fluorescence correlation spectroscopy. PMID:25404711

Park, Hweon; Han, Sung-Sik; Sako, Yasushi; Pack, Chan-Gi

2015-03-01

132

A comparative evaluation of Raman and fluorescence spectroscopy for optical diagnosis of oral neoplasia  

NASA Astrophysics Data System (ADS)

We report the results of a comparative evaluation of in vivo fluorescence and Raman spectroscopy for diagnosis of oral neoplasia. The study carried out at Tata Memorial Hospital, Mumbai, involved 26 healthy volunteers and 138 patients being screened for neoplasm of oral cavity. Spectral measurements were taken from multiple sites of abnormal as well as apparently uninvolved contra-lateral regions of the oral cavity in each patient. The different tissue sites investigated belonged to one of the four histopathology categories: 1) squamous cell carcinoma (SCC), 2) oral sub-mucous fibrosis (OSMF), 3) leukoplakia (LP) and 4) normal squamous tissue. A probability based multivariate statistical algorithm utilizing nonlinear Maximum Representation and Discrimination Feature for feature extraction and Sparse Multinomial Logistic Regression for classification was developed for direct multi-class classification in a leave-one-patient-out cross validation mode. The results reveal that the performance of Raman spectroscopy is considerably superior to that of fluorescence in stratifying the oral tissues into respective histopathologic categories. The best classification accuracy was observed to be 90%, 93%, 94%, and 89% for SCC, SMF, leukoplakia, and normal oral tissues, respectively, on the basis of leave-one-patient-out cross-validation, with an overall accuracy of 91%. However, when a binary classification was employed to distinguish spectra from all the SCC, SMF and leukoplakik tissue sites together from normal, fluorescence and Raman spectroscopy were seen to have almost comparable performances with Raman yielding marginally better classification accuracy of 98.5% as compared to 94% of fluorescence.

Majumder, S. K.; Krishna, H.; Sidramesh, M.; Chaturvedi, P.; Gupta, P. K.

2010-12-01

133

A comparative evaluation of Raman and fluorescence spectroscopy for optical diagnosis of oral neoplasia  

NASA Astrophysics Data System (ADS)

We report the results of a comparative evaluation of in vivo fluorescence and Raman spectroscopy for diagnosis of oral neoplasia. The study carried out at Tata Memorial Hospital, Mumbai, involved 26 healthy volunteers and 138 patients being screened for neoplasm of oral cavity. Spectral measurements were taken from multiple sites of abnormal as well as apparently uninvolved contra-lateral regions of the oral cavity in each patient. The different tissue sites investigated belonged to one of the four histopathology categories: 1) squamous cell carcinoma (SCC), 2) oral sub-mucous fibrosis (OSMF), 3) leukoplakia (LP) and 4) normal squamous tissue. A probability based multivariate statistical algorithm utilizing nonlinear Maximum Representation and Discrimination Feature for feature extraction and Sparse Multinomial Logistic Regression for classification was developed for direct multi-class classification in a leave-one-patient-out cross validation mode. The results reveal that the performance of Raman spectroscopy is considerably superior to that of fluorescence in stratifying the oral tissues into respective histopathologic categories. The best classification accuracy was observed to be 90%, 93%, 94%, and 89% for SCC, SMF, leukoplakia, and normal oral tissues, respectively, on the basis of leave-one-patient-out cross-validation, with an overall accuracy of 91%. However, when a binary classification was employed to distinguish spectra from all the SCC, SMF and leukoplakik tissue sites together from normal, fluorescence and Raman spectroscopy were seen to have almost comparable performances with Raman yielding marginally better classification accuracy of 98.5% as compared to 94% of fluorescence.

Majumder, S. K.; Krishna, H.; Sidramesh, M.; Chaturvedi, P.; Gupta, P. K.

2011-08-01

134

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

135

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

136

Analysis of green fluorescent protein bioluminescence in vivo and in vitro using a glow discharge  

NASA Astrophysics Data System (ADS)

The discovery of fluorescent proteins has been a revolution in cell biology and related sciences because of their many applications, mainly emphasizing their use as cellular markers. The green fluorescent protein (GFP) is one of the most used as it requires no cofactors to generate fluorescence and retains this property into any organism when it is expressed by recombinant DNA techniques, which is a great advantage. In this work, we analyze the emission spectra of recombinant green fluorescent protein in vivo and in vitro exposed to a glow discharge plasma of nitrogen in order to relate electron temperature to fluorescence intensity.

Hernández, L.; Mandujano, L. A.; Cuevas, J.; Reyes, P. G.; Osorio-González, D.

2015-03-01

137

Investigation of asphaltene association by front-face fluorescence spectroscopy.  

PubMed

The tendency of asphaltenes to aggregate and form clusters in solvents was studied by fluorescence spectroscopy. This was done by evaluating the relative fluorescence quantum yield of asphaltenes diluted at several concentrations in toluene and by studying the changes in the fluorescence spectra of asphaltene solutions as the composition of the solvent, toluene and cyclohexane, is changed. The asphaltene fraction (heptane insoluble) was collected from a Brazilian heavy crude oil, and solutions of this material varying from 0.016 g/L up to 10 g/L were prepared in toluene. Front-face emission spectra were obtained in two wavelength ranges, from 310 to 710 nm, excited at 300 nm (short range), and from 410 to 710 nm, excited at 400 nm (long range). Severe quenching was observed at concentrations above about 0.1 g/L. Stern-Volmer plots (reciprocal of quantum yield against concentration) exhibited nonlinear, downward-curved behavior, indicating that a more complex suppression mechanism, probably influenced by the association of the asphaltene molecules, is taking place. The same asphaltenes were dissolved (0.1 g/L) in binary mixtures of toluene and cyclohexane, and emission spectra in both the short range and long range were obtained. Fluorescence was progressively quenched at longer wavelengths of the spectra as the proportion of cyclohexane in the solvent grew. Cyclohexane, a poor asphaltene solvent, is probably inducing static quenching through association of asphaltenes. PMID:14658659

Albuquerque, Flávio Cortiñas; Nicodem, David E; Rajagopal, Krishnaswamy

2003-07-01

138

Fluorescence spectroscopy for endogenous porphyrins in human facial skin  

NASA Astrophysics Data System (ADS)

The activity of certain bacteria in skin is known to correlate to the presence of porphyrins. In particular the presence of coproporphyrin produced by P.acnes inside plugged pores has been correlated to acne vulgaris. Another porphyrin encountered in skin is protoporphyrin IX, which is produced by the body in the pathway for production of heme. In the present work, a fluorescence spectroscopy system was developed to measure the characteristic spectrum and quantify the two types of porphyrins commonly present in human facial skin. The system is comprised of a Xe lamp both for fluorescence excitation and broadband light source for diffuse reflectance measurements. A computer-controlled filter wheel enables acquisition of sequential spectra, first excited by blue light at 405 nm then followed by the broadband light source, at the same location. The diffuse reflectance spectrum was used to correct the fluorescence spectrum due to the presence of skin chromophores, such as blood and melanin. The resulting fluorescence spectra were employed for the quantification of porphyrin concentration in a population of healthy subjects. The results show great variability on the concentration of these porphyrins and further studies are being conducted to correlate them with skin conditions such as inflammation and acne vulgaris.

Seo, I.; Tseng, S. H.; Cula, G. O.; Bargo, P. R.; Kollias, N.

2009-02-01

139

Hazards and benefits of in-vivo Raman spectroscopy of human skin  

NASA Astrophysics Data System (ADS)

The resurgence of Raman spectroscopy, in the late 1980's has led to an increase in the use of the technique for the analysis of biological tissues. Consequently, Raman spectroscopy is now regarded to be a well-established non- invasive, non-destructive technique, which is used to obtain good quality spectra from biological tissues with minimal fluorescence. What is presently of interest to our group is to develop further and establish the technique for in vivo investigations of healthy and diseased skin. This presentation discusses some potentially valuable clinical applications of the technique, and also highlights some of the experimental difficulties that were encountered when examining patients who were receiving treatment for psoriasis.

Carter, Elizabeth A.; Williams, Adrian C.; Barry, Brian W.; Edwards, Howell G.

1999-04-01

140

Evaluation of a fiber-optic fluorescence spectroscopy system to assist neurosurgical tumor resections  

NASA Astrophysics Data System (ADS)

The highly malignant brain tumor, glioblastoma multiforme, is difficult to totally resect without aid due to its infiltrative way of growing and its morphological similarities to surrounding functioning brain under direct vision in the operating field. The need for an inexpensive and robust real-time visualizing system for resection guiding in neurosurgery has been formulated by research groups all over the world. The main goal is to develop a system that helps the neurosurgeon to make decisions during the surgical procedure. A compact fiber optic system using fluorescence spectroscopy has been developed for guiding neurosurgical resections. The system is based on a high power light emitting diode at 395 nm and a spectrometer. A fiber bundle arrangement is used to guide the excitation light and fluorescence light between the instrument and the tissue target. The system is controlled through a computer interface and software package especially developed for the application. This robust and simple instrument has been evaluated in vivo both on healthy skin but also during a neurosurgical resection procedure. Before surgery the patient received orally a low dose of 5-aminolevulinic acid, converted to the fluorescence tumor marker protoporphyrin IX in the malignant cells. Preliminary results indicate that PpIX fluorescence and brain tissue autofluorescence can be recorded with the help of the developed system intraoperatively during resection of glioblastoma multiforme.

Ilias, Michail A.; Richter, Johan; Westermark, Frida; Brantmark, Martin; Andersson-Engels, Stefan; Wårdell, Karin

2007-07-01

141

Fluorescence sensing systems: In vivo detection of biophysical variations in field corn due to nitrogen supply  

Microsoft Academic Search

Leaf and canopy fluorescence properties of field corn (Zea mays L.) grown under varying levels of nitrogen (N) fertilization were characterized to provide an improved N sensing capability which may assist growers in site-specific N management decisions. In vivo fluorescence emissions can occur in the wavelength region from 300 to 800 nm and are dependent on the wavelength of illumination.

Lawrence A Corp; James E McMurtrey; Elizabeth M Middleton; Charles L Mulchi; Emmett W Chappelle; Craig S. T Daughtry

2003-01-01

142

Preparation and Characterization of Highly Fluorescent, Glutathione-coated Near Infrared Quantum Dots for in Vivo Fluorescence Imaging  

PubMed Central

Fluorescent probes that emit in the near-infrared (NIR, 700–1,300 nm) region are suitable as optical contrast agents for in vivo fluorescence imaging because of low scattering and absorption of the NIR light in tissues. Recently, NIR quantum dots (QDs) have become a new class of fluorescent materials that can be used for in vivo imaging. Compared with traditional organic fluorescent dyes, QDs have several unique advantages such as size- and composition-tunable emission, high brightness, narrow emission bands, large Stokes shifts, and high resistance to photobleaching. In this paper, we report a facile method for the preparation of highly fluorescent, water-soluble glutathione (GSH)-coated NIR QDs for in vivo imaging. GSH-coated NIR QDs (GSH-QDs) were prepared by surface modification of hydrophobic CdSeTe/CdS (core/shell) QDs. The hydrophobic surface of the CdSeTe/CdS QDs was exchanged with GSH in tetrahydrofuran-water. The resulting GSH-QDs were monodisperse particles and stable in PBS (phosphate buffered saline, pH = 7.4). The GSH-QDs (800 nm emission) were highly fluorescent in aqueous solutions (quantum yield = 22% in PBS buffer), and their hydrodynamic diameter was less than 10 nm, which is comparable to the size of proteins. The cellular uptake and viability for the GSH-QDs were examined using HeLa and HEK 293 cells. When the cells were incubated with aqueous solutions of the GSH-QDs (10 nM), the QDs were taken into the cells and distributed in the perinuclear region of both cells. After 12 hrs incubation of 4 nM of GSH-QDs, the viabilities of HeLa and HEK 293 cells were ca. 80 and 50%, respectively. As a biomedical utility of the GSH-QDs, in vivo NIR-fluorescence imaging of a lymph node in a mouse is presented. PMID:19325735

Jin, Takashi; Fujii, Fumihiko; Komai, Yutaka; Seki, Junji; Seiyama, Akitoshi; Yoshioka, Yoshichika

2008-01-01

143

Fluorescence imaging with near-infrared light: new technological advances that enable in vivo molecular imaging  

Microsoft Academic Search

.   A recent development in biomedical imaging is the non-invasive mapping of molecular events in intact tissues using fluorescence.\\u000a Underpinning to this development is the discovery of bio-compatible, specific fluorescent probes and proteins and the development\\u000a of highly sensitive imaging technologies for in vivo fluorescent detection. Of particular interest are fluorochromes that\\u000a emit in the near infrared (NIR), a spectral

Vasilis Ntziachristos; Christoph Bremer; Ralph Weissleder

2003-01-01

144

Optical fiber fluorescence spectroscopy for detecting AFM1 in milk  

NASA Astrophysics Data System (ADS)

Fluorescence spectroscopy carried out by means of optical fibers was used for the rapid screening of M1 aflatoxin in milk, enabling the detection of concentrations up to the legal limit, which is 50 ppt. A compact fluorometric device equipped with a LED source, a miniaturized spectrometer, and optical fibers for illumination/detection of the measuring micro-cell was tested for measuring threshold values of AFM1 in pre-treated milk samples. Multivariate processing of the spectral data made it possible to obtain a preliminary screening at the earlier stages of the industrial process, as well as to discard contaminated milk stocks before their inclusion in the production chain.

Mignani, A. G.; Cucci, C.; Ciaccheri, L.; Dall'Asta, C.; Galaverna, G.; Dossena, A.; Marchelli, R.

2008-04-01

145

Dynamics of Large Semiflexible Chains Probed by Fluorescence Correlation Spectroscopy  

NASA Astrophysics Data System (ADS)

Fluorescence correlation spectroscopy was used to probe the dynamics of ?-phage DNA in aqueous solution labeled with the randomly intercalating dye TOTO. The linear macromolecules (i) carry more than one chromophore and (ii) are larger than the waist of the focal volume. The correlation function decays significantly faster than expected for a stiff globule of corresponding size but is in good agreement with the dynamic model of semiflexible chains including hydrodynamic interactions. As the chromophore density is lowered the correlation time decreases in accordance with this model.

Lumma, D.; Keller, S.; Vilgis, T.; Rädler, J. O.

2003-05-01

146

Identification of active fluorescence stained bacteria by Raman spectroscopy  

NASA Astrophysics Data System (ADS)

Microorganisms can be found everywhere e.g. in food both as useful ingredients or harmful contaminations causing food spoilage. Therefore, a fast and easy to handle analysis method is needed to detect bacteria in different kinds of samples like meat, juice or air to decide if the sample is contaminated by harmful microorganisms. Conventional identification methods in microbiology require always cultivation and therefore are time consuming. In this contribution we present an analysis approach to identify fluorescence stained bacteria on strain level by means of Raman spectroscopy. The stained bacteria are highlighted and can be localized easier against a complex sample environment e.g. in food. The use of Raman spectroscopy in combination with chemometrical methods allows the identification of single bacteria within minutes.

Krause, Mario; Beyer, Beatrice; Pietsch, Christian; Radt, Benno; Harz, Michaela; Rösch, Petra; Popp, Jürgen

2008-04-01

147

Detectors for single-molecule fluorescence imaging and spectroscopy  

PubMed Central

Single-molecule observation, characterization and manipulation techniques have recently come to the forefront of several research domains spanning chemistry, biology and physics. Due to the exquisite sensitivity, specificity, and unmasking of ensemble averaging, single-molecule fluorescence imaging and spectroscopy have become, in a short period of time, important tools in cell biology, biochemistry and biophysics. These methods led to new ways of thinking about biological processes such as viral infection, receptor diffusion and oligomerization, cellular signaling, protein-protein or protein-nucleic acid interactions, and molecular machines. Such achievements require a combination of several factors to be met, among which detector sensitivity and bandwidth are crucial. We examine here the needed performance of photodetectors used in these types of experiments, the current state of the art for different categories of detectors, and actual and future developments of single-photon counting detectors for single-molecule imaging and spectroscopy. PMID:20157633

MICHALET, X.; SIEGMUND, O.H.W.; VALLERGA, J.V.; JELINSKY, P.; MILLAUD, J.E.; WEISS, S.

2010-01-01

148

Identification of Atherosclerotic Plaques in Carotid Artery by Fluorescence Spectroscopy  

NASA Astrophysics Data System (ADS)

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

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

2008-04-01

149

In Vivo Dendritic Cell Tracking Using Fluorescence Lifetime Imaging and Near-Infrared-Emissive Polymersomes  

Microsoft Academic Search

Purpose  Noninvasive in vivo cell-tracking techniques are necessary to advance the field of cellular-based therapeutics as well as to elucidate mechanisms\\u000a governing in vivo cell biology. Fluorescence is commonly used for in vitro and postmortem biomedical studies but has been limited by autofluorescence at the whole-animal level.\\u000a \\u000a \\u000a \\u000a Procedures  In this report, we demonstrate the ability of in vivo fluorescent lifetime imaging to

Natalie A. Christian; Fabian Benencia; Michael C. Milone; Guizhi Li; Paul R. Frail; Michael J. Therien; George Coukos; Daniel A. Hammer

2009-01-01

150

Anatomy-Based Algorithms for Detecting Oral Cancer Using Reflectance and Fluorescence Spectroscopy  

E-print Network

OBJECTIVES: We used reflectance and fluorescence spectroscopy to noninvasively and quantitatively distinguish benign from dysplastic/malignant oral lesions. We designed diagnostic algorithms to account for differences in ...

McGee, Sasha

151

In Vivo Imaging with Fluorescent Smart Probes to Assess Treatment Strategies for Acute Pancreatitis  

PubMed Central

Background and Aims Endoprotease activation is a key step in acute pancreatitis and early inhibition of these enzymes may protect from organ damage. In vivo models commonly used to evaluate protease inhibitors require animal sacrifice and therefore limit the assessment of dynamic processes. Here, we established a non-invasive fluorescence imaging-based biomarker assay to assess real-time protease inhibition and disease progression in a preclinical model of experimental pancreatitis. Methods Edema development and trypsin activation were imaged in a rat caerulein-injection pancreatitis model. A fluorescent “smart” probe, selectively activated by trypsin, was synthesized by labeling with Cy5.5 of a pegylated poly-L-lysine copolymer. Following injection of the probe, trypsin activation was monitored in the presence or absence of inhibitors by in vivo and ex vivo imaging. Results We established the trypsin-selectivity of the fluorescent probe in vitro using a panel of endopeptidases and specific inhibitor. In vivo, the probe accumulated in the liver and a region attributed to the pancreas by necropsy. A dose dependent decrease of total pancreatic fluorescence signal occurred upon administration of known trypsin inhibitors. The fluorescence-based method was a better predictor of trypsin inhibition than pancreatic to body weight ratio. Conclusions We established a fluorescence imaging assay to access trypsin inhibition in real-time in vivo. This method is more sensitive and dynamic than classic tissue sample readouts and could be applied to preclinically optimize trypsin inhibitors towards intrapancreatic target inhibition. PMID:23409095

Agarwal, Abhiruchi; Boettcher, Andreas; Kneuer, Rainer; Sari-Sarraf, Farid; Donovan, Adriana; Woelcke, Julian; Simic, Oliver; Brandl, Trixi; Krucker, Thomas

2013-01-01

152

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

153

Fluorescence spectroscopy of polynuclear aromatic compounds in environmental monitoring.  

PubMed

The occurrence of polynuclear aromatic compounds (PAC) in the environment and experimental techniques suitable for the detection of PAC in environmental compartments are briefly reviewed. The specific requirements for on-site andin situ environmental analysis are outlined. Particular emphasis is given to fluorescence spectroscopic techniques for the investigation of humic acid- and soil-containing samples. Some examples of studies in the literature on Shpol'skii and jet spectroscopy and on laser-induced fluorescence (OF) measurements of PAC and mineral oils are highlighted. Contaminants in the environment are usually encountered as multicomponent mixtures in very complex matrices. Total fluorescence analysis in combination with the chemometrical technique of rank annihilation factor analysis (RAFA) was employed for the evaluation of a six-component PAC mixture in toluene. It was shown that even in the presence of strong spectral overlap the qualitative identification of all compounds and the reliable quantification of five substances was possible. Results are presented from our stationary and time-resolved fluorescence investigations of the interactions between pyrene and humic acid in water. The Stern-Volmer analysis showed a significant effect of pH on the static quenching efficiency which can be explained by the pH-dependent macromolecular structure of humic acids. Preliminary results from studies of the deactivation of triplet PAC and quenching of delayed fluorescence by humic acid are reported. LIF measurements of mineral oils directly from soil surfaces and of a model oil in a soil column were performed with a fiber-optic coupled multichannel spectrometer. The fluorescence intensity/ concentration relationships were established for a crude and a fuel oil; the corresponding lower limits of detection (LOD) were determined to be 0.025 and 0.125% m/m (mass/mass percentages). These detection limits are compared with realistic oil contaminations of soils. In a soil column designed to mimic fixed-bed bioreactors the distributions of fluorescence signal intensities from a perylene-doped model oil before and after water flooding were determined. These results fromin situ measurements can provide a quantitative basis for the modelling of temporal and spatial contaminants' distributions in reactor design. PMID:24226656

Kumke, M U; Löhmannsröben, H G; Roch, T

1995-06-01

154

Remote excitation fluorescence correlation spectroscopy using silver nanowires  

NASA Astrophysics Data System (ADS)

Fluorescence correlation spectroscopy (FCS), a powerful tool to resolve local properties, dynamical process of molecules, rotational and translational diffusion motions, relies on the fluctuations of florescence observables in the observation volume. In the case of rare transition events or small dynamical fluctuations, FCS requires few molecules or even single molecules in the observation volume at a time to minimize the background signals. Metal nanoparticle which possess unique localized surface plasmon resonance (LSPR) have been used to reduce the observation volume down to sub-diffraction limited scale while maintain at high analyst concentration up to tens of micromolar. Nevertheless, the applications of functionalized nanoparticles in living cell are limited due to the continuous diffusion after cell uptake, which makes it difficult to target the region of interests in the cell. In this work, we demonstrate the use of silver nanowires for remote excitation FCS on fluorescent molecules in solution. By using propagation surface plasmon polaritons (SPPs) which supported by the silver nanowire to excite the fluorescence, both illumination and observation volume can be reduced simultaneously. In such a way, less perturbation is induced to the target region, and this will broaden the application scope of silver nanowire as tip in single cell endoscopy.

Su, Liang; Yuan, Haifeng; Lu, Gang; Hofkens, Johan; Roeffaers, Maarten; Uji-i, Hiroshi

2014-11-01

155

Two-photon excited fluorescence microscopy application for ex vivo investigation of ocular fundus samples  

NASA Astrophysics Data System (ADS)

Two-photon excited fluorescence (TPEF) imaging of ocular tissue has recently become a promising tool in ophthalmology for diagnostic and research purposes. The feasibility and the advantages of TPEF imaging, namely deeper tissue penetration and improved high-resolution imaging of microstructures, have been demonstrated lately using human ocular samples. The autofluorescence properties of endogenous fluorophores in ocular fundus tissue are well known from spectrophotometric analysis. But fluorophores, especially when it comes to fluorescence lifetime, typically display a dependence of their fluorescence properties on local environmental parameters. Hence, a more detailed investigation of ocular fundus autofluorescence ideally in vivo is of utmost interest. The aim of this study is to determine space-resolved the stationary and time-resolved fluorescence properties of endogenous fluorophores in ex vivo porcine ocular fundus samples by means of two-photon excited fluorescence spectrum and lifetime imaging microscopy (FSIM/FLIM). By our first results, we characterized the autofluorescence of individual anatomical structures of porcine retina samples excited at 760 nm. The fluorescence properties of almost all investigated retinal layers are relatively homogenous. But as previously unknown, ganglion cell bodies show a significantly shorter fluorescence lifetime compared to the adjacent mueller cells. Since all retinal layers exhibit bi-exponential autofluorescence decays, we were able to achieve a more precise characterization of fluorescence properties of endogenous fluorophores compared to a present in vivo FLIM approach by confocal scanning laser ophthalmoscope (cSLO).

Peters, Sven; Hammer, Martin; Schweitzer, Dietrich

2011-07-01

156

Trimodal detection of early childhood caries using laser light scanning and fluorescence spectroscopy: clinical prototype  

PubMed Central

Abstract. There is currently a need for a safe and effective way to detect and diagnose early stages of childhood caries. A multimodal optical clinical prototype for diagnosing caries demineralization in vivo has been developed. The device can be used to quickly image and screen for any signs of demineralized enamel by obtaining high-resolution and high-contrast surface images using a 405-nm laser as the illumination source, as well as obtaining autofluorescence and bacterial fluorescence images. When a suspicious region of demineralization is located, the device also performs dual laser fluorescence spectroscopy using 405- and 532-nm laser excitation. An autofluorescence ratio of the two excitation lasers is computed and used to quantitatively diagnose enamel health. The device was tested on five patients in vivo as well as on 28 extracted teeth with clinically diagnosed carious lesions. The device was able to provide detailed images that highlighted the lesions identified by the clinicians. The autofluorescence spectroscopic ratios obtained from the extracted teeth successfully quantitatively discriminated between sound and demineralized enamel. PMID:23986369

Zhang, Liang; Kim, Amy S.; Ridge, Jeremy S.; Nelson, Leonard Y.; Berg, Joel H.; Seibel, Eric J.

2013-01-01

157

Spectral fluorescent properties of tissues in vivo with excitation in the red wavelength range  

NASA Astrophysics Data System (ADS)

The spectral fluorescence analysis is a promising method for differential tissue diagnostic. Usually the UV and visible light is used for fluorescence excitation with emission registration in the visible wavelength range. The light penetration length in this wavelength range is very small allowing one to analyze only the surface region of the tissue. Here we present the tissue fluorescent spectra in vivo excited in the red wavelength region. As excitation light source we used compact He-Ne laser (632.8 nm) and observed the fluorescence in 650 - 800 nm spectral range. The various tissues including normal skin, psoriasis, tumors, necrosis as well as photosensitized tissues have been measured.

Stratonnikov, Alexander A.; Loschenov, Victor B.; Klimov, D. V.; Edinac, N. E.; Wolnukhin, V. A.; Strashkevich, I. A.

1997-12-01

158

Microneedles rollers as a potential device to increase ALA diffusion and PpIX production: evaluations by wide-field fluorescence imaging and fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

One of the limitations of topical photodynamic therapy (PDT) using 5-aminolevulinic acid (ALA) is the poor ability to penetrate biological barriers of skin and the recurrence rates in treatments. This study aimed to identify possible signs of increased diffusion of ALA-induced PpIX by fluorescence images and fluorescence spectroscopy. The research was done using in vivo porcine skin model. Before the cream application, microholes was performed with microneedles rollers in only one direction, afterward the ALA cream was applied at a 2.5cm2 area in triplicate and an occlusive dressing was placed. PpIX production was monitored using fluorescence spectroscopy collected at skin surface after 70, 100, 140, and 180 minutes of ALA incubation. About 100 fluorescence spectra of each treatment were collected, distributed by about five points for each site. Wide-field fluorescence imaging was made after 70, 90, and 170 minutes after treatment. The results obtained by imaging analysis indicated increase of the PpIX diffusion in the skin surface using the microneedles rollers (MNs) before ALA application. Circular regions of red fluorescence around the microholes were observed. In addition, the fluorescence spectra showed a greater intensity (2 times as many) in groups microneedles rollers associated. In conclusion, our data shown greater homogeneity and PpIX production in the groups pre-treated with microneedles indicating that the technique can be used to greater uniformity of PpIX production throughout the area to be treated reducing the chances of recurrent tumor as well as has potential for decreasing the time of therapy. (FUNDING SUPPORT:CAPES, CNPq and FAPESP)

Gracielli Sousa, R. Phamilla; de Menezes, Priscila F. C.; Fujita, Alessandra K. L.; Requena, Michelle B.; Govone, Angelo Biassi; Escobar, André; de Nardi, Andrigo B.; Kurachi, Cristina; Bagnato, Vanderlei Salvador

2014-03-01

159

In vivo transfection of testicular germ cells and transgenesis by using the mitochondrially localized jellyfish fluorescent protein gene  

Microsoft Academic Search

We aimed to introduce foreign DNA into spermatogenic cells in the testis by injection of the DNA encoding jellyfish fluorescent proteins, green fluorescent protein (GFP) and yellow fluorescent protein (YFP) into the seminiferous tubules and in vivo electroporation. We obtained fluorescent spermatozoa only when using the gene of the YFP protein fused to a mitochondrial localization signal peptide. Intracytoplasmic injection

Zhenyong Huang; Masaru Tamura; Takayuki Sakurai; Shinichiro Chuma; Tetsuichiro Saito; Norio Nakatsuji

2000-01-01

160

Imaging tumor metabolism using in vivo magnetic resonance spectroscopy.  

PubMed

Magnetic resonance spectroscopy (MRS) is a powerful tool for noninvasively investigating normal and abnormal metabolism. When used in combination with imaging strategies, multinuclear MRS methods provide detailed biochemical information that can be directly correlated with anatomical features. Hyperpolarized C MRS is a new technology that reflects real-time metabolic conversion and is likely to be extremely valuable in managing patients with cancer. This article reviews the use of in vivo P, H, and C MRS for assessing cancer metabolism in order to provide information for diagnosis, planning treatment, assessing response to therapy, and predicting survival for patients with cancer. PMID:25815853

Li, Yan; Park, Ilwoo; Nelson, Sarah J

2015-01-01

161

In vivo near-infrared fluorescence three-dimensional positioning system with binocular stereovision  

NASA Astrophysics Data System (ADS)

Fluorescence is a powerful tool for in-vivo imaging in living animals. The traditional in-vivo fluorescence imaging equipment is based on single-view two-dimensional imaging systems. However, they cannot meet the needs for accurate positioning during modern scientific research. A near-infrared in-vivo fluorescence imaging system is demonstrated, which has the capability of deep source signal detecting and three-dimensional positioning. A three-dimensional coordinates computing (TDCP) method including a preprocess algorithm is presented based on binocular stereo vision theory, to figure out the solution for diffusive nature of light in tissue and the emission spectra overlap of fluorescent labels. This algorithm is validated to be efficient to extract targets from multispectral images and determine the spot center of biological interests. Further data analysis indicates that this TDCP method could be used in three-dimensional positioning of the fluorescent target in small animals. The study also suggests that the combination of a large power laser and deep cooling charge-coupled device will provide an attractive approach for fluorescent detection from deep sources. This work demonstrates the potential of binocular stereo vision theory for three-dimensional positioning for living animal in-vivo imaging.

Song, Bofan; Jin, Wei; Wang, Ying; Jin, Qinhan; Mu, Ying

2014-11-01

162

Frequently asked questions about in vivo chlorophyll fluorescence: practical issues.  

PubMed

The aim of this educational review is to provide practical information on the hardware, methodology, and the hands on application of chlorophyll (Chl) a fluorescence technology. We present the paper in a question and answer format like frequently asked questions. Although nearly all information on the application of Chl a fluorescence can be found in the literature, it is not always easily accessible. This paper is primarily aimed at scientists who have some experience with the application of Chl a fluorescence but are still in the process of discovering what it all means and how it can be used. Topics discussed are (among other things) the kind of information that can be obtained using different fluorescence techniques, the interpretation of Chl a fluorescence signals, specific applications of these techniques, and practical advice on different subjects, such as on the length of dark adaptation before measurement of the Chl a fluorescence transient. The paper also provides the physiological background for some of the applied procedures. It also serves as a source of reference for experienced scientists. PMID:25119687

Kalaji, Hazem M; Schansker, Gert; Ladle, Richard J; Goltsev, Vasilij; Bosa, Karolina; Allakhverdiev, Suleyman I; Brestic, Marian; Bussotti, Filippo; Calatayud, Angeles; D?browski, Piotr; Elsheery, Nabil I; Ferroni, Lorenzo; Guidi, Lucia; Hogewoning, Sander W; Jajoo, Anjana; Misra, Amarendra N; Nebauer, Sergio G; Pancaldi, Simonetta; Penella, Consuelo; Poli, DorothyBelle; Pollastrini, Martina; Romanowska-Duda, Zdzislawa B; Rutkowska, Beata; Serôdio, João; Suresh, Kancherla; Szulc, Wies?aw; Tambussi, Eduardo; Yanniccari, Marcos; Zivcak, Marek

2014-11-01

163

Longitudinal in vivo two-photon fluorescence imaging  

PubMed Central

Fluorescence microscopy is an essential technique for the basic sciences, especially biomedical research. Since the invention of laser scanning confocal microscopy in 1980s, that enabled imaging both fixed and living biological tissue with three-dimensional precision, high-resolution fluorescence imaging has revolutionized biological research. Confocal microscopy, by its very nature, has one fundamental limitation. Due to the confocal pinhole, deep tissue fluorescence imaging is not practical. In contrast (no pun intended), two-photon fluorescence microscopy allows, in principle, the collection of all emitted photons from fluorophores in the imaged voxel, dramatically extending our ability to see deep into living tissue. Since the development of transgenic mice with genetically encoded fluorescent protein in neocortical cells in 2000, two-photon imaging has enabled the dynamics of individual synapses to be followed for up to two years. Since the initial landmark contributions to this field in 2002, the technique has been used to understand how neuronal structure are changed by experience, learning and memory and various diseases. Here we provide a basic summary of the crucial elements that are required for such studies, and discuss many applications of longitudinal two-photon fluorescence microscopy that have appeared since 2002. PMID:24214350

Crowe, Sarah E.; Ellis-Davies, Graham C.R.

2014-01-01

164

Autofluorescence spectroscopy of colorectal carcinoma: ex vivo study  

NASA Astrophysics Data System (ADS)

Diagnosis established by means of fluorescence spectroscopy is currently used in the field of urology and bronchology. Its major advantage is that it allows the diagnosis of epithelial dysplasia or malignant proliferation even if routine diagnostic endoscopy fails to reveal any macroscopic changes. The authors present results of their observations that deal with fluorescence diagnosis of colorectal carcinoma. They examined the wet microscopic mounts of healthy colon mucosa and compared them to that prepared from colon mucosa affected by adenocarcinoma. The diagnosis of adenocarcinoma was verified by using clinical and histology means. Fluorescence spectra of tissue samples, excited by means of 488 and 514.5 nm lines of Ar ion laser and/or by He-Ne laser line 632.8 nm, have been studied. This study demonstrated differences in both the spectral shape and in the signal intensity (at unchanged spectral shape) of photoluminescence spectra emitted from tissue affected by adenocarcinoma as compared to that of healthy colon mucosa. The results encourage us to continue the study aimed at development of the diagnostic system usable in the clinical practice.

Horak, Ladislav; Svec, Alexandr; Lezal, Dimitrij; Zavadil, Jiri

2003-10-01

165

Photoresponsive fluorescent reduced graphene oxide by spiropyran conjugated hyaluronic acid for in vivo imaging and target delivery.  

PubMed

This present article demonstrates the strategy to prepare photoresponsive reduced graphene oxide with mussel inspired adhesive material dopamine (DN) and photochromic dye spiropyran (SP) conjugated to the backbone of the targeting ligand hyaluronic acid (HA; HA-SP). Graphene oxide (GO) was reduced by prepared HA-SP accepting the advantages of catechol chemistry under mildly alkaline condition enabling to achieve functionalized graphene (rGO/HA-SP) as fluorescent nanoparticles. Due to containing HA, rGO/HA-SP can bind to the CD44 cell receptors. The prepared rGO/HA-SP is able to retain its photochromic features and can be converted to merocyanine (MC) form upon irradiation with UV light (wavelength: 365 nm) displaying purple color. Photochromic behavior of rGO/HA-SP was monitored by UV-vis and fluorescence spectroscopy. In vitro fluorescence behavior, examined by confocal laser scanning microscope (CLSM), of rGO/HA-SP in cancerous A549 cell lines assured that efficient delivery of rGO/HA-SP was gained due to HA as targeting ligand. In this work, we have shown that in vivo fluorescence image of spiropyran is possible by administrating MC form solution of rGO/HA-SP using Balb/C mice as in vivo modal. Accumulation of rGO/HA-SP in tumor tissue from biodistribution analysis strongly supports the specific delivery of prepared graphene to the target destination. The well tuned drug release manner from the surface of rGO/HA-SP strongly recommends the developed material not only as fluorescent probe for diagnosis but also as a drug carrier in drug delivery system. PMID:24106989

Nahain, Abdullah-Al; Lee, Jung-Eun; Jeong, Ji Hoon; Park, Sung Young

2013-11-11

166

Real time monitoring of superoxide dynamics in vivo through fluorescent proteins using a sensitive fiber probe  

NASA Astrophysics Data System (ADS)

Superoxide anion is the primary oxygen free radical generated in mitochondria that causes intracellular oxidative stress. The lack of a method to directly monitor superoxide concentration in vivo in real time has severely hindered our understanding on its pathophysiology. We made transgenic zebrafish to specifically express fluorescent proteins, which are recently developed as reversible superoxide-specific indicators, in the liver. A fiber-optic fluorescent probe was used to noninvasively monitor superoxide generation in the liver in real time. The fish were placed in microfluidic channels for manipulation and reagents administration. Several superoxide-inducing and scavenging reagents were administrated onto the fish to investigate their effects on superoxide anion balancing. The biochemical dynamics of superoxide due to the application reagents were revealed in the transient behaviors of fluorescence time courses. With the ability to monitor superoxide dynamics in vivo in real time, this method can be used as an in vivo pharmaceutical screening platform.

Chang, Yu-Chung; Ken, Chuian-Fu; Hsu, Che-Wei; Liu, Ya-Ging

2014-03-01

167

Fluorescence and UV-vis Spectroscopy of Synovial Fluids  

NASA Astrophysics Data System (ADS)

Total joint arthroplasty involves replacing the worn cartilaginous surfaces of the joint with man-made materials that are designed to be biocompatible and to withstand mechanical stresses. Commonly these bearing materials consist of metallic alloys (TiAlV or CoCrMo) and UHMWPE. Following joint arthroplasty, the normal generation of micro-metallic wear debris particles that dislodge from the prosthesis has been shown to cause inflammatory aseptic osteolysis (bone loss) that ultimately results in the failure of the implant. Here we report our results on the novel use of Fluorescence and UV-vis spectroscopy to investigate the metallic content of synovial fluid specimens taken from postoperative total knee arthroplasties. Preliminary finding showed presence of alumina and chromium is some specimens. The ability to detect and monitor the wear rate of these implants could have far reaching implications in the prevention of metallic wear-debris induced osteolysis and impending implant failure.

