WorldWideScience

Sample records for two-photon time-lapse imaging

  1. Imaging zebrafish embryos by two-photon excitation time-lapse microscopy.

    Science.gov (United States)

    Carvalho, Lara; Heisenberg, Carl-Philipp

    2009-01-01

    The zebrafish is a favorite model organism to study tissue morphogenesis during development at a subcellular level. This largely results from the fact that zebrafish embryos are transparent and thus accessible to various imaging techniques, such as confocal and two-photon excitation (2PE) microscopy. In particular, 2PE microscopy has been shown to be useful for imaging deep cell layers within the embryo and following tissue morphogenesis over long periods. This chapter describes how to use 2PE microscopy to study morphogenetic movements during early zebrafish embryonic development, providing a general blueprint for its use in zebrafish.

  2. TimeLapseAnalyzer: Multi-target analysis for live-cell imaging and time-lapse microscopy

    DEFF Research Database (Denmark)

    Huth, Johannes; Buchholz, Malte; Kraus, Johann M.;

    2011-01-01

    , we developed TimeLapseAnalyzer. Apart from general purpose image enhancements and segmentation procedures, this extensible, self-contained, modular cross-platform package provides dedicated modalities for fast and reliable analysis of multi-target cell tracking, scratch wound healing analysis, cell...

  3. Time-lapse seismic imaging of the Reykjanes geothermal reservoir

    Science.gov (United States)

    Weemstra, Cornelis; Obermann, Anne; Blanck, Hanna; Verdel, Arie; Paap, Bob; Árni Guðnason, Egill; Páll Hersir, Gylfi; Jousset, Philippe; Sigurðsson, Ómar

    2016-04-01

    We report on the results obtained from a dense seismic deployment over a geothermal reservoir. The reservoir has been producing continuously for almost a decade and is located on the tip of the Reykjanes peninsula, SW Iceland. The seismic stations on top of the reservoir have continuously recorded the ambient seismic wavefield between April 2014 and September 2015. The density of the seismic network makes the data well suited for time-lapse seismic imaging of the reservoir. To that end we compute time-lapse responses through the application of seismic interferometry. These interferometric lapse responses are obtained by simple crosscorrelation of the seismic noise recorded by the different seismic stations. We subsequently evaluate the temporal variation of the coda of these crosscorrelations. The term coda refers to the later arriving, multiple scattered waves. The multiple scattering implies that these waves have sampled the subsurface very densely and hence become highly sensitive to tiny mechanical and structural changes in that subsurface. This sensitivity allows one, in principle at least, to monitor the geothermal reservoir. Preliminary results indeed suggest a relation between the temporal variation of the coda waves and the reservoir. Ultimately, this method may lead to a means to monitor a geothermal reservoir in both space and time.

  4. Two-photon time-lapse microscopy of BODIPY-cholesterol reveals anomalous sterol diffusion in chinese hamster ovary cells

    DEFF Research Database (Denmark)

    Lund, F. W.; Lomholt, M. A.; Solanko, L. M.

    2012-01-01

    Background: Cholesterol is an important membrane component, but our knowledge about its transport in cells is sparse. Previous imaging studies using dehydroergosterol (DHE), an intrinsically fluorescent sterol from yeast, have established that vesicular and non-vesicular transport modes contribute...... to sterol trafficking from the plasma membrane. Significant photobleaching, however, limits the possibilities for in-depth analysis of sterol dynamics using DHE. Co-trafficking studies with DHE and the recently introduced fluorescent cholesterol analog BODIPY-cholesterol (BChol) suggested that the latter...... probe has utility for prolonged live-cell imaging of sterol transport. Results: We found that BChol is very photostable under two-photon (2P)-excitation allowing the acquisition of several hundred frames without significant photobleaching. Therefore, long-term tracking and diffusion measurements...

  5. Two-photon time-lapse microscopy of BODIPY-cholesterol reveals anomalous sterol diffusion in chinese hamster ovary cells

    Directory of Open Access Journals (Sweden)

    Lund Frederik W

    2012-10-01

    Full Text Available Abstract Background Cholesterol is an important membrane component, but our knowledge about its transport in cells is sparse. Previous imaging studies using dehydroergosterol (DHE, an intrinsically fluorescent sterol from yeast, have established that vesicular and non-vesicular transport modes contribute to sterol trafficking from the plasma membrane. Significant photobleaching, however, limits the possibilities for in-depth analysis of sterol dynamics using DHE. Co-trafficking studies with DHE and the recently introduced fluorescent cholesterol analog BODIPY-cholesterol (BChol suggested that the latter probe has utility for prolonged live-cell imaging of sterol transport. Results We found that BChol is very photostable under two-photon (2P-excitation allowing the acquisition of several hundred frames without significant photobleaching. Therefore, long-term tracking and diffusion measurements are possible. Two-photon temporal image correlation spectroscopy (2P-TICS provided evidence for spatially heterogeneous diffusion constants of BChol varying over two orders of magnitude from the cell interior towards the plasma membrane, where D ~ 1.3 μm2/s. Number and brightness (N&B analysis together with stochastic simulations suggest that transient partitioning of BChol into convoluted membranes slows local sterol diffusion. We observed sterol endocytosis as well as fusion and fission of sterol-containing endocytic vesicles. The mobility of endocytic vesicles, as studied by particle tracking, is well described by a model for anomalous subdiffusion on short time scales with an anomalous exponent α ~ 0.63 and an anomalous diffusion constant of Dα = 1.95 x 10-3 μm2/sα. On a longer time scale (t > ~5 s, a transition to superdiffusion consistent with slow directed transport with an average velocity of v ~ 6 x 10-3 μm/s was observed. We present an analytical model that bridges the two regimes and fit this model to vesicle

  6. Seismic imaging of reservoir flow properties: Time-lapse pressurechanges

    Energy Technology Data Exchange (ETDEWEB)

    Vasco, Don W.

    2003-04-08

    Time-lapse fluid pressure and saturation estimates are sensitive to reservoir flow properties such as permeability. In fact, given time-lapse estimates of pressure and saturation changes, one may define a linear partial differential equation for permeability variations within the reservoir. The resulting linear inverse problem can be solved quite efficiently using sparse matrix techniques. An application to a set of crosswell saturation and pressure estimates from a CO{sub 2} flood at the Lost Hills field in California demonstrates the utility of this approach. From the crosswell estimates detailed estimates of reservoir permeability are produced. The resulting permeability estimates agree with a permeability log in an adjacent well and are in accordance with water and CO{sub 2} saturation changes in the interwell region.

  7. Time-lapse imaging of neural development: zebrafish lead the way into the fourth dimension.

    Science.gov (United States)

    Rieger, Sandra; Wang, Fang; Sagasti, Alvaro

    2011-07-01

    Time-lapse imaging is often the only way to appreciate fully the many dynamic cell movements critical to neural development. Zebrafish possess many advantages that make them the best vertebrate model organism for live imaging of dynamic development events. This review will discuss technical considerations of time-lapse imaging experiments in zebrafish, describe selected examples of imaging studies in zebrafish that revealed new features or principles of neural development, and consider the promise and challenges of future time-lapse studies of neural development in zebrafish embryos and adults.