Pinti, Marie J.; Stojilovic, Nenad; Kovacik, Mark W.

2009-10-01

168

Fluorescence spectroscopy, exciton dynamics, and photochemistry of single allophycocyanin trimers  

SciTech Connect

The authors report a study of the allophycocyanin trimer (APC), a light-harvesting protein complex from cyanobacteria, by room-temperature single-molecule measurements of fluorescence spectra, lifetimes, intensity trajectories, and polarization modulation. Emission spectra of individual APC trimers are found to be homogeneous on the time scale of seconds. In contrast, their emission lifetimes are found to be widely distributed because of generation of long-lived exciton traps during the course of measurements. The intensity trajectories and polarization modulation experiments indicate reversible exciton trap formation within the three quasi-independent pairs of strong interacting {alpha}84 and {beta}84 chromophores in APC, as well as photobleaching of individual chromophores. Comparison experiments under continuous-wave and pulsed excitation reveal a two-photon mechanism for generating exciton traps and/or photobleaching, which involves exciton-exciton annihilation. These single-molecule experiments provide new insights into the spectroscopy, exciton dynamics, and photochemistry of light-harvesting complexes.

Ying, L.; Sie, X.S. [Pacific Northwest National Lab., Richland, WA (United States). William R. Wiley Environmental Molecular Sciences Lab.] [Pacific Northwest National Lab., Richland, WA (United States). William R. Wiley Environmental Molecular Sciences Lab.

1998-12-10

169

Fluorescence Spectroscopy, Exciton Dynamics and Photochemistry of Single Allophycocyanin Trimers  

SciTech Connect

We report a study of the spectroscopy and exciton dynamics of the allophycocyanin trimer (APC), a light harvesting protein complex from cyanobacteria, by room-temperature single-molecule measurements of fluorescence spectra, lifetimes, intensity trajectories and polarization modulation. Emission spectra of individual APC trimers are found to be homogeneous on the time scale of seconds. In contrast, their emission lifetimes are found to be widely distributed, because of generation of exciton traps during the course of measurements. The intensity trajectories and polarization modulation experiments indicate reversible ixciton trap formation within the three quasi-independent pairs of strong interacting a84 and B84 chromophores in APC, as well a photobleaching of individual chromophores. Comparison experiments under continuous wave and pulsed excitation reveal a two-photon mechanism for generating exciton traps and/or photobleaching, which involves exciton-exciton annihilation. These single-molecule experiments provide new insights into exciton dynamics and photochemistry of light-harvesting complexes.

Ying, Liming (ASSOC WESTERN UNIVERSITY) [ASSOC WESTERN UNIVERSITY; Xie, Xiaoliang (HARVARD UNIVERSITY) [HARVARD UNIVERSITY

1998-01-01

170

Polymers and single molecule fluorescence spectroscopy, what can we learn?  

PubMed

This tutorial review summarizes the most important results and developments in the field of polymer science by means of single molecule fluorescence spectroscopy (SMFS) at ambient temperatures. A broad range of topics will be addressed and it will be discussed which single molecule methods are suitable to get the maximum amount of information about polymer structure, polymer dynamics and the photophysics of incorporated or embedded dye molecules. In particular, we will report on the use of polymer films for immobilization of molecules, the visualization of dynamics near the glass transition temperature Tg, the reptation of polymer chains, the conformation adopted by polymer chains and the in situ observation of the polymerization reaction itself. PMID:19169450

Wöll, Dominik; Braeken, Els; Deres, Ania; De Schryver, Frans C; Uji-i, Hiroshi; Hofkens, Johan

2009-02-01

171

Profiling the near field of nanoshells using surface enhanced Raman spectroscopy and fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Plasmon resonances in metal nanoparticles control the far field and near field optical properties of these metallic structures. The enhanced electromagnetic near field is strongest at the surface of the nanoparticles and rapidly decays away from the surface. This enhanced near field is exploited in surface enhanced spectroscopies including Surface Enhanced Raman Spectroscopy (SERS) and Metal Enhanced Fluorescence Spectroscopy (MEFS). A measurement of the decay profile of the fringing field is important both for further development of surface enhanced spectroscopy for sensor device application, and for understanding from a fundamental physics point of view. Gold nanoshells are spherical colloidal nanoparticles with a silica core covered by a thin gold shell. The plasmon resonance of nanoshells can be controllably tuned in the visible and infrared parts of the spectrum. The near field profile of nanoshells can be theoretically calculated on the basis of Mie scattering theory. The thesis describes a series of experiments designed to experimentally verify the near field profile of nanoshells. A scaffold of ss-DNA is used to place a fluorescein dye molecule at varying distances from the nanoshell surface. The SERS intensity from both the scaffold molecules and the fluorescein placed at the end of the tether is measured simultaneously and self consistently. The fluorescein-ss-DNA nanoshell conjugate structures are also used to study the distance dependence of the fluorescence emission from fluorescein. The thesis discusses the results of the SERS intensity profile agreement with the intensity profile calculated using Mie scattering theory. The quenching and enhancement of the fluorescence emission at varying distances from the nanoshell surface are also discussed.

Lal, Surbhi

172

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

173

Fluorescence correlation spectroscopy in biology, chemistry, and medicine.  

PubMed

This review describes the method of fluorescence correlation spectroscopy (FCS) and its applications. FCS is used for investigating processes associated with changes in the mobility of molecules and complexes and allows researchers to study aggregation of particles, binding of fluorescent molecules with supramolecular complexes, lipid vesicles, etc. The size of objects under study varies from a few angstroms for dye molecules to hundreds of nanometers for nanoparticles. The described applications of FCS comprise various fields from simple chemical systems of solution/micelle to sophisticated regulations on the level of living cells. Both the methodical bases and the theoretical principles of FCS are simple and available. The present review is concentrated preferentially on FCS applications for studies on artificial and natural membranes. At present, in contrast to the related approach of dynamic light scattering, FCS is poorly known in Russia, although it is widely employed in laboratories of other countries. The goal of this review is to promote the development of FCS in Russia so that this technique could occupy the position it deserves in modern Russian science. PMID:21639831

Perevoshchikova, I V; Kotova, E A; Antonenko, Y N

2011-05-01

174

Single molecule fluorescence correlation spectroscopy of single apoptotic cells using a red-fluorescent caspase probe.  

PubMed

The detection of single molecules in single cells has enabled biochemical analyses to be conducted with high sensitivity and high temporal resolution. In this work, detection of apoptosis was studied by single molecule fluorescence correlation spectroscopy (FCS) in single living cells. Caspase activity was assayed using a new red fluorogenic probe that avoids the spectral overlap of green fluorescent probes and cell autofluorescence. This new probe, 2SBPO-Casp, was synthesized by coupling a water-soluble Nile Blue derivative (2SBPO) to an aspartic acid residue. Upon apoptosis induction and caspase activation, free 2SBPO dye is shown to accumulate inside the cell after probe cleavage. In previous work in our lab, single molecule fluorescence in single apoptotic cells was detected 45 min after induction using a rhodamine 110-based probe. However, significant statistical analysis was needed to exclude false positives. The use of 2SBPO-Casp overcomes the autofluorescence problem and offers a steady fluorescence signal. In our single molecule FCS measurements, Ramos cells were determined apoptotic on the basis of their correlation coefficient value (R(2)). Cells that contain an R(2) ? 0.65 were identified as highly correlated and therefore determined to be apoptotic. Single apoptotic cells identified in this manner were found as early as 30 min after induction and the number of apoptotic cells reached a peak value at the 3rd hour, which is consistent with other techniques. Using single molecule techniques and a new apoptosis probe, the temporal dynamics were elucidated with better sensitivity and resolution than in previous studies. PMID:22314869

Dong, Meicong; Martinez, Michelle M; Mayer, Michael F; Pappas, Dimitri

2012-07-01

175

Inflammation Modulates Murine Venous Thrombosis Resolution In Vivo: Assessment by Multimodal Fluorescence Molecular Imaging  

PubMed Central

Objective Assessment of thrombus inflammation in vivo could provide new insights into deep vein thrombosis (DVT) resolution. Here we develop and evaluate two integrated fluorescence molecular-structural imaging strategies to quantify DVT-related inflammation and architecture, and to assess the effect of thrombus inflammation on subsequent DVT resolution in vivo. Methods and Results Murine DVT were created with topical 5% FeCl3 application to thigh or jugular veins (n=35). On day 3, mice received macrophage and matrix metalloproteinase (MMP) activity fluorescence imaging agents. On day 4, integrated assessment of DVT inflammation and architecture was performed using confocal fluorescence intravital microscopy (IVM). Day 4 analyses showed robust relationships among in vivo thrombus macrophages, MMP activity, and FITC-dextran deposition (r>0.70, p<0.01). In a serial two-timepoint study, mice with DVT underwent IVM at day 4 and at day 6. Analyses revealed that the intensity of thrombus inflammation at day 4 predicted the magnitude of DVT resolution at day 6 (p<0.05). In a second approach, noninvasive fluorescence molecular tomography-computed tomography (FMT-CT) was employed, and detected macrophages within jugular DVT (p<0.05 vs. sham-controls). Conclusions Integrated fluorescence molecular-structural imaging demonstrates that the DVT-induced inflammatory response can be readily assessed in vivo, and can inform the magnitude of thrombus resolution. PMID:22995524

Ripplinger, Crystal M.; Kessinger, Chase W.; Li, Chunqiang; Kim, Jin Won; McCarthy, Jason R.; Weissleder, Ralph; Henke, Peter K.; Lin, Charles P.; Jaffer, Farouc A.

2012-01-01

176

Lipidots: competitive organic alternative to quantum dots for in vivo fluorescence imaging  

NASA Astrophysics Data System (ADS)

The use of fluorescent nanostructures can bring several benefits on the signal to background ratio for in vitro microscopy, in vivo small animal imaging, and image-guided surgery. Fluorescent quantum dots (QDs) display outstanding optical properties, with high brightness and low photobleaching rate. However, because of their toxic element core composition and their potential long term retention in reticulo-endothelial organs such as liver, their in vivo human applications seem compromised. The development of new dye-loaded (DiO, DiI, DiD, DiR, and Indocyanine Green (ICG)) lipid nanoparticles for fluorescence imaging (lipidots) is described here. Lipidot optical properties quantitatively compete with those of commercial QDs (QTracker®705). Multichannel in vivo imaging of lymph nodes in mice is demonstrated for doses as low as 2 pmols of particles. Along with their optical properties, fluorescent lipidots display very low cytotoxicity (IC50 > 75 nM), which make them suitable tools for in vitro, and especially in vivo, fluorescence imaging applications.

Gravier, Julien; Navarro, Fabrice P.; Delmas, Thomas; Mittler, Frédérique; Couffin, Anne-Claude; Vinet, Françoise; Texier, Isabelle

2011-09-01

177

In vivo imaging of cerebral energy metabolism with two-photon fluorescence lifetime microscopy of NADH  

PubMed Central

Minimally invasive, specific measurement of cellular energy metabolism is crucial for understanding cerebral pathophysiology. Here, we present high-resolution, in vivo observations of autofluorescence lifetime as a biomarker of cerebral energy metabolism in exposed rat cortices. We describe a customized two-photon imaging system with time correlated single photon counting detection and specialized software for modeling multiple-component fits of fluorescence decay and monitoring their transient behaviors. In vivo cerebral NADH fluorescence suggests the presence of four distinct components, which respond differently to brief periods of anoxia and likely indicate different enzymatic formulations. Individual components show potential as indicators of specific molecular pathways involved in oxidative metabolism. PMID:23412419

Yaseen, Mohammad A.; Sakadži?, Sava; Wu, Weicheng; Becker, Wolfgang; Kasischke, Karl A.; Boas, David A.

2013-01-01

178

In vivo imaging of cerebral energy metabolism with two-photon fluorescence lifetime microscopy of NADH.  

PubMed

Minimally invasive, specific measurement of cellular energy metabolism is crucial for understanding cerebral pathophysiology. Here, we present high-resolution, in vivo observations of autofluorescence lifetime as a biomarker of cerebral energy metabolism in exposed rat cortices. We describe a customized two-photon imaging system with time correlated single photon counting detection and specialized software for modeling multiple-component fits of fluorescence decay and monitoring their transient behaviors. In vivo cerebral NADH fluorescence suggests the presence of four distinct components, which respond differently to brief periods of anoxia and likely indicate different enzymatic formulations. Individual components show potential as indicators of specific molecular pathways involved in oxidative metabolism. PMID:23412419

Yaseen, Mohammad A; Sakadži?, Sava; Wu, Weicheng; Becker, Wolfgang; Kasischke, Karl A; Boas, David A

2013-02-01

179

In vivo two-dimensional NMR correlation spectroscopy  

NASA Astrophysics Data System (ADS)

The poor resolution of in-vivo one- dimensional nuclear magnetic resonance spectroscopy (NMR) has limited its clinical potential. Currently, only the large singlet methyl resonances arising from N-acetyl aspartate (NAA), choline, and creatine are quantitated in a clinical setting. Other metabolites such as myo- inositol, glutamine, glutamate, lactate, and ?- amino butyric acid (GABA) are of clinical interest but quantitation is difficult due to the overlapping resonances and limited spectral resolution. To improve the spectral resolution and distinguish between overlapping resonances, a series of two- dimensional chemical shift correlation spectroscopy experiments were developed for a 1.5 Tesla clinical imaging magnet. Two-dimensional methods are attractive for in vivo spectroscopy due to their ability to unravel overlapping resonances with the second dimension, simplifying the interpretation and quantitation of low field NMR spectra. Two-dimensional experiments acquired with mix-mode line shape negate the advantages of the second dimension. For this reason, a new experiment, REVOLT, was developed to achieve absorptive mode line shape in both dimensions. Absorptive mode experiments were compared to mixed mode experiments with respect to sensitivity, resolution, and water suppression. Detailed theoretical and experimental calculations of the optimum spin lock and radio frequency power deposition were performed. Two-dimensional spectra were acquired from human bone marrow and human brain tissue. The human brain tissue spectra clearly reveal correlations among the coupled spins of NAA, glutamine, glutamate, lactate, GABA, aspartate and myo-inositol obtained from a single experiment of 23 minutes from a volume of 59 mL. (Copies available exclusively from MIT Libraries, Rm. 14-0551, Cambridge, MA 02139-4307. Ph. 617-253-5668; Fax 617-253-1690.)

Kraft, Robert A.

1999-10-01

180

Spectral features selection and classification for bimodal optical spectroscopy applied to bladder cancer in vivo diagnosis.  

PubMed

This paper describes an experimental study combining spatially resolved autofluorescence (AF) and diffuse reflectance (DR) fibred spectroscopies to discriminate in vivo between healthy and pathological tissues in a preclinical model of bladder cancer. Then, a detailed step-by-step analysis scheme is presented for the extraction and the selection of discriminative spectral features (correlation, linear discriminant, and logistic regression analysis), and for the spectroscopic data final classification algorithms (regularized discriminant analysis and support vector machines). Significant differences between healthy, inflammatory, and tumoral tissues were obtained by selecting a reasonable number of discriminant spectral features from AF, DR, and intrinsic fluorescence spectra, leading to improved sensitivity (87%) and specificity (77%) compared to monomodality (AF or DR alone). PMID:21216703

Péry, Emilie; Blondel, Walter C P M; Tindel, Samy; Ghribi, Maha; Leroux, Agnès; Guillemin, François

2014-01-01

181

Noninvasive imaging in vivo with fluorescent proteins from centimeters to micrometers  

NASA Astrophysics Data System (ADS)

Whole-body imaging with fluorescent proteins has been shown to be a powerful technology with many applications in small animals. Our laboratory pioneered in vivo imaging with fluorescent proteins (1) including noninvasive whole-body imaging (2). Whole-body imaging with fluorescent proteins depends in large part on the brightness of the protein. Brighter, red-shifted proteins can make whole-body imaging more sensitive due to reduced absorption by tissues and less scatter. Non-invasive imaging with fluorescent proteins has been shown to be able to quantitatively track tumor growth and metastasis, gene expression, angiogenesis, and bacterial infection (3) even at subcellular resolution depending on the position of the cells in the animal. Interference by skin autofluorescence is kept to a minimum with the use of proper filters. To noninvasively image cancer cell/stromal cell interaction in the tumor microenvironment and drug response at the cellular level in live animals in real time, we developed a new imageable three-color animal model. The model consists of green fluorescent protein (GFP)-expressing mice transplanted with dual-color cancer cells labeled with GFP in the nucleus and red fluorescent protein (RFP) in the cytoplasm. Various in vivo phenomena of tumor-host interaction and cellular dynamics were imaged, including mitotic and apoptotic tumor cells, stromal cells interacting with the tumor cells, tumor vasculature, and tumor blood flow as well as drug response. This imageable technology should lead to many new insights of in vivo cancer cell biology.

Yang, Meng; Jiang, Ping; Al-Zaid, Manal; Hoffman, Robert M.

2008-02-01

182

In Vivo and Ex Vivo Transcutaneous Glucose Detection Using Surface-Enhanced Raman Spectroscopy  

NASA Astrophysics Data System (ADS)

Diabetes mellitus is widely acknowledged as a large and growing health concern. The lack of practical methods for continuously monitoring glucose levels causes significant difficulties in successful diabetes management. Extensive validation work has been carried out using surface-enhanced Raman spectroscopy (SERS) for in vivo glucose sensing. This dissertation details progress made towards a Raman-based glucose sensor for in vivo, transcutaneous glucose detection. The first presented study combines spatially offset Raman spectroscopy (SORS) with SERS (SESORS) to explore the possibility of in vivo, transcutaneous glucose sensing. A SERS-based glucose sensor was implanted subcutaneously in Sprague-Dawley rats. SERS spectra were acquired transcutaneously and analyzed using partial least-squares (PLS). Highly accurate and consistent results were obtained, especially in the hypoglycemic range. Additionally, the sensor demonstrated functionality at least17 days after implantation. A subsequent study further extends the application of SESORS to the possibility of in vivo detection of glucose in brain through skull. Specifically, SERS nanoantennas were buried in an ovine tissue behind a bone with 8 mm thickness and detected by using SESORS. In addition, quantitative detection through bones by using SESORS was also demonstrated. A device that could measure glucose continuously as well as noninvasively would be of great use to patients with diabetes. The inherent limitation of the SESORS approach may prevent this technique from becoming a noninvasive method. Therefore, the prospect of using normal Raman spectroscopy for glucose detection was re-examined. Quantitative detection of glucose and lactate in the clinically relevant range was demonstrated by using normal Raman spectroscopy with low power and short acquisition time. Finally, a nonlinear calibration method called least-squares support vector machine regression (LS-SVR) was investigated for analyzing spectroscopic data sets of glucose detection. Comparison studies were demonstrated between LS-SVR and PLS. LS-SVR demonstrated significant improvements in accuracy over PLS for glucose detection, especially when a global calibration model was required. The improvements imparted by LS-SVR open up the possibility of developing an accurate prediction algorithm for Raman-based glucose sensing applicable to a large human population. Overall, these studies show the high promise held by the Raman-based sensor for the challenge of optimal glycemic control.

Ma, Ke

183

Fluorescence spectroscopy of atmospherically relevant bacterial and fungal spores and potential interferences  

NASA Astrophysics Data System (ADS)

Single-particle fluorescence spectroscopy was used to study fluorescence properties of fungal spores and bacteria, selected for their possible atmospheric relevance. In addition, aromatic organic acid aerosols, potentially interfering with laser induced fluorescence measurement, were studied. The results indicate that fungal spores and bacteria have dissimilar fluorescence spectra. The tested aromatic organic acids had fluorescence properties rather similar to common biological molecules. It may be possible to classify atmospheric bacterial and fungal spores through the dissimilar fluorescence properties, but the influence of the potential interferences must be taken into account.

Saari, S. E.; Putkiranta, M. J.; Keskinen, J.

2013-06-01

184

Biocompatible fluorescent nanoparticles for in vivo stem cell tracking  

NASA Astrophysics Data System (ADS)

Efficient application of stem cells to the treatment of neurodegenerative diseases requires safe cell tracking to follow stem cell fate over time in the host environment after transplantation. In this work, for the first time, fluorescent and biocompatible methyl methacrylate (MMA)-based nanoparticles (fluoNPs) were synthesized through a free-radical co-polymerization process with a fluorescent macromonomer obtained by linking Rhodamine B and hydroxyethyl methacrylate. We demonstrate that the fluoNPs produced by polymerization of MMA-Rhodamine complexes (1) were efficient for the labeling and tracking of multipotent human amniotic fluid cells (hAFCs); (2) did not alter the main biological features of hAFCs (such as viability, cell growth and metabolic activity); (3) enabled us to determine the longitudinal bio-distribution of hAFCs in different brain areas after graft in the brain ventricles of healthy mice by a direct fluorescence-based technique. The reliability of our approach was furthermore confirmed by magnetic resonance imaging analyses, carried out by incubating hAFCs with both superparamagnetic iron oxide nanoparticles and fluoNPs. Our data suggest that these finely tunable and biocompatible fluoNPs can be exploited for the longitudinal tracking of stem cells.

Cova, Lidia; Bigini, Paolo; Diana, Valentina; Sitia, Leopoldo; Ferrari, Raffaele; Pesce, Ruggiero Maria; Khalaf, Rushd; Bossolasco, Patrizia; Ubezio, Paolo; Lupi, Monica; Tortarolo, Massimo; Colombo, Laura; Giardino, Daniela; Silani, Vincenzo; Morbidelli, Massimo; Salmona, Mario; Moscatelli, Davide

2013-06-01

185

Fluorescence Spectroscopy for Characterization and Differentiation of Beers  

Microsoft Academic Search

J. Inst. Brew. 110(4), 267-275, 2004 Total luminescence and synchronous scanning fluorescence spectroscopic techniques were applied for characterization of the intrinsic fluorescence of eight different beers. Spectra were mea- sured using different geometries to reveal the presence of similar fluorescent components. The total luminescence and synchro- nous fluorescence spectra exhibit a relatively intense short- wavelength emission ascribed to aromatic amino

Ewa Sikorska; Tomasz Górecki; Igor V. Khmelinskii; Marek Sikorski; Denis De Keukeleire

186

Intradermal Indocyanine Green for In Vivo Fluorescence Laser Scanning Microscopy of Human Skin: A Pilot Study  

PubMed Central

Background In clinical diagnostics, as well as in routine dermatology, the increased need for non-invasive diagnosis is currently satisfied by reflectance laser scanning microscopy. However, this technique has some limitations as it relies solely on differences in the reflection properties of epidermal and dermal structures. To date, the superior method of fluorescence laser scanning microscopy is not generally applied in dermatology and predominantly restricted to fluorescein as fluorescent tracer, which has a number of limitations. Therefore, we searched for an alternative fluorophore matching a novel skin imaging device to advance this promising diagnostic approach. Methodology/Principal Findings Using a Vivascope®-1500 Multilaser microscope, we found that the fluorophore Indocyanine-Green (ICG) is well suited as a fluorescent marker for skin imaging in vivo after intradermal injection. ICG is one of few fluorescent dyes approved for use in humans. Its fluorescence properties are compatible with the application of a near-infrared laser, which penetrates deeper into the tissue than the standard 488 nm laser for fluorescein. ICG-fluorescence turned out to be much more stable than fluorescein in vivo, persisting for more than 48 hours without significant photobleaching whereas fluorescein fades within 2 hours. The well-defined intercellular staining pattern of ICG allows automated cell-recognition algorithms, which we accomplished with the free software CellProfiler, providing the possibility of quantitative high-content imaging. Furthermore, we demonstrate the superiority of ICG-based fluorescence microscopy for selected skin pathologies, including dermal nevi, irritant contact dermatitis and necrotic skin. Conclusions/Significance Our results introduce a novel in vivo skin imaging technique using ICG, which delivers a stable intercellular fluorescence signal ideal for morphological assessment down to sub-cellular detail. The application of ICG in combination with the near infrared laser opens new ways for minimal-invasive diagnosis and monitoring of skin disorders. PMID:21904601

Jonak, Constanze; Skvara, Hans; Kunstfeld, Rainer; Trautinger, Franz; Schmid, Johannes A.

2011-01-01

187

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

188

Cell labeling approaches for fluorescence-based in vivo flow cytometry*  

PubMed Central

We provide an overview of the methods used to label circulating cells for fluorescence detection by in vivo flow cytometry. These methods are useful for cell tracking in small animals without the need to draw blood samples, and are particularly useful for the detection of circulating cancer cells and quantification of circulating immune cells. PMID:21905206

Pitsillides, Costas M.; Runnels, Judith M.; Zhi, Liang; Wu, Meixiong; Lin, Charles P.

2011-01-01

189

In vivo fluorescence lifetime detection of an activatable probe in infarcted myocardium  

NASA Astrophysics Data System (ADS)

Activatable fluorescent molecular probes are predominantly nonfluorescent in their inactivated state due to intramolecular quenching, but increase fluorescence yield significantly after enzyme-mediated hydrolysis of peptides. Continuous wave in vivo detection of these protease-activatable fluorophores in the heart, however, is limited by the inability to differentiate between activated and nonactivated fractions of the probe and is frequently complicated by large background signal from probe accumulation in the liver. Using a cathepsin-activatable near-infrared probe (PGC-800), we demonstrate here that fluorescence lifetime (FL) significantly increases in infarcted murine myocardial tissue (0.67 ns) when compared with healthy myocardium (0.59 ns) after 24 h. Furthermore, we show that lifetime contrast can be used to distinguish in vivo cardiac fluorescence from background nonspecific liver signal. The results of this study show that lifetime contrast is a helpful addition to preclinical imaging of activatable fluorophores in the myocardium by reporting molecular activity in vivo due to changes in intramolecular quenching. This characterization of FL from activatable molecular probes will be helpful for advancing in vivo imaging of enzyme activity.

Goergen, Craig J.; Chen, Howard H.; Bogdanov, Alexei; Sosnovik, David E.; Kumar, Anand T. N.

2012-05-01

190

Intensity Dependence of the Fluorescence Lifetime of in vivo Chlorophyll Excited by a Picosecond Light Pulse  

Microsoft Academic Search

New data on intensity-dependent lifetimes indicate that all previous in vivo fluorescence studies of chlorophyll by picosecond techniques must be reinterpreted. Anomalously short lifetimes result from high-intensity effects due to exciton-exciton annihilation processes. Measurements in Chlorella pyrenoidosa with single-pulse, low-intensity excitation indicate a longer \\

A. J. Campillo; V. H. Kollman; S. L. Shapiro

1976-01-01

191

In Vivo Mitochondrial Oxygen Tension Measured by a Delayed Fluorescence Lifetime Technique  

PubMed Central

Mitochondrial oxygen tension (mitoPO2) is a key parameter for cellular function, which is considered to be affected under various pathophysiological circumstances. Although many techniques for assessing in vivo oxygenation are available, no technique for measuring mitoPO2 in vivo exists. Here we report in vivo measurement of mitoPO2 and the recovery of mitoPO2 histograms in rat liver by a novel optical technique under normal and pathological circumstances. The technique is based on oxygen-dependent quenching of the delayed fluorescence lifetime of protoporphyrin IX. Application of 5-aminolevulinic acid enhanced mitochondrial protoporphyrin IX levels and induced oxygen-dependent delayed fluorescence in various tissues, without affecting mitochondrial respiration. Using fluorescence microscopy, we demonstrate in isolated hepatocytes that the signal is of mitochondrial origin. The delayed fluorescence lifetime was calibrated in isolated hepatocytes and isolated perfused livers. Ultimately, the technique was applied to measure mitoPO2 in rat liver in vivo. The results demonstrate mitoPO2 values of ?30–40 mmHg. mitoPO2 was highly sensitive to small changes in inspired oxygen concentration around atmospheric oxygen level. Ischemia-reperfusion interventions showed altered mitoPO2 distribution, which flattened overall compared to baseline conditions. The reported technology is scalable from microscopic to macroscopic applications, and its reliance on an endogenous compound greatly enhances its potential field of applications. PMID:18641065

Mik, Egbert G.; Johannes, Tanja; Zuurbier, Coert J.; Heinen, Andre; Houben-Weerts, Judith H. P. M.; Balestra, Gianmarco M.; Stap, Jan; Beek, Johan F.; Ince, Can

2008-01-01

192

Instrument for fluorescence sensing of circulating cells with diffuse light in mice in vivo  

NASA Astrophysics Data System (ADS)

Accurate quantification of circulating cell populations in mice is important in many areas of preclinical biomedical research. Normally, this is done either by extraction and analysis of small blood samples or, more recently, by using microscopy-based in vivo fluorescence flow cytometry. We describe a new technological approach to this problem using detection of diffuse fluorescent light from relatively large blood vessels in vivo. The diffuse fluorescence flow cytometer (DFFC) uses a laser to illuminate a mouse limb and an array of optical fibers coupled to a high-sensitivity photomultiplier tube array operating in photon counting mode to detect weak fluorescence signals from cells. We first demonstrate that the DFFC instrument is capable of detecting fluorescent microspheres and Vybrant-DiD-labeled cells in a custom-made optical flow phantom with similar size, optical properties, linear flow rates, and autofluorescence as a mouse limb. We also present preliminary data demonstrating that the DFFC is capable of detecting circulating cells in nude mice in vivo. In principle, this device would allow interrogation of the whole blood volume of a mouse in minutes, with sensitivity improvement by several orders of magnitude compared to current approaches.

Zettergren, Eric; Vickers, Dwayne; Runnels, Judith; Murthy, Shashi K.; Lin, Charles P.; Niedre, Mark

2012-03-01

193

Instrument for fluorescence sensing of circulating cells with diffuse light in mice in vivo  

PubMed Central

Abstract. Accurate quantification of circulating cell populations in mice is important in many areas of preclinical biomedical research. Normally, this is done either by extraction and analysis of small blood samples or, more recently, by using microscopy-based in vivo fluorescence flow cytometry. We describe a new technological approach to this problem using detection of diffuse fluorescent light from relatively large blood vessels in vivo. The diffuse fluorescence flow cytometer (DFFC) uses a laser to illuminate a mouse limb and an array of optical fibers coupled to a high-sensitivity photomultiplier tube array operating in photon counting mode to detect weak fluorescence signals from cells. We first demonstrate that the DFFC instrument is capable of detecting fluorescent microspheres and Vybrant-DiD-labeled cells in a custom-made optical flow phantom with similar size, optical properties, linear flow rates, and autofluorescence as a mouse limb. We also present preliminary data demonstrating that the DFFC is capable of detecting circulating cells in nude mice in vivo. In principle, this device would allow interrogation of the whole blood volume of a mouse in minutes, with sensitivity improvement by several orders of magnitude compared to current approaches. PMID:22502573

Zettergren, Eric; Vickers, Dwayne; Runnels, Judith; Murthy, Shashi K.; Lin, Charles P.; Niedre, Mark

2012-01-01

194

Photoacoustic correlation spectroscopy for in vivo blood flow speed measurement  

NASA Astrophysics Data System (ADS)

Photoacoustic imaging has been widely used in structural and functional imaging. Because of its safety, high resolution, and high imaging depth, it has great potential for a variety of medical studies. Capillaries are the smallest blood vessels and enable the exchange of oxygen and nutrients. Noninvasive flow speed measurement of capillaries in vivo can benefit the study of vascular tone changes and rheological properties of blood cells in capillaries. Recently, there has been a growing interest in photoacoustic velocimetry, such as photoacoustic Doppler and M-mode photoacoustic flow imaging. Methods capable of high-resolution imaging and low-speed flow measurement are suitable to measure blood speeds in capillaries. Previously we proposed photoacoustic correlation spectroscopy (PACS) and shown its feasibility for lowspeed flow measurement. Here, in vivo measurement of blood speeds in capillaries in a chick embryo model by PACS technique is demonstrated. The laser-scanning photoacoustic microscopy system is used for fast imaging acquisition and high-resolution imaging. The measured speed in capillaries is similar to those found in literatures, which confirm the feasibility of the PACS method for blood velocimetry. This technique suggests a fairly simple way to study blood flow speeds in capillaries.

Chen, Sung-Liang; Xie, Zhixing; Carson, Paul L.; Wang, Xueding; Guo, L. Jay

2012-02-01

195

Infrared Fluorescent Imaging as a Potent Tool for In Vitro, Ex Vivo and In Vivo Models of Visceral Leishmaniasis  

PubMed Central

Background Visceral leishmaniasis (VL) is hypoendemic in the Mediterranean region, where it is caused by the protozoan Leishmania infantum. An effective vaccine for humans is not yet available and the severe side-effects of the drugs in clinical use, linked to the parenteral administration route of most of them, are significant concerns of the current leishmanicidal medicines. New drugs are desperately needed to treat VL and phenotype-based High Throughput Screenings (HTS) appear to be suitable to achieve this goal in the coming years. Methodology/Principal findings We generated two infrared fluorescent L. infantum strains, which stably overexpress the IFP 1.4 and iRFP reporter genes and performed comparative studies of their biophotonic properties at both promastigote and amastigote stages. To improve the fluorescence emission of the selected reporter in intracellular amastigotes, we engineered distinct constructs by introducing regulatory sequences of differentially-expressed genes (A2, AMASTIN and HSP70 II). The final strain that carries the iRFP gene under the control of the L. infantum HSP70 II downstream region (DSR), was employed to perform a phenotypic screening of a collection of small molecules by using ex vivo splenocytes from infrared-infected BALB/c mice. In order to further investigate the usefulness of this infrared strain, we monitored an in vivo infection by imaging BALB/c mice in a time-course study of 20 weeks. Conclusions/Significance The near-infrared fluorescent L. infantum strain represents an important step forward in bioimaging research of VL, providing a robust model of phenotypic screening suitable for HTS of small molecule collections in the mammalian parasite stage. Additionally, HSP70 II+L. infantum strain permitted for the first time to monitor an in vivo infection of VL. This finding accelerates the possibility of testing new drugs in preclinical in vivo studies, thus supporting the urgent and challenging drug discovery program against this parasitic disease. PMID:25826250

Calvo-Álvarez, Estefanía; Stamatakis, Kostantinos; Punzón, Carmen; Álvarez-Velilla, Raquel; Tejería, Ana; Escudero-Martínez, José Miguel; Pérez-Pertejo, Yolanda; Fresno, Manuel; Balaña-Fouce, Rafael; Reguera, Rosa M.

2015-01-01

196

In vivo magnetic resonance spectroscopy of liver tumors and metastases  

PubMed Central

Primary liver cancer is the fifth most common malignancy in men and the eighth in women worldwide. The liver is also the second most common site for metastatic spread of cancer. To assist in the diagnosis of these liver lesions non-invasive advanced imaging techniques are desirable. Magnetic resonance (MR) is commonly used to identify anatomical lesions, but it is a very versatile technique and also can provide specific information on tumor pathophysiology and metabolism, in particular with the application of MR spectroscopy (MRS). This may include data on the type, grade and stage of tumors, and thus assist in further management of the disease. The purpose of this review is to summarize and discuss the available literature on proton, phosphorus and carbon-13-MRS as performed on primary liver tumors and metastases, with human applications as the main perspective. Upcoming MRS approaches with potential applications to liver tumors are also included. Since knowledge of some technical background is indispensable to understand the results, a basic introduction of MRS and some technical issues of MRS as applied to tumors and metastases in the liver are described as well. In vivo MR spectroscopy of tumors in a metabolically active organ such as the liver has been demonstrated to provide important information on tumor metabolism, but it also is challenging as compared to applications on some other tissues, in particular in humans, mostly because of its abdominal location where movement may be a disturbing factor. PMID:22215937

ter Voert, EGW; Heijmen, L; van Laarhoven, HWM; Heerschap, A

2011-01-01

197

Multimodal Raman-fluorescence spectroscopy of formalin fixed samples is able to discriminate brain tumors from dysplastic tissue  

NASA Astrophysics Data System (ADS)

In the recent years, there has been a considerable surge in the application of spectroscopy for disease diagnosis. Raman and fluorescence spectra provide characteristic spectral profile related to biochemical and morphological changes when tissues progress from normal state towards malignancy. Spectroscopic techniques offer the advantage of being minimally invasive compared to traditional histopathology, real time and quantitative. In biomedical optical diagnostics, freshly excised specimens are preferred for making ex-vivo spectroscopic measurements. With regard to fresh tissues, if the lab is located far away from the clinic it could pose a problem as spectral measurements have to be performed immediately after dissection. Tissue samples are usually placed in a fixative agent such as 4% formaldehyde to preserve the samples before processing them for routine histopathological studies. Fixation prevents the tissues from decomposition by arresting autolysis. In the present study, we intend to investigate the possibility of using formalin fixed samples for discrimination of brain tumours from dysplastic tissue using Raman and fluorescence spectroscopy. Formalin fixed samples were washed with phosphate buffered saline for about 5 minutes in order to remove the effects of formalin during spectroscopic measurements. In case of fluorescence spectroscopy, changes in spectral profile have been observed in the region between 550-670 nm between dysplastic and tumor samples. For Raman measurements, we found significant differences in the spectral profiles between dysplasia and tumor. In conclusion, formalin fixed samples can be potentially used for the spectroscopic discrimination of tumor against dysplastic tissue in brain samples.

Anand, Suresh; Cicchi, Riccardo; Giordano, Flavio; Buccoliero, Anna Maria; Pavone, Francesco Saverio

2014-05-01

198

In Vivo Time-gated Fluorescence Imaging with Biodegradable Luminescent Porous Silicon Nanoparticles  

PubMed Central

Fluorescence imaging is one of the most versatile and widely used visualization methods in biomedical research. However, tissue autofluorescence is a major obstacle confounding interpretation of in vivo fluorescence images. The unusually long emission lifetime (5-13 ?s) of photoluminescent porous silicon nanoparticles can allow the time-gated imaging of tissues in vivo, completely eliminating shorter-lived (< 10 ns) emission signals from organic chromophores or tissue autofluorescence.Here, using a conventional animal imaging system not optimized for such long-lived excited states, we demonstrate improvement of signal to background contrast ratio by > 50-fold in vitro and by > 20-fold in vivo when imaging porous silicon nanoparticles. Time-gated imaging of porous silicon nanoparticles accumulated in a human ovarian cancer xenograft following intravenous injection is demonstrated in a live mouse. The potential for multiplexing of images in the time domain by using separate porous silicon nanoparticles engineered with different excited state lifetimes is discussed. PMID:23933660

Gu, Luo; Hall, David J.; Qin, Zhengtao; Anglin, Emily; Joo, Jinmyoung; Mooney, David J.; Howell, Stephen B.; Sailor, Michael J.

2014-01-01

199

Fluorescence Correlation Spectroscopy and Nonlinear Stochastic Reaction-Diffusion  

SciTech Connect

The currently existing theory of fluorescence correlation spectroscopy (FCS) is based on the linear fluctuation theory originally developed by Einstein, Onsager, Lax, and others as a phenomenological approach to equilibrium fluctuations in bulk solutions. For mesoscopic reaction-diffusion systems with nonlinear chemical reactions among a small number of molecules, a situation often encountered in single-cell biochemistry, it is expected that FCS time correlation functions of a reaction-diffusion system can deviate from the classic results of Elson and Magde [Biopolymers (1974) 13:1-27]. We first discuss this nonlinear effect for reaction systems without diffusion. For nonlinear stochastic reaction-diffusion systems there are no closed solutions; therefore, stochastic Monte-Carlo simulations are carried out. We show that the deviation is small for a simple bimolecular reaction; the most significant deviations occur when the number of molecules is small and of the same order. Extending Delbrück-Gillespie’s theory for stochastic nonlinear reactions with rapidly stirring to reaction-diffusion systems provides a mesoscopic model for chemical and biochemical reactions at nanometric and mesoscopic level such as a single biological cell.

Del Razo, Mauricio; Pan, Wenxiao; Qian, Hong; Lin, Guang

2014-05-30

200

Fluorescence Correlation Spectroscopy and Nonlinear Stochastic Reaction-Diffusion  

E-print Network

The currently existing theory of fluorescence correlation spectroscopy(FCS) is based on the linear fluctuation theory originally developed by Einstein, Onsager, Lax, and others as a phenomenological approach to equilibrium fluctuations in bulk solutions. For mesoscopic reaction-diffusion systems with nonlinear chemical reactions among a small number of molecules, a situation often encountered in single-cell biochemistry, it is expected that FCS time correlation functions of a reaction-diffusion system can deviate from the classic results of Elson and Magde. We first discuss this nonlinear effect for reaction systems without diffusion. For nonlinear stochastic reaction-diffusion systems here are no closed solutions; therefore, stochastic Monte-Carlo simulations are carried out. We show that the deviation is small for a simple bimolecular reaction; the most significant deviations occur when the number of molecules is small and of the same order. Our results show that current linear FCS theory could be adequate for measurements on biological systems that contain many other sources of uncertainties. At the same time it provides a framework for future measurements of nonlinear, fluctuating chemical reactions with high-precision FCS. Extending Delbr\\"uck-Gillespie's theory for stochastic nonlinear reactions with rapidly stirring to reaction-diffusion systems provides a mesoscopic model for chemical and biochemical reactions at nanometric and mesoscopic level such as a single biological cell.

Mauricio J. Del Razo; Wenxiao Pan; Hong Qian; Guang Lin

2014-06-16

201

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

202

Characterization of flow direction in microchannels and zebrafish blood vessels by scanning fluorescence correlation spectroscopy  

E-print Network

The investigation of flow profiles in microstructures and tissues by fluorescence correlation spectroscopy (FCS) has been a challenging topic in the past decade. Due to its inherent optical configuration, a circular focused ...

Pan, Xiaotao

203

Protein oligomerization monitored by fluorescence fluctuation spectroscopy: Self-assembly of Rubisco activase  

Technology Transfer Automated Retrieval System (TEKTRAN)

A methodology is presented to characterize complex protein assembly pathways by fluorescence correlation spectroscopy. We have derived the total autocorrelation function describing the behavior of mixtures of labeled and unlabeled protein under equilibrium conditions. Our modeling approach allows us...

204

A 32-channel photon counting module with embedded auto/cross-correlators for real-time parallel fluorescence correlation spectroscopy.  

PubMed

Fluorescence correlation spectroscopy (FCS) is a well-established technique to study binding interactions or the diffusion of fluorescently labeled biomolecules in vitro and in vivo. Fast FCS experiments require parallel data acquisition and analysis which can be achieved by exploiting a multi-channel Single Photon Avalanche Diode (SPAD) array and a corresponding multi-input correlator. This paper reports a 32-channel FPGA based correlator able to perform 32 auto/cross-correlations simultaneously over a lag-time ranging from 10 ns up to 150 ms. The correlator is included in a 32 × 1 SPAD array module, providing a compact and flexible instrument for high throughput FCS experiments. However, some inherent features of SPAD arrays, namely afterpulsing and optical crosstalk effects, may introduce distortions in the measurement of auto- and cross-correlation functions. We investigated these limitations to assess their impact on the module and evaluate possible workarounds. PMID:25362365

Gong, S; Labanca, I; Rech, I; Ghioni, M

2014-10-01

205

A 32-channel photon counting module with embedded auto/cross-correlators for real-time parallel fluorescence correlation spectroscopy  

SciTech Connect

Fluorescence correlation spectroscopy (FCS) is a well-established technique to study binding interactions or the diffusion of fluorescently labeled biomolecules in vitro and in vivo. Fast FCS experiments require parallel data acquisition and analysis which can be achieved by exploiting a multi-channel Single Photon Avalanche Diode (SPAD) array and a corresponding multi-input correlator. This paper reports a 32-channel FPGA based correlator able to perform 32 auto/cross-correlations simultaneously over a lag-time ranging from 10 ns up to 150 ms. The correlator is included in a 32 × 1 SPAD array module, providing a compact and flexible instrument for high throughput FCS experiments. However, some inherent features of SPAD arrays, namely afterpulsing and optical crosstalk effects, may introduce distortions in the measurement of auto- and cross-correlation functions. We investigated these limitations to assess their impact on the module and evaluate possible workarounds.

Gong, S.; Labanca, I.; Rech, I.; Ghioni, M. [Dipartimento di Elettronica, Informazione e Bioingegneria, Politecnico di Milano, Piazza Leonardo da Vinci 32, 20133 Milano (Italy)

2014-10-15

206

Intestine pH measurements using fluorescence imaging: an in-vivo preliminary study  

NASA Astrophysics Data System (ADS)

Measurement of gastrointestinal intramucosal pH has been recognized as an important factor in the detection of hypoxia-induced dysfunctions. However, current pH measurement techniques are limited in terms of time and spatial resolution. A major advance in accurate pH measurement was the development of the ratiometric fluorescent indicator dye, 2',7'-bis(carboxyethyl)-4,5- carboxyfluorescein (BCECF). This study aimed to demonstrate the feasibility of fluorescence imaging technique to measure in vivo the pH of intestine. The intestine was inserted in an optical chamber placed under a microscope. Animals were injected i.v. with the pH-sensitive fluorescent dye BCECF. Fluorescence was visualized by illuminating the intestine alternately at 490 and 470 nm. The emitted fluorescence was directed to an intensified camera. The ratio of emitted fluorescence at excitation wavelengths of 490 and 470 nm was measured, corrected and converted to pH by constructing a calibration curve. The pH controls were performed with a pH microelectrode correlated with venous blood gas sampling. We concluded that accurate pH measurements of rat intestine can be obtained by fluorescence imaging using BCECF. This technology could be easily adapted for endoscopic pH measurement.

Marechal, Xavier-Marie; Mordon, Serge R.; Devoisselle, Jean-Marie; Begu, Sylvie; Mathieu, D.; Buys, Bruno; Dhelin, Guy; Lesage, Jean C.; Neviere, Remi; Chopin, Claude

1999-02-01

207

Investigation of efflorescence of inorganic aerosols using fluorescence spectroscopy.  

PubMed

The phase transition is one of the most fundamental phenomena affecting the physical and chemical properties of atmospheric aerosols. Efflorescence, in particular, is not well understood, partly because the molecular interactions between the solute and water molecules of saturated or supersaturated solution droplets have not been well characterized. Recently, we developed a technique that combines the use of an electrodynamic balance and a fluorescence dye, 8-hydroxyl-1,3,6-pyrenetrisulfonate (pyranine), to study the distributions of solvated and free water in aqueous droplets (Choi, M. Y.; Chan, C. K.; Zhang, Y. H. J. Phys. Chem. A 2004, 108, 1133). We found that the equality of the amounts of solvated and free water is a necessary but not sufficient condition for efflorescence. For efflorescing compounds such as Na2SO4, (NH4)2SO4, and a mixture of NaCl and Na2SO4, the amount of free water decreases, while that of solvated water is roughly constant in bulk measurements and decreases less dramatically than that of free water in single-particle measurements as the relative humidity (RH) decreases. Efflorescence of the supersaturated droplets of these solutions occurs when the amounts of free and solvated water are equal, which is consistent with our previous observation for NaCl. For nonefflorescing compounds in single-particle levitation experiments such as MgSO4 and Mg(NO3)2, the amounts of free and solvated water are equal at a water-to-solute molar ratio of about 6, at which spectral changes due to the formation of contact ion pairs between magnesium and the anions occur as shown by Raman spectroscopy. Fluorescence imaging shows that the droplets of diluted Mg(NO3)2 (at 80% RH) and MgSO4 are homogeneous but those of NaCl, Na2SO4, (NH4)2SO4, and supersaturated Mg(NO3)2 (at 10% RH) are heterogeneous in terms of the solvated-to-free water distribution. The solvated-to-free water ratios in NaCl, Na2SO4, and (NH4)2SO4 droplets are higher in the outer regions by about half a radius deep than at the center of the droplets. PMID:16833412

Choi, Man Yee; Chan, Chak K

2005-02-17

208

In vivo fluorescence imaging of exogenous enzyme activity in the gastrointestinal tract  

PubMed Central

Exogenous enzymes are administered orally to treat several diseases, such as pancreatic insufficiency and lactose intolerance. Due to the proteinaceous nature of enzymes, they are subject to inactivation and/or digestion in the gastrointestinal (GI) tract. Here we describe a convenient fluorescence-based assay to monitor the activity of therapeutic enzymes in real time in vivo in the GI tract. To establish the proof of principle, the assay was applied to proline-specific endopeptidases (PEPs), a group of enzymes recently proposed as adjuvant therapy for celiac disease (a highly prevalent immunogenetic enteropathy). A short PEP-specific peptide sequence which is part of larger immunotoxic sequences of gluten was labeled with a fluorescent dye and a corresponding quencher. Upon enzymatic cleavage, the fluorescence emission was dequenched and detected with an in vivo imaging system. PEPs originating from Flavobacterium meningosepticum (FM) and Myxococcus xanthus (MX) were evaluated after oral administration in rats. While MX PEP could not cleave the peptide in the stomach, FM PEP showed significant gastric activity reaching 40–60% of the maximal in vivo signal intensity. However, both enzymes produced comparable fluorescence signals in the small intestine. Coadministration of an antacid drug significantly enhanced MX PEP’s gastric activity due to increased pH and/or inhibition of stomach proteases. With this simple procedure, differences in the in vivo performance of PEPs, which could not be identified under in vitro conditions, were detected. This imaging assay could be used to study other oral enzymes in vivo and therefore be instrumental in improving their therapeutic efficiency. PMID:21576491

Fuhrmann, Gregor; Leroux, Jean-Christophe

2011-01-01

209

Prediction of myocardial damage depth induced by extracellular photosensitization reaction using fluorescence measurement in vivo  

NASA Astrophysics Data System (ADS)

We experimentally studied the correlation between myocardial damage depth due to the extracellular photosensitization reaction (PR) using talaporfin sodium and fluorescence-fall amount (FA), which is calculated from the measured backscattering fluorescence intensity via a manipulatable 7 Fr. laser catheter during the PR operation in vivo to establish treatment depth predictor for a non-thermal tachyarrhythmia treatment. The PR was performed to left and/or right ventricle in the open-chest canine heart. The laser irradiation of 663+/-2 nm in wavelength via the laser catheter was operated 15 min after the intravenous administration of talaporfin sodium with concentration of 36.2+/-8.0 ?g/ml in plasma. The irradiation was operated with irradiance of 5, 10, 20 W/cm2, and duration of 5, 10, 20 s. Backscattering fluorescence of 710+/-2 nm in wavelength was measured via the laser catheter during the PR. The FA was calculated multiplying the irradiation duration by the fluorescence-fall, which is subtraction of the fluorescence intensity at the kickoff and end of the irradiation. The canine heart was extracted 1 week after the PR and HE stained specimen was histologically evaluated. The correlation of the myocardial damage depth and FA was investigated. We found that FA obtained a logarithmic relation to the myocardial damage depth. We think that the FA might be available to predict the PR induced myocardial damage depth for the application of tachyarrhythmia treatment under catheterization in vivo.

Takahashi, M.; Ogawa, E.; Nakamura, T.; Kawakami, H.; Machida, N.; Yajima, M.; Kurotsu, M.; Ito, A.; Kimura, T.; Arai, T.

2014-03-01

210

Biocompatible near-infrared fluorescent nanoparticles for macro and microscopic in vivo functional bioimaging.  

PubMed

Near-infrared (NIR) imaging technology has been widely used for biomedical research and applications, since it can achieve deep penetration in biological tissues due to less absorption and scattering of NIR light. In our research, polymer nanoparticles with NIR fluorophores doped were synthesized. The morphology, absorption/emission features and chemical stability of the fluorescent nanoparticles were characterized, separately. NIR fluorescent nanoparticles were then utilized as bright optical probes for macro in vivo imaging of mice, including sentinel lymph node (SLN) mapping, as well as distribution and excretion monitoring of nanoparticles in animal body. Furthermore, we applied the NIR fluorescent nanoparticles in in vivo microscopic bioimaging via a confocal microscope. Under the 635 nm-CW excitation, the blood vessel architecture in the ear and the brain of mice, which were administered with nanoparticles, was visualized very clearly. The imaging depth of our one-photon microscopy, which was assisted with NIR fluorescent nanoprobes, can reach as deep as 500 ?m. Our experiments show that NIR fluorescent nanoparticles have great potentials in various deep-tissue imaging applications. PMID:25426331

Chu, Liliang; Wang, Shaowei; Li, Kanghui; Xi, Wang; Zhao, Xinyuan; Qian, Jun

2014-11-01

211

Biocompatible near-infrared fluorescent nanoparticles for macro and microscopic in vivo functional bioimaging  

PubMed Central

Near-infrared (NIR) imaging technology has been widely used for biomedical research and applications, since it can achieve deep penetration in biological tissues due to less absorption and scattering of NIR light. In our research, polymer nanoparticles with NIR fluorophores doped were synthesized. The morphology, absorption/emission features and chemical stability of the fluorescent nanoparticles were characterized, separately. NIR fluorescent nanoparticles were then utilized as bright optical probes for macro in vivo imaging of mice, including sentinel lymph node (SLN) mapping, as well as distribution and excretion monitoring of nanoparticles in animal body. Furthermore, we applied the NIR fluorescent nanoparticles in in vivo microscopic bioimaging via a confocal microscope. Under the 635 nm-CW excitation, the blood vessel architecture in the ear and the brain of mice, which were administered with nanoparticles, was visualized very clearly. The imaging depth of our one-photon microscopy, which was assisted with NIR fluorescent nanoprobes, can reach as deep as 500 ?m. Our experiments show that NIR fluorescent nanoparticles have great potentials in various deep-tissue imaging applications. PMID:25426331

Chu, Liliang; Wang, Shaowei; Li, Kanghui; Xi, Wang; Zhao, Xinyuan; Qian, Jun

2014-01-01

212

Single-Molecule Fluorescence Spectroscopy: New Probes of Protein Function and Dynamics  

NSDL National Science Digital Library

Single-molecule fluorescence methods provide new tools for the study of biological systems. Single-pair fluorescence resonance energy transfer has provided detailed information about dynamics and structure of the Ca2+-signaling protein calmodulin. Single-molecule polarization modulation spectroscopy has probed the mechanism by which calmodulin activates the plasma membrane Ca2+ pump

PhD Carey K. Johnson (University of Kansas Department of Chemistry)

2005-02-01

213

Fluorescence spectroscopy of single-walled carbon nanotubes synthesized from alcohol  

E-print Network

I&EC 221 Fluorescence spectroscopy of single-walled carbon nanotubes synthesized from alcohol fluorescence measurements of single-walled carbon nanotubes (SWNTs) catalytically synthesized from alcohol (Alcohol catalytic CVD method, ACCVD) in various experimental conditions were performed. The chirality

Maruyama, Shigeo

214

Micro-Raman Spectroscopy of Algae: Composition Analysis and Fluorescence Background Behavior  

E-print Network

ARTICLE Micro-Raman Spectroscopy of Algae: Composition Analysis and Fluorescence Background performed using Stokes Raman scattering for compositional analysis of algae. Two algal species, Chlorella while acquiring Raman signals from the algae. The time dependence of fluorescence background is char

215

Triplet fraction buildup effect of the DNA–YOYO complex studied with fluorescence correlation spectroscopy  

Microsoft Academic Search

DNA fragments of various lengths and YOYO-1 iodide (YOYO) were mixed at various ratios, and fluorescence was measured using fluorescence correlation spectroscopy. The number of substantially emitting YOYO molecules binding to the DNA and the binding intervals between the YOYO molecules were estimated for DNA–YOYO complexes of various lengths. In the present study, we found an interesting phenomenon: triplet buildup.

Masafumi Shimizu; Satoshi Sasaki; Masataka Kinjo

2007-01-01

216

The effects of daunomycin antibiotic on histone H 1: thermal denaturation and fluorescence spectroscopy studies  

Microsoft Academic Search

Using thermal denaturation and fluorescence spectroscopy, we have investigated the interaction of antitumor antibiotic, daunomycin, with calf thymus histone H1 under several ionic strengths. The results show that daunomycin binds to histone H1 and increases its melting temperature. Increasing ionic strength elevates this effect. Fluorescence emission data show that the interaction of daunomycin with histone H1 decreases the emission intensity

Seyed Jalal Zargar; Azra Rabbani

2000-01-01

217

A method for the study of turbulent mixing using fluorescence spectroscopy  

Microsoft Academic Search

The mixing of two feed streams in a reactor, one with a fluorescent tracer, the other without, results in a fluctuating concentration field, due to the turbulent flow. Fluorescence spectroscopy allows the characterization of the fluctuations at small scale and high frequencies. Measurements have been made with a spatial resolution of about 30 µm and up to a frequency of

S. Gaskey; P. Vacus; R. David; J. Villermaux; J. C. André

1990-01-01

218

In vivo PDT dosimetry: singlet oxygen emission and photosensitizer fluorescence  

NASA Astrophysics Data System (ADS)

Photodynamic therapy (PDT) is a light activated chemotherapy that is dependent on three parameters: photosensitizer (PS) concentration; oxygen concentration; and light dosage. Due to highly variable treatment response, the development of an accurate dosimeter to optimize PDT treatment outcome is an important requirement for practical applications. Singlet oxygen is an active species in PDT, and we are developing two instruments, an ultra-sensitive singlet oxygen point sensor and a 2D imager, with the goal of a real-time dosimeter for PDT researchers. The 2D imaging system can visualize spatial maps of both the singlet oxygen production and the location of the PS in a tumor during PDT. We have detected the production of singlet oxygen during PDT treatments with both in-vitro and in-vivo studies. Effects of photobleaching have also been observed. These results are promising for the development of the sensor as a real-time dosimeter for PDT which would be a valuable tool for PDT research and could lead to more effective treatment outcome. We summarize recent results in this paper.

Lee, Seonkyung; Galbally-Kinney, Kristin L.; Murphy, Brian A.; Davis, Steven J.; Hasan, Tayyaba; Spring, Bryan; Tu, Yupeng; Pogue, Brian W.; Isabelle, Martin E.; O'Hara, Julia A.

2010-02-01

219

Real-time Raman spectroscopy for in vivo, online gastric cancer diagnosis during clinical endoscopic examination  

NASA Astrophysics Data System (ADS)

Optical spectroscopic techniques including reflectance, fluorescence and Raman spectroscopy have shown promising potential for in vivo precancer and cancer diagnostics in a variety of organs. However, data-analysis has mostly been limited to post-processing and off-line algorithm development. In this work, we develop a fully automated on-line Raman spectral diagnostics framework integrated with a multimodal image-guided Raman technique for real-time in vivo cancer detection at endoscopy. A total of 2748 in vivo gastric tissue spectra (2465 normal and 283 cancer) were acquired from 305 patients recruited to construct a spectral database for diagnostic algorithms development. The novel diagnostic scheme developed implements on-line preprocessing, outlier detection based on principal component analysis statistics (i.e., Hotelling's T2 and Q-residuals) for tissue Raman spectra verification as well as for organ specific probabilistic diagnostics using different diagnostic algorithms. Free-running optical diagnosis and processing time of < 0.5 s can be achieved, which is critical to realizing real-time in vivo tissue diagnostics during clinical endoscopic examination. The optimized partial least squares-discriminant analysis (PLS-DA) models based on the randomly resampled training database (80% for learning and 20% for testing) provide the diagnostic accuracy of 85.6% [95% confidence interval (CI): 82.9% to 88.2%] [sensitivity of 80.5% (95% CI: 71.4% to 89.6%) and specificity of 86.2% (95% CI: 83.6% to 88.7%)] for the detection of gastric cancer. The PLS-DA algorithms are further applied prospectively on 10 gastric patients at gastroscopy, achieving the predictive accuracy of 80.0% (60/75) [sensitivity of 90.0% (27/30) and specificity of 73.3% (33/45)] for in vivo diagnosis of gastric cancer. The receiver operating characteristics curves further confirmed the efficacy of Raman endoscopy together with PLS-DA algorithms for in vivo prospective diagnosis of gastric cancer. This work successfully moves biomedical Raman spectroscopic technique into real-time, on-line clinical cancer diagnosis, especially in routine endoscopic diagnostic applications.

Duraipandian, Shiyamala; Sylvest Bergholt, Mads; Zheng, Wei; Yu Ho, Khek; Teh, Ming; Guan Yeoh, Khay; Bok Yan So, Jimmy; Shabbir, Asim; Huang, Zhiwei

2012-08-01

220

In-vivo Fluorescent X-ray CT Imaging of Mouse Brain  

SciTech Connect

Using a non-radioactive iodine-127 labeled cerebral perfusion agent (I-127 IMP), fluorescent X-ray computed tomography (FXCT) clearly revealed the cross-sectional distribution of I-127 IMP in normal mouse brain in-vivo. Cerebral perfusion of cortex and basal ganglion was depicted with 1 mm in-plane spatial resolution and 0.1 mm slice thickness. Degree of cerebral perfusion in basal ganglion was about 2-fold higher than that in cortical regions. This result suggests that in-vivo cerebral perfusion imaging is realized quantitatively by FXCT at high volumetric resolution.

Takeda, T.; Wu, J.; Lwin, Thet-Thet; Huo, Q.; Minami, M. [Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8575 (Japan); Sunaguchi, N.; Murakami, T.; Mouri, S.; Nasukawa, S.; Yuasa, T.; Akatsuka, T. [Faculty of Engineering, Yamagata University, Yonezawa, Yamagata 992-8510 (Japan); Hyodo, K. [Institute of Material Science, High Energy Accelerator Research Organization, Tsukuba, Ibaraki 305-0801 (Japan); Hontani, H. [Department of Computer Science and Engineering, Nagoya Institute of Technology, Nagoya, Aichi 466-8555 (Japan)

2007-01-19

221

Development of an Immunologically Tolerated Combination of Fluorescent Proteins for In vivo Two-photon Imaging  

PubMed Central

Combinations of fluorescent proteins (FPs) are routinely used for multi-parameter in vivo imaging experiments to visualize tagged proteins or cell populations of interest. Studies involving FPs are often limited by spectral overlap, toxicity, relative quantum efficiency, and the potential for immunological rejection upon transfer into a non-tolerant recipient. Here we evaluate the immunologic visibility of several commonly used FPs by the murine immune system and identify a spectrally compatible, immunologically tolerated combination of FPs well suited for in vivo two-photon imaging. PMID:25322934

Gossa, Selamawit; Nayak, Debasis; Zinselmeyer, Bernd H.; McGavern, Dorian B.

2014-01-01

222

In vivo stepwise multi-photon activation fluorescence imaging of melanin in human skin  

NASA Astrophysics Data System (ADS)

The stepwise multi-photon activated fluorescence (SMPAF) of melanin is a low cost and reliable method of detecting melanin because the activation and excitation can be a continuous-wave (CW) mode near infrared (NIR) laser. Our previous work has demonstrated the melanin SMPAF images in sepia melanin, mouse hair, and mouse skin. In this study, we show the feasibility of using SMPAF to detect melanin in vivo. in vivo melanin SMPAF images of normal skin and benign nevus are demonstrated. SMPAF images add specificity for melanin detection than MPFM images and CRM images. Melanin SMPAF is a promising technology to enable early detection of melanoma for dermatologists.

Lai, Zhenhua; Gu, Zetong; Abbas, Saleh; Lowe, Jared; Sierra, Heidy; Rajadhyaksha, Milind; DiMarzio, Charles

2014-03-01

223

A fresh look at the validity of the diffusion approximation for modeling fluorescence spectroscopy in biological tissue.  

PubMed

Fluorescence has become a widely used technique for applications in noninvasive diagnostic tissue spectroscopy. The standard model used for characterizing fluorescence photon transport in biological tissue is based on the diffusion approximation. On the premise that the total energy of excitation and fluorescent photon flows must be conserved, we derive the widely used diffusion equations in fluorescence spectroscopy and show that there must be an additional term to account for the transport of fluorescent photons. The significance of this additional term in modeling fluorescence spectroscopy in biological tissue is assessed. PMID:23366185

Handapangoda, Chintha Chamalie; Premaratne, Malin; Nahavandi, Saeid

2012-01-01

224

In vivo impedance spectroscopy of deep brain stimulation electrodes  

NASA Astrophysics Data System (ADS)

Deep brain stimulation (DBS) represents a powerful clinical technology, but a systematic characterization of the electrical interactions between the electrode and the brain is lacking. The goal of this study was to examine the in vivo changes in the DBS electrode impedance that occur after implantation and during clinically relevant stimulation. Clinical DBS devices typically apply high-frequency voltage-controlled stimulation, and as a result, the injected current is directly regulated by the impedance of the electrode-tissue interface. We monitored the impedance of scaled-down clinical DBS electrodes implanted in the thalamus and subthalamic nucleus of a rhesus macaque using electrode impedance spectroscopy (EIS) measurements ranging from 0.5 Hz to 10 kHz. To further characterize our measurements, equivalent circuit models of the electrode-tissue interface were used to quantify the role of various interface components in producing the observed electrode impedance. Following implantation, the DBS electrode impedance increased and a semicircular arc was observed in the high-frequency range of the EIS measurements, commonly referred to as the tissue component of the impedance. Clinically relevant stimulation produced a rapid decrease in electrode impedance with extensive changes in the tissue component. These post-operative and stimulation-induced changes in impedance could play an important role in the observed functional effects of voltage-controlled DBS and should be considered during clinical stimulation parameter selection and chronic animal research studies.

Lempka, Scott F.; Miocinovic, Svjetlana; Johnson, Matthew D.; Vitek, Jerrold L.; McIntyre, Cameron C.

2009-08-01

225

Excitation-emission matrices (EEMs) and synchronous fluorescence spectroscopy (SFS) investigations of gastrointestinal tissues  

NASA Astrophysics Data System (ADS)

In this report we will present our recent investigations of the fluorescence properties of lower part gastrointestinal tissues using excitation-emission matrix and synchronous fluorescence spectroscopy measurement modalities. The spectral peculiarities observed will be discussed and the endogenous sources of the fluorescence signal will be addressed. For these fluorescence spectroscopy measurements the FluoroLog 3 system (HORIBA Jobin Yvon, France) was used. It consists of a Xe lamp (300 W, 200-650 nm), a double mono-chromators, and a PMT detector with a work region at 220- 850 nm. Autofluorescence signals were detected in the form of excitation-emission matrices for the samples of normal mucosa, dysphasia and colon carcinoma and specific spectral features for each tissue were found. Autofluorescence signals from the same samples are observed through synchronous fluorescence spectroscopy, which is a novel promising modality for fluorescence spectroscopy measurements of bio-samples. It is one of the most powerful techniques for multicomponent analysis, because of its sensitivity. In the SFS regime, the fluorescence signal is recorded while both excitation ?exc and emission wavelengths ?em are simultaneously scanned. A constant wavelength interval is maintained between the ?exc and ?em wavelengths throughout the spectrum. The resulted fluorescence spectrum shows narrower peak widths, in comparison with EEMs, which are easier for identification and minimizes the chance for false determinations or pretermission of specific spectral feature. This modality is also faster, than EEMs, a much smaller number of data points are required.1 In our measurements we use constant wavelength interval ?? in the region of 10-200 nm. Measurements are carried out in the terms of finding ??, which results in a spectrum with most specific spectral features for comparison with spectral characteristics observed in EEMs. Implementing synchronous fluorescence spectroscopy in optical methods for analyzing biological tissues could result in a better differentiation between normal and dysplastic tissue. Thus could establish fluorescence imaging as a diagnostic modality among optical techniques applied in clinical practice.

Genova, Ts.; Borisova, E.; Zhelyazkova, Al.; Semyachkina-Glushkovskaya, O.; Penkov, N.; Keremedchiev, M.; Vladimirov, B.; Avramov, L.

2015-01-01

226

Single gold nanoparticles to enhance the detection of single fluorescent molecules at micromolar concentration using fluorescence correlation spectroscopy  

NASA Astrophysics Data System (ADS)

Single nanoparticles made of noble metals are strongly appealing to develop practical applications to detect fluorescent molecules in solution. Here, we detail the use of a single gold nanoparticle of 100 nm diameter to enhance the detection of single Alex Fluor 647 fluorescent molecules at high concentrations of several micromolar. We discuss the implementation of fluorescence correlation spectroscopy, and provide a new method to reliably extract the enhanced fluorescence signal stemming from the nanoparticle near-field from the background generated in the confocal volume. The applicability of our method is checked by reporting the invariance of the single molecule results as function of the molecular concentration, and the experimental data is found in good agreement with numerical simulations.

Punj, Deep; Rigneault, Hervé; Wenger, Jérôme

2014-05-01

227

In Vivo Photoacoustic and Fluorescence Cystography Using Clinically Relevant Dual Modal Indocyanine Green  

PubMed Central

Conventional X-ray-based cystography uses radio-opaque materials, but this method uses harmful ionizing radiation and is not sensitive. In this study, we demonstrate nonionizing and noninvasive photoacoustic (PA) and fluorescence (FL) cystography using clinically relevant indocyanine green (ICG) in vivo. After transurethral injection of ICG into rats through a catheter, their bladders were photoacoustically and fluorescently visualized. A deeply positioned bladder below the skin surface (i.e., ?1.5–5 mm) was clearly visible in the PA and FL image using a laser pulse energy of less than 2 mJ/cm2 (1/15 of the safety limit). Then, the in vivo imaging results were validated through in situ studies. Our results suggest that dual modal cystography can provide a nonionizing and noninvasive imaging tool for bladder mapping. PMID:25337743

Park, Sungjo; Kim, Jeesu; Jeon, Mansik; Song, Jaewon; Kim, Chulhong

2014-01-01

228

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

229

Reliable Assessment and Quantification of the Fluorescence-Labeled Antisense Oligonucleotides In Vivo  

PubMed Central

The availability of fluorescent dyes and the advances in the optical systems for in vivo imaging have stimulated an increasing interest in developing new methodologies to study and quantify the biodistribution of labeled agents. However, despite these great achievements, we are facing significant challenges in determining if the observed fluorescence does correspond to the quantity of the dye in the tissues. In fact, although the far-red and near-infrared lights can propagate through several centimetres of tissue, they diffuse within a few millimetres as consequence of the elastic scattering of photons. In addition, when dye-labeled oligonucleotides form stable complex with cationic carriers, a large change in the fluorescence intensity of the dye is observed. Therefore, the measured fluorescence intensity is altered by the tissue heterogeneity and by the fluctuation of dye intensity. Hence, in this study a quantification strategy for fluorescence-labeled oligonucleotides was developed to solve these disadvantageous effects. Our results proved that upon efficient homogenization and dilution with chaotropic agents, such as guanidinium thiocyanate, it is possible to achieve a complete fluorescence intensity recovery. Furthermore, we demonstrated that this method has the advantage of good sensitivity and reproducibility, as well as easy handling of the tissue samples. PMID:24967340

Chiara Munisso, Maria; Yamaoka, Tetsuji

2014-01-01

230

In vivo vascular imaging with adaptive optics confocal scanning fluorescence microscopy  

NASA Astrophysics Data System (ADS)

Adaptive optics is implemented in a confocal scanning fluorescence microscope with wavefront sensorless scheme. Using the image sharpness as the optimization metric, aberration correction is performed to compensate both system- and specimen-induced aberrations by using stochastic parallel gradient descent algorithm based upon Zernike polynomial modes. In vivo vascular imaging of mice ear is completed and the results revealed the improved signal and resolution leading to in substantially enhanced image contrast with aberration correction which allowed us to detect clearer vasculature structures.

Wang, Zhibin; Wei, Lin; He, Yi; Li, Xiqi; Shi, Guohua; Zhang, Yudong

2014-09-01

231

Fluorescence diagnosis of the status of the human lens in vivo  

NASA Astrophysics Data System (ADS)

We have studied fluorescence spectra of the human lens in vivo for healthy eyes and in different stages of senile cataract development. We propose a spectral criterion, the lens opacity index, allowing us to differentiate between stages of cataract development. We show a high correlation between the stage of cataract development and the opacity index. We propose an empirical expression for determining the stage of senile cataract development from the value of the lens opacity index. The technique has been clinically tested.

Vladimirova, E. S.; Salmin, V. V.; Salmina, A. B.; Oskirko, S. A.; Lazarenko, V. I.; Provorov, A. S.

2012-03-01

232

In vivo optical imaging of brain tumors and arthritis using fluorescent SapC-DOPS nanovesicles.  

PubMed

We describe a multi-angle rotational optical imaging (MAROI) system for in vivo monitoring of physiopathological processes labeled with a fluorescent marker. Mouse models (brain tumor and arthritis) were used to evaluate the usefulness of this method. Saposin C (SapC)-dioleoylphosphatidylserine (DOPS) nanovesicles tagged with CellVue Maroon (CVM) fluorophore were administered intravenously. Animals were then placed in the rotational holder (MARS) of the in vivo imaging system. Images were acquired in 10° steps over 380°. A rectangular region of interest (ROI) was placed across the full image width at the model disease site. Within the ROI, and for every image, mean fluorescence intensity was computed after background subtraction. In the mouse models studied, the labeled nanovesicles were taken up in both the orthotopic and transgenic brain tumors, and in the arthritic sites (toes and ankles). Curve analysis of the multi angle image ROIs determined the angle with the highest signal. Thus, the optimal angle for imaging each disease site was characterized. The MAROI method applied to imaging of fluorescent compounds is a noninvasive, economical, and precise tool for in vivo quantitative analysis of the disease states in the described mouse models. PMID:24837630

Chu, Zhengtao; LaSance, Kathleen; Blanco, Victor; Kwon, Chang-Hyuk; Kaur, Balveen; Frederick, Malinda; Thornton, Sherry; Lemen, Lisa; Qi, Xiaoyang

2014-01-01

233

In Vivo Optical Imaging of Brain Tumors and Arthritis Using Fluorescent SapC-DOPS Nanovesicles  

PubMed Central

We describe a multi-angle rotational optical imaging (MAROI) system for in vivo monitoring of physiopathological processes labeled with a fluorescent marker. Mouse models (brain tumor and arthritis) were used to evaluate the usefulness of this method. Saposin C (SapC)-dioleoylphosphatidylserine (DOPS) nanovesicles tagged with CellVue Maroon (CVM) fluorophore were administered intravenously. Animals were then placed in the rotational holder (MARS) of the in vivo imaging system. Images were acquired in 10° steps over 380°. A rectangular region of interest (ROI) was placed across the full image width at the model disease site. Within the ROI, and for every image, mean fluorescence intensity was computed after background subtraction. In the mouse models studied, the labeled nanovesicles were taken up in both the orthotopic and transgenic brain tumors, and in the arthritic sites (toes and ankles). Curve analysis of the multi angle image ROIs determined the angle with the highest signal. Thus, the optimal angle for imaging each disease site was characterized. The MAROI method applied to imaging of fluorescent compounds is a noninvasive, economical, and precise tool for in vivo quantitative analysis of the disease states in the described mouse models. PMID:24837630

Chu, Zhengtao; LaSance, Kathleen; Blanco, Victor; Kwon, Chang-Hyuk; Kaur, Balveen; Frederick, Malinda; Thornton, Sherry; Lemen, Lisa; Qi, Xiaoyang

2014-01-01

234

Exploiting post-transcriptional regulation to probe RNA structures in vivo via fluorescence  

PubMed Central

While RNA structures have been extensively characterized in vitro, very few techniques exist to probe RNA structures inside cells. Here, we have exploited mechanisms of post-transcriptional regulation to synthesize fluorescence-based probes that assay RNA structures in vivo. Our probing system involves the co-expression of two constructs: (i) a target RNA and (ii) a reporter containing a probe complementary to a region in the target RNA attached to an RBS-sequestering hairpin and fused to a sequence encoding the green fluorescent protein (GFP). When a region of the target RNA is accessible, the area can interact with its complementary probe, resulting in fluorescence. By using this system, we observed varied patterns of structural accessibility along the length of the Tetrahymena group I intron. We performed in vivo DMS footprinting which, along with previous footprinting studies, helped to explain our probing results. Additionally, this novel approach represents a valuable tool to differentiate between RNA variants and to detect structural changes caused by subtle mutations. Our results capture some differences from traditional footprinting assays that could suggest that probing in vivo via oligonucleotide hybridization facilitates the detection of folding intermediates. Importantly, our data indicate that intracellular oligonucleotide probing can be a powerful complement to existing RNA structural probing methods. PMID:25416800

Sowa, Steven W.; Vazquez-Anderson, Jorge; Clark, Chelsea A.; De La Peña, Ricardo; Dunn, Kaitlin; Fung, Emily K.; Khoury, Mark J.; Contreras, Lydia M.