  8. Long-term time-lapse live imaging reveals extensive cell migration during annelid regeneration.

    Science.gov (United States)

    Zattara, Eduardo E; Turlington, Kate W; Bely, Alexandra E

    2016-03-23

    Time-lapse imaging has proven highly valuable for studying development, yielding data of much finer resolution than traditional "still-shot" studies and allowing direct examination of tissue and cell dynamics. A major challenge for time-lapse imaging of animals is keeping specimens immobile yet healthy for extended periods of time. Although this is often feasible for embryos, the difficulty of immobilizing typically motile juvenile and adult stages remains a persistent obstacle to time-lapse imaging of post-embryonic development. Here we describe a new method for long-duration time-lapse imaging of adults of the small freshwater annelid Pristina leidyi and use this method to investigate its regenerative processes. Specimens are immobilized with tetrodotoxin, resulting in irreversible paralysis yet apparently normal regeneration, and mounted in agarose surrounded by culture water or halocarbon oil, to prevent dehydration but allowing gas exchange. Using this method, worms can be imaged continuously and at high spatial-temporal resolution for up to 5 days, spanning the entire regeneration process. We performed a fine-scale analysis of regeneration growth rate and characterized cell migration dynamics during early regeneration. Our studies reveal the migration of several putative cell types, including one strongly resembling published descriptions of annelid neoblasts, a cell type suggested to be migratory based on "still-shot" studies and long hypothesized to be linked to regenerative success in annelids. Combining neurotoxin-based paralysis, live mounting techniques and a starvation-tolerant study system has allowed us to obtain the most extensive high-resolution longitudinal recordings of full anterior and posterior regeneration in an invertebrate, and to detect and characterize several cell types undergoing extensive migration during this process. We expect the tetrodotoxin paralysis and time-lapse imaging methods presented here to be broadly useful in studying

  9. Seismic time-lapse imaging using Interferometric least-squares migration

    KAUST Repository

    Sinha, Mrinal

    2016-09-06

    One of the problems with 4D surveys is that the environmental conditions change over time so that the experiment is insufficiently repeatable. To mitigate this problem, we propose the use of interferometric least-squares migration (ILSM) to estimate the migration image for the baseline and monitor surveys. Here, a known reflector is used as the reference reflector for ILSM. Results with synthetic and field data show that ILSM can eliminate artifacts caused by non-repeatability in time-lapse surveys.

  10. 3D Time-lapse Imaging and Quantification of Mitochondrial Dynamics

    Science.gov (United States)

    Sison, Miguel; Chakrabortty, Sabyasachi; Extermann, Jérôme; Nahas, Amir; James Marchand, Paul; Lopez, Antonio; Weil, Tanja; Lasser, Theo

    2017-02-01

    We present a 3D time-lapse imaging method for monitoring mitochondrial dynamics in living HeLa cells based on photothermal optical coherence microscopy and using novel surface functionalization of gold nanoparticles. The biocompatible protein-based biopolymer coating contains multiple functional groups which impart better cellular uptake and mitochondria targeting efficiency. The high stability of the gold nanoparticles allows continuous imaging over an extended time up to 3000 seconds without significant cell damage. By combining temporal autocorrelation analysis with a classical diffusion model, we quantify mitochondrial dynamics and cast these results into 3D maps showing the heterogeneity of diffusion parameters across the whole cell volume.

  11. Noise-free accurate count of microbial colonies by time-lapse shadow image analysis.

    Science.gov (United States)

    Ogawa, Hiroyuki; Nasu, Senshi; Takeshige, Motomu; Funabashi, Hisakage; Saito, Mikako; Matsuoka, Hideaki

    2012-12-01

    Microbial colonies in food matrices could be counted accurately by a novel noise-free method based on time-lapse shadow image analysis. An agar plate containing many clusters of microbial colonies and/or meat fragments was trans-illuminated to project their 2-dimensional (2D) shadow images on a color CCD camera. The 2D shadow images of every cluster distributed within a 3-mm thick agar layer were captured in focus simultaneously by means of a multiple focusing system, and were then converted to 3-dimensional (3D) shadow images. By time-lapse analysis of the 3D shadow images, it was determined whether each cluster comprised single or multiple colonies or a meat fragment. The analytical precision was high enough to be able to distinguish a microbial colony from a meat fragment, to recognize an oval image as two colonies contacting each other, and to detect microbial colonies hidden under a food fragment. The detection of hidden colonies is its outstanding performance in comparison with other systems. The present system attained accuracy for counting fewer than 5 colonies and is therefore of practical importance.

  12. A method to investigate radial glia cell behavior using two-photon time-lapse microscopy in an ex vivo model of spinal cord development.

    Directory of Open Access Journals (Sweden)

    Janelle M.P. Pakan

    2014-04-01

    Full Text Available The mammalian central nervous system (CNS develops from multipotent progenitor cells, which proliferate and differentiate into the various cell types of the brain and spinal cord. Despite the wealth of knowledge from progenitor cell culture studies, there is a significant lack of understanding regarding dynamic progenitor cell behavior over the course of development. This is in part due to shortcomings in the techniques available to study these processes in living tissues as they are occurring. In order to investigate cell behavior under physiologically relevant conditions we established an ex vivo model of the developing rat spinal cord. This method allows us to directly observe specific populations of cells ex vivo in real time and over extended developmental periods as they undergo proliferation, migration and differentiation in the CNS. Previous investigations of progenitor cell behavior have been limited in either spatial or temporal resolution (or both due to the necessity of preserving tissue viability and avoiding phototoxic effects of fluorescent imaging. The method described here overcomes these obstacles. Using two-photon and confocal microscopy and transfected organotypic spinal cord slice cultures we have undertaken detailed imaging of a unique population of neural progenitors, radial glial cells. This method uniquely enables analysis of large populations as well as individual cells; ultimately resulting in a 4D dataset of progenitor cell behavior for up to seven days during embryonic development. This approach can be adapted to study a variety of cell populations at different stages of development using appropriate promoter driven fluorescent protein expression. The ability to control the tissue micro-environment makes this ex vivo method a powerful tool to elucidate the underlying molecular mechanisms regulating cell behavior during embryonic development.

  13. Time-Lapse Electrical Resistivity Investigations for Imaging the Grouting Injection in Shallow Subsurface Cavities

    Directory of Open Access Journals (Sweden)

    Muhammad Farooq

    2014-01-01

    Full Text Available The highway of Yongweol-ri, Muan-gun, south-western part of the South Korean Peninsula, is underlain by the abandoned of subsurface cavities, which were discovered in 2005. These cavities lie at shallow depths with the range of 5∼15 meters below the ground surface. Numerous subsidence events have repeatedly occurred in the past few years, damaging infrastructure and highway. As a result of continuing subsidence issues, the Korean Institute of Geosciences and Mineral Resources (KIGAM was requested by local administration to resolve the issue. The KIGAM used geophysical methods to delineate subsurface cavities and improve more refined understanding of the cavities network in the study area. Cement based grouting has been widely employed in the construction industry to reinforce subsurface ground. In this research work, time-lapse electrical resistivity surveys were accomplished to monitor the grouting injection in the subsurface cavities beneath the highway, which have provided a quasi-real-time monitoring for modifying the subsurface cavities related to ground reinforcement, which would be difficult with direct methods. The results obtained from time-lapse electrical resistivity technique have satisfactory imaged the grouting injection experiment in the subsurface cavities beneath the highway. Furthermore, the borehole camera confirmed the presence of grouting material in the subsurface cavities, and hence this procedure increases the mechanical resistance of subsurface cavities below the highway.

  14. Denoising two-photon calcium imaging data.

    Science.gov (United States)

    Malik, Wasim Q; Schummers, James; Sur, Mriganka; Brown, Emery N

    2011-01-01

    Two-photon calcium imaging is now an important tool for in vivo imaging of biological systems. By enabling neuronal population imaging with subcellular resolution, this modality offers an approach for gaining a fundamental understanding of brain anatomy and physiology. Proper analysis of calcium imaging data requires denoising, that is separating the signal from complex physiological noise. To analyze two-photon brain imaging data, we present a signal plus colored noise model in which the signal is represented as harmonic regression and the correlated noise is represented as an order autoregressive process. We provide an efficient cyclic descent algorithm to compute approximate maximum likelihood parameter estimates by combing a weighted least-squares procedure with the Burg algorithm. We use Akaike information criterion to guide selection of the harmonic regression and the autoregressive model orders. Our flexible yet parsimonious modeling approach reliably separates stimulus-evoked fluorescence response from background activity and noise, assesses goodness of fit, and estimates confidence intervals and signal-to-noise ratio. This refined separation leads to appreciably enhanced image contrast for individual cells including clear delineation of subcellular details and network activity. The application of our approach to in vivo imaging data recorded in the ferret primary visual cortex demonstrates that our method yields substantially denoised signal estimates. We also provide a general Volterra series framework for deriving this and other signal plus correlated noise models for imaging. This approach to analyzing two-photon calcium imaging data may be readily adapted to other computational biology problems which apply correlated noise models.

  15. Imaging Rainfall Infiltration Processes with the Time-Lapse Electrical Resistivity Imaging Method

    Science.gov (United States)

    Zhang, Gang; Zhang, Gui-Bin; Chen, Chien-chih; Chang, Ping-Yu; Wang, Tzu-Pin; Yen, Horng-Yuan; Dong, Jia-Jyun; Ni, Chuen-Fa; Chen, Su-Chin; Chen, Chao-Wei; Jia, Zheng-yuan

    2016-06-01

    Electrical Resistivity Imaging (ERI) was carried out continuously for 10 days to map the subsurface resistivity distribution along a potentially hazardous hillslope at the Jieshou Junior High School in Taoyuan, Taiwan. The reliability of the inverted resistivity structures down to about 25 m depth was examined with synthetic modeling using the same electrode arrangements installed on land surface as in field surveys, together with a DOI (depth-of-investigation) index calculated from the ERI data. The subsurface resistivity distribution is consistent with results from well logging. These ERI recordings were taken daily and provided highly resolved imagery of the resistivity distribution underground and illustrated the dynamical fluid-flow behavior due to heavy rainfall infiltration. Using Archie's law, the resistivity distribution was transformed into a map of relative water saturation (RWS), which is strongly correlated with the rainfall infiltration process. We then found that the averaged RWS is significantly correlated with daily precipitation. Our observations indicate that time-lapse ERI is effective in monitoring subterraneous rainfall infiltration; moreover, the preferential flow paths can be delineated according to the changes in averaged RWS derived from the ERI data.

  16. Tracking neural stem cells in time-lapse microscopy image sequences

    Science.gov (United States)

    Althoff, Karin; Degerman, Johan; Gustavsson, Tomas

    2005-04-01

    This paper describes an algorithm for tracking neural stem/progenitor cells in a time-lapse microscopy image sequence. The cells were segmented in a semiautomatic way using dynamic programming. Since the interesting cells were identified by fluorescent staining at the end of the sequence, the tracking was performed backwards. The number of detected cells varied throughout the sequence: cells could appear or disappear at the image boundaries or at cell clusters, some cells split, and the segmentation was not always correct. To solve this asymmetric assignment problem, a modified version of the auction algorithm by Bertsekas was used. The assignment weights were calculated based on distance, correlation and size between possible matching cells. Cell splits are of special interest, therefore tracks without a matching cell were divided into two groups: 1. Merging cells (splitting cells, moving forward in time) and 2. Non-merging cells. These groups were separated based on difference in size of the involved cells, and difference in image intensity of the contour and interior of the possibly merged cell. The tracking algorithm was evaluated using a sequence consisting of 57 images, each image containing approximately 50 cells. The evaluation showed that 99% of the cell-to-cell associations were correct. In most cases, only one association per track was incorrect so in total 55 out of 78 different tracks in the sequence were tracked correctly. Further improvements will be to apply interleaved segmentation and tracking to produce a more reliable segmentation as well as better tracking results.

  17. Distortion effects in a switch array UWB radar for time-lapse imaging of human heartbeats

    Science.gov (United States)

    Brovoll, Sverre; Berger, Tor; Aardal, Åyvind; Lande, Tor S.; Hamran, Svein-Erik

    2014-05-01

    Cardiovascular diseases (CVD) are a major cause of deaths all over the world. Microwave radar can be an alternative sensor for heart diagnostics and monitoring in modern healthcare that aids early detection of CVD symptoms. In this paper measurements from a switch array radar system are presented. This UWB system operates below 3 GHz and does time-lapse imaging of the beating heart inside the human body. The array consists of eight fat dipole elements. With a switch system, every possible sequence of transmit/receive element pairs can be selected to build a radar image from the recordings. To make the radar waves penetrate the human tissue, the antenna array is placed in contact with the body. Removal of the direct signal leakage through the antennas and body surface are done by high-pass (HP) filtering of the data prior to image processing. To analyze the results, measurements of moving spheres in air and simulations are carried out. We see that removal of the direct signal introduces amplitude distortion in the images. In addition, the effect of small target motion between the collection times of data from the individual elements is analyzed. With low pulse repetition frequency (PRF) this motion will distort the image. By using data from real measurements of heart motion in simulations, we analyze how the PRF and the antenna geometry influence this distortions.