2015-01-01

235

A Ga(3+)self-assembled fluorescent probe for ATP imaging in vivo.  

PubMed

Adenosine 5'-triphosphate (ATP) is a functional molecule associated with many important biological processes. Fluorescent detection methods for ATP with facile performance and high selectivity are in demand. One of the possible multi-membered arrays assembled between DHBO and Ga(3+) ions was conducted in aqueous solution, which can selectively recognize ATP with fluorescence enhancement from ADP, AMP and other structurally similar nucleoside triphosphates in vitro and in vivo. ATP facilitates the interaction between DHBO and Ga(3+) ions, resulting in the fluorescence increase. The detection limit for ATP was calculated to be 5.49×10(-7)M, which is much lower than that of intracellular concentrations (1-10mM). In addition, DHBO-Ga(3+) can be applied to detect ATP-relevant enzyme activity. PMID:25461153

Xiao, Liangliang; Sun, Shiguo; Pei, Zhichao; Pei, Yuxin; Pang, Yi; Xu, Yongqian

2014-10-18

236

Photoacoustic imaging of the near-infrared fluorescent protein iRFP in vivo  

NASA Astrophysics Data System (ADS)

Genetically encoded probes powerfully and non-invasively target specific tissues, cells, and subcellular locations. iRFP, a novel near-infrared fluorescent protein with low quantum yield whose absorption and fluorescence maxima are located at wavelengths longer than the Q-band of hemoglobin absorption, is ideal for PAT. Here, we report on an in vitro comparison of iRFP with other far-red fluorescent proteins, and its use in imaging a mouse tumor xenograft model. In an in vivo experiment, we stably transfected iRFP into MTLn3 adenocarcinoma cells and injected them into the mammary fat pad of female SCID/NCr mice, then imaged the resulting tumors two and three weeks post injection. The contrast increase from the protein expression was high enough to clearly separate the tumor region from the rest of the animal.

Krumholz, Arie; Filonov, Grigory S.; Xia, Jun; Yao, Junjie; Verkhusha, Vladislav V.; Wang, Lihong V.

2012-02-01

237

In vivo self-bio-imaging of tumors through in situ biosynthesized fluorescent gold nanoclusters  

PubMed Central

Fluorescence imaging in vivo allows non-invasive tumor diagnostic thus permitting a direct monitoring of cancer therapies progresses. It is established herein that fluorescent gold nanoclusters are spontaneously biosynthesized by cancerous cell (i.e., HepG2, human hepatocarcinoma cell line; K562, leukemia cell line) incubated with micromolar chloroauric acid solutions, a biocompatible molecular Au(III) species. Gold nanoparticles form by Au(III) reduction inside cells cytoplasms and ultimately concentrate around their nucleoli, thus affording precise cell imaging. Importantly, this does not occur in non-cancerous cells, as evidenced with human embryo liver cells (L02) used as controls. This dichotomy is exploited for a new strategy for in vivo self-bio-imaging of tumors. Subcutaneous injections of millimolar chloroauric acid solution near xenograft tumors of the nude mouse model of hepatocellular carcinoma or chronic myeloid leukemia led to efficient biosynthesis of fluorescent gold nanoclusters without significant dissemination to the surrounding normal tissues, hence allowing specific fluorescent self-bio-marking of the tumors. PMID:23362457

Wang, Jianling; Zhang, Gen; Li, Qiwei; Jiang, Hui; Liu, Chongyang; Amatore, Christian; Wang, Xuemei

2013-01-01

238

MR and fluorescence imaging of doxorubicin loaded nanoparticles using a novel in vivo model  

PubMed Central

We report here the in vivo combined-modality imaging of multifunctional drug delivery nanoparticles. These dextran core-based stealth liposomal nanoparticles (nanosomes) contained doxorubicin, iron oxide for MRI contrast, and Bodipy for fluorescence. The particles were long-lived in vivo due to surface decoration with polyethylene glycol (PEG) and the incorporation of acetylated lipids which were UV cross-linked for physical stability. We developed a rodent dorsal skinfold window chamber which facilitated both MRI and non-destructive optical imaging of nanoparticle accumulation in the same tumors. Chamber tumors were genetically labeled with DsRed-2 that enabled the MR images, the red fluorescence of the tumor, and the blue fluorescence of the nanoparticles to be co-localized. The nanoparticle design and MR imaging developed with the window chamber were then extended to orthotopic pancreatic tumors expressing DsRed-2. The tumors were MR imaged using iron oxide-dextran liposomes and by fluorescence to demonstrate the deep imaging capability of these nanoparticles. PMID:20599526

Erten, Ahmet; Wrasidlo, Wolf; Scadeng, Miriam; Esener, Sadik; Hoffman, Robert; Bouvet, Michael; Makale, Milan

2010-01-01

239

In vivo self-bio-imaging of tumors through in situ biosynthesized fluorescent gold nanoclusters.  

PubMed

Fluorescence imaging in vivo allows non-invasive tumor diagnostic thus permitting a direct monitoring of cancer therapies progresses. It is established herein that fluorescent gold nanoclusters are spontaneously biosynthesized by cancerous cell (i.e., HepG2, human hepatocarcinoma cell line; K562, leukemia cell line) incubated with micromolar chloroauric acid solutions, a biocompatible molecular Au(III) species. Gold nanoparticles form by Au(III) reduction inside cells cytoplasms and ultimately concentrate around their nucleoli, thus affording precise cell imaging. Importantly, this does not occur in non-cancerous cells, as evidenced with human embryo liver cells (L02) used as controls. This dichotomy is exploited for a new strategy for in vivo self-bio-imaging of tumors. Subcutaneous injections of millimolar chloroauric acid solution near xenograft tumors of the nude mouse model of hepatocellular carcinoma or chronic myeloid leukemia led to efficient biosynthesis of fluorescent gold nanoclusters without significant dissemination to the surrounding normal tissues, hence allowing specific fluorescent self-bio-marking of the tumors. PMID:23362457

Wang, Jianling; Zhang, Gen; Li, Qiwei; Jiang, Hui; Liu, Chongyang; Amatore, Christian; Wang, Xuemei

2013-01-01

240

In vivo self-bio-imaging of tumors through in situ biosynthesized fluorescent gold nanoclusters  

NASA Astrophysics Data System (ADS)

Fluorescence imaging in vivo allows non-invasive tumor diagnostic thus permitting a direct monitoring of cancer therapies progresses. It is established herein that fluorescent gold nanoclusters are spontaneously biosynthesized by cancerous cell (i.e., HepG2, human hepatocarcinoma cell line; K562, leukemia cell line) incubated with micromolar chloroauric acid solutions, a biocompatible molecular Au(III) species. Gold nanoparticles form by Au(III) reduction inside cells cytoplasms and ultimately concentrate around their nucleoli, thus affording precise cell imaging. Importantly, this does not occur in non-cancerous cells, as evidenced with human embryo liver cells (L02) used as controls. This dichotomy is exploited for a new strategy for in vivo self-bio-imaging of tumors. Subcutaneous injections of millimolar chloroauric acid solution near xenograft tumors of the nude mouse model of hepatocellular carcinoma or chronic myeloid leukemia led to efficient biosynthesis of fluorescent gold nanoclusters without significant dissemination to the surrounding normal tissues, hence allowing specific fluorescent self-bio-marking of the tumors.

Wang, Jianling; Zhang, Gen; Li, Qiwei; Jiang, Hui; Liu, Chongyang; Amatore, Christian; Wang, Xuemei

2013-01-01

241

Fluorescent Nanorods and Nanospheres for Real-Time In Vivo Probing of Nanoparticle Shape-Dependent Tumor Penetration  

E-print Network

Shape dependent: Fluorescent quantum-dot-based nanospheres and nanorods with identical hydrodynamic size and surface properties but different aspect ratios were developed for real-time in vivo tumor imaging. The nanorods ...

Chauhan, Vikash P.

242

A fluorescence spectroscopy study of traditional Chinese medicine Angelica  

NASA Astrophysics Data System (ADS)

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

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

2013-10-01

243

Quantitation of ten 30S ribosomal assembly intermediates using fluorescence triple correlation spectroscopy  

PubMed Central

The self-assembly of bacterial 30S ribosomes involves a large number of RNA folding and RNA-protein binding steps. The sequence of steps determines the overall assembly mechanism and the structure of the mechanism has ramifications for the robustness of biogenesis and resilience against kinetic traps. Thermodynamic interdependencies of protein binding inferred from omission-reconstitution experiments are thought to preclude certain assembly pathways and thus enforce ordered assembly, but this concept is at odds with kinetic data suggesting a more parallel assembly landscape. A major challenge is deconvolution of the statistical distribution of intermediates that are populated during assembly at high concentrations approaching in vivo assembly conditions. To specifically resolve the intermediates formed by binding of three ribosomal proteins to the full length 16S rRNA, we introduce Fluorescence Triple-Correlation Spectroscopy (F3CS). F3CS identifies specific ternary complexes by detecting coincident fluctuations in three-color fluorescence data. Triple correlation integrals quantify concentrations and diffusion kinetics of triply labeled species, and F3CS data can be fit alongside auto-correlation and cross-correlation data to quantify the populations of 10 specific ribosome assembly intermediates. The distribution of intermediates generated by binding three ribosomal proteins to the entire native 16S rRNA included significant populations of species that were not previously thought to be thermodynamically accessible, questioning the current interpretation of the classic omission-reconstitution experiments. F3CS is a general approach for analyzing assembly and function of macromolecular complexes, especially those too large for traditional biophysical methods. PMID:22869699

Ridgeway, William K.; Millar, David P.; Williamson, James R.

2012-01-01

244

Determination of the PSI/PSII ratio in living plant cells at room temperature by spectrally resolved fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Leaf cells of living plants exhibit strong fluorescence from chloroplasts, the reaction centers of photosynthesis. Mutations in the photosystems change their structure and can, thus, be monitored by recording the fluorescence spectra of the emitted chlorophyll light. These measurements have, up to now, mostly been carried out at low temperatures (77 K), as these conditions enable the differentiation between the fluorescence of Photosystem I (PSI) and Photosystem II (PSII). In contrast, at room temperature, energy transfer processes between the various photosynthetic complexes result in very similar fluorescence emissions, which mainly consist of fluorescence photons emitted by PSII hindering a discrimination based on spectral ROIs (regions of interest). However, by statistical analysis of high resolution fluorescence spectra recorded at room temperature, it is possible to draw conclusions about the relative PSI/PSII ratio. Here, the possibility of determining the relative PSI/PSII ratio by fluorescence spectroscopy is demonstrated in living maize plants. Bundle-sheath chloroplasts of mature maize plants have a special morphologic characteristic; they are agranal, or exhibit only rudimentary grana, respectively. These chloroplasts are depleted in PSII activity and it could be shown that PSII is progressively reduced during leaf differentiation. A direct comparison of PSII activity in isolated chloroplasts is nearly impossible, since the activity of PSII in both mesophyll- and bundle-sheath chloroplasts decays with time after isolation and it takes significantly longer to isolate bundle-sheath chloroplasts. Considering this fact the measurement of PSI/PSII ratios with the 77K method, which includes taking fluorescence spectra from a diluted suspension of isolated chloroplasts at 77K, is questionable. These spectra are then used to analyze the distribution of energy between PSI and PSII. After rapid cooling to 77K secondary biochemical influences, which attenuate the fluorescence emanated from PSI, are frozen out. Due to their characteristic morphology, maize chloroplasts of mesophyll and bundle-sheath cells are an appropriate system for demonstrating the applicability of our in vivo method which, unlike the common 77K method, does not require the isolation of chloroplasts. In mesophyll chloroplasts of higher land plants, the thylakoids have a heterogenic morphology of appressed and non-appressed membrane domains, called the grana and the stroma lamellae. PSII is enriched in the grana, whereas PSI is enriched in the stroma lamellae. Changes in chloroplast membrane structure and composition, according to changes in the PSI/ PSII ratio, can be triggered by light quality and carbon source deficiency. Here, we demonstrate the applicability of statistical analysis of fluorescence spectra to detect changes in the PSI/PSII ratio resulting from structure changes in the thylakoid membrane.

Elgass, Kirstin; Zell, Martina; Maurino, Veronica G.; Schleifenbaum, Frank

2011-02-01

245

Subcellular In Vivo 1 H MR Spectroscopy of Xenopus laevis Oocytes  

E-print Network

to allow the recording of the first compartment-selective in vivo MR spectra from the animal and vegetal properties, diffusion or flow inside an animal or human body can be obtained with magnetic resonance imagingSubcellular In Vivo 1 H MR Spectroscopy of Xenopus laevis Oocytes Seung-Cheol Lee,* Jee-Hyun Cho

Hammerton, James

246

In vivo tomographic imaging of deep-seated cancer using fluorescence lifetime contrast.  

PubMed

Preclinical cancer research would benefit from noninvasive imaging methods that allow tracking and visualization of early-stage metastasis in vivo. Although fluorescent proteins revolutionized intravital microscopy, two major challenges that still remain are tissue autofluorescence and hemoglobin absorption, which act to limit intravital optical techniques to large or subcutaneous tumors. Here, we use time-domain (TD) technology for the effective separation of tissue autofluorescence from extrinsic fluorophores, based on their distinct fluorescence lifetimes. In addition, we use cancer cells labeled with near infrared fluorescent proteins (iRFP) to allow deep-tissue imaging. Our results demonstrate that TD imaging allows the detection of metastasis in deep-seated organs of living mice with a more than 20-fold increase in sensitivity compared with conventional continuous wave techniques. Furthermore, the distinct fluorescence lifetimes of iRFPs enable lifetime multiplexing of three different tumors, each expressing unique iRFP labels in the same animal. Fluorescence tomographic reconstructions reveal three-dimensional distributions of iRFP720-expressing cancer cells in lungs and brain of live mice, allowing ready longitudinal monitoring of cancer cell fate with greater sensitivity than otherwise currently possible. Cancer Res; 75(7); 1236-43. ©2015 AACR. PMID:25670171

Rice, William L; Shcherbakova, Daria M; Verkhusha, Vladislav V; Kumar, Anand T N

2015-04-01

247

Sensitive ?-galactosidase-targeting fluorescence probe for visualizing small peritoneal metastatic tumours in vivo  

PubMed Central

Fluorescence-guided diagnostics is one of the most promising approaches for facile detection of cancer in situ. Here we focus on ?-galactosidase, which is overexpressed in primary ovarian cancers, as a molecular target for visualizing peritoneal metastases from ovarian cancers. As existing fluorescence probes are unsuitable, we have designed membrane-permeable HMRef-?Gal, in which the optimized intramolecular spirocyclic function affords >1,400-fold fluorescence enhancement on activation. We confirm that HMRef-?Gal sensitively detects intracellular ?-galactosidase activity in several ovarian cancer lines. In vivo, this probe visualizes metastases as small as <1?mm in diameter in seven mouse models of disseminated human peritoneal ovarian cancer (SHIN3, SKOV3, OVK18, OVCAR3, OVCAR4, OVCAR5 and OVCAR8). Because of its high brightness, real-time detection of metastases with the naked eye is possible. Endoscopic fluorescence detection of metastases is also demonstrated. The results clearly indicate preclinical potential value of the probe for fluorescence-guided diagnosis of peritoneal metastases from ovarian cancers. PMID:25765713

Asanuma, Daisuke; Sakabe, Masayo; Kamiya, Mako; Yamamoto, Kyoko; Hiratake, Jun; Ogawa, Mikako; Kosaka, Nobuyuki; Choyke, Peter L.; Nagano, Tetsuo; Kobayashi, Hisataka; Urano, Yasuteru

2015-01-01

248

Sensitive ?-galactosidase-targeting fluorescence probe for visualizing small peritoneal metastatic tumours in vivo.  

PubMed

Fluorescence-guided diagnostics is one of the most promising approaches for facile detection of cancer in situ. Here we focus on ?-galactosidase, which is overexpressed in primary ovarian cancers, as a molecular target for visualizing peritoneal metastases from ovarian cancers. As existing fluorescence probes are unsuitable, we have designed membrane-permeable HMRef-?Gal, in which the optimized intramolecular spirocyclic function affords >1,400-fold fluorescence enhancement on activation. We confirm that HMRef-?Gal sensitively detects intracellular ?-galactosidase activity in several ovarian cancer lines. In vivo, this probe visualizes metastases as small as <1?mm in diameter in seven mouse models of disseminated human peritoneal ovarian cancer (SHIN3, SKOV3, OVK18, OVCAR3, OVCAR4, OVCAR5 and OVCAR8). Because of its high brightness, real-time detection of metastases with the naked eye is possible. Endoscopic fluorescence detection of metastases is also demonstrated. The results clearly indicate preclinical potential value of the probe for fluorescence-guided diagnosis of peritoneal metastases from ovarian cancers. PMID:25765713

Asanuma, Daisuke; Sakabe, Masayo; Kamiya, Mako; Yamamoto, Kyoko; Hiratake, Jun; Ogawa, Mikako; Kosaka, Nobuyuki; Choyke, Peter L; Nagano, Tetsuo; Kobayashi, Hisataka; Urano, Yasuteru

2015-01-01

249

Fluorescence spectra of blood plasma treated with ultraviolet irradiation in vivo  

NASA Astrophysics Data System (ADS)

We have studied the fluorescence spectra of blood plasma from patients with acute coronary syndrome, and also the effect of therapeutic doses of in vivo ultraviolet blood irradiation (UBI) on the spectra. We have established that the maxima in the fluorescence spectra of the original plasma samples, obtained from unirradiated blood, are located in the wavelength interval 330-340 nm, characteristic for the fluorescence of tryptophan residues. In extracorporeal UBI ( ? = 254 nm), we observed changes in the shape and also both a blue and a red shift in the maxima of the fluorescence spectra, differing in magnitude for blood plasma samples from different patients in the test group. We show that UBI-initiated changes in the fluorescence spectra of the plasma depend on the original pathological disturbances of metabolite levels, and also on the change in the oxygen-transport function of the blood and the acid-base balance, affecting the oxidative stability of the plasma. We have concluded that UV irradiation, activating buffer systems in the blood, has an effect on the universal and specific interactions of the tryptophan residue with the amino acid residues and water surrounding it.

Zalesskaya, G. A.; Maslova, T. O.

2010-09-01

250

In vivo fluorescence enhanced optical tomography reconstruction of lung cancer of non immersed small animals  

NASA Astrophysics Data System (ADS)

Fluorescence enhanced diffuse optical tomography (fDOT) is envisioned to be useful to collect functional information from small animal models. For oncology applications, cancer-targeted fluorescent markers can be used as a surrogate of the cancer activity. We are developing a continuous wave fDOT bench intended to be integrated in systems dedicated to whole body small animal fluorescence analyses. The focus is currently put on the reconstruction of non immersed small animals imaged by a CCD camera. The reconstruction stage already corrects the tissue heterogeneity artifacts through the computation of an optical heterogeneity map. We will show how this formalism coupled with the determination of the animal boundaries performed by a laser scanner, can be used to manage non contact acquisitions. The time of reconstruction for a 10 × 9 laser source positions, 45 × 40 detector elements and 14 × 11 × 14 mesh voxels is typically 10 minutes on a 3GHz PCs corresponding to the acquisition time allowing the two tasks to be performed in parallel. The system is validated on an in vivo experiment performed on three healthy nude mice and a mouse bearing a lung tumor at 10, 12 and 14 days after implantation allowing the follow up of the disease. The 3D fluorescence reconstructions of this mouse are presented and the total fluorescence amounts are compared.

Hervé, L.; Koenig, A.; Da Silva, A.; Berger, M.; Boutet, J.; Dinten, J. M.; Peltié, P.; Rizo, P.

2007-02-01

251

Anabaena cell ageing monitored with confocal fluorescence spectroscopy.  

PubMed

Cyanobacteria use a sophisticated system of pigments to collect light energy across the visible spectrum for photosynthesis. The pigments are assembled in structures called phycobilisomes, composed of phycoerythrocyanin, phycocyanin and allophycocyanin, which absorb energy and transfer it to chlorophyll in photosystem II reaction centres. All of the components of this system are fluorescent, allowing sensitive measurements of energy transfer using single cell confocal fluorescence microscopy. The native pigments can be interrogated without the use of reporters. Here, we use confocal fluorescence microscopy to monitor changes in the efficiency of energy transfer as single cells age, between the time they are born at cell division until they are ready to divide again. Alteration of fluorescence was demonstrated to change with the age of the cyanobacterial cell. PMID:25378560

Ke, Shan; Bindokas, Vytas; Haselkorn, Robert

2015-01-01

252

Laser-induced fluorescence spectroscopy for combustion diagnostics  

NASA Technical Reports Server (NTRS)

The types of spectroscopic and collisional measurements that are needed to develop laser-induced fluorescence as a diagnostic technique are discussed, with emphasis placed on combustion measurements. The spectroscopic measurements under collision-free conditions include production of radicals, excitation scan studies, lifetime measurements, and fluorescence scans. The collisional studies discussed here are quenching, energy transfer, and polarization phenomena. The results of recent laboratory experiments are presented.

Crosley, D. R.; Smith, G. P.

1983-01-01

253

Ultrafast Fluorescence Spectroscopy via Upconversion: Applications to Biophysics  

PubMed Central

This chapter reviews basic concepts of nonlinear fluorescence upconversion, a technique whose temporal resolution is essentially limited only by the pulse width of the ultrafast laser. Design aspects for upconversion spectrophotofluorometers are discussed, and a recently developed system is described. We discuss applications in biophysics, particularly the measurement of time-resolved fluorescence spectra of proteins (with subpicosecond time resolution). Application of this technique to biophysical problems such as dynamics of tryptophan, peptides, proteins, and nucleic acids is reviewed. PMID:19152860

Xu, Jianhua; Knutson, Jay R.

2012-01-01

254

Fluorescence spectroscopy of anisole at elevated temperatures and pressures  

NASA Astrophysics Data System (ADS)

Laser-induced fluorescence of anisole as tracer of isooctane at an excitation wavelength of 266 nm was investigated for conditions relevant to rapid compression machine studies and for more general application of internal combustion engines regarding temperature, pressure, and ambient gas composition. An optically accessible high pressure and high temperature chamber was operated by using different ambient gases (Ar, N2, CO2, air, and gas mixtures). Fluorescence experiments were investigated at a large range of pressure and temperature (0.2-4 MPa and 473-823 K). Anisole fluorescence quantum yield decreases strongly with temperature for every considered ambient gas, due to efficient radiative mechanisms of intersystem crossing. Concerning the pressure effect, the fluorescence signal decreases with increasing pressure, because increasing the collisional rate leads to more important non-radiative collisional relaxation. The quenching effect is strongly efficient in oxygen, with a fluorescence evolution described by Stern-Volmer relation. The dependence of anisole fluorescence versus thermodynamic parameters suggests the use of this tracer for temperature imaging in specific conditions detailed in this paper. The calibration procedure for temperature measurements is established for the single-excitation wavelength and two-color detection technique.

Tran, K. H.; Morin, C.; Kühni, M.; Guibert, P.

2014-06-01

255

Probing the extracellular diffusion of antibodies in brain using in vivo integrative optical imaging and ex vivo fluorescence imaging.  

PubMed

Antibody-based therapeutics exhibit great promise in the treatment of central nervous system (CNS) disorders given their unique customizable properties. Although several clinical trials have evaluated therapeutic antibodies for treatment of CNS disorders, success to date has likely been limited in part due to complex issues associated with antibody delivery to the brain and antibody distribution within the CNS compartment. Major obstacles to effective CNS delivery of full length immunoglobulin G (IgG) antibodies include transport across the blood-brain and blood-cerebrospinal fluid barriers. IgG diffusion within brain extracellular space (ECS) may also play a role in limiting central antibody distribution; however, IgG transport in brain ECS has not yet been explored using established in vivo methods. Here, we used real-time integrative optical imaging to measure the diffusion properties of fluorescently labeled, non-targeted IgG after pressure injection in both free solution and in adult rat neocortex in vivo, revealing IgG diffusion in free medium is ~10-fold greater than in brain ECS. The pronounced hindered diffusion of IgG in brain ECS is likely due to a number of general factors associated with the brain microenvironment (e.g. ECS volume fraction and geometry/width) but also molecule-specific factors such as IgG size, shape, charge and specific binding interactions with ECS components. Co-injection of labeled IgG with an excess of unlabeled Fc fragment yielded a small yet significant increase in the IgG effective diffusion coefficient in brain, suggesting that binding between the IgG Fc domain and endogenous Fc-specific receptors may contribute to the hindered mobility of IgG in brain ECS. Importantly, local IgG diffusion coefficients from integrative optical imaging were similar to those obtained from ex vivo fluorescence imaging of transport gradients across the pial brain surface following controlled intracisternal infusions in anesthetized animals. Taken together, our results confirm the importance of diffusive transport in the generation of whole brain distribution profiles after infusion into the cerebrospinal fluid, although convective transport in the perivascular spaces of cerebral blood vessels was also evident. Our quantitative in vivo diffusion measurements may allow for more accurate prediction of IgG brain distribution after intrathecal or intracerebroventricular infusion into the cerebrospinal fluid across different species, facilitating the evaluation of both new and existing strategies for CNS immunotherapy. PMID:25449807

Wolak, Daniel J; Pizzo, Michelle E; Thorne, Robert G

2015-01-10

256

In vivo optical imaging of dihydroethidium oxidation in the mouse brain employing fluorescence intensity and lifetime contrast  

Microsoft Academic Search

Reactive oxygen species (ROS) are believed to be involved in many diseases and injuries to the brain, but the molecular processes are not well understood due to a lack of in vivo imaging techniques to evaluate ROS. The fluorescent oxidation products of dihydroethidium (DHE) can monitor ROS production in vivo. Here we demonstrate the novel optical imaging of brain in

David J. Hall; Sung-Ho Han; Laura Dugan

2009-01-01

257

Quantification of in vivo Magnetic Resonance Spectroscopy signals with baseline and lineshape  

E-print Network

Quantification of in vivo Magnetic Resonance Spectroscopy signals with baseline and lineshape Nuclear, Magnetic Resonance Unit, Katholieke Universiteit Leuven, Leuven, Belgium Email:maria.osorio@esat.kuleuven.be Abstract--Quantification of Magnetic Resonance Spectroscopy (MRS) signals is a method to estimate

258

Elimination of autofluorescence in fluorescence Correlation spectroscopy by using the AzaDiOxaTriAngulenium (ADOTA) fluorophore in combination with time correlated single photon counting (TCSPC)  

PubMed Central

Fluorescence Correlation Spectroscopy (FCS) is a frequently applied technique that allows for precise and sensitive analyses of molecular diffusion and interactions. However, the potential of FCS for in vitro or ex vivo studies has not been fully realized due in part to artifacts originating from autofluorescence (fluorescence of inherent components and fixative-induced fluorescence). Here, we propose the azadioxatriangulenium (ADOTA) dye as a solution to this problem. The lifetime of the ADOTA probe, about 19.4 ns, is much longer than most components of autofluorescence. Thus, it can be easily separated by time correlated single photon counting (TCSPC) methods. Here, we demonstrate the suppression of autofluorescence in FCS by using ADOTA labeled Hyaluronan macromolecules (HAs) with Rhodamine 123 added to simulate diffusing fluorescent background components. The emission spectrum and decay rate of Rhodamine 123 overlap with the usual sources of autofluorescence, and its diffusion behavior is well known. We show that the contributions from Rhodamine 123 can be eliminated by time-gating or by fluorescence lifetime correlation spectroscopy (FLCS). While the pairing of ADOTA and time-gating is an effective strategy for the removal of autofluorescence from fluorescence imaging, the loss of photons leads to erroneous concentration values with FCS. On the other hand, FLCS eliminates autofluorescence without such errors. We then show that both time gating and FLCS may be used successfully with ADOTA-labeled HA to detect the presence of hyaluronidase, the over-expression of which has been observed in many types of cancer. PMID:23564284

Rich, Ryan M.; Mummert, Mark; Gryczynski, Zygmunt; Borejdo, Julian; Sørensen, Thomas Just; Laursen, Bo W.; Foldes-Papp, Zeno; Gryczynski, Ignacy; Fudala, Rafal

2013-01-01

259

Europium Uptake and Partitioning in Oat (Avena sativa) Roots as studied By Laser-Induced Fluorescence Spectroscopy and Confocal Microscopy Profiling Technique  

SciTech Connect

The uptake of Eu3+ by elongating oat plant roots was studied by fluorescence spectroscopy, fluorescence lifetime measurement, as well as laser excitation time-resolved confocal fluorescence profiling technique. The results of this work indicated that the initial uptake of Eu(III) by oat root was most evident within the apical meristem of the root just proximal to the root cap. Distribution of assimilated Eu(III) within the roots differentiation and elongation zone was non-uniform. Higher concentrations were observed within the vascular cylinder, specifically in the phloem and developing xylem parenchyma. Elevated levels of the metal were also observed in the root hairs of the mature root. The concentration of assimilated Eu3+ dropped sharply from the apical meristem to the differentiation and elongation zone and then gradually decreased as the distance from the root cap increased. Fluorescence spectroscopic characteristics of the assimilated Eu3+ suggested that the Eu3+ exists a s inner-sphere mononuclear complexes inside the root. This work has also demonstrated the effectiveness of a time-resolved Eu3+ fluorescence spectroscopy and confocal fluorescence profiling techniques for the in vivo, real-time study of metal[Eu3+] accumulation by a functioning intact plant root. This approach can prove valuable for basic and applied studies in plant nutrition and environmental uptake of actinide radionuclides.

Fellows, Robert J.; Wang, Zheming; Ainsworth, Calvin C.

2003-11-15

260

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

NASA Astrophysics Data System (ADS)

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

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

2014-09-01

261

In vitro and in vivo demonstrations of Fluorescence by Unbound Excitation from Luminescence (FUEL).  

PubMed

Bioluminescence imaging is a powerful technique that allows for deep-tissue analysis in living, intact organisms. However, in vivo optical imaging is compounded by difficulties due to light scattering and absorption. While light scattering is relatively difficult to overcome and compensate, light absorption by biological tissue is strongly dependent upon wavelength. For example, light absorption by mammalian tissue is highest in the blue-yellow part of the visible energy spectrum. Many natural bioluminescent molecules emit photonic energy in this range, thus in vivo optical detection of these molecules is primarily limited by absorption. This has driven efforts for probe development aimed to enhance photonic emission of red light that is absorbed much less by mammalian tissue using either direct genetic manipulation, and/or resonance energy transfer methods. Here we describe a recently identified alternative approach termed Fluorescence by Unbound Excitation from Luminescence (FUEL), where bioluminescent molecules are able to induce a fluorescent response from fluorescent nanoparticles through an epifluorescence mechanism, thereby significantly increasing both the total number of detectable photons as well as the number of red photons produced. PMID:24166383

Dragavon, Joe; Rekiki, Abdessalem; Theodorou, Ioanna; Samson, Chelsea; Blazquez, Samantha; Rogers, Kelly L; Tournebize, Régis; Shorte, Spencer

2014-01-01

262

An in vivo study of the microlymphatics in psoriasis using fluorescence microlymphography.  

PubMed

Angiogenesis is a recognized event in psoriasis. Previous ultrastructural studies have demonstrated lymphatic capillaries extending high into the dermal papillae. Using the novel method of fluorescence microlymphography which permits visualization of upper dermal initial lymphatics in vivo we tested the hypothesis that lymphangiogenesis exists within plaque psoriasis. Six patients underwent fluorescence microlymphography with fluorescein isothiocyanate-dextran administered intracutaneously within a psoriatic plaque on the leg. Stereological analysis permitted quantification of the lymphatic network opacified both within (lesional) and without (perilesional) the plaque. Results showed a greater spread of tracer from the depot into perilesional skin than into the plaque (P < 0.006). The mean length of lymphatics per unit area at increasing distance from the centre of the depot was also increased for the perilesional skin, 10.5 +/- 1.9/cm2 (mean +/- SEM), compared with lesional skin, 3.06 +/- 0.8/cm (P < 0.001). The cumulative lymphatic length was also greater in perilesional, 22 +/- 7.3 cm2, compared with lesional skin, 3.6 +/- 0.3 cm (P < 0.006). Fluorescence microlymphography has proved to be an effective in vivo technique for the assessment of the dermal microlymphatics in psoriasis. Stereology provided quantitative analysis of the lymphatic network visualized. Overall, there is a greater network of lymphatics in perilesional compared with lesional skin in patients with plaque psoriasis. This finding is at odds with the accepted view that the lymphatic dermal vessels are increased within the psoriatic plaque. PMID:10215769

Cliff, S; Bedlow, A J; Stanton, A W; Mortimer, P S

1999-01-01

263

Improved diffuse fluorescence flow cytometer prototype for high sensitivity detection of rare circulating cells in vivo  

PubMed Central

Abstract. Detection and enumeration of rare circulating cells in mice are important problems in many areas of preclinical biomedical research. Recently, we developed a new method termed “diffuse fluorescence flow cytometry” (DFFC) that uses diffuse photons to increase the blood sampling volume and sensitivity versus existing in vivo flow cytometry methods. In this work, we describe a new DFFC prototype with approximately an order-of-magnitude improvement in sensitivity compared to our previous work. This sensitivity improvement is enabled by a number of technical innovations, which include a method for the removal of motion artifacts (allowing interrogation of mouse hindlegs that was less optically attenuating versus the tail) and improved collection optics and signal preamplification. We validated our system first in limb mimicking optical flow phantoms with fluorescent microspheres and then in nude mice with fluorescently labeled mesenchymal stem cells at injected concentrations of 5×103??cells/mL. In combination, these improvements resulted in an overall cell counting sensitivity of about 1??cell/mL or better in vivo. PMID:23831714

Pestana, Noah; Mortensen, Luke J.; Runnels, Judith M.; Vickers, Dwayne; Murthy, Shashi K.; Lin, Charles P.; Niedre, Mark

2013-01-01

264

A computer vision approach to rare cell in vivo fluorescence flow cytometry  

PubMed Central

Non-invasive enumeration of rare circulating cell populations in small animals is of great importance in many areas of biomedical research. In this work we describe a macroscopic fluorescence imaging system and automated computer vision algorithm that allows in vivo detection, enumeration and tracking of circulating fluorescently-labeled cells from multiple large blood vessels in the ear of a mouse. This imaging system uses a 660 nm laser and a high sensitivity electron-multiplied charge coupled device camera (EMCCD) to acquire fluorescence image sequences from relatively large (~5 × 5 mm2) imaging areas. The primary technical challenge was developing an automated method for identifying and tracking rare cell events in image sequences with substantial autofluorescence and noise content. To achieve this, we developed a two-step image analysis algorithm that first identified cell candidates in individual frames, and then merged cell candidates into tracks by dynamic analysis of image sequences. The second step was critical since it allowed rejection of >97% of false positive cell counts. Overall, our computer vision IVFC (CV-IVFC) approach allows single-cell detection sensitivity at estimated concentrations of 20 cells per mL of peripheral blood. In addition to simple enumeration, the technique recovers the cell’s trajectory, which in the future could be used to automatically identify, for example, in vivo homing and docking events. PMID:24273157

Markovic, Stacey; Li, Binlong; Pera, Vivian; Sznaier, Mario; Camps, Octavia; Niedre, Mark

2014-01-01

265

Noninvasive and quantitative assessment of in vivo fetomaternal interface angiogenesis using RGD-based fluorescence.  

PubMed

Angiogenesis is a key process for proper placental development and for the success of pregnancy. Although numerous in vitro methods have been developed for the assessment of this process, relatively few reliable in vivo methods are available to evaluate this activity throughout gestation. Here we report an in vivo technique that specifically measures placental neovascularization. The technique is based on the measurement of a fluorescent alpha v beta 3 (?v?3) integrin-targeting molecule called Angiolone-Alexa-Fluor 700. The ?v?3 integrin is highly expressed by endothelial cells during the neovascularization and by trophoblast cells during their invasion of the maternal decidua. Angiolone was injected to gravid mice at 6.5 and 11.5 days post coitus (dpc). The fluorescence was analyzed one day later at 7.5 and 12.5?dpc, respectively. We demonstrated that (i) Angiolone targets ?v?3 protein in the placenta with a strong specificity, (ii) this technique is quantitative as the measurement was correlated to the increase of the placental size observed with increasing gestational age, and (iii) information on the outcome is possible, as abnormal placentation could be detected early on during gestation. In conclusion, we report the validation of a new noninvasive and quantitative method to assess the placental angiogenic activity, in vivo. PMID:25110672

Keramidas, M; Lavaud, J; Sergent, F; Hoffmann, P; Brouillet, S; Feige, J-J; Coll, J-L; Alfaidy, N

2014-01-01

266

In vivo imaging of choroidal angiogenesis using fluorescence-labeled cationic liposomes  

PubMed Central

Purpose Precise monitoring of active angiogenesis in neovascular eye diseases such as age-related macular degeneration (AMD) enables sensitive use of antiangiogenic drugs and reduces adverse side effects. So far, no in vivo imaging methods are available to specifically label active angiogenesis. Here, we report such a technique using fluorophore-labeled cationic liposomes (CL) detected with a standard clinical in vivo scanning laser ophthalmoscope (SLO). Methods C57Bl/6 mice underwent laser coagulations at day 0 (d0) to induce choroidal neovascularization (CNV). Liposomes labeled with Oregon green, rhodamine (Rh), or indocyanine green (ICG) were injected into the tail vein at various time points after laser coagulation, and their fluorescence was observed in vivo 60 min later using an SLO, or afterwards in choroidal flatmounts or cryosections. Results SLO detected accumulated fluorescence only in active CNV lesions with insignificant background noise. The best signal was obtained with CL-ICG. Choroidal flatmounts and cryosections of the eye confirmed the location of retained CL in CNV lesions. Neutral liposomes, in contrast, showed no accumulation. Conclusions These results establish fluorophore-labeled CL as high affinity markers to selectively stain active CNV. This novel, non-invasive SLO imaging technique could improve risk assessment and indication for current intraocular antiangiogenic drugs in neovascular eye diseases, as well as monitor therapeutic outcomes. Labeling of angiogenic vessels using CL can be of interest not only for functional imaging in ophthalmology but also for other conditions where localization of active angiogenesis is desirable. PMID:22605917

Hua, Jing; Gross, Nikolai; Schulze, Brita; Michaelis, Uwe; Bohnenkamp, Hermann; Guenzi, Eric; Hansen, Lutz L.; Martin, Gottfried

2012-01-01

267

In vivo metabolic imaging of insulin with multiphoton fluorescence of human insulin-Au nanodots.  