  18. Image-based characterization of thrombus formation in time-lapse DIC microscopy.

    Science.gov (United States)

    Brieu, Nicolas; Navab, Nassir; Serbanovic-Canic, Jovana; Ouwehand, Willem H; Stemple, Derek L; Cvejic, Ana; Groher, Martin

    2012-05-01

    The characterization of thrombus formation in time-lapse DIC microscopy is of increased interest for identifying genes which account for atherothrombosis and coronary artery diseases (CADs). In particular, we are interested in large-scale studies on zebrafish, which result in large amount of data, and require automatic processing. In this work, we present an image-based solution for the automatized extraction of parameters quantifying the temporal development of thrombotic plugs. Our system is based on the joint segmentation of thrombotic and aortic regions over time. This task is made difficult by the low contrast and the high dynamic conditions observed in vivo DIC microscopic scenes. Our key idea is to perform this segmentation by distinguishing the different motion patterns in image time series rather than by solving standard image segmentation tasks in each image frame. Thus, we are able to compensate for the poor imaging conditions. We model motion patterns by energies based on the idea of dynamic textures, and regularize the model by two prior energies on the shape of the aortic region and on the topological relationship between the thrombus and the aorta. We demonstrate the performance of our segmentation algorithm by qualitative and quantitative experiments on synthetic examples as well as on real in vivo microscopic sequences.

  19. Segmentation Method of Time-Lapse Microscopy Images with the Focus on Biocompatibility Assessment.

    Science.gov (United States)

    Soukup, Jindřich; Císař, Petr; Šroubek, Filip

    2016-06-01

    Biocompatibility testing of new materials is often performed in vitro by measuring the growth rate of mammalian cancer cells in time-lapse images acquired by phase contrast microscopes. The growth rate is measured by tracking cell coverage, which requires an accurate automatic segmentation method. However, cancer cells have irregular shapes that change over time, the mottled background pattern is partially visible through the cells and the images contain artifacts such as halos. We developed a novel algorithm for cell segmentation that copes with the mentioned challenges. It is based on temporal differences of consecutive images and a combination of thresholding, blurring, and morphological operations. We tested the algorithm on images of four cell types acquired by two different microscopes, evaluated the precision of segmentation against manual segmentation performed by a human operator, and finally provided comparison with other freely available methods. We propose a new, fully automated method for measuring the cell growth rate based on fitting a coverage curve with the Verhulst population model. The algorithm is fast and shows accuracy comparable with manual segmentation. Most notably it can correctly separate live from dead cells.

  20. Two-photon imaging of stem cells

    Science.gov (United States)

    Uchugonova, A.; Gorjup, E.; Riemann, I.; Sauer, D.; König, K.

    2008-02-01

    A variety of human and animal stem cells (rat and human adult pancreatic stem cells, salivary gland stem cells, dental pulpa stem cells) have been investigated by femtosecond laser 5D two-photon microscopy. Autofluorescence and second harmonic generation have been imaged with submicron spatial resolution, 270 ps temporal resolution, and 10 nm spectral resolution. In particular, NADH and flavoprotein fluorescence was detected in stem cells. Major emission peaks at 460nm and 530nm with typical mean fluorescence lifetimes of 1.8 ns and 2.0 ns, respectively, were measured using time-correlated single photon counting and spectral imaging. Differentiated stem cells produced the extracellular matrix protein collagen which was detected by SHG signals at 435 nm.

  1. A Modular and Affordable Time-Lapse Imaging and Incubation System Based on 3D Printed Parts, a Smartphone, and Off-The-Shelf Electronics

    OpenAIRE

    Hernández Vera, Rodrigo; Schwan, Emil; Fatsis-Kavalopoulos, Nikos; Kreuger, Johan

    2016-01-01

    Time-lapse imaging is a powerful tool for studying cellular dynamics and cell behavior over long periods of time to acquire detailed functional information. However, commercially available time-lapse imaging systems are expensive and this has limited a broader implementation of this technique in low-resource environments. Further, the availability of time-lapse imaging systems often present workflow bottlenecks in well-funded institutions. To address these limitations we have designed a modul...

  2. Microfluidics-integrated time-lapse imaging for analysis of cellular dynamics.

    Science.gov (United States)

    Albrecht, Dirk R; Underhill, Gregory H; Resnikoff, Joshua; Mendelson, Avital; Bhatia, Sangeeta N; Shah, Jagesh V

    2010-06-01

    An understanding of the mechanisms regulating cellular responses has recently been augmented by innovations enabling the observation of phenotypes at high spatio-temporal resolution. Technologies such as microfluidics have sought to expand the throughput of these methods, although assimilation with advanced imaging strategies has been limited. Here, we describe the pairing of high resolution time-lapse imaging with microfluidic multiplexing for the analysis of cellular dynamics, utilizing a design selected for facile fabrication and operation, and integration with microscopy instrumentation. This modular, medium-throughput platform enables the long-term imaging of living cells at high numerical aperture (via oil immersion) by using a conserved 96-well, approximately 6 x 5 mm(2) imaging area with a variable input/output channel design chosen for the number of cell types and microenvironments under investigation. In the validation of this system, we examined fundamental features of cell cycle progression, including mitotic kinetics and spindle orientation dynamics, through the high-resolution parallel analysis of model cell lines subjected to anti-mitotic agents. We additionally explored the self-renewal kinetics of mouse embryonic stem cells, and demonstrate the ability to dynamically assess and manipulate stem cell proliferation, detect rare cell events, and measure extended time-scale correlations. We achieved an experimental throughput of >900 cells/experiment, each observed at >40x magnification for up to 120 h. Overall, these studies illustrate the capacity to probe cellular functions and yield dynamic information in time and space through the integration of a simple, modular, microfluidics-based imaging platform.

  3. Three dimensional time lapse imaging of live cell mitochondria with photothermal optical lock-in optical coherence microscopy (Conference Presentation)

    Science.gov (United States)

    Sison, Miguel; Chakrabortty, Sabyasachi; Extermann, Jerome; Nahas, Amir; Pache, Christophe; Weil, Tanja; Lasser, Theo

    2016-03-01

    The photothermal optical lock-in optical coherence microscope (poli-OCM) introduced molecular specificity to OCM imaging, which is conventionally, a label-free technique. Here we achieve three-dimensional live cell and mitochondria specific imaging using ~4nm protein-functionalized gold nanoparticles (AuNPs). These nanoparticles do not photobleach and we demonstrate they're suitability for long-term time lapse imaging. We compare the accuracy of labelling with these AuNPs using classical fluorescence confocal imaging with a standard mitochondria specific marker. Furthermore, time lapse poli-OCM imaging every 5 minutes over 1.5 hours period was achieved, revealing the ability for three-dimensional monitoring of mitochondria dynamics.

  4. Using electromagnetic conductivity imaging to generate time-lapse soil moisture estimates.

    Science.gov (United States)

    Huang, Jingyi; Scuderio, Elia; Corwin, Dennis; Triantafilis, John

    2015-04-01

    Irrigated agriculture is crucial to the agricultural productivity of the Moreno valley. To maintain profitability, more will need to be done by irrigators with less water, owing to competing demands from rapidly expanding urbanisation in southern California. In this regard, irrigators need to understand the spatial and temporal variation of soil moisture to discern inefficiencies. However, soil moisture is difficult to measure and monitor, unless a large bank of soil sensors are installed and at various depths in the profile. In order to value add to the limited amount of information, geophysical techniques, such as direct current resisivity (DCR) arrays are used to develop electrical resistivity images (ERI). Whilst successful the approach is time consuming and labour intensive. In this research we describe how equivalent data can be collected using a proximal sensing electromagnetic (EM) induction instrument (i.e. DUALEM-421) and inversion software (EM4Soil) to generate EM conductivity images (EMCI). Figure 1 shows the EMCI generated from DUALEM-421 data acquired at various days of a time-lapse experiment and including; day a) 0, b) 1, c) 2, d) 3, e) 5, f) 7 and g) 11. We calibrate the estimates of true electrical conductivity (sigma - mS/m) with volumetric moisture content and show with good accuracy the spatial and temporal variation of soil moisture status and over 12 day period. The results show clearly that the pivot sprinkler irrigation system is effective at providing sufficient amounts of water to the top 0.5 m of a Lucerne crop (i.e. red shaded areas of high sigma). However, in some places faulty sprinklers are evident owing to the lack of wetting (i.e. blue shaded areas of low sigma). In addition, and over time, our approach shows clearly the effect the Lucerne crop has in drying the soil profile and using the soil moisture.

  5. Subcellular in vivo time-lapse imaging and optical manipulation of Caenorhabditis elegans in standard multiwell plates.

    Science.gov (United States)

    Rohde, Christopher B; Yanik, Mehmet Fatih

    2011-01-01

    High-resolution in vivo time-lapse assays require repeated immobilization and imaging of whole animals. Here we report a technology for screening Caenorhabditis elegans at cellular resolution over its entire lifespan inside standard multiwell plates using repeated immobilization, imaging and optical manipulation. Our system does not use any fluidic or mechanical components, and can operate for tens of thousands of cycles without failure. It is also compatible with industrial high-throughput screening platforms and robotics, and it allows both chemical, and forward and reverse genetic screens. We used this technology to perform subcellular-resolution femtosecond laser microsurgery of single neurons in vivo, and to image the subsequent regeneration dynamics at subcellular resolution. Our single-neuron in vivo time-lapse analysis shows that neurite regrowth occurring over short time windows is significantly greater than that predicted by ensemble averaging over many animals.

  6. Pixel timing correction in time-lapsed calcium imaging using point scanning microscopy.

    Science.gov (United States)

    Boiroux, Dimitri; Oke, Yoshihiko; Miwakeichi, Fumikazu; Oku, Yoshitaka

    2014-11-30

    In point scanning imaging, data are acquired by sequentially scanning each pixel of a predetermined area. This way of scanning leads to time delays between pixels, especially for lower scanning speed or large scanned areas. Therefore, experiments are often performed at lower framerates in order to ensure a sufficient signal-to-noise ratio, even though framerates above 30 frames per second are technically feasible. For these framerates, we suggest that it becomes crucial to correct the time delay between image pixels prior to analyses. In this paper, we apply temporal interpolation (or pixel timing correction) for calcium imaging in two-photon microscopy as an example of fluorescence imaging. We present and compare three interpolation methods (linear, Lanczos and cubic B-spline). We test these methods on a simulated network of coupled bursting neurons at different framerates. In this network, we introduce a time delay to simulate a scanning by point scanning microscopy. We also assess these methods on actual microscopic calcium imaging movies recorded at usual framerates. Our numerical results suggest that point scanning microscopy imaging introduces statistically significant time delays between image pixels at low frequency. However, we demonstrate that pixel timing correction compensates for these time delays, regardless of the used interpolation method.

  7. OPTICAL FLOW APPLIED TO TIME-LAPSE IMAGE SERIES TO ESTIMATE GLACIER MOTION IN THE SOUTHERN PATAGONIA ICE FIELD

    OpenAIRE

    E. Lannutti; Lenzano, M. G.; Toth, C; L. Lenzano; Rivera, A.

    2016-01-01

    In this work, we assessed the feasibility of using optical flow to obtain the motion estimation of a glacier. In general, former investigations used to detect glacier changes involve solutions that require repeated observations which are many times based on extensive field work. Taking into account glaciers are usually located in geographically complex and hard to access areas, deploying time-lapse imaging sensors, optical flow may provide an efficient solution at good spatial and te...

  8. Optical Flow Applied to Time-Lapse Image Series to Estimate Glacier Motion in the Southern Patagonia Ice Field

    Science.gov (United States)

    Lannutti, E.; Lenzano, M. G.; Toth, C.; Lenzano, L.; Rivera, A.

    2016-06-01

    In this work, we assessed the feasibility of using optical flow to obtain the motion estimation of a glacier. In general, former investigations used to detect glacier changes involve solutions that require repeated observations which are many times based on extensive field work. Taking into account glaciers are usually located in geographically complex and hard to access areas, deploying time-lapse imaging sensors, optical flow may provide an efficient solution at good spatial and temporal resolution to describe mass motion. Several studies in computer vision and image processing community have used this method to detect large displacements. Therefore, we carried out a test of the proposed Large Displacement Optical Flow method at the Viedma Glacier, located at South Patagonia Icefield, Argentina. We collected monoscopic terrestrial time-lapse imagery, acquired by a calibrated camera at every 24 hour from April 2014 until April 2015. A filter based on temporal correlation and RGB color discretization between the images was applied to minimize errors related to changes in lighting, shadows, clouds and snow. This selection allowed discarding images that do not follow a sequence of similarity. Our results show a flow field in the direction of the glacier movement with acceleration in the terminus. We analyzed the errors between image pairs, and the matching generally appears to be adequate, although some areas show random gross errors related to the presence of changes in lighting. The proposed technique allowed the determination of glacier motion during one year, providing accurate and reliable motion data for subsequent analysis.

  9. Near real-time imaging of molasses injections using time-lapse electrical geophysics at the Brandywine DRMO, Brandywine, Maryland

    Science.gov (United States)

    Versteeg, R. J.; Johnson, T.; Major, B.; Day-Lewis, F. D.; Lane, J. W.

    2010-12-01

    Enhanced bioremediation, which involves introduction of amendments to promote biodegradation, increasingly is used to accelerate cleanup of recalcitrant compounds and has been identified as the preferred remedial treatment at many contaminated sites. Although blind introduction of amendments can lead to sub-optimal or ineffective remediation, the distribution of amendment throughout the treatment zone is difficult to measure using conventional sampling. Because amendments and their degradation products commonly have electrical properties that differ from those of ambient soil, time-lapse electrical geophysical monitoring has the potential to verify amendment emplacement and distribution. In order for geophysical monitoring to be useful, however, results of the injection ideally should be accessible in near real time. In August 2010, we demonstrated the feasibility of near real-time, autonomous electrical geophysical monitoring of amendment injections at the former Defense Reutilization and Marketing Office (DRMO) in Brandywine, Maryland. Two injections of about 1000 gallons each of molasses, a widely used amendment for enhanced bioremediation, were monitored using measurements taken with borehole and surface electrodes. During the injections, multi-channel resistance data were recorded; data were transmitted to a server and processed using a parallel resistivity inversion code; and results in the form of time-lapse imagery subsequently were posted to a website. This process occurred automatically without human intervention. The resulting time-lapse imagery clearly showed the evolution of the molasses plume. The delay between measurements and online delivery of images was between 45 and 60 minutes, thus providing actionable information that could support decisions about field procedures and a check on whether amendment reached target zones. This experiment demonstrates the feasibility of using electrical imaging as a monitoring tool both during amendment emplacement

  10. Analysis of Zebrafish Kidney Development with Time-lapse Imaging Using a Dissecting Microscope Equipped for Optical Sectioning.