PubMed

Functional human insulin-Au nanodots (NDs) are synthesized for the in vivo imaging of insulin metabolism. Benefiting from its efficient red to near infrared fluorescence, deep tissue subcellular uptake of insulin-Au NDs can be clearly resolved through a least-invasive harmonic generation and two-photon fluorescence (TPF) microscope. In vivo investigations on mice ear and ex vivo assays on human fat tissues conclude that cells with rich insulin receptors have higher uptake of administrated insulin. Interestingly, the insulin-Au NDs can even permeate into lipid droplets (LDs) of adipocytes. Using this newly discovered metabolic phenomenon of insulin, it is found that enlarged adipocytes in type II diabetes mice have higher adjacent/LD concentration contrast with small-sized ones in wild type mice. For human clinical samples, the epicardial adipocytes of patients with diabetes and coronary artery disease (CAD) also show elevated adjacent/LD concentration contrast. As a result, human insulin-Au nanodots provide a new approach to explore subcellular insulin metabolism in model animals or patients with metabolic or cardiovascular diseases. PMID:23172627

Liu, Chien-Liang; Liu, Tzu-Ming; Hsieh, Tsung-Yuan; Liu, Han-Wen; Chen, Yu-Shing; Tsai, Cheng-Kun; Chen, Hsieh-Chih; Lin, Jong-Wei; Hsu, Ron-Bin; Wang, Tzung-Dau; Chen, Chien-Cheng; Sun, Chi-Kuang; Chou, Pi-Tai

2013-06-24

268

Amplitude and Phase Fluorescence-Spectroscopy Methods for Dissolved Oxygen Concentration Evaluation: Comparative Practical Results  

Microsoft Academic Search

This paper shows the practical results from a detailed comparative study of amplitude and phase fluorescence-spectroscopy methods for dissolved oxygen concentration evaluation. These results were obtained with an implemented optoelectronic measurement system that guarantees near-optimal operation conditions for both methods and a commercial fluorescence optical-fiber sensor, which is excited by a continuous-regulated sinusoidal-amplitude modulated light beam. The comparison was made

Gustavo J. Grillo; Miguel A. Pérez; Marta Valledor; Rubén Ramos

2005-01-01

269

Surface intrinsic fluorescence spectroscopy of proteins using a UV linearly polarized pulsed laser beam  

NASA Astrophysics Data System (ADS)

The proposed new interfacial spectroscopic method allows to measure the fluorescence emission spectra of the two tryptophans of the Bovine Serum Albumin (BSA) during its adsorption at the air/water and egg-lecithin (egg-PC)/water interfaces, using an UV excimer laser. Surface fluorescence spectroscopy shows changes in the spectroscopic properties of adsorbed BSA, spread BSA and mixed egg-PC/BSA films. These results are related to the surface pressure measurements which characterize the different BSA surface organizations.

Yapoudjian, S.; Ivanova, M.; Uteza, Olivier P.; Marine, Vladimir I.; Sentis, Marc L.

2000-04-01

270

Attenuation-corrected fluorescence spectra unmixing for spectroscopy and microscopy.  

PubMed

In fluorescence measurements, light is often absorbed and scattered by a sample both for excitation and emission, resulting in the measured spectra to be distorted. Conventional linear unmixing methods computationally separate overlapping spectra but do not account for these effects. We propose a new algorithm for fluorescence unmixing that accounts for the attenuation-related distortion effect on fluorescence spectra. Using a matrix representation, we derive forward measurement formation and a corresponding inverse method; the unmixing algorithm is based on nonnegative matrix factorization. We also demonstrate how this method can be extended to a higher-dimensional tensor form, which is useful for unmixing overlapping spectra observed under the attenuation effect in spectral imaging microscopy. We evaluate the proposed methods in simulation and experiments and show that it outperforms a conventional, linear unmixing method when absorption and scattering contributes to the measured signals, as in deep tissue imaging. PMID:25321030

Ikoma, Hayato; Heshmat, Barmak; Wetzstein, Gordon; Raskar, Ramesh

2014-08-11

271

Challenges of Using MR Spectroscopy to Detect Neural Progenitor Cells In Vivo  

PubMed Central

SUMMARY A recent report of detection of neural progenitor cells (NPCs) in living human brain by using in vivo proton MR spectroscopy (1H-MR spectroscopy) has sparked great excitement in the field of biomedicine because of its potential influence and utility in clinical neuroscience research. On the other hand, the method used and the findings described in the report also caused heated debate and controversy. In this article, we will briefly detail the reasons for the debate and controversy from the point of view of the in vivo 1H-MR spectroscopy methodology and will propose some technical strategies in both data acquisition and data processing to improve the feasibility of detecting NPCs in future studies by using in vivo 1H-MR spectroscopy. PMID:19357383

Dong, Z.; Dreher, W.; Leibfritz, D.; Peterson, B.S.

2010-01-01

272

In vivo fluorescence and photodynamic activity of zinc phthalocyanine administered in liposomes.  

PubMed Central

Zinc(II) phthalocyanine, a hydrophobic photosensitiser, was incorporated in unilamellar liposomes and studied in vivo for fluorescence kinetics and photodynamic activity. An observation chamber mounted in a dorsal skinfold of female WAG/Rij rats was used as a model system. In the chamber, an isogeneic mammary carcinoma was transplanted in the subcutaneous tissue. Phthalocyanine fluorescence was excited at 610 nm with a power density of 0.25 mW cm-2 and was detected above 665 nm through a high-pass filter using a two-stage image intensifier coupled to a charge-coupled device (CCD) camera. Following i.v. administration of 0.14 mg kg-1 of the drug, the fluorescence pharmacokinetics of the dye in vasculature, normal tissue and tumour tissue was determined as a function of time. Tumour fluorescence increased slowly to a maximum about 3 h post injection (p.i.), and remained well above the normal tissue fluorescence till 24 h p.i. Fluorescence in the circulation was always stronger than in the tissues. A treatment light dose at a wavelength of 675 nm was delivered 24 h p.i. One group of six animals received a total light dose of 150 J cm-2 (100 mW cm-2). A second group of six animals received a total light dose of 450 J cm-2 at the same dose rate. Vascular damage resulting from treatment was observed only at the final stages of the irradiation, despite the relatively high levels of fluorescence in the circulation. Immediate post-treatment (re)transplantation of the content of the chamber into the flank always resulted in tumour regrowth, confirming the presence of viable tumour cells following photodynamic therapy (PDT). When the chamber was left intact, the light dose of 450 J cm-2 yielded complete tissue necrosis. The role of the dye-carrier complex in shielding the vascular surrounding from photoproducts was studied in a third group of animals. The presence of peroxides was demonstrated in the serum of these animals after PDT with zinc phthalocyanine in liposomes (ZnPc-lip) using a total light dose of 450 J cm-2. This ex vivo observation supports the previously reported observations in vitro that the carrier complex is able to quench the photoproducts resulting from photoactivation of the photosensitiser which is present in the circulation. Images Figure 2 PMID:8180012

van Leengoed, H. L.; Cuomo, V.; Versteeg, A. A.; van der Veen, N.; Jori, G.; Star, W. M.

1994-01-01

273

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

274

Short communication: rapid detection of milk fat adulteration with vegetable oil by fluorescence spectroscopy.  

PubMed

This study assessed the potential application of fluorescence spectroscopy in detecting adulteration of milk fat with vegetable oil and characterizing the samples according to the source of the fat. Pure butterfat was adulterated with different vegetable oils at various concentrations (0, 5, 10, 15, 20, 30, and 40%). Nonfat and reduced-fat milk were also adulterated with vegetable oils to simulate full-fat milk (3.2%). The 2- and 3-dimensional front-face fluorescence spectroscopy and gas chromatography were used to obtain the fluorescence spectra and fatty acid profile, respectively. Principal component analysis and 3-way partial least squares regression analysis were applied to analyze the data. The pure and adulterated samples were discriminated based on the total concentration of saturated fatty acids and unsaturated fatty acids, and also on the 3 major fluorophores: tryptophan, tocopherols, and riboflavin. Fluorescence spectroscopy was able to detect up to 5% of adulteration of vegetable oil into the butterfat. The saturated fatty acids showed higher predictability than the unsaturated fatty acids (R(2) = 0.73-0.92 vs. 0.20-0.65, respectively). The study demonstrated the high potential of fluorescence spectroscopy to rapidly detect adulteration of milk fat with vegetable oil, and discriminate commercial butter and milk according to the source of the fat. PMID:23415535

Ntakatsane, M P; Liu, X M; Zhou, P

2013-04-01

275

High-Resolution In Vivo Imaging of Fluorescent Proteins Using Window Chamber Models  

PubMed Central

Fluorescent proteins enable in vivo characterization of a wide and growing array of morphological and functional biomarkers. To fully capitalize on the spatial and temporal information afforded by these reporter proteins, a method for imaging these proteins at high resolution longitudinally is required. This chapter describes the use of window chamber models as a means of imaging fluorescent proteins and other optical parameters. Such models essentially involve surgically implanting a window through which tumor or normal tissue can be imaged using existing microscopy techniques. This enables acquisition of high-quality images down to the cellular or subcellular scale, exploiting the diverse array of optical contrast mechanisms, while also maintaining the native microenvironment of the tissue of interest. This makes these techniques applicable to a wide array of problems in the biomedical sciences. PMID:22700402

Palmer, Gregory M.; Fontanella, Andrew N.; Shan, Siqing; Dewhirst, Mark W.

2013-01-01

276

Compact low-cost detector for in vivo assessment of microphytobenthos using laser induced fluorescence  

NASA Astrophysics Data System (ADS)

The development of a compact low-cost detector for non-destructive assessment of microphytobenthos using laser induced fluorescence was described. The detector was built from a specially modified commercial miniature fiber optic spectrometer (Ocean Optics USB4000). Its usefulness is experimentally verified by the study of diatom-dominated biofilms inhabiting the upper layers of intertidal sediments of the Tagus Estuary, Portugal. It is demonstrated that, operating with a laser emitter producing 30 mJ pulses at the wavelength of 532 nm, the detector is capable to record fluorescence signals with sufficient intensity for the quantitative biomass characterization of the motile epipelic microphytobenthic communities and to monitor their migratory activity. This paves the way for building an entire emitter-detector LIF system for microphytobenthos monitoring, which will enable microalgae communities occupying hardly accessible intertidal flats to be monitored in vivo at affordable cost.

Utkin, A. B.; Vieira, S.; Marques da Silva, J.; Lavrov, A.; Leite, E.; Cartaxana, P.

2013-03-01

277

Correlative electron and fluorescence microscopy of magnetotactic bacteria in liquid: toward in vivo imaging.  

PubMed

Magnetotactic bacteria biomineralize ordered chains of uniform, membrane-bound magnetite or greigite nanocrystals that exhibit nearly perfect crystal structures and species-specific morphologies. Transmission electron microscopy (TEM) is a critical technique for providing information regarding the organization of cellular and magnetite structures in these microorganisms. However, conventional TEM can only be used to image air-dried or vitrified bacteria removed from their natural environment. Here we present a correlative scanning TEM (STEM) and fluorescence microscopy technique for imaging viable cells of Magnetospirillum magneticum strain AMB-1 in liquid using an in situ fluid cell TEM holder. Fluorescently labeled cells were immobilized on microchip window surfaces and visualized in a fluid cell with STEM, followed by correlative fluorescence imaging to verify their membrane integrity. Notably, the post-STEM fluorescence imaging indicated that the bacterial cell wall membrane did not sustain radiation damage during STEM imaging at low electron dose conditions. We investigated the effects of radiation damage and sample preparation on the bacteria viability and found that approximately 50% of the bacterial membranes remained intact after an hour in the fluid cell, decreasing to ~30% after two hours. These results represent a first step toward in vivo studies of magnetite biomineralization in magnetotactic bacteria. PMID:25358460

Woehl, Taylor J; Kashyap, Sanjay; Firlar, Emre; Perez-Gonzalez, Teresa; Faivre, Damien; Trubitsyn, Denis; Bazylinski, Dennis A; Prozorov, Tanya

2014-01-01

278

Correlative Electron and Fluorescence Microscopy of Magnetotactic Bacteria in Liquid: Toward In Vivo Imaging  

PubMed Central

Magnetotactic bacteria biomineralize ordered chains of uniform, membrane-bound magnetite or greigite nanocrystals that exhibit nearly perfect crystal structures and species-specific morphologies. Transmission electron microscopy (TEM) is a critical technique for providing information regarding the organization of cellular and magnetite structures in these microorganisms. However, conventional TEM can only be used to image air-dried or vitrified bacteria removed from their natural environment. Here we present a correlative scanning TEM (STEM) and fluorescence microscopy technique for imaging viable cells of Magnetospirillum magneticum strain AMB-1 in liquid using an in situ fluid cell TEM holder. Fluorescently labeled cells were immobilized on microchip window surfaces and visualized in a fluid cell with STEM, followed by correlative fluorescence imaging to verify their membrane integrity. Notably, the post-STEM fluorescence imaging indicated that the bacterial cell wall membrane did not sustain radiation damage during STEM imaging at low electron dose conditions. We investigated the effects of radiation damage and sample preparation on the bacteria viability and found that approximately 50% of the bacterial membranes remained intact after an hour in the fluid cell, decreasing to ~30% after two hours. These results represent a first step toward in vivo studies of magnetite biomineralization in magnetotactic bacteria. PMID:25358460

Woehl, Taylor J.; Kashyap, Sanjay; Firlar, Emre; Perez-Gonzalez, Teresa; Faivre, Damien; Trubitsyn, Denis; Bazylinski, Dennis A.; Prozorov, Tanya

2014-01-01

279

Multispectral opto-acoustic tomography of deep-seated fluorescent proteins in vivo  

NASA Astrophysics Data System (ADS)

Fluorescent proteins have become essential reporter molecules for studying life at the cellular and sub-cellular level, re-defining the ways in which we investigate biology. However, because of intense light scattering, most organisms and tissues remain inaccessible to current fluorescence microscopy techniques at depths beyond several hundred micrometres. We describe a multispectral opto-acoustic tomography technique capable of high-resolution visualization of fluorescent proteins deep within highly light-scattering living organisms. The method uses multiwavelength illumination over multiple projections combined with selective-plane opto-acoustic detection for artifact-free data collection. Accurate image reconstruction is enabled by making use of wavelength-dependent light propagation models in tissue. By performing whole-body imaging of two biologically important and optically diffuse model organisms, Drosophila melanogaster pupae and adult zebrafish, we demonstrate the facility to resolve tissue-specific expression of eGFP and mCherrry fluorescent proteins for precise morphological and functional observations in vivo.

Razansky, Daniel; Distel, Martin; Vinegoni, Claudio; Ma, Rui; Perrimon, Norbert; Köster, Reinhard W.; Ntziachristos, Vasilis

2009-07-01

280

Correlative Electron and Fluorescence Microscopy of Magnetotactic Bacteria in Liquid: Toward In Vivo Imaging  

NASA Astrophysics Data System (ADS)

Magnetotactic bacteria biomineralize ordered chains of uniform, membrane-bound magnetite or greigite nanocrystals that exhibit nearly perfect crystal structures and species-specific morphologies. Transmission electron microscopy (TEM) is a critical technique for providing information regarding the organization of cellular and magnetite structures in these microorganisms. However, conventional TEM can only be used to image air-dried or vitrified bacteria removed from their natural environment. Here we present a correlative scanning TEM (STEM) and fluorescence microscopy technique for imaging viable cells of Magnetospirillum magneticum strain AMB-1 in liquid using an in situ fluid cell TEM holder. Fluorescently labeled cells were immobilized on microchip window surfaces and visualized in a fluid cell with STEM, followed by correlative fluorescence imaging to verify their membrane integrity. Notably, the post-STEM fluorescence imaging indicated that the bacterial cell wall membrane did not sustain radiation damage during STEM imaging at low electron dose conditions. We investigated the effects of radiation damage and sample preparation on the bacteria viability and found that approximately 50% of the bacterial membranes remained intact after an hour in the fluid cell, decreasing to ~30% after two hours. These results represent a first step toward in vivo studies of magnetite biomineralization in magnetotactic bacteria.

Woehl, Taylor J.; Kashyap, Sanjay; Firlar, Emre; Perez-Gonzalez, Teresa; Faivre, Damien; Trubitsyn, Denis; Bazylinski, Dennis A.; Prozorov, Tanya

2014-10-01

281

Interactions of Indocyanine Green and Lipid in Enhancing Near-Infrared Fluorescence Properties: The Basis for Near-Infrared Imaging in Vivo  

PubMed Central

Indocyanine green (ICG) is a near-infrared (NIR) contrast agent commonly used for in vivo cardiovascular and eye imaging. For medical diagnosis, ICG is limited by its aqueous instability, concentration-dependent aggregation, and rapid degradation. To overcome these limitations, scientists have formulated ICG in various liposomes, which are spherical lipid membrane vesicles with an aqueous core. Some encapsulate ICG, while others mix it with liposomes. There is no clear understanding of lipid–ICG interactions. Therefore, we investigated lipid–ICG interactions by fluorescence and photon correlation spectroscopy. These data were used to design stable and maximally fluorescent liposomal ICG nanoparticles for NIR optical imaging of the lymphatic system. We found that ICG binds to and is incorporated completely and stably into the lipid membrane. At a lipid:ICG molar ratio of 250:1, the maximal fluorescence intensity was detected. ICG incorporated into liposomes enhanced the fluorescence intensity that could be detected across 1.5 cm of muscle tissue, while free ICG only allowed 0.5 cm detection. When administered subcutaneously in mice, lipid-bound ICG in liposomes exhibited a higher intensity, NIR image resolution, and enhanced lymph node and lymphatic vessel visualization. It also reduced the level of fluorescence quenching due to light exposure and degradation in storage. Lipid-bound ICG could provide additional medical diagnostic value with NIR optical imaging for early intervention in cases of lymphatic abnormalities. PMID:24512123

2015-01-01

282

Multiphoton microscopy and fluorescence lifetime imaging provide a novel method in studying drug distribution and metabolism in the rat liver in vivo  

NASA Astrophysics Data System (ADS)

Multiphoton microscopy has been shown to be a useful tool in studying drug distribution in biological tissues. In addition, fluorescence lifetime imaging provides information about the structure and dynamics of fluorophores based on their fluorescence lifetimes. Fluorescein, a commonly used fluorescent probe, is metabolized within liver cells to fluorescein mono-glucuronide, which is also fluorescent. Fluorescein and its glucuronide have similar excitation and emission spectra, but different fluorescence lifetimes. In this study, we employed multiphoton fluorescence lifetime imaging to study the distribution and metabolism of fluorescein and its metabolite in vivo in rat liver. Fluorescence lifetime values in vitro were used to interpret in vivo data. Our results show that the mean fluorescence lifetimes of fluorescein and its metabolite decrease over time after injection of fluorescein in three different regions of the liver. In conclusion, we have demonstrated a novel method to study a fluorescent compound and metabolite in vivo using multiphoton fluorescence lifetime imaging.

Thorling, Camilla A.; Dancik, Yuri; Hupple, Clinton W.; Medley, Gregory; Liu, Xin; Zvyagin, Andrei V.; Robertson, Tom A.; Burczynski, Frank J.; Roberts, Michael S.

2011-08-01

283

Comparison of fluorescence spectroscopy and FTIR in differentiation of plant pollens  

NASA Astrophysics Data System (ADS)

Spectroscopic techniques are under investigation on possibility of differentiation of airborne particles. This paper describes pollen discrimination among others bio-particles in laboratory conditions. Pollen samples were characterized with UV-Vis fluorescence, drift and KBr pellet techniques of infrared spectroscopy. Principal Component Analysis of UV-Vis fluorescence and FTIR spectra revealed that pollens can be distinguished from other bio-materials with use of these methods. Both methods resulted in similar classification capability. Combined FTIR and fluorescence data analysis did not improve the discrimination between pollen allergens and other airborne biological materials.

Mularczyk-Oliwa, Monika; Bombalska, Aneta; Kaliszewski, Miron; W?odarski, Maksymilian; Kopczy?ski, Krzysztof; Kwa?ny, Miros?aw; Szpakowska, Ma?gorzata; Trafny, El?bieta A.

2012-11-01

284

Tailor-made dyes for fluorescence correlation spectroscopy (FCS).  

PubMed

Two new fluorescent labels are presented that are optimized for excitation with He/Ne laser and red diode lasers. Application in FCS and labeling of proteins and oligomers are demonstrated. A strong rise of quantum yield and emission life time upon binding to biomolecules are characteristic features of the dyes. PMID:11347900

Czerney, P; Lehmann, F; Wenzel, M; Buschmann, V; Dietrich, A; Mohr, G J

2001-03-01

285

Fluorescence spectroscopy of polynuclear aromatic compounds in environmental monitoring  

Microsoft Academic Search

The occurrence of polynuclear aromatic compounds (PAC) in the environment and experimental techniques suitable for the detection of PAC in environmental compartments are briefly reviewed. The specific requirements for on-site andin situ environmental analysis are outlined. Particular emphasis is given to fluorescence spectroscopic techniques for the investigation of humic acid- and soil-containing samples. Some examples of studies in the literature

M. U. Kumke; H.-G. Löhmannsröben; Th. Roch

1995-01-01

286

Two-photon fluorescence excitation spectroscopy of biological molecules  

NASA Astrophysics Data System (ADS)

The UV fluorescence spectra of aromatic amino-acids and some proteins at two photon excitation by second harmonic of Nd:YAG laser are received. Two-photon absorption cross sections of tryptophan, tyrosine, phenylalanine and proteins: bovine serum albumin, lysozyme, trypsin, (alpha) - chymotrypsinogen and pepsin at wavelength 532 nm were measured by means of the two-quantum standard method.

Meshalkin, Yuri P.; Alfimov, E. E.; Groshev, D. E.; Makukha, V. K.

1996-06-01

287

Fluorescence Spectroscopy of Conformational Changes of Single LH2 Complexes  

PubMed Central

We have investigated the energy landscape of the bacterial photosynthetic peripheral light-harvesting complex LH2 of purple bacterium Rhodopseudomonas acidophila by monitoring sequences of fluorescence spectra of single LH2 assemblies, at room temperature, with different excitation intensities as well as at elevated temperatures, utilizing a confocal microscope. The fluorescence peak wavelength of individual LH2 complexes was found to abruptly move between long-lived quasi-stable levels differing by up to 30 nm. The frequency and size of these fluorescence peak movements were found to increase linearly with the excitation intensity. These spectral shifts either to the blue or to the red were accompanied by a broadening and decrease of the intensity of the fluorescence spectrum. The probability for a particle to undergo significant spectral shift in either direction was found to be roughly the same. Using the modified Redfield theory, the observed changes in spectral shape and intensity were accounted for by changes in the realization of the static disorder. Long lifetimes of the quasi-stable states suggest large energetic barriers between the states characterized by different emission spectra. PMID:15501944

Rutkauskas, Danielis; Novoderezhkin, Vladimir; Cogdell, Richard J.; van Grondelle, Rienk

2005-01-01

288

Fluorescence Spectroscopy, Exciton Dynamics, and Photochemistry of Single Allophycocyanin Trimers  

E-print Network

that the photobleaching of a single B-phycoerythrin molecule occurs in a single step. With confocal microscopy, two-harvesting protein complex from cyanobacteria, by room-temperature single-molecule measurements of fluorescence/or photobleaching, which involves exciton-exciton annihilation. These single-molecule experiments provide new

Xie, Xiaoliang Sunney

289

The daily cycle of fluorescence in vivo in the Central Equatorial Pacific  

E-print Network

and towed fish. Vertical lines indicate time of switching. Anomalously low values at 2145L and 2314L are due to air bubbles in line. . . . . . . . . . . . . . . 10 3. Underway sampling scheme used for data collection during Cruise 79-G-6 aboard R... not totally filter out the light effect (Fig. 13). Clear indication remains of a daily cycle in the in vivo fluorescence. Looking for a more effective, strictly empirical equation to adjust for the daily cycle, I tried Eq. 3 rewritten as: Fm = Fi - k sin...

Setser, Patrick James

1980-01-01

290

Micro-endoscope for in vivo widefield high spatial resolution fluorescent imaging  

PubMed Central

In this paper we report the design, testing and use of a scannerless probe specifically for minimally invasive imaging of deep tissue in vivo with an epi-fluorescence modality. The probe images a 500 ?m diameter field of view through a 710 ?m outer diameter probe with a maximum tissue penetration depth of 15 mm specifically configured for eGFP imaging. Example results are given from imaging the pituitary gland of rats and zebrafish hearts with lateral resolution of 2.5 ?m. PMID:22741074

Saunter, C D; Semprini, S.; Buckley, C.; Mullins, J; Girkin, J M

2012-01-01

291

Multimodal in vivo imaging of oral cancer using fluorescence lifetime, photoacoustic and ultrasound techniques  

PubMed Central

This work reports a multimodal system for label-free tissue diagnosis combining fluorescence lifetime imaging (FLIm), ultrasound backscatter microscopy (UBM), and photoacoustic imaging (PAI). This system provides complementary biochemical, structural and functional features allowing for enhanced in vivo detection of oral carcinoma. Results from a hamster oral carcinoma model (normal, precancer and carcinoma) are presented demonstrating the ability of FLIm to delineate biochemical composition at the tissue surface, UBM and related radiofrequency parameters to identify disruptions in the tissue microarchitecture and PAI to map optical absorption associated with specific tissue morphology and physiology. PMID:24049693

Fatakdawala, Hussain; Poti, Shannon; Zhou, Feifei; Sun, Yang; Bec, Julien; Liu, Jing; Yankelevich, Diego R.; Tinling, Steven P.; Gandour-Edwards, Regina F.; Farwell, D. Gregory; Marcu, Laura

2013-01-01

292

Monitoring cellular movement in vivo with photoconvertible fluorescence protein "Kaede" transgenic mice.  

PubMed

Kaede is a photoconvertible fluorescence protein that changes from green to red upon exposure to violet light. The photoconversion of intracellular Kaede has no effect on cellular function. Using transgenic mice expressing the Kaede protein, we demonstrated that movement of cells with the photoconverted Kaede protein could be monitored from lymphoid organs to other tissues as well as from skin to the draining lymph node. Analysis of the kinetics of cellular movement revealed that each subset of cells in the lymph node, such as CD4(+) T, CD8(+) T, B, and dendritic cells, has a distinct migration pattern in vivo. Thus, the Kaede transgenic mouse system would be an ideal tool to monitor precise cellular movement in vivo at different stages of immune response to pathogens as well as in autoimmune diseases. PMID:18663225

Tomura, Michio; Yoshida, Naoki; Tanaka, Junko; Karasawa, Satoshi; Miwa, Yoshihiro; Miyawaki, Atsushi; Kanagawa, Osami

2008-08-01

293

In vivo analysis of a fluorescent SUMO fusion in transgenic Drosophila.  

PubMed

Sumoylation, the covalent attachment of SUMO, a 90 amino acid peptide related to ubiquitin, is a major modulator of protein functions. Fluorescent SUMO protein fusions have been used in cell cultures to visualize SUMO in vivo but not in multicellular organisms. We generated a transgenic line of Drosophila expressing an mCherry-SUMO fusion. We analyzed its pattern in vivo in salivary gland nuclei expressing Venus-HP1 to recognize the different chromatin components (Chromocenter, chromosome IV). We compared it to SUMO immunostaining on squashed polytene chromosomes and observed similar patterns. In addition to the previously reported SUMO localizations (chromosome arms and chromocenter), we identify 2 intense binding sites: the fourth chromosome telomere and the DAPI-bright band in the region 81F. PMID:25483255

Bocksberger, Marion; Karch, François; Gibert, Jean-Michel

2014-01-01

294

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

295

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

296

Fluorescence spectroscopy of kerosene vapour at high temperatures and pressures: potential for gas turbines measurements  

NASA Astrophysics Data System (ADS)

Laser-induced fluorescence spectroscopy of kerosene vapour was performed in a heated test cell operating between 450 and 900 K, at pressure from 0.1 to 3.0 MPa, for oxygen molar fraction between 0 and 21 %, with different laser excitation wavelengths (248, 266, 282 and 308 nm). Results show that, depending on the laser excitation scheme, kerosene fluorescence spectrum exhibits one or two fluorescence bands in the UV-visible range (attributed to aromatics naturally present in kerosene fuel). Fluorescence intensity of these bands decreases with increasing temperature, pressure and oxygen molar fraction. Different imaging strategies were derived from spectroscopic findings to simultaneously measure temperature and equivalence ratio fields in kerosene/air sprays, or flame structure and fuel spatial distribution in kerosene/air aeronautical combustors, by means of planar laser-induced fluorescence on kerosene vapour (K-PLIF).

Orain, M.; Baranger, P.; Ledier, C.; Apeloig, J.; Grisch, F.

2014-09-01

297

An individually coated near-infrared fluorescent protein as a safe and robust nanoprobe for in vivo imaging  

NASA Astrophysics Data System (ADS)

A prerequisite for in vivo fluorescence imaging is the safety of fluorescent probes. Among all fluorescent probes, fluorescent proteins (FPs) might be the safest ones, which have been widely used in biological sciences at the gene level. But FPs have not been used in vivo in the purified form yet due to the instability of proteins. Here, we individually coat near-infrared (NIR) FPs (NIRFPs) with a silica nanoshell, resulting in NIRFP@silica, one of the safest and brightest NIR fluorescent nanoprobes with a quantum yield of 0.33 for in vivo imaging. The silica shell not only protects NIRFPs from denaturation and metabolic digestion, but also enhances the quantum yield and photostability of the coated NIRFPs. When injected via the tail vein, NIRFP@silica NPs can distribute all over the mouse body, and then can be efficiently eliminated through urine in 24 h, demonstrating its potential applications as a safe and robust NIR fluorescence probe for whole body imaging.A prerequisite for in vivo fluorescence imaging is the safety of fluorescent probes. Among all fluorescent probes, fluorescent proteins (FPs) might be the safest ones, which have been widely used in biological sciences at the gene level. But FPs have not been used in vivo in the purified form yet due to the instability of proteins. Here, we individually coat near-infrared (NIR) FPs (NIRFPs) with a silica nanoshell, resulting in NIRFP@silica, one of the safest and brightest NIR fluorescent nanoprobes with a quantum yield of 0.33 for in vivo imaging. The silica shell not only protects NIRFPs from denaturation and metabolic digestion, but also enhances the quantum yield and photostability of the coated NIRFPs. When injected via the tail vein, NIRFP@silica NPs can distribute all over the mouse body, and then can be efficiently eliminated through urine in 24 h, demonstrating its potential applications as a safe and robust NIR fluorescence probe for whole body imaging. Electronic supplementary information (ESI) available: A chromatogram of APTS-NIRFP, a TEM image of 40 nm NIRFP@silica, dispersion stability of NIRFP@silica, more whole body fluorescent images, serum biochemical parameters, and optical images of HE stained organ slices. See DOI: 10.1039/c3nr02508j

Yang, Yu; Xiang, Kun; Yang, Yi-Xin; Wang, Yan-Wen; Zhang, Xin; Cui, Yangdong; Wang, Haifang; Zhu, Qing-Qing; Fan, Liqiang; Liu, Yuanfang; Cao, Aoneng

2013-10-01

298

Laser Induced Fluorescence Spectroscopy of a Langmuir Monolayer of C-16 Fluorescent Dipyrrinone Liquid Crystal  

NASA Astrophysics Data System (ADS)

A C-16 Fluorescent Dipyrrinone Liquid Crystal synthesized by the Chemistry department, University of West Florida, has been prepared in a Langmuir monolayer using a Nima Langmuir-Blodgett Trough. DeLuca et al. [1] studied how the length of the hydrocarbon tail influences the behavior of the pressure-area isotherm of the Langmuir film. The C-16 Fluorescent Dipyrrinone Liquid Crystal film produced a stable film at 20 mN/m and a stable, optical quality film at 40 mN/m. We present a study of the fluorescence properties of the C-16 fluorescent dipyrrinone liquid crystal film. Once the monolayer is compressed the sample is excited using a 410 nm wavelength laser and the fluorescence is measured using an Oriel MS260i 1/4 m Spectrograph. [4pt] [1] Deluca, Giovanni; Carroll, Alexander; Prayaga, Chandra; Wade, Aaron; Heath, Christopher; Renaud, Amy; Huggins, Michael. ``Preparation and Characterization of C-16 and C-10 Fluorescent Dipyrrinone Liquid Crystal Langmuir-Blodgett Films.'' American Physical Society, APS March Meeting 2012, 02/2012.

Struebing, Christian; Deluca, Giovanni; Prayaga, Chandra; Wade, Aaron; Huggins, Michael; Renaud, Amy; Chandler, Rebecca

2013-03-01

299

In vivo monitoring of toxic metals: assessment of neutron activation and x-ray fluorescence techniques  

SciTech Connect

To date, cadmium, lead, aluminum, and mercury have been measured in vivo in humans. The possibilities of monitoring other toxic metals have also been demonstrated, but no human studies have been performed. Neutron activation analysis appears to be most suitable for Cd and Al measurements, while x-ray fluorescence is ideally suited for measurement of lead in superficial bone. Filtered neutron beams and polarized x-ray sources are being developed which will improve in vivo detection limits. Even so, several of the current facilities are already suitable for use in epidemiological studies of selected populations with suspected long-term low-level ''environmental'' exposures. Evaluation and diagnosis of patients presenting with general clinical symptoms attributable to possible toxic metal exposure may be assisted by in vivo examination. Continued in vivo monitoring of industrial workers, especially follow-up measurements, will provide the first direct assessment of changes in body burden and a direct measure of the biological life-times of these metals in humans. 50 refs., 4 figs., 2 tabs.

Ellis, K.J.

1986-01-01

300

Tracking dynamics of muscle engraftment in small animals by in vivo fluorescent imaging.  

PubMed

Muscular dystrophies are a group of degenerative muscle diseases characterized by progressive loss of contractile muscle cells. Currently, there is no curative treatment available. Recent advances in stem cell biology have generated new hopes for the development of effective cell based therapies to treat these diseases. Transplantation of various types of stem cells labeled with fluorescent proteins into muscles of dystrophic animal models has been used broadly in the field. A non-invasive technique with the capability to track the transplanted cell fate longitudinally can further our ability to evaluate muscle engraftment by transplanted cells more accurately and efficiently. In the present study, we demonstrate that in vivo fluorescence imaging is a sensitive and reliable method for tracking transplanted GFP (Green Fluorescent Protein)-labeled cells in mouse skeletal muscles. Despite the concern about background due to the use of an external light necessary for excitation of fluorescent protein, we found that by using either nude mouse or eliminating hair with hair removal reagents much of this problem is eliminated. Using a CCD camera, the fluorescent signal can be detected in the tibialis anterior (TA) muscle after injection of 5 x 10(5) cells from either GFP transgenic mice or eGFP transduced myoblast culture. For more superficial muscles such as the extensor digitorum longus (EDL), injection of fewer cells produces a detectable signal. Signal intensity can be measured and quantified as the number of emitted photons per second in a region of interest (ROI). Since the acquired images show clear boundaries demarcating the engrafted area, the size of the ROI can be measured as well. If the legs are positioned consistently every time, the changes in total number of photons per second per muscle and the size of the ROI reflect the changes in the number of engrafted cells and the size of the engrafted area. Therefore the changes in the same muscle over time are quantifiable. In vivo fluorescent imaging technique has been used primarily to track the growth of tumorogenic cells, our study shows that it is a powerful tool that enables us to track the fate of transplanted stem cells. PMID:19770816

Yang, Zhong; Zeng, Qing; Ma, Zhiyuan; Wang, Yaming; Xu, Xiaoyin

2009-01-01

301

Digitally synthesized beat frequency-multiplexed fluorescence lifetime spectroscopy  

PubMed Central

Frequency domain fluorescence lifetime imaging is a powerful technique that enables the observation of subtle changes in the molecular environment of a fluorescent probe. This technique works by measuring the phase delay between the optical emission and excitation of fluorophores as a function of modulation frequency. However, high-resolution measurements are time consuming, as the excitation modulation frequency must be swept, and faster low-resolution measurements at a single frequency are prone to large errors. Here, we present a low cost optical system for applications in real-time confocal lifetime imaging, which measures the phase vs. frequency spectrum without sweeping. Deemed Lifetime Imaging using Frequency-multiplexed Excitation (LIFE), this technique uses a digitally-synthesized radio frequency comb to drive an acousto-optic deflector, operated in a cat’s-eye configuration, to produce a single laser excitation beam modulated at multiple beat frequencies. We demonstrate simultaneous fluorescence lifetime measurements at 10 frequencies over a bandwidth of 48 MHz, enabling high speed frequency domain lifetime analysis of single- and multi-component sample mixtures. PMID:25574449

Chan, Jacky C. K.; Diebold, Eric D.; Buckley, Brandon W.; Mao, Sien; Akbari, Najva; Jalali, Bahram

2014-01-01

302

Impurity studies in fusion devices using laser-fluorescence-spectroscopy  

SciTech Connect

Resonance fluorescence excitation of neutral atoms using tunable radiation from dye lasers offers a number of unique advantages for impurity studies in fusion devices. Using this technique, it is possible to perform local, time-resolved measurements of the densities and velocity distributions of metallic impurities in fusion devices without disturbing the plasma. Velocities are measured by monitoring the fluorescence intensity while tuning narrow bandwidth laser radiation through the Doppler - broadened absorbtion spectrum of the transition. The knowledge of the velocity distribution of neutral impurities is particularly useful for the determination of impurity introduction mechanisms. The laser fluorescence technique will be described in terms of its application to metallic impurities in fusion devices and related laboratory experiments. Particular attention will be given to recent results from the ISX-B tokamak using pulsed dye lasers where detection sensitivities for neutral Fe of 10/sup 6/ atoms/cm/sup 3/ with a velocity resolution of 600 m/sec (0.1 eV) have been achieved. Techniques for exciting plasma particles (H,D) will also be discussed.

Husinsky, W.R.