    Science.gov (United States)

    Perner, Birgit; Schnerwitzki, Danny; Graf, Michael; Englert, Christoph

    2016-04-07

    In order to understand organogenesis, the spatial and temporal alterations that occur during development of tissues need to be recorded. The method described here allows time-lapse analysis of normal and impaired kidney development in zebrafish embryos by using a fluorescence dissecting microscope equipped for structured illumination and z-stack acquisition. To visualize nephrogenesis, transgenic zebrafish (Tg(wt1b:GFP)) with fluorescently labeled kidney structures were used. Renal defects were triggered by injection of an antisense morpholino oligonucleotide against the Wilms tumor gene wt1a, a factor known to be crucial for kidney development. The advantage of the experimental setup is the combination of a zoom microscope with simple strategies for re-adjusting movements in x, y or z direction without additional equipment. To circumvent focal drift that is induced by temperature variations and mechanical vibrations, an autofocus strategy was applied instead of utilizing a usually required environmental chamber. In order to re-adjust the positional changes due to a xy-drift, imaging chambers with imprinted relocation grids were employed. In comparison to more complex setups for time-lapse recording with optical sectioning such as confocal laser scanning or light sheet microscopes, a zoom microscope is easy to handle. Besides, it offers dissecting microscope-specific benefits such as high depth of field and an extended working distance. The method to study organogenesis presented here can also be used with fluorescence stereo microscopes not capable of optical sectioning. Although limited for high-throughput, this technique offers an alternative to more complex equipment that is normally used for time-lapse recording of developing tissues and organ dynamics.

  11. Time-lapse Confocal Imaging of Migrating Neurons in Organotypic Slice Culture of Embryonic Mouse Brain Using In Utero Electroporation.

    Science.gov (United States)

    Wiegreffe, Christoph; Feldmann, Svenja; Gaessler, Simeon; Britsch, Stefan

    2017-07-25

    In utero electroporation is a rapid and powerful approach to study the process of radial migration in the cerebral cortex of developing mouse embryos. It has helped to describe the different steps of radial migration and characterize the molecular mechanisms controlling this process. To directly and dynamically analyze migrating neurons they have to be traced over time. This protocol describes a workflow that combines in utero electroporation with organotypic slice culture and time-lapse confocal imaging, which allows for a direct examination and dynamic analysis of radially migrating cortical neurons. Furthermore, detailed characterization of migrating neurons, such as migration speed, speed profiles, as well as radial orientation changes, is possible. The method can easily be adapted to perform functional analyses of genes of interest in radially migrating cortical neurons by loss and gain of function as well as rescue experiments. Time-lapse imaging of migrating neurons is a state-of-the-art technique that once established is a potent tool to study the development of the cerebral cortex in mouse models of neuronal migration disorders.

  12. Imaging subsurface migration of dissolved CO2 in a shallow aquifer using 3-D time-lapse electrical resistivity tomography

    DEFF Research Database (Denmark)

    Auken, Esben; Doetsch, Joseph; Fiandaca, Gianluca

    2014-01-01

    of aeolian and glacial sands near the surface and marine sands below 10m depth. 3-D time-lapse ERT inversions clearly image the dissolved CO2 plume with decreased electrical resistivity values. We can image the geochemical changes induced by the dissolved CO2 until the end of the acquisition, 120days after......Contamination of groundwater by leaking CO2 is a potential risk of carbon sequestration. With the help of a field experiment in western Denmark, we investigate to what extent surface electrical resistivity tomography (ERT) can detect and image dissolved CO2 in a shallow aquifer. For this purpose......, we injected CO2 at a depth of 5 and 10m and monitored its migration using 320 electrodes on a 126m×25m surface grid. A fully automated acquisition system continuously collected data and uploaded it into an online database. The large amount of data allows for time-series analysis using geostatistical...

  13. Time-lapse walkaway VSP imaging using reverse-time migration in the angle domain for monitoring CO2 injection at the SACROC EOR field

    Science.gov (United States)

    Huang, L.; Huang, H.

    2012-12-01

    Time-lapse walkaway vertical seismic profiling (VSP) surveys can reveal important reservoir changes caused by CO2 injection. We study the capability of time-lapse walkaway VSP imaging using reverse-time migration in the angle-domain for monitoring CO2 injection. During the Phase II project of the Southwest Regional Partnership on Carbon Sequestration, one baseline and one repeat walkaway VSP surveys were conducted in 2008 and 2009, respectively, at the SACROC enhanced oil recovery (EOR) field for monitoring CO2 injection. The datasets were acquired by Baker Atlas in collaboration with Kinder Morgan. In this study, we apply reverse-time migration in the angle domain to the time-lapse walkaway VSP datasets from the SACROC EOR field, and conduct detailed analyses of common-image gathers. Our migration results demonstrate that reverse-time migration in the angle domain produces images of time-lapse walkaway VSP data with a better image quality compared to those obtained using conventional reverse-time migration. The time-lapse image difference along the bottom of the reservoir where CO2 is injected is much more significant than that along the top of the reservoir. This is partially because we use the same baseline velocity model for migrations of both datasets. The reservoir velocity decreases during CO2 injection, leading to slightly change in the migration image location along the bottom of the reservoir for the repeat VSP data.

  14. A Modular and Affordable Time-Lapse Imaging and Incubation System Based on 3D-Printed Parts, a Smartphone, and Off-The-Shelf Electronics.

    Science.gov (United States)

    Hernández Vera, Rodrigo; Schwan, Emil; Fatsis-Kavalopoulos, Nikos; Kreuger, Johan

    2016-01-01

    Time-lapse imaging is a powerful tool for studying cellular dynamics and cell behavior over long periods of time to acquire detailed functional information. However, commercially available time-lapse imaging systems are expensive and this has limited a broader implementation of this technique in low-resource environments. Further, the availability of time-lapse imaging systems often present workflow bottlenecks in well-funded institutions. To address these limitations we have designed a modular and affordable time-lapse imaging and incubation system (ATLIS). The ATLIS enables the transformation of simple inverted microscopes into live cell imaging systems using custom-designed 3D-printed parts, a smartphone, and off-the-shelf electronic components. We demonstrate that the ATLIS provides stable environmental conditions to support normal cell behavior during live imaging experiments in both traditional and evaporation-sensitive microfluidic cell culture systems. Thus, the system presented here has the potential to increase the accessibility of time-lapse microscopy of living cells for the wider research community.

  15. A Modular and Affordable Time-Lapse Imaging and Incubation System Based on 3D-Printed Parts, a Smartphone, and Off-The-Shelf Electronics

    Science.gov (United States)

    Schwan, Emil; Fatsis-Kavalopoulos, Nikos; Kreuger, Johan

    2016-01-01

    Time-lapse imaging is a powerful tool for studying cellular dynamics and cell behavior over long periods of time to acquire detailed functional information. However, commercially available time-lapse imaging systems are expensive and this has limited a broader implementation of this technique in low-resource environments. Further, the availability of time-lapse imaging systems often present workflow bottlenecks in well-funded institutions. To address these limitations we have designed a modular and affordable time-lapse imaging and incubation system (ATLIS). The ATLIS enables the transformation of simple inverted microscopes into live cell imaging systems using custom-designed 3D-printed parts, a smartphone, and off-the-shelf electronic components. We demonstrate that the ATLIS provides stable environmental conditions to support normal cell behavior during live imaging experiments in both traditional and evaporation-sensitive microfluidic cell culture systems. Thus, the system presented here has the potential to increase the accessibility of time-lapse microscopy of living cells for the wider research community. PMID:28002463

  16. Grain-scale deformation in granular materials: time-lapse XCT-imaging of a deforming reservoir sandstone

    Science.gov (United States)

    Hangx, Suzanne; Cordonnier, Benoît; Pijnenburg, Ronald; Renard, François; Spiers, Christopher

    2017-04-01

    Relating macroscopic deformation of granular media to grain-scale processes, such as grain fracturing, has been a focus of many studies. Understanding these processes is key for predicting surface subsidence and induced seismicity caused by hydrocarbon depletion, the hydraulic fracturing response of geothermal reservoirs, and post-seismic crustal deformation. With the development of state-of-the-art techniques, such as time-lapse X-ray tomography imaging during triaxial deformation, new avenues to investigate the operating mechanisms have opened up. As a first step to understanding grain-scale deformation processes, we performed a deformation experiment on highly porous sandstone, obtained from a depleting gas reservoir, using a novel small-scale triaxial deformation apparatus coupled to high-resolution 4D X-ray tomography, available at the European Synchrotron Radiation Facility (ESRF, Grenoble) and Université Grenoble Alpes. This state-of-the-art apparatus allows for 3D time-lapse imaging of samples, while deforming at pressure, temperature and fluid flow conditions relevant for geological reservoirs. We performed our experiment at relevant in-situ reservoir conditions (T = 100˚ C, 10 MPa pore pressure, 40 MPa effective confining pressure). Axial stress was increased step-wise until failure occurred, while continuously imaging deformation. This enables us to monitor progressive grain failure, and strain localisation, during deformation in real-time. Though the vast amount of data obtained from even a single test poses challenges for data analysis, this presentation will address the first results obtained from this experiment.

  17. Composite time-lapse computed tomography and micro finite element simulations: A new imaging approach for characterizing cement flows and mechanical benefits of vertebroplasty.

    Science.gov (United States)

    Stadelmann, Vincent A; Zderic, Ivan; Baur, Annick; Unholz, Cynthia; Eberli, Ursula; Gueorguiev, Boyko

    2016-02-01

    Vertebroplasty has been shown to reinforce weak vertebral bodies and reduce fracture risks, yet cement leakage is a major problem that can cause severe complications. Since cement flow is nearly impossible to control during surgery, small volumes of cement are injected, but then mechanical benefits might be limited. A better understanding of cement flows within bone structure is required to further optimize vertebroplasty and bone augmentation in general. We developed a novel imaging method, composite time-lapse CT, to characterize cement flow during injection. In brief, composite-resolution time-lapse CT exploits the qualities of microCT and clinical CT. The method consists in overlaying low-resolution time-lapse CT scans acquired during injection onto pre-operative high-resolution microCT scans, generating composite-resolution time-lapse CT series of cement flow within bone. In this in vitro study, composite-resolution time-lapse CT was applied to eight intact and five artificially fractured cadaveric vertebrae during vertebroplasty. The time-lapse scans were acquired at one-milliliter cement injection steps until a total of 10 ml cement was injected. The composite-resolution series were then converted into micro finite element models to compute strains distribution under virtual axial loading. Relocation of strain energy density within bone structure was observed throughout the progression of the procedure. Interestingly, the normalized effect of cement injection on the overall stiffness of the vertebrae was similar between intact and fractured specimens, although at different orders of magnitude. In conclusion, composite time-lapse CT can picture cement flows during bone augmentation. The composite images can also be easily converted into finite element models to compute virtual strain distributions under loading at every step of an injection, providing deeper understanding on the biomechanics of vertebroplasty.

  18. Time-Lapse Observation of Electrolysis of Copper Sulfate with a Full-Field X-ray Fluorescence Imaging Microscope

    Science.gov (United States)

    Ohigashi, Takuji; Aota, Tatsuya; Watanabe, Norio; Takano, Hidekazu; Yokosuka, Hiroki; Aoki, Sadao

    2008-06-01

    The time-lapse observation of the electrodeposition of copper in copper sulfate solution was performed by imaging X-ray fluorescence from the copper deposition. The X-ray fluorescence was directly imaged with a full-field Wolter mirror microscope, which was constructed at the Photon Factory. Controlling the electric current in the solution from 0 to 71.7 µA, the deposition of copper on a Pt cathode was directly observed by imaging its X-ray fluorescence. One exposure time for obtaining an X-ray fluorescence image was 80 s. Then, it was 17 min later from the beginning of the electrolysis when the X-ray fluorescence image of the electrodeposition is observed for the first time. At this exposure time, the detection limit of the mass of copper was estimated to be 0.60 pg/image, which was calculated using test samples of 1.00×10-3-1.00 mol/l copper sulfate solutions.

  19. Analysis of the temperature dynamics of a proglacial river using time-lapse thermal imaging and energy balance modeling

    Science.gov (United States)

    Cardenas, M. Bayani; Doering, Michael; Rivas, Denny S.; Galdeano, Carlos; Neilson, Bethany T.; Robinson, Christopher T.

    2014-11-01

    Understanding the temperature dynamics of rivers is critical for their management and for ecological and biogeochemical aquatic processes. In proglacial rivers, there is typically a paucity of thermal observations which in turn limits the understanding of these sensitive and evolving environments. Here we collected ground-based thermal images, with approximately meter resolution and imaged every half hour for 24 h, of a proglacial river and 2 km2 of its floodplain and interpret the observations using a numerical energy balance model. The images revealed the longitudinal thermal pattern of the Urbach River in Switzerland - there was gradual cooling in the upstream half of the study section and then warming in the remaining downstream portion. This pattern persisted through the diurnal warming and cooling cycle. The spatio-temporal thermal pattern was explained by a model that included distributed thermal inputs of cooler water in the upstream half coming from alluvial fans and warmer water in the downstream half running off steep cliffs that warm snowmelt. The warm inputs from the cliffs were confirmed by the thermal imaging. These data and the associated modeling illustrated that distributed inflows can overwhelm the influence of atmospheric fluxes, and that their knowledge is critical for understanding stream temperatures. The combination of modeling and detailed time-lapse thermal imaging allowed for identification and quantification of processes critical to in-stream temperature dynamics in a proglacial river.