1980-08-01

303

In vivo fluorescence imaging of the reticuloendothelial system using quantum dots in combination with bioluminescent tumour monitoring  

Microsoft Academic Search

Purpose  We characterised in vivo fluorescence imaging (FLI) of the reticuloendothelial system using quantum dots (QD) and investigated\\u000a its use in combination with in vivo bioluminescence imaging (BLI).\\u000a \\u000a \\u000a \\u000a Materials and methods  In vivo FLI was performed in five mice repeatedly after the intravenous administration of QD without conjugation to targeting\\u000a ligands. Ex vivo FLI of the excised organs was performed 24 h after

Yusuke Inoue; Kiyoko Izawa; Kohki Yoshikawa; Haruyasu Yamada; Arinobu Tojo; Kuni Ohtomo

2007-01-01

304

Multicontrast photoacoustic in vivo imaging using near-infrared fluorescent proteins  

NASA Astrophysics Data System (ADS)

Non-invasive imaging of biological processes in vivo is invaluable in advancing biology. Photoacoustic tomography is a scalable imaging technique that provides higher resolution at greater depths in tissue than achievable by purely optical methods. Here we report the application of two spectrally distinct near-infrared fluorescent proteins, iRFP670 and iRFP720, engineered from bacterial phytochromes, as photoacoustic contrast agents. iRFPs provide tissue-specific contrast without the need for delivery of any additional substances. Compared to conventional GFP-like red-shifted fluorescent proteins, iRFP670 and iRFP720 demonstrate stronger photoacoustic signals at longer wavelengths, and can be spectrally resolved from each other and hemoglobin. We simultaneously visualized two differently labeled tumors, one with iRFP670 and the other with iRFP720, as well as blood vessels. We acquired images of a mouse as 2D sections of a whole animal, and as localized 3D volumetric images with high contrast and sub-millimeter resolution at depths up to 8 mm. Our results suggest iRFPs are genetically-encoded probes of choice for simultaneous photoacoustic imaging of several tissues or processes in vivo.

Krumholz, Arie; Shcherbakova, Daria M.; Xia, Jun; Wang, Lihong V.; Verkhusha, Vladislav V.

2014-02-01

305

Dynamic dual-tracer MRI-guided fluorescence tomography to quantify receptor density in vivo  

PubMed Central

The up-regulation of cell surface receptors has become a central focus in personalized cancer treatment; however, because of the complex nature of contrast agent pharmacokinetics in tumor tissue, methods to quantify receptor binding in vivo remain elusive. Here, we present a dual-tracer optical technique for noninvasive estimation of specific receptor binding in cancer. A multispectral MRI-coupled fluorescence molecular tomography system was used to image the uptake kinetics of two fluorescent tracers injected simultaneously, one tracer targeted to the receptor of interest and the other tracer a nontargeted reference. These dynamic tracer data were then fit to a dual-tracer compartmental model to estimate the density of receptors available for binding in the tissue. Applying this approach to mice with deep-seated gliomas that overexpress the EGF receptor produced an estimate of available receptor density of 2.3 ± 0.5 nM (n = 5), consistent with values estimated in comparative invasive imaging and ex vivo studies. PMID:23671066

Davis, Scott C.; Samkoe, Kimberley S.; Tichauer, Kenneth M.; Sexton, Kristian J.; Gunn, Jason R.; Deharvengt, Sophie J.; Hasan, Tayyaba; Pogue, Brian W.

2013-01-01

306

In vivo fluorescence confocal microscopy: indocyanine green enhances the contrast of epidermal and dermal structures  

NASA Astrophysics Data System (ADS)

In recent years, in vivo skin imaging devices have been successfully implemented in skin research as well as in clinical routine. Of particular importance is the use of reflectance confocal microscopy (RCM) and fluorescence confocal microscopy (FCM) that enable visualization of the tissue with a resolution comparable to histology. A newly developed commercially available multi-laser device in which both technologies are integrated now offers the possibility to directly compare RCM with FCM. The fluorophore indocyanine green (ICG) was intradermally injected into healthy forearm skin of 10 volunteers followed by in vivo imaging at various time points. In the epidermis, accurate assessment of cell morphology with FCM was supplemented by identification of pigmented cells and structures with RCM. In dermal layers, only with FCM connective tissue fibers were clearly contoured down to a depth of more than 100 ?m. The fluorescent signal still provided a favorable image contrast 24 and 48 hours after injection. Subsequently, ICG was applied to different types of skin diseases (basal cell carcinoma, actinic keratosis, seborrhoeic keratosis, and psoriasis) in order to demonstrate the diagnostic benefit of FCM when directly compared with RCM. Our data suggest a great impact of FCM in combination with ICG on clinical and experimental dermatology in the future.

Skvara, Hans; Kittler, Harald; Schmid, Johannes A.; Plut, Ulrike; Jonak, Constanze

2011-09-01

307

Ultrafast fluorescence imaging in vivo with conjugated polymer fluorophores in the second near-infrared window  

NASA Astrophysics Data System (ADS)

In vivo fluorescence imaging in the second near-infrared window (1.0-1.7??m) can afford deep tissue penetration and high spatial resolution, owing to the reduced scattering of long-wavelength photons. Here we synthesize a series of low-bandgap donor/acceptor copolymers with tunable emission wavelengths of 1,050-1,350?nm in this window. Non-covalent functionalization with phospholipid-polyethylene glycol results in water-soluble and biocompatible polymeric nanoparticles, allowing for live cell molecular imaging at >1,000?nm with polymer fluorophores for the first time. Importantly, the high quantum yield of the polymer allows for in vivo, deep-tissue and ultrafast imaging of mouse arterial blood flow with an unprecedented frame rate of >25 frames per second. The high time-resolution results in spatially and time resolved imaging of the blood flow pattern in cardiogram waveform over a single cardiac cycle (~200?ms) of a mouse, which has not been observed with fluorescence imaging in this window before.

Hong, Guosong; Zou, Yingping; Antaris, Alexander L.; Diao, Shuo; Wu, Di; Cheng, Kai; Zhang, Xiaodong; Chen, Changxin; Liu, Bo; He, Yuehui; Wu, Justin Z.; Yuan, Jun; Zhang, Bo; Tao, Zhimin; Fukunaga, Chihiro; Dai, Hongjie

2014-06-01

308

Reaction-based epoxide fluorescent probe for in vivo visualization of hydrogen sulfide.  

PubMed

Hydrogen sulfide (H2S) has emerged as the most important biosynthetic gasotransmitters along with nitric oxide (NO) and carbon monoxide (CO). In this study, we report the design and the synthesis of a new epoxide fluorescent probe 7-glycidyloxy-9-(2-glycidyloxycarbonylphenyl)-2-xanthone (FEPO) for use in in vivo visualization of hydrogen sulfide. The probe employs a fluorescein as a fluorophore, and is equipped with an operating epoxide unit. FEPO functions via epoxide ring opening upon nucleophilic attack of H2S. This ring opening strategy may open a new avenue for the development of various H2S fluorescent sensors. FEPO showed high selectivity and high sensitivity for H2S. FEPO's cytotoxicity was tested using MTT (2-(4,5-dimethyl-2-thiazolyl)-3,5-diphenyl-2H-tetrazolium bromide) assay. Furthermore, the use of confocal imaging of H2S and in vivo imaging in live zebra fish demonstrated FEPO's potential biological applications. We anticipate that, owing to their ideal properties, probes of this type will find great uses in exploring the role of H2S in biology. PMID:25660659

Sathyadevi, Palanisamy; Chen, Yu-Jen; Wu, Shou-Cheng; Chen, Yen-Hao; Wang, Yun-Ming

2015-06-15

309

Three-Dimensional In Vivo Imaging of Green Fluorescent Protein-Expressing T Cells in Mice with Noncontact Fluorescence Molecular Tomography  

Microsoft Academic Search

Given that optical tomography is capable of quantitatively imaging the distribution of several important chromophores and fluorophores in vivo, there has been a great deal of interest in developing optical imaging systems with increased numbers of measurements under optimal experimental conditions. In this article, we present a novel system that enables three-dimensional imaging of fluorescent probes in whole animals using

Anikitos Garofalakis; Giannis Zacharakis; Heiko Meyer; Eleftherios N. Economou; Clio Mamalaki; Joseph Papamatheakis; Dimitris Kioussis; Vasilis Ntziachristos; Jorge Ripoll

2007-01-01

310

Hyperspectral Imaging and Spectroscopy of Fluorescently Coupled Acyl-CoA: Cholesterol Acyltransferase in Insect Cells  

NASA Technical Reports Server (NTRS)

A high-performance hyperspectral imaging module with high throughput of light suitable for low-intensity fluorescence microscopic imaging and subsequent analysis, including single-pixel-defined emission spectroscopy, was tested on Sf21 insect cells expressing green fluorescence associated with recombinant green fluorescent protein linked or not with the membrane protein acyl-CoA:cholesterol acyltransferase. The imager utilized the phenomenon of optical activity as a new technique providing information over a spectral range of 220-1400 nm, and was inserted between the microscope and an 8-bit CCD video-rate camera. The resulting fluorescence image did not introduce observable image aberrations. The images provided parallel acquisition of well resolved concurrent spatial and spectral information such that fluorescence associated with green fluorescent protein alone was demonstrated to be diffuse within the Sf21 insect cell, and that green fluorescence associated with the membrane protein was shown to be specifically concentrated within regions of the cell cytoplasm. Emission spectra analyzed from different regions of the fluorescence image showed blue shift specific for the regions of concentration associated with the membrane protein.

Malak, H.; Mahtani, H.; Herman, P.; Vecer, J.; Lu, X.; Chang, T. Y.; Richmond, Robert C.; Whitaker, Ann F. (Technical Monitor)

2001-01-01

311

Methods of single-molecule fluorescence spectroscopy and microscopy  

Microsoft Academic Search

Optical spectroscopy at the ultimate limit of a single molecule has grown over the past dozen years into a powerful technique for exploring the individual nanoscale behavior of molecules in complex local environments. Observing a single molecule removes the usual ensemble average, allowing the exploration of hidden heterogeneity in complex condensed phases as well as direct observation of dynamical state

W. E. Moerner; David P. Fromm

2003-01-01

312

Application of Fluorescence Spectroscopy for Rapid Detection of Pathogens in Food  

Technology Transfer Automated Retrieval System (TEKTRAN)

The potential of fluorescence spectroscopy was investigated for the detection food bone pathogens. E coli, Salmonella and Campylobactor, the most commonly present in food, were selectively identified. Each pathogen, grown in agar plate, was diluted in saline and prepared in different concentrations....

313

Analysis of the optical properties of the Orinoco River plume by absorption and fluorescence spectroscopy  

Microsoft Academic Search

The discharge of the Orinoco River significantly affects the optical properties of the water in the Caribbean Sea by increasing primary productivity and introducing large amounts of colored dissolved organic matter (CDOM) to the region. The optical characteristics of the CDOM in the Orinoco River plume were investigated during two cruises to the eastern Caribbean using absorption and fluorescence spectroscopy.

Carlos E. Del Castillo; Paula G. Coble; Julio M. Morell; José M. López; Jorge E. Corredor

1999-01-01

314

Binding of the Alzheimer amyloid ?-peptide to neuronal cell membranes by fluorescence correlation spectroscopy  

Microsoft Academic Search

The deposition of the Alzheimer amyloid ?-peptide (A?) fibrils in brain is a key step in Alzheimer's disease. The aggregated A? is found to be toxic to neurons since cells die when the aggregated A? is added to the cell culture medium. However, target of action of A? to cells is unknown. We have applied the fluorescence correlation spectroscopy (FCS)

Shakil Hossain; Mirtha Grande; Galib Ahmadkhanov; Aladdin Pramanik

2007-01-01

315

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

316

En route to traceable reference standards for surface group quantifications by XPS, NMR and fluorescence spectroscopy.  

PubMed

The fluorine content of polymer particles labelled with 2,2,2-trifluoroethylamine was reliably quantified with overlapping sensitivity ranges by XPS and solid-state NMR. This provides a first step towards reference materials for the metrological traceability of surface group quantifications. The extension of this concept to fluorescence spectroscopy is illustrated. PMID:25652135

Hennig, Andreas; Dietrich, Paul M; Hemmann, Felix; Thiele, Thomas; Borcherding, Heike; Hoffmann, Angelika; Schedler, Uwe; Jäger, Christian; Resch-Genger, Ute; Unger, Wolfgang E S

2015-03-21

317

Capillary Electrophoresis and Fluorescence Excitation-Emission Matrix Spectroscopy for Characterization of Humic Substances  

Technology Transfer Automated Retrieval System (TEKTRAN)

Capillary electrophoresis (CE) and fluorescence spectroscopy have been used in natural organic matter (NOM) studies. In this study, we characterized five fulvic acids, six humic acids and two unprocessed NOM samples obtained from the International Humic Substances Society (IHSS) using these two ana...

318

Fluorescence spectroscopy and molecular weight distribution of extracellular polymers from full-scale activated sludge biomass  

Microsoft Academic Search

Two fractions of extracellular polymer substances (EPSs), soluble and readily extractable (RE), were characterised in terms of their molecular weight distributions (MWD) and 3-D excitation-emission- matrix (EEM) fluorescence spectroscopy signatures. The EPS fractions were different: the soluble EPSs were composed mainly of high molecular weight compounds, while the RE EPSs were composed of small molecular weight compounds. Contrary to previous

M. Esparza-Soto; P. K. Westerhoff

2001-01-01

319

A quantitative study of the intracellular dynamics of fluorescently labelled glyco-gold nanoparticles via fluorescence correlation spectroscopy.  

PubMed

The dynamic behaviour of gold nanoparticles functionalised with glucose (Glc-Au NPs) has been studied here by means of fluorescence correlation spectroscopy (FCS). Meaningful data on the state of aggregation and dynamics of Glc-Au NPs fluorescently-labelled with HiLyte Fluor647 (Glc-Au-Hi NPs) in the intracellular environment were obtained. Moreover, the work presented here shows that FCS can be used to visualise the presence of single NPs or NP aggregates following uptake and to estimate, locally, NP concentrations within the cell. FCS measurements become possible after applying a "prebleaching" methodology, when the immobile NP fraction has been effectively removed and thus significant FCS data has been recorded. In this study, Glc-Au-Hi NPs have been incubated with HepG2 cells and their diffusion time in the intracellular environment has been measured and compared with their diffusion value in water and cell media. PMID:24639360

Murray, Richard A; Qiu, Yuan; Chiodo, Fabrizio; Marradi, Marco; Penadés, Soledad; Moya, Sergio E

2014-07-01

320

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

321

Dual-color fluorescence cross-correlation spectroscopy for multicomponent diffusional analysis in solution.  

PubMed Central

The present paper describes a new experimental scheme for following diffusion and chemical reaction systems of fluorescently labeled molecules in the nanomolar concentration range by fluorescence correlation analysis. In the dual-color fluorescence cross-correlation spectroscopy provided here, the concentration and diffusion characteristics of two fluorescent species in solution as well as their reaction product can be followed in parallel. By using two differently labeled reaction partners, the selectivity to investigate the temporal evolution of reaction product is significantly increased compared to ordinary one-color fluorescence autocorrelation systems. Here we develop the theoretical and experimental basis for carrying out measurements in a confocal dual-beam fluorescence correlation spectroscopy setup and discuss conditions that are favorable for cross-correlation analysis. The measurement principle is explained for carrying out DNA-DNA renaturation kinetics with two differently labeled complementary strands. The concentration of the reaction product can be directly determined from the cross-correlation amplitude. Images FIGURE 2 FIGURE 3 PMID:9083691

Schwille, P; Meyer-Almes, F J; Rigler, R

1997-01-01

322

Effect of indocyanin green formulation on blood clearance and in vivo fluorescence kinetic profile of skin  

NASA Astrophysics Data System (ADS)

Indocyanine green has been used to measure cardiac and liver functions. More recently, it has been proposed as a contrast agent in ophthalmic angiography, tumor imaging and as an infrared absorbing dye in the context of laser-induced thermal damage of blood vessels. The aim of the study is to overcome the disadvantage of a very short blood half-time and to participate to a better confinement in blood vessels. Indocyanine green was administered intravenously to Wistar rats at a 7.5 mg/kg dose. Formulations consist in indocyanine green aqueous solution and o/w emulsion. Blood samples were collected and analyzed by spectrophotometry. Fluorescence was recorded in vivo by spectrofluorometry using an optic fiber coupled to an optical multichannel analyzer. The fiber optic was placed at a 4 mm distance from the skin surface. Results show that aqueous solution of indocyanine green leads to a rapid blood clearance. To the administration of ICG emulsion belongs the advantage of increasing the half-time and the residence time of indocyanine green in skin. It may be noted that however the formulation is, the observed blood clearance profiles are quite different from the tissue fluorescence kinetic profiles. The dye could have a longer residence time (20 - 60 min. plateau phase). Moreover, a shift of the maximum emission peak is noted after i.v. administration. The study of ICG fluorescence in the presence of model membranes shows that ICG is able to interact with phospholipid bilayers. These findings may be interesting for therapeutic applications of indocyanine green requiring a high level of dye in tissues for a great period of time and participate to the knowledge of ICG behavior in vivo.

Devoisselle, Jean-Marie; Soulie-Begu, Sylvie; Mordon, Serge R.; Mestres, G.; Desmettre, Thomas; Maillols, H.

1995-12-01

323

Development of in-vivo fluorescence imaging with the Matrix-Free method  

NASA Astrophysics Data System (ADS)

Non-contact Fluorescence Molecular Tomography is an emerging technique for imaging of fluorescent probes or proteins in live animals. One of the main characteristics of the non-contact acquisition systems in comparison to the usual fibre-based systems is the much denser boundary data sets that are created. When model-based reconstruction methods are used that rely on the inversion of a derivative operator, the large number of measurements poses a challenge since the explicit formulation and storage of the Jacobian matrix could be in general not feasible. In this paper we test a matrix-free method that addresses the problems of large data sets and reduces the computational cost and memory requirements for the reconstruction. More specifically we challenged the Matrix-Free method with in-vivo measurements from mice where fluorescence tubes of different but controlled concentrations are inserted, to assess the quantification performance of the method. We extended the test with simulations, using realistic geometries extracted from a mouse-atlas and including prior known information about the optical properties of tissue into the forward model.

Zacharopoulos, Athanasios; Garofalakis, Anikitos; Ripoll, Jorge; Arridge, Simon

2010-11-01

324

Development of a noncontact 3-D fluorescence tomography system for small animal in vivo imaging  

NASA Astrophysics Data System (ADS)

Fluorescence imaging is an important tool for tracking molecular-targeting probes in preclinical studies. It offers high sensitivity, but nonetheless low spatial resolution compared to other leading imaging methods such CT and MRI. We demonstrate our methodological development in small animal in vivo whole-body imaging using fluorescence tomography. We have implemented a noncontact fluid-free fluorescence diffuse optical tomography system that uses a raster-scanned continuous-wave diode laser as the light source and an intensified CCD camera as the photodetector. The specimen is positioned on a motorized rotation stage. Laser scanning, data acquisition, and stage rotation are controlled via LabVIEW applications. The forward problem in the heterogeneous medium is based on a normalized Born method, and the sensitivity function is determined using a Monte Carlo method. The inverse problem (image reconstruction) is performed using a regularized iterative algorithm, in which the cost function is defined as a weighted sum of the L-2 norms of the solution image, the residual error, and the image gradient. The relative weights are adjusted by two independent regularization parameters. Our initial tests of this imaging system were performed with an imaging phantom that consists of a translucent plastic cylinder filled with tissue-simulating liquid and two thin-wall glass tubes containing indocyanine green. The reconstruction is compared to the output of a finite element method-based software package NIRFAST and has produced promising results.

Zhang, Xiaofeng; Badea, Cristian; Jacob, Mathews; Johnson, G. Allan

2009-02-01

325

In vivo interstitial glucose characterization and monitoring in the skin by ATR-FTIR spectroscopy  

Microsoft Academic Search

Successful development of real-time non-invasive glucose monitoring would represent a major advancement not only in the treatment and management of patients with diabetes mellitus and carbohydrate metabolism disorders, but also for understanding in those biochemical, metabolic and (patho-)physiological processes of glucose at the molecular level in vivo. Here, ATR-FTIR spectroscopy technique has been challenged not only for in vivo measurement

Natalja Skrebova Eikje

2011-01-01

326

MRS Sparse-FFT: Reducing Acquisition Time and Artifacts for In Vivo 2D Correlation Spectroscopy  

E-print Network

MRS Sparse-FFT: Reducing Acquisition Time and Artifacts for In Vivo 2D Correlation Spectroscopy Sparse FFT (N1=64) FFT (N1=64) Full FFT (N1=180) CS (N1=64) Fig. 1. Phantom Data Result Sparse FFT (N1=60) FFT (N1=60) Full FFT (N1=160) CS (N1=60) Fig. 4. In Vivo Data Result Fig. 3. Phantom Data (f2=4.4 ppm

327

Environmental Laboratory Exercise: Analysis of Hydrogen Peroxide by Fluorescence Spectroscopy  

NASA Astrophysics Data System (ADS)

In this experiment, students analyze precipitation samples for trace concentrations of H2O2 using a newly developed fluorescence technique (Fenton method). Analysis is based on the production of OH radical by the reaction between H2O2 and ferrous ion (Fenton's reagent) with subsequent radical scavenging by benzoic acid. Unlike standard spectroscopic and titrimetric techniques, Fenton--hydrobenzoic acid analysis is sensitive enough for the determination of ambient peroxide in precipitation. In contrast to a more sensitive enzyme-based method, the reagents required are inexpensive and easily available. Reagent solutions are stable, making this experiment well-suited for the undergraduate laboratory.

Weinstein--Lloyd, Judith; Lee, Jai H.

1995-11-01

328

Classification evaluation of tobaccos using LED-induced fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Tobacco is one of the most important economic crops in the world, assessment of its quality has a very important business significance. A compact, low-cost, and maneuverable optical sensor system for classification evaluation of different tobaccos was described in this paper using light-emitting-diodes (LEDs)-induced fluorescence. The principal components analysis (PCA) method is used to extract the dominant features of the tobaccos for identifying the classification of tobaccos. The technique is suitable for practical identification due to the use of a straightforward data evaluation method and compact system.

Zhong, Weijia; Dong, Yongjiang; Liu, Xuan; Lin, Hongze; Mei, Liang; Yan, Chunsheng

2014-02-01

329

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

330

Synchronous fluorescence spectroscopy: tool for monitoring thermally stressed edible oils.  

PubMed

Total synchronous fluorescence spectra are proposed for monitoring edible oils during thermal stress. Synchronous fluorescence spectra obtained at an 80 nm wavelength interval combined with principal component analysis are suitable for classification of vegetable oil deterioration. Spectroscopic features in the range of 300-500 nm have been used for extra virgin olive, olive pomace, and sesame oil and the range of 320-520 nm has been used for corn, soybean, sunflower, and a commercial blend of oils. The score in the first two principal components explains 91.1% of the data matrix variance for extra virgin olive, sesame, and olive pomace oil and 89.3% for corn, soybean, sunflower, and the commercial blend of oils. The objective of this study is to develop a rapid method for the prediction of edible oil quality during thermal stress. Spectroscopic changes are indicative of oxidative deterioration as measured through wet chemistry methods: peroxide value, p-anisidine value, totox value, and radical-scavenging capacity. PMID:19722493

Poulli, Konstantina I; Chantzos, Nickolaos V; Mousdis, George A; Georgiou, Constantinos A

2009-09-23

331

Laser-induced fluorescence spectroscopy of human normal and cancerous tissues  

NASA Astrophysics Data System (ADS)

Fluorescence properties of stomach tissues have been investigated to determine whether malignant specimens can be discriminated in vitro. Differences between normal and tumor tissues are found that concern both the intensity distribution and spectral shape of the autofluorescence emission. According to the distinctions tumor tissues would be differed from the normal tissues. The resulting spectra could be differentiate histologically stomach abnormal tissues from normal tissues with a sensitivity and specificity value of 90 percent and 92 percent. Furthermore, fluorescence spectra from human stomach normal and malignant tissues have been measured in real time in vivo.

Chen, Weimin; He, Bin; Wei, Guang Hui; Gao, Ge; Li, Shirong

1996-09-01

332

Fluorescence and absorption spectroscopy of the weakly fluorescent chlorophyll a in cytochrome b6f of Synechocystis PCC6803.  

PubMed Central

A spectroscopic characterization of the chlorophyll a (Chl) molecule in the monomeric cytochrome b6f complex (Cytb6f) isolated from the cyanobacterium Synechocystis PCC6803 is presented. The fluorescence lifetime and quantum yield have been determined, and it is shown that Chl in Cytb6f has an excited-state lifetime that is 20 times smaller than that of Chl in methanol. This shortening of the Chl excited state lifetime is not caused by an increased rate of intersystem crossing. Most probably it is due to quenching by a nearby amino acid. It is suggested that this quenching is a mechanism for preventing the formation of Chl triplets, which can lead to the formation of harmful singlet oxygen. Using site-selected fluorescence spectroscopy, detailed information on vibrational frequencies in both the ground and Qy excited states has been obtained. The vibrational frequencies indicate that the Chl molecule has one axial ligand bound to its central magnesium and accepts a hydrogen bond to its 13(1)-keto carbonyl. The results show that the Chl binds to a well-defined pocket of the protein and experiences several close contacts with nearby amino acids. From the site-selected fluorescence spectra, it is further concluded that the electron-phonon coupling is moderately strong. Simulations of both the site-selected fluorescence spectra and the temperature dependence of absorption and fluorescence spectra are presented. These simulations indicate that the Huang-Rhys factor characterizing the electron-phonon coupling strength is between 0.6 and 0.9. The width of the Gaussian inhomogeneous distribution function is 210 +/- 10 cm-1. PMID:9649396

Peterman, E J; Wenk, S O; Pullerits, T; Pâlsson, L O; van Grondelle, R; Dekker, J P; Rögner, M; van Amerongen, H

1998-01-01

333

Detection of macrophage activity in atherosclerosis in vivo using multichannel, high-resolution laser scanning fluorescence microscopy  

Microsoft Academic Search

Molecular and cellular mechanisms of atherogenesis and its treatment are largely being unraveled by in vitro techniques. We describe methodology to directly image macrophage cell activity in vivo in a murine model of atherosclerosis using laser scanning fluorescence microscopy (LSFM) and a macrophage-targeted, near-infrared fluorescent (NIRF) magnetofluorescent nanoparticle (MFNP). Atherosclerotic apolipoprotein E deficient (apoE -\\/-) mice (n=10) are injected with

Ashvin N. Pande; Rainer H. Kohler; Elena Aikawa; Ralph Weissleder; Farouc A. Jaffer

2006-01-01

334

Organ transplant tissue rejection: detection and staging by fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

Patients receiving heart or other organ transplants usually require some level of anti-rejection drug therapy, most commonly cyclosporine. The rejection status of the organ must be monitored to determine the optimal anti-rejection drug therapy. The current method for monitoring post-transplant rejection status of heart transplant patients consists of taking biopsies from the right ventricle. In this work we have developed a system employing optical and signal-processing techniques that will allow a cardiologist to measure spectral changes associated with tissue rejection using an optical catheter probe. The system employs time gated illumination and detection systems to deal with the dynamic signal acquisition problems associated with in vivo measurements of a beating heart. Spectral data processing software evaluates and processes the data to produce a simple numerical score. Results of measurements made on 100 excised transplanted isograft and allograft rat hearts have demonstrated the ability of the system to detect the presence of rejection and to accurately correlate the spectroscopic results with the ISHLT (International Society for Heart and Lung Transplantation) stage of rejection determined by histopathology. In vivo measurements using a pig transplant model are now in process.

MacAulay, Calum E.; Whitehead, Peter D.; McManus, Bruce; Zeng, Haishan; Wilson-McManus, Janet; MacKinnon, Nick; Morgan, David C.; Dong, Chunming; Gerla, Paul; Kenyon, Jennifer

1998-07-01

335

Use of a Microscope Photometer To Analyze In Vivo Fluorescence Intensity of Epilithic Microalgae Grown on Artificial Substrata  

PubMed Central

An epifluorescence microscope photometer was used to develop a new, in vivo fluorimetric method for analyzing fluorescence intensities of epilithic microalgae grown on clay tiles in the field. This enabled a nondestructive, direct quantification of algal biomass on the substratum surface. Measurements of a chlorophyll a standard in ethanol (90%) with our fluorimetric method (exitation at 546 nm; emission, >590 nm) correlated well with those from conventional spectrofluorimetric and spectrophotometric methods. Biofilms were analyzed with the microscope photometer by measuring the in vivo fluorescence intensity of 70 spots distributed randomly over the tile surface. They were then analyzed by the two in vitro methods after photopigment extraction. Chlorophyll a content and in vivo fluorescence intensity correlated well. The regression curves were linear up to 6 (mu)g cm(sup-2) but were quadratic or hyperbolic at higher concentrations of up to 28 (mu)g cm(sup-2). The degree of scatter among individual measurements was higher in biofilms than chlorophyll a standards. This in vivo analysis is well suited to ecological experiments and has the advantage of measuring on an extremely small scale, which enables direct analysis of the microdistribution of epilithic microalgae in live biofilms. We demonstrated this by comparing fluorescence intensities of the grazing tracks of the snail Ancylus fluviatilis with those of ungrazed areas. Our in vivo analysis is also unique in enabling biofilms on artificial substrata to be removed, analyzed, and then returned intact in field or laboratory experiments. PMID:16535568

Becker, G.; Holfeld, H.; Hasselrot, A. T.; Fiebig, D. M.; Menzler, D. A.

1997-01-01

336

High-content data evaluation by means of confocal fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

With the development of fluorescence correlation spectroscopy (FCS) in the 1990s, a fundamental milestone was set in the field of highly resolved and quantitative fluorescence detection. Moreover, the increasing knowledge about the meaning of confocal fluorescence detection and its experienced handling enabled unrivalled degrees of detection sensitivity. In the end of the decade, hence the possibility of detecting single fluorescent molecules initiated a productive scientific rush for a comprehensive exploitation of fluorescence properties on the single molecule level. Meanwhile, confocal fluorescence spectroscopy has overcome its predominantly scientific meaning in basic research, and rather found wide applications even in the life science industry. However, biological assay systems relevant for industrial dedication mainly require reagent concentrations above those of classical single molecule detection. They rather lie within the "molecular fluctuation range", which means that in the temporal average a plurality of fluorescent particles rather than only one is present within the confocal detection area at a time. Thus, although individual molecules may not longer be resolved, their diffusive fluctuations are furthermore visible and contain a valuable amount of information. On the other hand FCS, a typical fluctuation evaluation technique, is restricted to the quantification of translational molecular diffusion, which in a number of cases is not sufficient to characterize the biological system of interest. Hence, a series of additional complementary fluctuation and non-fluctuation based confocal spectroscopic techniques was developed during recent years and named FCS+plus. They provide simultaneous access to a multitude of molecular properties like concentration, translational and rotational diffusion, molecular brightness, coincidence and fluorescence lifetime and thus meet the needs of both scientific research and industrial application. In the following particular aspects of molecular polarization will be shortly described and illustrated by a comparison of stationary and time-resolved anisotropy. Another valuable subject in especially industrial application of fluorescence is that of artificially interfering effects. The most prominent of these disturbances is given with the potential "auto-fluorescence" of non-labeled biological molecules. In these - frequently appearing - cases the wanted signal of fluorescently labeled material will be superimposed by artifacts, making a proper data interpretation rather difficult. However, the FCS+plus' abilities of decomposing a fluorescence signal into the molecular species' fractional contributions enables a sophisticated consideration of unwanted interferences.

Brand, L.; Schroers, A.

2005-03-01

337

Assessing gene expression in vivo: magnetic resonance imaging and spectroscopy  

Microsoft Academic Search

Recent developments in magnetic resonance imaging and spectroscopy afford the possibility of detecting and assessing transfer, expression and subsequent therapeutic changes of effector or marker transgenes noninvasively. In the field of MR imaging, ‘smart’ MR contrast agents are being developed, so called because they change their conformational structure and in so doing induce MR detectable changes in a given tissue.

JD Bell; SD Taylor-Robinson

2000-01-01

338

Rapid detection of authenticity and adulteration of walnut oil by FTIR and fluorescence spectroscopy: A comparative study.  

PubMed

Fourier transform infrared (FTIR) and fluorescence spectroscopy combined with soft independent modeling of class analogies (SIMCA) and partial least square (PLS) were used to detect the authenticity of walnut oil and adulteration amount of soybean oil in walnut oil. A SIMCA model of FTIR spectra could differentiate walnut oil and other oils into separate categories; the classification limit of soybean oil in walnut oil was 10%. Fluorescence spectroscopy could differentiate oil composition by the peak position and intensity of emission spectrum without multivariate analysis. The classification limit of soybean oil adulterated in walnut oil by fluorescence spectroscopy was below 5%. The deviation of the prediction model for fluorescence spectra was lower than that for FTIR spectra. Fluorescence spectroscopy was more applicable than FTIR in the adulteration detection of walnut oil, both from the determination limit and prediction deviation. PMID:25794716

Li, Bingning; Wang, Haixia; Zhao, Qiaojiao; Ouyang, Jie; Wu, Yanwen

2015-08-15

339

Objective Assessment of Endogenous Collagen In Vivo during Tissue Repair by Laser Induced Fluorescence  

PubMed Central

Collagen, a triple helical protein with the primary role of mechanical function, provides tensile strength to the skin, and plays a pivotal task in tissue repair. During tissue regeneration, collagen level increases gradually and therefore, monitoring of such changes in vivo by laser induced fluorescence was the main objective behind the present study. In order to accomplish this, 15 mm diameter excisional wounds were created on six to eight week old Swiss albino mice. The collagen deposition accelerated upon irradiation of single exposure of 2 J/cm2 He-Ne laser dose immediately after wounding was recorded by laser induced autofluorescence in vivo along with un-illuminated and un-wounded controls. Autofluorescence spectra were recorded for each animal of the experimental groups on 0, 5, 10, 30, 45 and 60 days post-wounding, by exciting the granulation tissue/skin with 325 nm He-Cd laser. The variations in the average collagen intensities from the granulation tissue/skin of mice were inspected as a function of age and gender. Further, the spectral findings of the collagen synthesis in wound granulation tissue/un-wounded skin tissues were validated by Picro-Sirius red- polarized light microscopy in a blinded manner through image analysis of the respective collagen birefringence. The in vivo autofluorescence studies have shown a significant increase in collagen synthesis in laser treated animals as compared to the un-illuminated controls. Image analysis of the collagen birefringence further authenticated the ability of autofluorescence in the objective monitoring of collagen in vivo. Our results clearly demonstrate the potential of laser induced autofluorescence in the monitoring of collegen synthesis during tissue regeneration, which may have clinical implications. PMID:24874229

Prabhu, Vijendra; Rao, Satish B. S.; Fernandes, Edward Mark; Rao, Anuradha C. K.; Prasad, Keerthana; Mahato, Krishna K.

2014-01-01

340

Noninvasive Fluorescence Resonance Energy Transfer Imaging of in vivo Premature Drug Release from Polymeric Nanoparticles  

PubMed Central

Understanding in vivo drug release kinetics is critical for the development of nanoparticle-based delivery systems. In this study, we developed a fluorescence resonance energy transfer (FRET) imaging approach to noninvasively monitor in vitro and in vivo cargo release from polymeric nanoparticles. The FRET donor dye (DiO or DiD) and acceptor dye (DiI or DiR) were individually encapsulated into poly(ethylene oxide)-b-polystyrene (PEO-PS) nanoparticles. When DiO (donor) nanoparticles and DiI (acceptor) nanoparticles were co-incubated with cancer cells for 2 h, increased FRET signals were observed from cell membranes, suggesting rapid release of DiO and DiI to cell membranes. Similarly, increased FRET ratios were detected in nude mice after intravenous co-administration of DiD (donor) nanoparticles and DiR (acceptor) nanoparticles. In contrast, another group of nude mice i.v. administrated with DiD/DiR co-loaded nanoparticles showed decreased FRET ratios. Based on the difference in FRET ratios between the two groups, in vivo DiD/DiR release half-life from PEO-PS nanoparticles was determined to be 9.2 min. In addition, it was observed that the presence of cell membranes facilitated burst release of lipophilic cargos while incorporation of oleic acid-coated iron oxide into PEO-PS nanoparticles slowed the release of DiD/DiR to cell membranes. The developed in vitro and in vivo FRET imaging techniques can be used to screening stable nano-formulations for lipophilic drug delivery. PMID:24033270

Zou, Peng; Chen, Hongwei; Paholak, Hayley J.; Sun, Duxin

2013-01-01

341

Application of fluorescence spectroscopy using a novel fluoroionophore for quantification of zinc in urban runoff.  

PubMed

Fluorescence spectroscopy has great potential for on-site and real-time monitoring of pollutants in aquatic environments; however, its application to environmental aquatic samples has been extremely limited. In this study, a novel fluoroionophore based on a BODIPY-terpyridine conjugate was developed and applied to determine Zn concentrations in urban runoff. The fluoroionophore selectively bound to Zn(2+) in water, which led to an instant red-shift of the fluorescence peak of the fluoroionophore from 539 nm to 567 nm that could be seen by the naked eye. Zn concentrations could be quantified using the ratio of fluorescence intensities, and the detection limit was 9 ?g/L, which is sufficiently low for environmental aquatic samples. To demonstrate applicability of the method to environmental samples, we measured Zn concentrations in urban runoff samples with a complex matrix (?60 mg/L dissolved organic carbon and ?20 mS/cm electrical conductivity). The total and dissolved fractions of Zn in the samples could be determined by fluorescence spectroscopy and its relative error was estimated to be less than 30% by inductively coupled plasma-atomic emission spectroscopy analysis. The proposed method is rapid and easy-to-use with simple pretreatment for Zn determination in environmental aquatic samples with complex matrices. PMID:24531076

Hafuka, Akira; Yoshikawa, Hiroaki; Yamada, Koji; Kato, Tsuyoshi; Takahashi, Masahiro; Okabe, Satoshi; Satoh, Hisashi

2014-05-01

342

Towards in situ fluorescence spectroscopy and microscopy investigations of asphaltene precipitation kinetics.  

PubMed

We perform a spectroscopic analysis of asphaltene in solution and in crude oil with the goal of designing an optical probe of asphaltene precipitation inside high-pressure cells. Quantitative analysis of steady-state spectroscopic data is employed to identify fluorescence and Raman contributions to the observed signals. Time-resolved fluorescence spectroscopy indicates that fluorescence lifetime can be used as a spectroscopic probe of asphaltene in crude oil. Quantitative confocal laser-scanning microscopy studies of asphaltene in n-heptane are used to calculate particle-size distributions as a function of time, both at the sample surface and asphaltene interior. The resulting precipitation kinetics is well described by stochastic numerical simulations of diffusion-limited aggregation. Based on these results, we present the design and construction of an apparatus to optically probe the in situ precipitation of asphaltene suitable for studies inside high pressure cells. Design considerations include the use of a spatial light modulator for aberration correction in microscopy measurements, together with the design of epi-fluorescence spectrometer, both fiber-based and for remote sensing fluorescence spectroscopy. PMID:24514660

Franco, Juliana C; Gonçalves, Grasiele; Souza, Monique S; Rosa, Samantha B C; Thiegue, Larissa M; Atvars, Teresa D Z; Rosa, Paulo T V; Nome, René A

2013-12-16

343

Fluorescence spectroscopy as a tool for determination of coumarins by multivariate calibration.  

PubMed

At present, it is necessary to check the quality of many food products in which the content of coumarins is limited. Since a rapid and simple method for the determination of coumarin (COU), 4-hydroxycoumarin (4HC) and dicoumarol (DC) in tea samples was needed, we developed an alternative option to chromatography, i.e., fluorescence spectroscopy with multivariate calibration. The synchronous fluorescence spectra were recorded at constant wavelength differences 70, 80 and 90 nm from 200 to 400 nm. The different experimental parameters affecting the synchronous fluorescence intensities of the analytes were carefully studied and optimized. Partial least squares (PLS) method and multi linear regression (MLR) were compared on determining the concentrations. The best results were obtained by the PLS method on synchronous fluorescence spectra at ???=?90 nm. The results from the analysis of herbal tea Melilotus officinalis by synchronous fluorescence spectroscopy with PLS model are equivalent with the results from HPLC. Fisher F- test and Student's t- test confirmed this finding. PMID:25595059

Polá?ek, Roman; Májek, Pavel; Hrobo?ová, Katarína; Sádecká, Jana

2015-03-01

344

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

345

Strengths and Weaknesses of Recently Engineered Red Fluorescent Proteins Evaluated in Live Cells Using Fluorescence Correlation Spectroscopy  

PubMed Central

The scientific community is still looking for a bright, stable red fluorescent protein (FP) as functional as the current best derivatives of green fluorescent protein (GFP). The red FPs exploit the reduced background of cells imaged in the red region of the visible spectrum, but photophysical short comings have limited their use for some spectroscopic approaches. Introduced nearly a decade ago, mCherry remains the most often used red FP for fluorescence correlation spectroscopy (FCS) and other single molecule techniques, despite the advent of many newer red FPs. All red FPs suffer from complex photophysics involving reversible conversions to a dark state (flickering), a property that results in fairly low red FP quantum yields and potential interference with spectroscopic analyses including FCS. The current report describes assays developed to determine the best working conditions for, and to uncover the shortcoming of, four recently engineered red FPs for use in FCS and other diffusion and spectroscopic studies. All five red FPs assayed had potential shortcomings leading to the conclusion that the current best red FP for FCS is still mCherry. The assays developed here aim to enable the rapid evaluation of new red FPs and their smooth adaptation to live cell spectroscopic microscopy and nanoscopy. PMID:24129172

Siegel, Amanda P.; Baird, Michelle A.; Davidson, Michael W.; Day, Richard N.

2013-01-01

346

Identification of a Novel Indoline Derivative for in Vivo Fluorescent Imaging of Blood-Brain Barrier Disruption in Animal Models  

PubMed Central

Disruption of the blood-brain barrier (BBB) can occur in various pathophysiological conditions. Administration of extraneous tracers that can pass the disrupted, but not the intact, BBB and detection of the extravasation have been widely used to assess BBB disruption in animal models. Although several fluorescent tracers have been successfully used, the administration of these tracers basically requires intravascular injection, which can be laborious when using small animals such as zebrafish. To identify fluorescent tracers that could be easily administered into various animal models and visualize the BBB disruption in vivo, we prepared nine structurally related indoline derivatives (IDs) as a minimum set of diverse fluorescent compounds. We found that one ID, ZMB741, had the highest affinity for serum albumin and emitted the strongest fluorescence in the presence of serum albumin of the nine IDs tested. The affinity to serum albumin and the fluorescence intensity was superior to those of Evans blue and indocyanine green that have been conventionally used to assess the BBB disruption. We showed that ZMB741 could be administered into zebrafish by static immersion or mice by intraperitoneal injection and visualizes the active disruption of their BBB. These results suggest that ZMB741 can be a convenient and versatile tool for in vivo fluorescent imaging of BBB disruption in various animal models. The strategy used in this study can also be applied to diversity-oriented libraries to identify novel fluorescent tracers that may be superior to ZMB741. PMID:23668665

2013-01-01

347

Two-photon-excited fluorescence spectroscopy of atomic fluorine at 170 nm  

NASA Astrophysics Data System (ADS)

Two-photon-excited fluorescence spectroscopy of atomic fluorine is reported. A doubled dye laser at 286-nm is Raman shifted in H2 to 170 nm (sixth anti-Stokes order) to excite ground-state 2P(0)J fluorine atoms to the 2D(0)J level. The fluorine atoms are detected by one of two methods: observing the fluorescence decay to the 2PJ level or observing F(+) production through the absorption of an additional photon by the excited atoms. Relative two-photon absorption cross sections to and the radiative lifetimes of the 2D(0)J states are measured.

Herring, G. C.; Dyer, Mark J.; Jusinski, Leonard E.; Bischel, William K.

1988-05-01

348

Highly sensitive real-time detection of DNA hybridization by using nanoporous waveguide fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

In this Letter, we report highly sensitive fluorescence spectroscopy using a nanoporous waveguide (NPWG) comprising a porous anodic alumina (PAA) layer and an Al layer. Simulations show that the TE0 waveguide mode excited in the PAA layer produces an electromagnetic field whose intensity is 40-fold higher than that of the incident light, and which yields enhanced intensity when used to excite fluorophores. We demonstrate the sensing ability of the NPWG by incorporating it into a fluorescent sensor to monitor duplex DNA formation in real-time, with a detection limit as low as 20 pM.

Fan, Yong; Hotta, Kazuhiro; Yamaguchi, Akira; Ding, Yu; He, Yonghong; Teramae, Norio; Sun, Shuqing; Ma, Hui

2014-07-01

349

Integrated fluorescence correlation spectroscopy device for point-of-care clinical applications  

PubMed Central

We describe an optical system which reduces the cost and complexity of fluorescence correlation spectroscopy (FCS), intended to increase the suitability of the technique for clinical use. Integration of the focusing optics and sample chamber into a plastic component produces a design which is simple to align and operate. We validate the system by measurements on fluorescent dye, and compare the results to a commercial instrument. In addition, we demonstrate its application to measurements of concentration and multimerization of the clinically relevant protein von Willebrand factor (vWF) in human plasma. PMID:23847733

Olson, Eben; Torres, Richard; Levene, Michael J.

2013-01-01

350

Two-photon-excited fluorescence spectroscopy of atomic fluorine at 170 nm  

NASA Technical Reports Server (NTRS)

Two-photon-excited fluorescence spectroscopy of atomic fluorine is reported. A doubled dye laser at 286-nm is Raman shifted in H2 to 170 nm (sixth anti-Stokes order) to excite ground-state 2P(0)J fluorine atoms to the 2D(0)J level. The fluorine atoms are detected by one of two methods: observing the fluorescence decay to the 2PJ level or observing F(+) production through the absorption of an additional photon by the excited atoms. Relative two-photon absorption cross sections to and the radiative lifetimes of the 2D(0)J states are measured.

Herring, G. C.; Dyer, Mark J.; Jusinski, Leonard E.; Bischel, William K.

1988-01-01

351

Portable fluorescence spectroscopy platform for Huanglongbing (HLB) citrus disease in situ detection  

NASA Astrophysics Data System (ADS)

In this work, the development of a portable fluorescence spectroscopy platform for Huanglongbing (HLB) citrus disease in situ detection is presented. The equipment consists of an excitation blue LED light source, a commercial miniature spectrometer and embedded software. Measurements of healthy, HLB-symptomatic and HLB-asymptomatic citrus leafs were performed. Leafs were excited with the blue LED and their fluorescence spectra collected. Embedded electronics and software were responsible for the spectrum processing and classification via partial least squares regression. Global success rates above 80% and 100% distinction of healthy and HLB-symptomatic leafs were obtained.

Mota, Alessandro D.; Rossi, Giuliano; de Castro, Guilherme Cunha; Ortega, Tiago A.; de Castro N., Jarbas C.

2014-02-01

352

Subcellular In Vivo 1 HM R Spectroscopy ofXenopus laevis Oocytes  

Microsoft Academic Search

In vivo magnetic resonance (MR) spectra are typically obtained from voxels whose spatial dimensions far exceed those of the cells they contain. This study was designed to evaluate the potential of localized MR spectroscopy to investigate subcellular phenomena. Using a high magnetic field and a home-built microscopy probe with large gradient field strengths, we achieved voxel sizes of (180 mm)3.

Seung-Cheol Lee; Jee-Hyun Cho; Daniel Mietchen; Young-Sook Kim; Kwan Soo Hong; Chulhyun Lee; Dongmin Kang; Byong-Seok Choi; Chaejoon Cheong

2006-01-01

353

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

354

In Vivo X-Ray Fluorescence Microtomographic Imaging of Elements in Single-Celled Fern Spores  

SciTech Connect

We have observed in vivo three-dimensional distributions of constituent elements of single-celled spores of the fern Adiantum capillus-veneris using an X-ray fluorescence computed microtomography method. The images of these distributions are generated from a series of slice data, each of which is acquired by a sample translation-rotation method. An incident X-ray microbeam irradiates the sample with a spot size of 1 {mu}m. The high Ca concentration in the testa and the localized and overlapping Fe and Zn concentrations inside the spore are shown in three-dimensional images. The K concentration is high throughout the cell, and there are localized regions of higher density. The atomic number densities of these elements in the testa and inside the cell in a tomographic slice are estimated with a resolution of about 1 {mu}m.

Hirai, Yasuharu; Yoneyama, Akio; Hisada, Akiko [Advanced Research Laboratory, Hitachi, Ltd., Hatoyama, Saitama 350-0395 (Japan); Uchida, Kenko [Central Research Laboratory, Hitachi, Ltd., Kokubunji, Tokyo 185-8601 (Japan)

2007-01-19

355

Imaging leukocyte trafficking in vivo with two-photon-excited endogenous tryptophan fluorescence  

PubMed Central

We describe a new method for imaging leukocytes in vivo by exciting the endogenous protein fluorescence in the ultraviolet (UV) spectral region where tryptophan is the major fluorophore. Two-photon excitation near 590 nm allows noninvasive optical sectioning through the epidermal cell layers into the dermis of mouse skin, where leukocytes can be observed by video-rate microscopy to interact dynamically with the dermal vascular endothelium. Inflammation significantly enhances leukocyte rolling, adhesion, and tissue infiltration. After exiting the vasculature, leukocytes continue to move actively in tissue as observed by time-lapse microscopy, and are distinguishable from resident autofluorescent cells that are not motile. Because the new method alleviates the need to introduce exogenous labels, it is potentially applicable for tracking leukocytes and monitoring inflammatory cellular reactions in humans. PMID:20173920

Li, Chunqiang; Pastila, Riikka K.; Pitsillides, Costas; Runnels, Judith M.; Puoris’haag, Mehron; Côté, Daniel; Lin, Charles P.

2010-01-01

356

Optimization of holographic optical tweezers for multiplexed fluorescence spectroscopy  

NASA Astrophysics Data System (ADS)

We are developing a multiplexed spectroscopy technique that employs holographic optical tweezers to trap and excite multiple sensor particles. Our goal is to develop a lab-on-a-chip measurement platform for monitoring pH and other ion concentrations with high spatial resolution in a microfluidic device or within biological cells. To ensure efficient use of the available laser power required to trap multiple particles, we address three aspects of the spatial light modulator (SLM) used in the holographic technique. We measure and optimize the input and output polarizations used before and after the birefringent SLM. We reduce optical aberrations by adding appropriate Zernike polynomials to the computed hologram. We optimize the diffraction efficiency of the SLM by adjusting the gray scale input-to-output table to account for the nonlinear phase response of the SLM.

Cibula, Matthew; McIntyre, David

2012-10-01

357

Electron multiplying charge-coupled device-based fluorescence cross-correlation spectroscopy for blood velocimetry on zebrafish embryos  

NASA Astrophysics Data System (ADS)

Biomedical issues in vasculogenesis and cardiogenesis require methods to follow hemodynamics with high spatial (micrometers) and time (milliseconds) resolution. At the same time, we need to follow relevant morphogenetic processes on large fields of view. Fluorescence cross-correlation spectroscopy coupled to scanning or wide-field microscopy meets these needs but has limited flexibility in the excitation pattern. To overcome this limitation, we develop here a two-photon two-spots setup coupled to an all-reflective near-infrared (NIR) optimized scanning system and to an electron multiplying charge-coupled device. Two NIR laser spots are spaced at adjustable micron-size distances (1 to 50 ?m) by means of a Twyman-Green interferometer and repeatedly scanned on the sample, allowing acquisition of information on flows at 4 ms-3 ?m time-space resolution in parallel on an extended field of view. We analyze the effect of nonhomogeneous and variable flow on the cross-correlation function by numerical simulations and show exemplary application of this setup in studies of blood flow in zebrafish embryos in vivo. By coupling the interferometer with the scanning mirrors and by computing the cross-correlation function of fluorescent red blood cells, we are able to map speed patterns in embryos' vessels.

Pozzi, Paolo; Sironi, Laura; D'Alfonso, Laura; Bouzin, Margaux; Collini, Maddalena; Chirico, Giuseppe; Pallavicini, Piersandro; Cotelli, Franco; Foglia, Efrem A.

2014-06-01

358

31P NMR spectroscopy of in vivo tumors  

NASA Astrophysics Data System (ADS)

A probe, suitable for any wide-bore NMR spectrometer, was constructed for monitoring high-resolution spectra of in vivo subcutaneously implanted tumors in mice. Preliminary studies of a variety of murine tumors (MOPC 104E myeloma, Dunn osteosarcoma, colon-26, ovarian M5, and mammary adenocarcinoma as well as human colon, mammary, and lung tumors in athymic mice) indicate that the 31P NMR spectrum is a sensitive monitor of progressive metabolic changes that occur during untreated tumor growth and an early indicator of tumor response to chemotherapy, hyperthermia, and X radiation. Response to each of these therapeutic modalities is accompanied by distinctly different spectral changes.

Ng, T. C.; Evanochko, W. T.; Hiramoto, R. N.; Ghanta, V. K.; Lilly, M. B.; Lawson, A. J.; Corbett, T. H.; Durant, J. R.; Glickson, J. D.

359

Detection of Non-Melanoma Skin Cancer by in vivo Fluorescence Imaging with Fluorocoxib A.  

PubMed

Non-melanoma skin cancer (NMSC) is the most common form of cancer in the US and its incidence is increasing. The current standard of care is visual inspection by physicians and/or dermatologists, followed by skin biopsy and pathologic confirmation. We have investigated the use of in vivo fluorescence imaging using fluorocoxib A as a molecular probe for early detection and assessment of skin tumors in mouse models of NMSC. Fluorocoxib A targets the cyclooxygenase-2 (COX-2) enzyme that is preferentially expressed by inflamed and tumor tissue, and therefore has potential to be an effective broadly active molecular biomarker for cancer detection. We tested the sensitivity of fluorocoxib A in a BCC allograft SCID hairless mouse model using a wide-field fluorescence imaging system. Subcutaneous allografts comprised of 1000 BCC cells were detectable above background. These BCC allograft mice were imaged over time and a linear correlation (R(2) = 0.8) between tumor volume and fluorocoxib A signal levels was observed. We also tested fluorocoxib A in a genetically engineered spontaneous BCC mouse model (Ptch1(+/-) K14-Cre-ER2 p53(fl/fl)), where sequential imaging of the same animals over time demonstrated that early, microscopic lesions (100 ?m size) developed into visible macroscopic tumor masses over 11 to 17 days. Overall, for macroscopic tumors, the sensitivity was 88% and the specificity was 100%. For microscopic tumors, the sensitivity was 85% and specificity was 56%. These results demonstrate the potential of fluorocoxib A as an in vivo imaging agent for early detection, margin delineation and guided biopsies of NMSCs. PMID:25748239

Ra, Hyejun; González-González, Emilio; Uddin, Md Jashim; King, Bonnie L; Lee, Alex; Ali-Khan, Irfan; Marnett, Lawrence J; Tang, Jean Y; Contag, Christopher H

2015-02-01

360

Detection of Non-Melanoma Skin Cancer by in vivo Fluorescence Imaging with Fluorocoxib A  

PubMed Central

Non-melanoma skin cancer (NMSC) is the most common form of cancer in the US and its incidence is increasing. The current standard of care is visual inspection by physicians and/or dermatologists, followed by skin biopsy and pathologic confirmation. We have investigated the use of in vivo fluorescence imaging using fluorocoxib A as a molecular probe for early detection and assessment of skin tumors in mouse models of NMSC. Fluorocoxib A targets the cyclooxygenase-2 (COX-2) enzyme that is preferentially expressed by inflamed and tumor tissue, and therefore has potential to be an effective broadly active molecular biomarker for cancer detection. We tested the sensitivity of fluorocoxib A in a BCC allograft SCID hairless mouse model using a wide-field fluorescence imaging system. Subcutaneous allografts comprised of 1000 BCC cells were detectable above background. These BCC allograft mice were imaged over time and a linear correlation (R2 = 0.8) between tumor volume and fluorocoxib A signal levels was observed. We also tested fluorocoxib A in a genetically engineered spontaneous BCC mouse model (Ptch1+/? K14-Cre-ER2 p53fl/fl), where sequential imaging of the same animals over time demonstrated that early, microscopic lesions (100 ?m size) developed into visible macroscopic tumor masses over 11 to 17 days. Overall, for macroscopic tumors, the sensitivity was 88% and the specificity was 100%. For microscopic tumors, the sensitivity was 85% and specificity was 56%. These results demonstrate the potential of fluorocoxib A as an in vivo imaging agent for early detection, margin delineation and guided biopsies of NMSCs. PMID:25748239

Ra, Hyejun; González-González, Emilio; Uddin, Md. Jashim; King, Bonnie L.; Lee, Alex; Ali-Khan, Irfan; Marnett, Lawrence J.; Tang, Jean Y.; Contag, Christopher H.

2015-01-01

361

Quantifying receptor density in vivo using a dual probe approach with fluorescence molecular imaging  

NASA Astrophysics Data System (ADS)

Molecular imaging technologies are advancing rapidly and optical techniques in particular are set to play a large role in preclinical pharmaceutical testing. These approaches, however, are generally unable to quantify the level of expression of imaging probe reporters. In this study a novel method of quantification is presented using dual-probe fluorescence imaging, where an endothelial growth factor receptor (EGFR) fluorescent probe was paired with a non-targeted probe before being injected, and tracer kinetic compartmental modeling was used to determine EGFR expression in a region of interest from the uptake curves of the two drugs in that region. The approach was tested out in a simulation experiment and then applied in an in vivo study in one mouse to investigate EGFR expression in various tissue types (pancreas, pancreas tumor, and leg). The binding potentials (a unitless correlate of target availability) of EGFR expression in the various tissue types were 8.57, 25.64, and 0.11 for the pancreas, pancreas tumor, respectively. For the pancreas and leg, these results correlate well with expected levels of EGFR expression, with the pancreas demonstrating a much higher expression than the skin and also as expected, the tumor expressed much more EGFR than either healthy tissue.

Tichauer, Kenneth M.; Samkoe, Kimberley S.; O'Hara, Julia; Sexton, Kristian J.; Davis, Scott C.; Pogue, Brian W.

2011-03-01

362

Assessment of humification degree of dissolved organic matter from different composts using fluorescence spectroscopy technology.  

PubMed

This study was conducted to assess the degree of humification in dissolved organic matter (DOM) from different composts, and their environmental impact after soil amending based on fluorescence measurements (emission, excitation, synchronous scan, and excitation-emission matrix [EEM]). The compost sources studied included dairy cattle manure (DCM), kitchen waste (KW), cabbage waste (CW), tomato stem waste (TSW), municipal solid waste (MSW), green waste (GW), chicken manure (CM), and peat (P). Conventional and EEM fluorescence spectroscopy indicated that the DOM of these composts contained compounds similar in structure but comparisons between conventional fluorescence parameters and fluorescence regional integration of EEM fluorescence spectra showed that the DOM was different in degree of humification. Regression analysis demonstrated significant corrections between major fluorescence parameters. In hierarchical cluster analysis, these composts were clustered into 2 groups and 4 subgroups, and projection pursuit regression analysis further ranked the compost sources as KW, CW, P>CM, DCM, TW, GW>MSW in their degree of humification in DOM. PMID:24188626

Wei, Zimin; Zhao, Xinyu; Zhu, Chaowei; Xi, Beidou; Zhao, Yue; Yu, Xue

2014-01-01

363

Characterization of the stratum corneum lipid matrix using fluorescence spectroscopy.  

PubMed

Using fluorescence techniques, we studied the dynamics of the lipid bilayer matrix of human stratum corneum (SC) and compared the results with that of distearoyl-phosphatidylcholine (DSPC). We employed a series of 9-anthroyloxy fatty acids (AF) that partitioned into the bilayer, enabling us to evaluate this structure as a function of depth within the lamellae. With AF probes, the re-orientation of the fluorophore is known to be affected by the polarity, hydrogen bonding, and rigidity of the surrounding medium, altering the emission maximum and lifetime in the excited state. In addition, we evaluated quenching, in which iodide collides with the fluorophore, revealing information on the accessibility of the fluorophore located in the bilayer. The emission and lifetime data showed that the reorientation of the fluorophore in SC was more hindered than in DSPC, indicating that SC bilayers were more rigid than DSPC bilayers. Quenching data of both SC and DSPC indicated that the deeper the fluorophore was positioned in the bilayer, the less accessible it was to iodide, pointing to a gradient in accessibility. In addition, the quenching results also showed that the SC is less accessible to iodide than in DSPC. The observed differences in bilayer rigidity and quencher accessibility between the two systems can be explained by differences in lipid composition and hydration. Whereas the DSPC bilayer consists of phospholipids, SC bilayers are composed of more anhydrous lipids like cholesterol and ceramides, which form a tight bilayer packing. In this way SC lipids exist in a relatively anhydrous and rigid environment, forming an effective diffusion barrier to water and ions. PMID:9734822

Pechtold, L A; Abraham, W; Potts, R O

1998-08-01

364

Intensity dependence of the fluorescence lifetime of in vivo chlorophyll excited by a picosecond light pulse. [Chlorella pyrenoidosa  

Microsoft Academic Search

New data on intensity-dependent lifetimes indicate that all previous in vivo fluorescence studies of chlorophyll by picosecond techniques must be reinterpreted. Anomalously short lifetimes result from high-intensity effects due to exciton-exciton annihilation processes. Measurements in Chlorella pyrenoidosa with single-pulse, low-intensity excitation indicate a longer ''true'' lifetime of 650 +- 150 picoseconds.

A. J. Campillo; V. H. Kollman; S. L. Shapiro

1976-01-01

365

Laser-Assisted Cryosurgery in ex vivo Mice Hepatic Tissue: Viability Assays Using Green Fluorescent Protein  

PubMed Central

An experimental investigation is carried out to develop a novel approach to cryosurgery, where laser heating counteracts tissue freezing to better confine damage to the targeted cancerous tissue within a lethal low-temperature isothermal boundary—an approach we refer to as laser-assisted cryosurgery (LAC). The advantage of this procedure relative to conventional cryosurgery assisted with urethral warmers or cryoheaters is that laser heating provides volumetric rather than superficial heating, which leads to deeper penetration, more homogeneous tissue protection and better demarcation of the destructive freezing effect to a well-defined targeted volume. Tissue viability assays are performed using green fluorescence protein (GFP) as a viability marker and correlated with temperature history after performing LAC procedures on ex vivo mice hepatic tissue. The limit for cell denaturation at the irradiated surface predicted by GFP analysis is further confirmed using reverse transcription polymerase chain reaction (RT-PCR). In addition, the correlation between GFP fluorescence and cell viability and loss of GFP fluorescence in non-viable cells has been tested and validated by histological analysis using a standard cell viability measuring method (hematoxylin and eosin staining). Analysis of our experimental measurements show that reproducible thermal gradients (of 236 °C/cm) and predictable tissue necrosis can be reliably produced by LAC without exceeding temperature thresholds for cell denaturation (of Tsurf ? 48 °C) beyond preset tissue boundaries (with resolution of 0.1 °C/mm). The results have shown the feasibility of controlling temperatures at specified tissue locations to prevent hyperthermal or freezing damage. PMID:20963494

Duperray, B.; Godinez, F.; Guillén, G.; Slade, A.; Aguilar, G.

2010-01-01

366

Combined fluorescence and X-Ray tomography for quantitative in vivo detection of fluorophore.  

PubMed

Initial results from a novel dual modality preclinical imager which combines non-contact fluorescence tomography (FT) and x-ray computed tomography (CT) for preclinical functional and anatomical in vivo imaging are presented. The anatomical data from CT provides a priori information to the FT reconstruction to create overlaid functional and anatomical images with accurate localization and quantification of fluorophore distribution. Phantoms with inclusions containing Indocyanine-Green (ICG), and with heterogeneous backgrounds including iodine in compartments at different concentrations for CT contrast, have been imaged with the dual modality FT/CT system. Anatomical information from attenuation maps and optical morphological information from absorption and scattering maps are used as a priori information in the FT reconstruction. Although ICG inclusions can be located without the a priori information, the recovered ICG concentration shows 75% error. When the a priori information is utilized, the ICG concentration can be recovered with only 15% error. Developing the ability to accurately quantify fluorophore concentration in anatomical regions of interest may provide a powerful tool for in vivo small animal imaging. PMID:20082529

Barber, W C; Lin, Y; Nalcioglu, O; Iwanczyk, J S; Hartsough, N E; Gulsen, G

2010-02-01

367

In vivo wound healing diagnosis with second harmonic and fluorescence lifetime imaging  

NASA Astrophysics Data System (ADS)

Skin wounds heal when a series of cell lineages are triggered, followed by collagen deposition, to reconstruct damaged tissues. This study evaluates the regeneration of collagen and change in cellular metabolic rate in vivo during wound healing in rats, with second harmonic generation (SHG) and fluorescence lifetime imaging microscopy respectively. The metabolic rate of cells is reflected through the lifetime of the autofluorescence from the co-enzyme protein, reduced nicotinamide adenine dinucleotide, due to its change in the relative concentration of bound and free forms. A higher than normal cellular metabolic rate is observed during the first week of healing, which decreases gradually after eight days of wound formation. SHG signal intensity change indicates the net degradation of collagen during the inflammatory phase, and net regeneration begins on day five. Eventually, the quantity of collagen increases gradually to form a scar tissue as the final product. Importantly, this work demonstrates the feasibility of an in vivo imaging approach for a normal wound on rat skin, which has the potential to supplement the noninvasive clinical diagnosis of wounds.

Deka, Gitanjal; Wu, Wei-Wen; Kao, Fu-Jen

2013-06-01

368

In Vivo Tracking of Mesechymal Stem Cells Using Fluorescent Nanoparticles in an Osteochondral Repair Model  

PubMed Central

We devised and tested an in vivo system to monitor the migration of mesenchymal stem cells (MSCs) within the marrow cavity. In vitro studies confirmed that platelet-derived growth factor (PDGF)-AA had the most potent chemotactic effect of the tested factors, and possessed the greatest number of receptors in MSCs. MSCs were labeled with fluorescent nanoparticles and injected into the marrow cavity of nude rats through osteochondral defects created in the distal femur. The defects were sealed with HCF (heparin-conjugated fibrin) or PDGF-AA-loaded HCF. In the HCF-only group, the nanoparticle-labeled MSCs dispersed outside the marrow cavity within 3 days after injection. In the PDGF-AA-loaded HCF group, the labeled cells moved time-dependently for 14 days toward the osteochondral defect. HCF-PDGF in low dose (LD; 8.5 ng/µl) was more effective than HCF-PDGF in high dose (HD; 17 ng/µl) in recruiting the MSCs to the osteochondral defect. After 21 days, the defects treated with PDGF and transforming growth factor (TGF)-?1-loaded HCF showed excellent cartilage repair compared with other groups. Further studies confirmed that this in vivo osteochondral MSCs tracking system (IOMTS) worked for other chemoattractants (chemokine (C-C motif) ligand 2 (CCL2) and PDGF-BB). IOMTS can provide a useful tool to examine the effect of growth factors or chemokines on endogenous cartilage repair. PMID:22491215

Lee, Jong Min; Kim, Byung-Soo; Lee, Haeshin; Im, Gun-Il

2012-01-01

369

In vivo deep tissue fluorescence imaging of the murine small intestine and colon  

NASA Astrophysics Data System (ADS)

Recently we described a novel technical approach with enhanced fluorescence detection capabilities in two-photon microscopy that achieves deep tissue imaging, while maintaining micron resolution. This technique was applied to in vivo imaging of murine small intestine and colon. Individuals with Inflammatory Bowel Disease (IBD), commonly presenting as Crohn's disease or Ulcerative Colitis, are at increased risk for developing colorectal cancer. We have developed a Gi?2 gene knock out mouse IBD model that develops colitis and colon cancer. The challenge is to study the disease in the whole animal, while maintaining high resolution imaging at millimeter depth. In the Gi?2-/- mice, we have been successful in imaging Lgr5-GFP positive stem cell reporters that are found in crypts of niche structures, as well as deeper structures, in the small intestine and colon at depths greater than 1mm. In parallel with these in vivo deep tissue imaging experiments, we have also pursued autofluorescence FLIM imaging of the colon and small intestine-at more shallow depths (roughly 160?m)- on commercial two photon microscopes with excellent structural correlation (in overlapping tissue regions) between the different technologies.

Crosignani, Viera; Dvornikov, Alexander; Aguilar, Jose S.; Stringari, Chiara; Edwards, Roberts; Mantulin, Williams; Gratton, Enrico

2012-03-01

370

Combined Fluorescence and X-Ray Tomography for Quantitative In Vivo Detection of Fluorophore  

PubMed Central

Initial results from a novel dual modality preclinical imager which combines non-contact fluorescence tomography (FT) and x-ray computed tomography (CT) for preclinical functional and anatomical in vivo imaging are presented. The anatomical data from CT provides a priori information to the FT reconstruction to create overlaid functional and anatomical images with accurate localization and quantification of fluorophore distribution. Phantoms with inclusions containing Indocyanine-Green (ICG), and with heterogeneous backgrounds including iodine in compartments at different concentrations for CT contrast, have been imaged with the dual modality FT/CT system. Anatomical information from attenuation maps and optical morphological information from absorption and scattering maps are used as a priori information in the FT reconstruction. Although ICG inclusions can be located without the a priori information, the recovered ICG concentration shows 75% error. When the a priori information is utilized, the ICG concentration can be recovered with only 15% error. Developing the ability to accurately quantify fluorophore concentration in anatomical regions of interest may provide a powerful tool for in vivo small animal imaging. PMID:20082529

Barber, W. C.; Lin, Y.; Nalcioglu, O.; Iwanczyk, J. S.; Hartsough, N. E.; Gulsen, G.

2010-01-01

371

Spot Variation Fluorescence Correlation Spectroscopy Allows for Superresolution Chronoscopy of Confinement Times in Membranes  

PubMed Central

Resolving the dynamical interplay of proteins and lipids in the live-cell plasma membrane represents a central goal in current cell biology. Superresolution concepts have introduced a means of capturing spatial heterogeneity at a nanoscopic length scale. Similar concepts for detecting dynamical transitions (superresolution chronoscopy) are still lacking. Here, we show that recently introduced spot-variation fluorescence correlation spectroscopy allows for sensing transient confinement times of membrane constituents at dramatically improved resolution. Using standard diffraction-limited optics, spot-variation fluorescence correlation spectroscopy captures signatures of single retardation events far below the transit time of the tracer through the focal spot. We provide an analytical description of special cases of transient binding of a tracer to pointlike traps, or association of a tracer with nanodomains. The influence of trap mobility and the underlying binding kinetics are quantified. Experimental approaches are suggested that allow for gaining quantitative mechanistic insights into the interaction processes of membrane constituents. PMID:21641330

Ruprecht, Verena; Wieser, Stefan; Marguet, Didier; Schütz, Gerhard J.

2011-01-01

372

Classification and characterization of beef muscles using front-face fluorescence spectroscopy.  

PubMed

The objective of this study was to evaluate the potential of fluorescence spectroscopy to identify different muscles and to predict some physicochemical and rheological parameters. Samples were taken from three muscles (Semitendinosus, Rectus abdominis and Infraspinatus) of Charolais breed. Dry matter content, fat content, protein content, texture and collagen content were determined. Moreover emission spectra were recorded in the range of 305-400nm, 340-540nm and 410-700nm by fixing the excitation wavelength at 290, 322 and 382nm, respectively. The results obtained were evaluated by partial least square discriminant analysis and partial least square regression. Results of our research work show that front-face fluorescence spectroscopy and chemometrics offer significant potential for the development of rapid and non-destructive methods for the identification and characterization of muscles. PMID:25306513

Sahar, Amna; Dufour, Eric

2015-02-01

373

Retrograde fluorescent labeling allows for targeted extracellular single-unit recording from identified neurons in vivo.  

PubMed

The overall goal of this method is to record single-unit responses from an identified population of neurons. In vivo electrophysiological recordings from individual neurons are critical for understanding how neural circuits function under natural conditions. Traditionally, these recordings have been performed 'blind', meaning the identity of the recorded cell is unknown at the start of the recording. Cellular identity can be subsequently determined via intracellular(1), juxtacellular(2) or loose-patch(3) iontophoresis of dye, but these recordings cannot be pre-targeted to specific neurons in regions with functionally heterogeneous cell types. Fluorescent proteins can be expressed in a cell-type specific manner permitting visually-guided single-cell electrophysiology(4-6). However, there are many model systems for which these genetic tools are not available. Even in genetically accessible model systems, the desired promoter may be unknown or genetically homogenous neurons may have varying projection patterns. Similarly, viral vectors have been used to label specific subgroups of projection neurons(7), but use of this method is limited by toxicity and lack of trans-synaptic specificity. Thus, additional techniques that offer specific pre-visualization to record from identified single neurons in vivo are needed. Pre-visualization of the target neuron is particularly useful for challenging recording conditions, for which classical single-cell recordings are often prohibitively difficult(8-11). The novel technique described in this paper uses retrograde transport of a fluorescent dye applied using tungsten needles to rapidly and selectively label a specific subset of cells within a particular brain region based on their unique axonal projections, thereby providing a visual cue to obtain targeted electrophysiological recordings from identified neurons in an intact circuit within a vertebrate CNS. The most significant novel advancement of our method is the use of fluorescent labeling to target specific cell types in a non-genetically accessible model system. Weakly electric fish are an excellent model system for studying neural circuits in awake, behaving animals(12). We utilized this technique to study sensory processing by "small cells" in the anterior exterolateral nucleus (ELa) of weakly electric mormyrid fish. "Small cells" are hypothesized to be time comparator neurons important for detecting submillisecond differences in the arrival times of presynaptic spikes(13). However, anatomical features such as dense myelin, engulfing synapses, and small cell bodies have made it extremely difficult to record from these cells using traditional methods(11, 14). Here we demonstrate that our novel method selectively labels these cells in 28% of preparations, allowing for reliable, robust recordings and characterization of responses to electrosensory stimulation. PMID:23928906

Lyons-Warren, Ariel M; Kohashi, Tsunehiko; Mennerick, Steven; Carlson, Bruce A

2013-01-01

374

Retrograde Fluorescent Labeling Allows for Targeted Extracellular Single-unit Recording from Identified Neurons In vivo  

PubMed Central

The overall goal of this method is to record single-unit responses from an identified population of neurons. In vivo electrophysiological recordings from individual neurons are critical for understanding how neural circuits function under natural conditions. Traditionally, these recordings have been performed 'blind', meaning the identity of the recorded cell is unknown at the start of the recording. Cellular identity can be subsequently determined via intracellular1, juxtacellular2 or loose-patch3 iontophoresis of dye, but these recordings cannot be pre-targeted to specific neurons in regions with functionally heterogeneous cell types. Fluorescent proteins can be expressed in a cell-type specific manner permitting visually-guided single-cell electrophysiology4-6. However, there are many model systems for which these genetic tools are not available. Even in genetically accessible model systems, the desired promoter may be unknown or genetically homogenous neurons may have varying projection patterns. Similarly, viral vectors have been used to label specific subgroups of projection neurons7, but use of this method is limited by toxicity and lack of trans-synaptic specificity. Thus, additional techniques that offer specific pre-visualization to record from identified single neurons in vivo are needed. Pre-visualization of the target neuron is particularly useful for challenging recording conditions, for which classical single-cell recordings are often prohibitively difficult8-11. The novel technique described in this paper uses retrograde transport of a fluorescent dye applied using tungsten needles to rapidly and selectively label a specific subset of cells within a particular brain region based on their unique axonal projections, thereby providing a visual cue to obtain targeted electrophysiological recordings from identified neurons in an intact circuit within a vertebrate CNS. The most significant novel advancement of our method is the use of fluorescent labeling to target specific cell types in a non-genetically accessible model system. Weakly electric fish are an excellent model system for studying neural circuits in awake, behaving animals12. We utilized this technique to study sensory processing by "small cells" in the anterior exterolateral nucleus (ELa) of weakly electric mormyrid fish. "Small cells" are hypothesized to be time comparator neurons important for detecting submillisecond differences in the arrival times of presynaptic spikes13. However, anatomical features such as dense myelin, engulfing synapses, and small cell bodies have made it extremely difficult to record from these cells using traditional methods11, 14. Here we demonstrate that our novel method selectively labels these cells in 28% of preparations, allowing for reliable, robust recordings and characterization of responses to electrosensory stimulation. PMID:23928906

Lyons-Warren, Ariel M.; Kohashi, Tsunehiko; Mennerick, Steven; Carlson, Bruce A.

2013-01-01

375

Picosecond-resolution fluorescence lifetime imaging microscopy: a useful tool for sensing molecular interactions in vivo via FRET.  

PubMed

Fluorescence lifetime imaging microscopy (FLIM) provides a promising, robust method of detecting molecular interaction not not nots in vivo via fluorescence/Förster resonance energy transfer (FRET), by monitoring the variation of donor fluorescence lifetime, which is insensitive to many artifacts influencing convential intensity-based measurements, e.g. fluorophore concentration, photobleaching, and spectral bleed-through. As proof of principle, we demonstrate the capability of a novel picosecond-resolution FLIM system to detect molecular interactions in a well-established FRET assay. We then apply the FLIM system to detect the molecular interaction of a transforming oncogene RhoC with a binding partner RhoGDIgamma in vivo, which is critical to understand and interfere with Rho signaling for cancer therapeutics. PMID:19551120

Zhong, Wei; Wu, Mei; Chang, Ching-Wei; Merrick, Karl A; Merajver, Sofia D; Mycek, Mary-Ann

2007-12-24

376

Optical phantoms with variable properties and geometries for diffuse and fluorescence optical spectroscopy  

NASA Astrophysics Data System (ADS)

Growing interest in optical instruments for biomedical applications has increased the use of optically calibrated phantoms. Often associated with tissue modeling, phantoms allow the characterization of optical devices for clinical purposes. Fluorescent gel phantoms have been developed, mimicking optical properties of healthy and tumorous brain tissues. Specific geometries of dedicated molds offer multiple-layer phantoms with variable thicknesses and monolayer phantoms with cylindrical inclusions at various depths and diameters. Organic chromophores are added to allow fluorescence spectroscopy. These phantoms are designed to be used with 405 nm as the excitation wavelength. This wavelength is then adapted to excite large endogenous molecules. The benefits of these phantoms in understanding fluorescence tissue analysis are then demonstrated. In particular, detectability aspects as a function of geometrical and optical parameters are presented and discussed.

Leh, Barbara; Siebert, Rainer; Hamzeh, Hussein; Menard, Laurent; Duval, Marie-Alix; Charon, Yves; Abi Haidar, Darine

2012-10-01

377

Analysis of RNA Folding and Ribonucleoprotein Assembly by Single-Molecule Fluorescence Spectroscopy  

PubMed Central

Summary To execute their diverse range of biological functions, RNA molecules must fold into specific tertiary structures and/or associate with one or more proteins to form ribonucleoprotein (RNP) complexes. Single-molecule fluorescence spectroscopy is a powerful tool for the study of RNA folding and RNP assembly processes, directly revealing different conformational subpopulations that are hidden in conventional ensemble measurements. Moreover, kinetic processes can be observed without the need to synchronize a population of molecules. In this chapter, we describe the fluorescence spectroscopic methods used for single-molecule measurements of freely diffusing or immobilized RNA molecules or RNA-protein complexes. We also provide practical protocols to prepare the fluorescently labeled RNA and protein molecules required for such studies. Finally, we provide two examples of how these various preparative and spectroscopic methods are employed in the study of RNA folding and RNP assembly processes. PMID:22573447

Pljevalj?i?, Goran; Robertson-Anderson, Rae; van der Schans, Edwin; Millar, David

2013-01-01

378

A conformational study of corneal dermatan sulfate proteoglycan using fluorescence spectroscopy.  

PubMed

DSPG is a major proteoglycan of the corneal stroma and is thought to be important for the transparency of the tissue. We have studied its conformation by exploring the microenvironment and dynamics of its lone Tryptophan (Trp) residue using steady state and time resolved fluorescence spectroscopy. DSPG exhibits a doublet Trp fluorescence emission. Such a doublet emission has been observed earlier in the copper protein azurin and in avian lens delta-crystallin. Unlike the above cases where the doublet emission is thought to arise due to vibronic structure or the location of Trp at the interface of interacting subunits, fluorescence quenching, denaturation studies and ANS binding with DSPG indicate the location of Trp at two different environments. Such a situation could arise from the differential glycosylation of the core protein or due to duplexation and aggregation of the glycosaminoglycan chains. PMID:8782723

Uma, L; Sharma, Y; Balasubramanian, D

1996-07-01

379

Fluorescence Spectroscopy as a Promising Tool for a Polyphasic Approach to Pseudomonad Taxonomy  

Microsoft Academic Search

Fluorescence spectroscopy is an emerging tool for the analysis of biomolecules from complex matrices. We explored the potentialities\\u000a of the method for the pseudomonad taxonomic purpose at the genus and species level. Emission spectra of three intrinsic fluorophores\\u000a (namely, NADH, tryptophan, and the complex of aromatic amino acids and nucleic acid) were collected from whole bacterial cells.\\u000a Their comparisons were

Belal Tourkya; Tahar Boubellouta; Eric Dufour; Françoise Leriche

2009-01-01

380

Real-Time Enzyme Kinetics Monitored by Dual-Color Fluorescence Cross-Correlation Spectroscopy  

Microsoft Academic Search

A method for sensitively monitoring enzyme kinetics and activities by using dual-color fluorescence cross-correlation spectroscopy is described. This universal method enables the development of highly sensitive and precise assays for real-time kinetic analyses of any catalyzed cleavage or addition reaction, where a chemical linkage is formed or cleaved through an enzyme's action between two fluorophores that can be discriminated spectrally.

Ulrich Kettling; Andre Koltermann; Petra Schwille; Manfred Eigen

1998-01-01

381

New energy levels of atomic niobium by laser induced fluorescence spectroscopy in the near infrared  

NASA Astrophysics Data System (ADS)

Laser-induced fluorescence spectroscopy was applied in order to find new energy levels of the niobium atom. A continuous wave tuneable titanium–sapphire laser in the wavelength range from 750 to 865 nm and a hollow-cathode lamp were used. We discovered four energy levels of even parity, three lying levels below 19 000 cm?1 and one at much higher energy. Additionally hyperfine structure data of six levels of odd parity were determined.

Öztürk, I. K.; Ba?ar, Gö; Er, A.; Güzelçimen, F.; Ba?ar, Gü; Kröger, S.

2015-01-01

382

Optimizing light dosimetry in photodynamic therapy of the bronchi by fluorescence spectroscopy  

Microsoft Academic Search

Under identical conditions (drug and light dose, timing), the results of photodynamic therapy (PDT) of carcinomas of the bronchi with tetra(meta-hydroxyphenyl)chlorin (mTHPC) show large variations between patients. Before patients underwent PDT treatment, the mTHPC level was measured in the lesion, the normal surrounding tissue and the oral cavity, with an apparatus based on fluorescence spectroscopy. The fluctuations in degree of

D. Braichotte; J.-F. Savary; T. Glanzmann; P. Monnier; G. Wagnières; H. Bergh

1996-01-01

383

TOTAL INTERNAL REFLECTION WITH FLUORESCENCE CORRELATION SPECTROSCOPY: APPLICATIONS TO SUBSTRATE-SUPPORTED PLANAR MEMBRANES  

PubMed Central

In this review paper, the conceptual basis and experimental design of total internal reflection with fluorescence correlation spectroscopy (TIR-FCS) is described. The few applications to date of TIR-FCS to supported membranes are discussed, in addition to a variety of applications not directly involving supported membranes. Methods related, but not technically equivalent, to TIR-FCS are also summarized. Future directions for TIR-FCS are outlined. PMID:19269331

Thompson, Nancy L.; Wang, Xiang; Navaratnarajah, Punya

2009-01-01

384

Cation binding characteristics of tetrandrine studied by UV-Vis absorption and fluorescence spectroscopies  

Microsoft Academic Search

The binding of monovalent (Li+, Na+, K+) and bivalent (Mg2+, Ca2+) metal cations to a macrocyclic alkaloid tetrandrine has been studied by UV-Vis absorption and fluorescence spectroscopies. The calculated binding constants are between 3654 and 282613dm3mol?1 and the corresponding free energies from 4.777 to 7.308kcalmol?1. The effect of the solvent on the complexation strength as well as the role of

Ioana Stanculescu; Cristina Mandravel; François Delattre; David Landy; Patrice Woisel; Gheorghe Surpateanu

2003-01-01

385

Multimodal Mn-doped I-III-VI quantum dots for near infrared fluorescence and magnetic resonance imaging: from synthesis to in vivo application  

NASA Astrophysics Data System (ADS)

The development of sensitive multimodal contrast agents is a key issue to provide better global, multi-scale images for diagnostic or therapeutic purposes. Here we present the synthesis of Zn-Cu-In-(S, Se)/Zn1-xMnxS core-shell quantum dots (QDs) that can be used as markers for both near-infrared fluorescence imaging and magnetic resonance imaging (MRI). We first present the synthesis of Zn-Cu-In-(S, Se) cores coated with a thick ZnS shell doped with various proportions of Mn. Their emission wavelengths can be tuned over the NIR optical window suitable for deep tissue imaging. The incorporation of manganese ions (up to a few thousand ions per QD) confers them a paramagnetic character, as demonstrated by structural analysis and electron paramagnetic resonance spectroscopy. These QDs maintain their optical properties after transfer to water using ligand exchange. They exhibit T1-relaxivities up to 1400 mM-1 [QD] s-1 at 7 T and 300 K. We finally show that these QDs are suitable multimodal in vivo probes and demonstrate MRI and NIR fluorescence detection of regional lymph nodes in mice.The development of sensitive multimodal contrast agents is a key issue to provide better global, multi-scale images for diagnostic or therapeutic purposes. Here we present the synthesis of Zn-Cu-In-(S, Se)/Zn1-xMnxS core-shell quantum dots (QDs) that can be used as markers for both near-infrared fluorescence imaging and magnetic resonance imaging (MRI). We first present the synthesis of Zn-Cu-In-(S, Se) cores coated with a thick ZnS shell doped with various proportions of Mn. Their emission wavelengths can be tuned over the NIR optical window suitable for deep tissue imaging. The incorporation of manganese ions (up to a few thousand ions per QD) confers them a paramagnetic character, as demonstrated by structural analysis and electron paramagnetic resonance spectroscopy. These QDs maintain their optical properties after transfer to water using ligand exchange. They exhibit T1-relaxivities up to 1400 mM-1 [QD] s-1 at 7 T and 300 K. We finally show that these QDs are suitable multimodal in vivo probes and demonstrate MRI and NIR fluorescence detection of regional lymph nodes in mice. Electronic supplementary information (ESI) available: Determination of Mn content; magnetization measurements; additional TEM and spectroscopic data; additional NIR fluorescence image; MTT assay results. See DOI: 10.1039/c4nr02239d

Sitbon, Gary; Bouccara, Sophie; Tasso, Mariana; Francois, Aurélie; Bezdetnaya, Lina; Marchal, Frédéric; Beaumont, Marine; Pons, Thomas

2014-07-01

386

New photon-counting detectors for single-molecule fluorescence spectroscopy and imaging  

PubMed Central

Solution-based single-molecule fluorescence spectroscopy is a powerful new experimental approach with applications in all fields of natural sciences. Two typical geometries can be used for these experiments: point-like and widefield excitation and detection. In point-like geometries, the basic concept is to excite and collect light from a very small volume (typically femtoliter) and work in a concentration regime resulting in rare burst-like events corresponding to the transit of a single-molecule. Those events are accumulated over time to achieve proper statistical accuracy. Therefore the advantage of extreme sensitivity is somewhat counterbalanced by a very long acquisition time. One way to speed up data acquisition is parallelization. Here we will discuss a general approach to address this issue, using a multispot excitation and detection geometry that can accommodate different types of novel highly-parallel detector arrays. We will illustrate the potential of this approach with fluorescence correlation spectroscopy (FCS) and single-molecule fluorescence measurements. In widefield geometries, the same issues of background reduction and single-molecule concentration apply, but the duration of the experiment is fixed by the time scale of the process studied and the survival time of the fluorescent probe. Temporal resolution on the other hand, is limited by signal-to-noise and/or detector resolution, which calls for new detector concepts. We will briefly present our recent results in this domain. PMID:24729836

Michalet, X.; Colyer, R. A.; Scalia, G.; Weiss, S.; Siegmund, Oswald H. W.; Tremsin, Anton S.; Vallerga, John V.; Villa, F.; Guerrieri, F.; Rech, I.; Gulinatti, A.; Tisa, S.; Zappa, F.; Ghioni, M.; Cova, S.

2013-01-01

387

Exploring the binding mechanism of Guaijaverin to human serum albumin: fluorescence spectroscopy and computational approach.  

PubMed

The Guaijaverin (Gua) is a polyphenolic substance which exhibits some pharmacological activities such as antibacterial and antioxidant activities. Here we have investigated the binding of Gua with human serum albumin (HSA) at physiological pH 7.0. In this study, the fluorescence spectroscopy, ab initio and molecular modeling calculations were applied. The Stern-Volmer quenching constant (K(SV)) and its modified form (K(a)) were calculated at 298, 303 and 308 K, with the corresponding thermodynamic parameters ?H, ?G and ?S as well. The fluorescence quenching method was used to determine the number of binding sites (n) and binding constants (K(b)) values at 298, 303 and 308 K. The distance between donor (HSA) and acceptor (Gua) was estimated according to fluorescence resonance energy transfer. The geometry optimization of Gua was performed in its ground state by using ab initio DFT/B3LYP functional with a 6-31G(d,p) basis set used in calculations. Molecular modeling calculation indicated that the Gua is located within the hydrophobic pocket of the subdomain IIA of HSA. The theoretical results obtained by molecular modeling were corroborated by fluorescence spectroscopy data. PMID:22820048

Caruso, Icaro P; Vilegas, Wagner; Fossey, Marcelo A; Cornélio, Marinônio L

2012-11-01

388

Exploring the binding mechanism of Guaijaverin to human serum albumin: Fluorescence spectroscopy and computational approach  

NASA Astrophysics Data System (ADS)

The Guaijaverin (Gua) is a polyphenolic substance which exhibits some pharmacological activities such as antibacterial and antioxidant activities. Here we have investigated the binding of Gua with human serum albumin (HSA) at physiological pH 7.0. In this study, the fluorescence spectroscopy, ab initio and molecular modeling calculations were applied. The Stern-Volmer quenching constant (KSV) and its modified form (Ka) were calculated at 298, 303 and 308 K, with the corresponding thermodynamic parameters ?H, ?G and ?S as well. The fluorescence quenching method was used to determine the number of binding sites (n) and binding constants (Kb) values at 298, 303 and 308 K. The distance between donor (HSA) and acceptor (Gua) was estimated according to fluorescence resonance energy transfer. The geometry optimization of Gua was performed in its ground state by using ab initio DFT/B3LYP functional with a 6-31G(d,p) basis set used in calculations. Molecular modeling calculation indicated that the Gua is located within the hydrophobic pocket of the subdomain IIA of HSA. The theoretical results obtained by molecular modeling were corroborated by fluorescence spectroscopy data.

Caruso, Ícaro P.; Vilegas, Wagner; Fossey, Marcelo A.; Cornélio, Marinônio L.

2012-11-01

389

X-ray microprobe for micro x-ray fluorescence and absorption spectroscopies at GSECARS  

NASA Astrophysics Data System (ADS)

The hard x-ray microprobe for x-ray fluorescence and absorption spectroscopy at GeoSoilEnviroCARS is presented. Using focused synchrotron radiation from an undulator beamline at the Advanced Photon Source at Argonne National Lab, the x-ray microprobe provides bright, monochromatic x-rays with typical spot sizes down to 1x1 ?m for x-ray fluorescence and absorption spectroscopies. Quantitative x-ray fluorescence (XRF) analysis gives precise elemental composition and correlations, while x-ray absorption spectroscopy (XAS) gives the chemical state and local atomic coordination for a selected atomic species. These two techniques can be used in conjunction with one another on a wide range of samples, including minerals, glasses, fluid inclusions, soils, sediments, and plant tissue. This x-ray microprobe is part of the GeoSoilEnviroCARS user facility, available for use in all areas geological, soil, and environmental sciences, and selected examples from these fields will be given.

Newville, M.; Sutton, S.; Rivers, M.

2002-12-01

390

DETECTION OF MERCURIC BROMIDE IN A GAS PHASE FLOW CELL BY LASER PHOTOFRAGMENT FLUORESCENCE SPECTROSCOPY. (R825380)  

EPA Science Inventory

Photofragment fluorescence (PFF) spectroscopy offers real-time monitoring capability with high-analytical sensitivity and selectivity for volatile mercury compounds found in process gas streams, such as incinerator stacks. In this work, low concentrations (6 ppb to...

391

Bayesian Approach to the Analysis of Fluorescence Correlation Spectroscopy Data II: Application to Simulated and In Vitro Data  

E-print Network

Fluorescence correlation spectroscopy (FCS) is a powerful approach to characterizing the binding and transport dynamics of macromolecules. The unbiased interpretation of FCS data relies on the evaluation of multiple competing ...

Guo, Syuan-Ming

392

Localizing the chemical forms of sulfur in vivo using X-ray fluorescence spectroscopic imaging: application to onion (Allium cepa) tissues.  

PubMed

Sulfur has a particularly rich biochemistry and fills a number of important roles in biology. In situ information on sulfur biochemistry is generally difficult to obtain because of a lack of biophysical techniques that have sufficient sensitivity to molecular form. We have recently reported that sulfur K-edge X-ray absorption spectroscopy can be used as a direct probe of the sulfur biochemistry of living mammalian cells [Gnida, M., et al. (2007) Biochemistry 46, 14735-14741]. Here we report an extension of this work and develop sulfur K-edge X-ray fluorescence spectroscopic imaging as an in vivo probe of sulfur metabolism in living cells. For this work, we have chosen onion (Allium cepa) as a tractable model system with well-developed sulfur biochemistry and present evidence of the localization of a number of different chemical forms. X-ray absorption spectroscopy of onion sections showed increased levels of lachrymatory factor (LF) and thiosulfinate and decreased levels of sulfoxide (LF precursor) following cell breakage. In intact cells, X-ray fluorescence spectroscopic imaging showed elevated levels of sulfoxides in the cytosol and elevated levels of reduced sulfur in the central transport vessels and bundle sheath cells. PMID:19463015

Pickering, Ingrid J; Sneeden, Eileen Yu; Prince, Roger C; Block, Eric; Harris, Hugh H; Hirsch, Gregory; George, Graham N

2009-07-28

393

Raman spectroscopy for the detection of cancers and precancers  

Microsoft Academic Search

Optical spectroscopy has been extensively studied as a potential in vivo diagnostic tool that can provide information about both the chemical and morphological structure of tissue in near real time. Most in vivo studies have concentrated on elastic scattering and fluorescence spectroscopies since these signals can be obtained with a good signal-to-noise ratio quickly. However, Raman spectroscopy, an inelastic scattering

Anita Mahadevan-Jansen; Rebecca R. Richards-Kortum

1996-01-01

394

In vivo spatial frequency domain spectroscopy of two layer media  

NASA Astrophysics Data System (ADS)

Monitoring of tissue blood volume and local oxygen saturation can inform the assessment of tissue health, healing, and dysfunction. These quantities can be estimated from the contribution of oxyhemoglobin and deoxyhemoglobin to the absorption spectrum of the dermis. However, estimation of blood related absorption in skin can be confounded by the strong absorption of melanin in the epidermis and epidermal thickness and pigmentation varies with anatomic location, race, gender, and degree of disease progression. Therefore, a method is desired that decouples the effect of melanin absorption in the epidermis from blood absorption in the dermis for a large range of skin types and thicknesses. A previously developed inverse method based on a neural network forward model was applied to simulated spatial frequency domain reflectance of skin for multiple wavelengths in the near infrared. It is demonstrated that the optical thickness of the epidermis and absorption and reduced scattering coefficients of the dermis can be determined independently and with minimal coupling. Then, the same inverse method was applied to reflectance measurements from a tissue simulating phantom and in vivo human skin. Oxygen saturation and total hemoglobin concentrations were estimated from the volar forearms of weakly and strongly pigmented subjects using a standard homogeneous model and the present two layer model.

Yudovsky, Dmitry; Nguyen, John Quan M.; Durkin, Anthony J.

2012-10-01

395

Photoacoustics and fluorescence based nanoprobes towards functional and structural imaging in vivo  

NASA Astrophysics Data System (ADS)

Imaging of chemical analytes and structural properties related to physiological activities within biological systems is of great bio-medical interest; it can contribute to the fundamental understanding of biological systems and can be applied to the diagnosis and prognosis of diseases, especially tumors. The work presented in this thesis focuses on the development and application of polymeric nanoprobe aided optical imaging of chemical analytes (Oxygen, pH) and structural properties in live cells and animal models. To this end, specific nanoprobes, based on the polyacrylamide nanoplatform, bearing both appropriate targeting functionalities, and high concentrations of sensing and contrast agents, have been developed. The nanoprobes presented here are biodegradable, biocompatible and non-toxic, rendering them safe for in vivo use. Furthermore the nanoprobes are designed to have variable optical properties that are dependent on the local concentration of the specific analyte of interest. Optical imaging techniques that are particularly suited for deep tissue applications, such as two-photon fluorescence and photoacoustics, were applied for non-invasive real-time imaging and sensing in cancer cells, tumor spheroids and animal models. Our results demonstrate that this technique enables high sensitive detection of chemical analytes with a sensitivity of <5 Torr for oxygen and <0.1 pH units in vivo, which is better than the currently available in vivo functional imaging techniques. This non-invasive and non-ionizing, yet low cost, method will enable morphological and functional evaluation across any tissue, with both high spatial and temporal resolution but without eliciting short- or long-term tissue damage. Currently no gold standard exists for such xii functional imaging. The approach presented here can be used for early detection and diagnosis of tumors, as well as for monitoring the progression of disease and therapy. This technique will also enable observing phenomena at the cellular level in vivo that would lead to a better understanding of the pathophysiology of diseases as well as the disease onset, progression, and response to therapy.

Ray, Aniruddha

396

Evaluation of the uncertainties associated with in vivo X-ray fluorescence bone lead calibrations  

NASA Astrophysics Data System (ADS)

An anthropometric leg phantom developed at the University of Cincinnati (UC) was used to evaluate the effects that changes in leg position and variation between subjects has on in vivo x-ray fluorescence (XRF) measurements of stable lead in bone. The changes in leg position that were evaluated include changes in source-phantom distance ranging between 0.0 mm and 30.0 mm and phantom rotation over 40 degrees. Source-phantom distance was determined to have a significant effect on XRF measurement results particularly at source-phantom distances greater than 10.0 mm. Rotation of the leg phantom through 40 degrees was determined to have no significant effect on XRF measurement results. Between subject factors that were evaluated include bone calcium content and overlying tissue thickness. Bone calcium content was determined to have a significant effect on XRF measurements when measuring lead in micrograms per gram bone material. However, if measurement results of micrograms of lead per gram calcium (or per gram bone mineral) is used the normalization method makes the change in calcium content not significant. Overlying tissue thickness was determined to have no significant effect on XRF measurement results with tissue thickness ranging between 5.7 and 11.62 mm. The UC leg phantom was modified to include a fibula bone phantom so that the effect that the fibula has on XRF measurement results could be evaluated. The fibula was determined to have no significant effect on XRF measurement results and in the future need not be incorporated into in vivo XRF calibration phantoms. A knee phantom was also developed for purposes of calibrations of in vivo XRF measurement of lead in the patella. XRF measurement results using this phantom were compared to results of XRF measurements made using the plaster-of-Paris (POP) phantoms. A significant difference was observed between the normalized count rates of the two phantom types when either micrograms of lead per gram of bone material or micrograms of lead per gram calcium (bone mineral) is used as the lead content. This difference is consistent with what is observed in real in vivo XRF measurements and indicates the need for the correction factors that are used.

Lodwick, Jeffrey C.

397

Fluorescent magnetic nanoparticle-labeled mesenchymal stem cells for targeted imaging and hyperthermia therapy of in vivo gastric cancer  

PubMed Central

How to find early gastric cancer cells in vivo is a great challenge for the diagnosis and therapy of gastric cancer. This study is aimed at investigating the feasibility of using fluorescent magnetic nanoparticle (FMNP)-labeled mesenchymal stem cells (MSCs) to realize targeted imaging and hyperthermia therapy of in vivo gastric cancer. The primary cultured mouse marrow MSCs were labeled with amino-modified FMNPs then intravenously injected into mouse model with subcutaneous gastric tumor, and then, the in vivo distribution of FMNP-labeled MSCs was observed by using fluorescence imaging system and magnetic resonance imaging system. After FMNP-labeled MSCs arrived in local tumor tissues, subcutaneous tumor tissues in nude mice were treated under external alternating magnetic field. The possible mechanism of MSCs targeting gastric cancer was investigated by using a micro-multiwell chemotaxis chamber assay. Results show that MSCs were labeled with FMNPs efficiently and kept stable fluorescent signal and magnetic properties within 14?days, FMNP-labeled MSCs could target and image in vivo gastric cancer cells after being intravenously injected for 14?days, FMNP-labeled MSCs could significantly inhibit the growth of in vivo gastric cancer because of hyperthermia effects, and CCL19/CCR7 and CXCL12/CXCR4 axis loops may play key roles in the targeting of MSCs to in vivo gastric cancer. In conclusion, FMNP-labeled MSCs could target in vivo gastric cancer cells and have great potential in applications such as imaging, diagnosis, and hyperthermia therapy of early gastric cancer in the near future. PMID:22709686

2012-01-01

398

Fluorescent magnetic nanoparticle-labeled mesenchymal stem cells for targeted imaging and hyperthermia therapy of in vivo gastric cancer  

NASA Astrophysics Data System (ADS)

How to find early gastric cancer cells in vivo is a great challenge for the diagnosis and therapy of gastric cancer. This study is aimed at investigating the feasibility of using fluorescent magnetic nanoparticle (FMNP)-labeled mesenchymal stem cells (MSCs) to realize targeted imaging and hyperthermia therapy of in vivo gastric cancer. The primary cultured mouse marrow MSCs were labeled with amino-modified FMNPs then intravenously injected into mouse model with subcutaneous gastric tumor, and then, the in vivo distribution of FMNP-labeled MSCs was observed by using fluorescence imaging system and magnetic resonance imaging system. After FMNP-labeled MSCs arrived in local tumor tissues, subcutaneous tumor tissues in nude mice were treated under external alternating magnetic field. The possible mechanism of MSCs targeting gastric cancer was investigated by using a micro-multiwell chemotaxis chamber assay. Results show that MSCs were labeled with FMNPs efficiently and kept stable fluorescent signal and magnetic properties within 14 days, FMNP-labeled MSCs could target and image in vivo gastric cancer cells after being intravenously injected for 14 days, FMNP-labeled MSCs could significantly inhibit the growth of in vivo gastric cancer because of hyperthermia effects, and CCL19/CCR7 and CXCL12/CXCR4 axis loops may play key roles in the targeting of MSCs to in vivo gastric cancer. In conclusion, FMNP-labeled MSCs could target in vivo gastric cancer cells and have great potential in applications such as imaging, diagnosis, and hyperthermia therapy of early gastric cancer in the near future.

Ruan, Jing; Ji, Jiajia; Song, Hua; Qian, Qirong; Wang, Kan; Wang, Can; Cui, Daxiang

2012-06-01

399

In vivo laser-induced fluorescence detection of pyrene in nematodes and determination of pyrene binding constants for humic substances by fluorescence quenching and bioconcentration experiments.  

PubMed

The bioconcentration of pyrene by bacterivorous thread worms (nematodes) of the species Caenorhabditis elegans was studied with laser-induced fluorescence (LIF) spectroscopy, fluorescence imaging and a radiotracer method. The vibronic band intensities of the LIF spectra indicated that the microenvironment of pyrene in the nematodes was similar to a low-polarity solvent, and thus provided direct evidence that pyrene was accumulated in lipid-rich areas inside the nematodes. The concentration of pyrene in the nematodes was estimated from the monomer/excimer fluorescence intensity ratio. Results from this method were in fair agreement with results using 14C labeled pyrene for measuring pyrene bioconcentration. Preliminary results indicated that LIF measurements of pyrene may be possible even in single nematodes. Fluorescence microscopic observations revealed that pyrene was not adsorbed on the outside of the organisms, but was strongly concentrated in restricted areas inside the worms. In the second part of the study, the effects of six different humic substances (HS) on the bioconcentration of pyrene were investigated and sorption coefficients (KDOC) calculated from reductions in bioconcentration (KDOC(biol)) were compared with sorption coefficients measured with a fluorescence quenching technique (KDOC(flu)). The results of these two different experimental methods agreed well (with KDOC(biol) being slightly lower than KDOC(flu), indicating that the fraction of pyrene that was determined as freely dissolved by the fluorescence quenching method was comparable to the bioavailable fraction. PMID:11253034

Haitzer, M; Löhmannsröben, H G; Steinberg, C E; Zimmermann, U

2000-04-01

400

Construction, figures of merit, and testing of a single-cell fluorescence excitation spectroscopy system  

PubMed Central

Characterization of phytoplankton community composition is critical to understanding the ecology and biogeochemistry of the oceans. One approach to taxonomic characterization takes advantage of differing pigmentation between algal taxa and thus differences in fluorescence excitation spectra. Analyses of bulk water samples, however, may be confounded by interference from chromophoric dissolved organic matter or suspended particulate matter. Here, we describe an instrument that uses a laser trap based on a Nikon TE2000-U microscope to position individual phytoplankton cells for confocal fluorescence excitation spectroscopy, thus avoiding interference from the surrounding medium. Quantitative measurements of optical power give data in the form of photons emitted per photon of exposure for an individual phytoplankton cell. Residence times for individual phytoplankton in the instrument can be as long as several minutes with no substantial change in their fluorescence excitation spectra. The laser trap was found to generate two-photon fluorescence from the organisms so a modification was made to release the trap momentarily during data acquisition. Typical signal levels for an individual cell are in the range of 106 photons?s of fluorescence using a monochromated 75 W Xe arc lamp excitation source with a 2% transmission neutral density filter. PMID:20113077

Hill, Laura S.; Richardson, Tammi L.; Profeta, Luisa T. M.; Shaw, Timothy J.; Hintz, Christopher J.; Twining, Benjamin S.; Lawrenz, Evelyn; Myrick, Michael L.

2010-01-01

401

The use of fluorescence correlation spectroscopy (FCS) as an alternative biomarker detection technique: a preliminary study  

PubMed Central

Abstract Biomarkers are essential part of daily medical practice. Currently, biomarkers are being used both for diagnostic and prognostic purposes. There are many approaches e.g. ELISA by which biomarker levels are detected from patient samples. However, all these approaches are laborious, time consuming and expensive. There is therefore a general need for exploring new technique which can overcome these drawbacks. Here, we present a preliminary study for detection of serum biomarkers by fluorescence correlation spectroscopy (FCS) based diagnostic technique. FCS is a technique basically used for spatial and temporal analysis of molecular interactions of extremely low-concentration biomolecules in solution. FCS is able to measure diffusion time of the fluorescent molecules passing through the open detection volume and it can also measure the average number of fluorescent molecules passing through the detection volume. Because diffusion speed is correlated with shape and molecular mass of the fluorescent molecule, this property makes it possible to study the complex formation between a small fluorescently labelled and a large unlabelled molecule. In this preliminary study, we utilize this FCS property for detection of serum biomarker. Further studies on various pathological serum samples are warranted to explore further aspects of this technique. PMID:21306559

Shahzad, Aamir; Knapp, Martin; Lang, Irene; Köhler, Gottfried

2011-01-01

402

Applications of fluorescence spectroscopy for predicting percent wastewater in an urban stream  

USGS Publications Warehouse

Dissolved organic carbon (DOC) is a significant organic carbon reservoir in many ecosystems, and its characteristics and sources determine many aspects of ecosystem health and water quality. Fluorescence spectroscopy methods can quantify and characterize the subset of the DOC pool that can absorb and re-emit electromagnetic energy as fluorescence and thus provide a rapid technique for environmental monitoring of DOC in lakes and rivers. Using high resolution fluorescence techniques, we characterized DOC in the Tualatin River watershed near Portland, Oregon, and identified fluorescence parameters associated with effluent from two wastewater treatment plants and samples from sites within and outside the urban region. Using a variety of statistical approaches, we developed and validated a multivariate linear regression model to predict the amount of wastewater in the river as a function of the relative abundance of specific fluorescence excitation/emission pairs. The model was tested with independent data and predicts the percentage of wastewater in a sample within 80% confidence. Model results can be used to develop in situ instrumentation, inform monitoring programs, and develop additional water quality indicators for aquatic systems.

Goldman, Jami H.; Rounds, Stewart A.; Needoba, Joseph A.

2012-01-01

403

Fluorescence spectroscopy for the detection of potentially malignant disorders of the oral cavity: analysis of 30 cases  

NASA Astrophysics Data System (ADS)

Oral cancer is a major health problem worldwide and although early diagnosis of potentially malignant and malignant diseases is associated with better treatment results, a large number of cancers are initially misdiagnosed, with unfortunate consequences for long-term survival. Fluorescence spectroscopy is a noninvasive modality of diagnostic approach using induced fluorescence emission in tumors that can improve diagnostic accuracy. The objective of this study was to determine the ability to discriminate between normal oral mucosa and potentially malignant disorders by fluorescence spectroscopy. Fluorescence investigation under 408 and 532 nm excitation wavelengths was performed on 60 subjects, 30 with potentially malignant disorders and 30 volunteers with normal mucosa. Data was analyzed to correlate fluorescence patterns with clinical and histopathological diagnostics. Fluorescence spectroscopy used as a point measurement technique resulted in a great variety of spectral information. In a qualitative analysis of the fluorescence spectral characteristics of each type of injury evaluated, it was possible to discriminate between normal and abnormal oral mucosa. The results show the potential use of fluorescence spectroscopy for an improved discrimination of oral disorders.

Francisco, A. L. N.; Correr, W. R.; Azevedo, L. H.; Galletta, V. K.; Pinto, C. A. L.; Kowalski, L. P.; Kurachi, C.

2014-01-01

404

Quantitative study of Co(II) complexation by synchronous fluorescence spectroscopy with Sundarban mangrove habitat humic substances  

Microsoft Academic Search

Attempt has been made to isolate and characterize humic substances and their relative role for complexation of Co (II) in the mangrove sediment. Conditional stability constant (Kc) for Co (II) complexes with humic and fulvic acids were de- termined by studying quenching of fluorescence intensity of humic substances with Co (II) using synchronous fluorescence spectroscopy. Fulvic acid forms more stable

H. Ghatak; S. K. Mukhopadhyaya; H. Biswas; T. K. Jana

405

A high-resolution large-acceptance analyzer for X-ray fluorescence and Raman spectroscopy  

SciTech Connect

A newly designed multi-crystal X-ray spectrometer and its applications in the fields of X-ray fluorescence and X-ray Raman spectroscopy are described. The instrument is based on 8 spherically curved Si crystals, each with a 3.5 inch diameter form bent to a radius of 86 cm. The crystals are individually aligned in the Rowland geometry capturing a total solid angle of 0.07 sr. The array is arranged in a way that energy scans can be performed by moving the whole instrument, rather than scanning each crystal by itself. At angles close to back scattering the energy resolution is between 0.3 and 1 eV depending on the beam dimensions at the sample. The instrument is mainly designed for X-ray absorption and fluorescence spectroscopy of transition metals in dilute systems such as metalloproteins. First results of the Mn K{beta} (3p -> 1s) emission in photosystem II are shown. An independent application of the instrument is the technique of X-ray Raman spectroscopy which can address problems similar to those in traditional soft X-ray absorption spectroscopies, and initial results are presented.

Bergmann, Uwe; Cramer, Stephen P.

2001-08-02

406

Native fluorescence spectroscopy of blood plasma of rats with experimental diabetes: identifying fingerprints of glucose-related metabolic pathways.  

PubMed

We present the results of a native fluorescence spectroscopy study of blood plasma of rats with experimental diabetes. It was shown that the fluorescence emission band shape at 320 nm excitation is the most indicative of hyperglycemia in the blood plasma samples. We provide the interpretation of this fact based on the changes in reduced nicotinamide adenine dinucleotide phosphate concentration due to glucose-related metabolic pathways and protein fluorescent cross-linking formation following nonenzymatic glycation. PMID:25692658

Shirshin, Evgeny; Cherkasova, Olga; Tikhonova, Tatiana; Berlovskaya, Elena; Priezzhev, Alexander; Fadeev, Victor

2015-05-01

407

In vivo Raman spectroscopy for breast cancer: diagnosis in animal model  

NASA Astrophysics Data System (ADS)

Raman spectroscopy has been well established as a powerful method for studying biological tissues and diagnosing diseases. In this study we have developed a breast cancer animal model and collected in vivo Raman spectra of mammary glands of 27 Sprague-Dawley female rats treated with DMBA and 5 non-treated used as control group. A dispersive Raman spectrometer with a @785 nm laser excitation coupled a fiber optic probe and a CCD detector was used to obtain the spectra. The obtained in vivo transcutaneous Raman spectra have shown important differences between normal and abnormal tissues when acquired from one side to the other side of the lesion.

Bitar, R.; Martins, M. A.; Ribeiro, D.; Carvalho, C.; Santos, E. A. P.; Ramalho, L. N. Z.; Ramalho, F.; Martinho, H.; Martin, A. A.

2008-02-01

408

Characterization of dissolved organic matter in fogwater by excitation-emission matrix fluorescence spectroscopy  

USGS Publications Warehouse

Dissolved organic matter (DOM) present in fogwater samples collected in southeastern Louisiana and central-eastern China has been characterized using excitation-emission matrix fluorescence spectroscopy. The goal of the study was to illustrate the utility of fluorescence for obtaining information on the large fraction of organic carbon in fogwaters (typically >40% by weight) that defies characterization in terms of specific chemical compounds without the difficulty inherent in obtaining sufficient fogwater volume to isolate DOM for assessment using other spectroscopic and chemical analyses. Based on the findings of previous studies using other characterization methods, it was anticipated that the unidentified organic carbon fraction would have characteristic peaks associated with humic substances and fluorescent amino acids. Both humic- and protein-like fluorophores were observed in the fogwater spectra and fluorescence-derived indices for the fogwater had similar values to those of soil and sediment porewater. Greater biological character was observed in samples with higher organic carbon concentrations. Fogwaters are shown to contain a mixture of terrestrially- and microbially-derived fluorescent organic material, which is expected to be derived from an array of different sources, such as suspended soil and dust particles, biogenic emissions and organic substances generated by atmospheric processes. The fluorescence results indicate that much of the unidenti