  20. Automatic Tracking and Motility Analysis of Human Sperm in Time-Lapse Images.

    Science.gov (United States)

    Urbano, Leonardo F; Masson, Puneet; VerMilyea, Matthew; Kam, Moshe

    2017-03-01

    We present a fully automated multi-sperm tracking algorithm. It has the demonstrated capability to detect and track simultaneously hundreds of sperm cells in recorded videos while accurately measuring motility parameters over time and with minimal operator intervention. Algorithms of this kind may help in associating dynamic swimming parameters of human sperm cells with fertility and fertilization rates. Specifically, we offer an image processing method, based on radar tracking algorithms, that detects and tracks automatically the swimming paths of human sperm cells in timelapse microscopy image sequences of the kind that is analyzed by fertility clinics. Adapting the well-known joint probabilistic data association filter (JPDAF), we automatically tracked hundreds of human sperm simultaneously and measured their dynamic swimming parameters over time. Unlike existing CASA instruments, our algorithm has the capability to track sperm swimming in close proximity to each other and during apparent cell-to-cell collisions. Collecting continuously parameters for each sperm tracked without sample dilution (currently impossible using standard CASA systems) provides an opportunity to compare such data with standard fertility rates. The use of our algorithm thus has the potential to free the clinician from having to rely on elaborate motility measurements obtained manually by technicians, speed up semen processing, and provide medical practitioners and researchers with more useful data than are currently available.

  1. Segmentation of Environmental Time Lapse Image Sequences for the Determination of Shore Lines Captured by Hand-Held Smartphone Cameras

    Science.gov (United States)

    Kröhnert, M.; Meichsner, R.

    2017-09-01

    The relevance of globally environmental issues gains importance since the last years with still rising trends. Especially disastrous floods may cause in serious damage within very short times. Although conventional gauging stations provide reliable information about prevailing water levels, they are highly cost-intensive and thus just sparsely installed. Smartphones with inbuilt cameras, powerful processing units and low-cost positioning systems seem to be very suitable wide-spread measurement devices that could be used for geo-crowdsourcing purposes. Thus, we aim for the development of a versatile mobile water level measurement system to establish a densified hydrological network of water levels with high spatial and temporal resolution. This paper addresses a key issue of the entire system: the detection of running water shore lines in smartphone images. Flowing water never appears equally in close-range images even if the extrinsics remain unchanged. Its non-rigid behavior impedes the use of good practices for image segmentation as a prerequisite for water line detection. Consequently, we use a hand-held time lapse image sequence instead of a single image that provides the time component to determine a spatio-temporal texture image. Using a region growing concept, the texture is analyzed for immutable shore and dynamic water areas. Finally, the prevalent shore line is examined by the resultant shapes. For method validation, various study areas are observed from several distances covering urban and rural flowing waters with different characteristics. Future work provides a transformation of the water line into object space by image-to-geometry intersection.

  2. Long-term, high-resolution confocal time lapse imaging of Arabidopsis cotyledon epidermis during germination.

    Science.gov (United States)

    Peterson, Kylee M; Torii, Keiko U

    2012-12-31

    Imaging in vivo dynamics of cellular behavior throughout a developmental sequence can be a powerful technique for understanding the mechanics of tissue patterning. During animal development, key cell proliferation and patterning events occur very quickly. For instance, in Caenorhabditis elegans all cell divisions required for the larval body plan are completed within six hours after fertilization, with seven mitotic cycles(1); the sixteen or more mitoses of Drosophila embryogenesis occur in less than 24 hr(2). In contrast, cell divisions during plant development are slow, typically on the order of a day (3,4,5) . This imposes a unique challenge and a need for long-term live imaging for documenting dynamic behaviors of cell division and differentiation events during plant organogenesis. Arabidopsis epidermis is an excellent model system for investigating signaling, cell fate, and development in plants. In the cotyledon, this tissue consists of air- and water-resistant pavement cells interspersed with evenly distributed stomata, valves that open and close to control gas exchange and water loss. Proper spacing of these stomata is critical to their function, and their development follows a sequence of asymmetric division and cell differentiation steps to produce the organized epidermis (Fig. 1). This protocol allows observation of cells and proteins in the epidermis over several days of development. This time frame enables precise documentation of stem-cell divisions and differentiation of epidermal cells, including stomata and epidermal pavement cells. Fluorescent proteins can be fused to proteins of interest to assess their dynamics during cell division and differentiation processes. This technique allows us to understand the localization of a novel protein, POLAR(6), during the proliferation stage of stomatal-lineage cells in the Arabidopsis cotyledon epidermis, where it is expressed in cells preceding asymmetric division events and moves to a characteristic area of

  3. Two-photon imaging through a multimode fiber

    CERN Document Server

    Morales-Delgado, Edgar E; Moser, Christophe

    2015-01-01

    In this work we demonstrate 3D imaging using two-photon excitation through a 20 cm long multimode optical fiber (MMF) of 350 micrometers diameter. The imaging principle is similar to single photon fluorescence through a MMF, except that a focused femtosecond pulse is delivered and scanned over the sample. In our approach, focusing and scanning through the fiber is accomplished by digital phase conjugation using mode selection by time gating with an ultra-fast reference pulse. The excited two-photon emission is collected through the same fiber. We demonstrate depth sectioning by scanning the focused pulse in a 3D volume over a sample consisting of fluorescent beads suspended in a polymer. The achieved resolution is 1 micrometer laterally and 15 micrometers axially. Scanning is performed over an 80x80 micrometers field of view. To our knowledge, this is the first demonstration of high-resolution three-dimensional imaging using two-photon fluorescence through a multimode fiber.

  4. In vivo time-lapse imaging of skin burn wound healing using second-harmonic generation microscopy

    Science.gov (United States)

    Yasui, Takeshi; Tanaka, Ryosuke; Hase, Eiji; Fukushima, Shu-ichiro; Araki, Tsutomu

    2014-02-01

    Wound healing is a process to repair the damaged tissue caused by thermal burn, incised wound, or stab wound. Although the wound healing has many aspects, it is common for dynamics of collagen fiber, such as decomposition, production, or growth, to be closely related with wound healing. If such the healing process can be visualized as a timelapse image of the collagen fiber in the same subject, one may obtain new findings regarding biological repairing mechanisms in the healing process. In this article, to investigate the temporal modoification of dermal collagen fiber in the burn wound healing, we used second-harmonic-generation (SHG) microscopy, showing high selectivity and good image contrast to collagen molecules as well as high spatial resolution, optical three-dimensional sectioning, minimal invasiveness, deep penetration, the absence of interference from background light, and in vivo measurement without additional staining. Since SHG light arises from a non-centrosymmetric triple helix of three polypeptide chains in the collagen molecule, SHG intensity sensitively reflects the structure maturity of collagen molecule and its aggregates. A series of time-lapse SHG images during the wound healing process of 2 weeks clearly indicated that condensation and melting of dermal collagen fibers by the deep dermal burn, decomposition of the damaged collagen fibers in the inflammation phase, production of new collagen fibers in the proliferation phase, and the growth of the new collagen fibers in the remodeling phase. These results show a high potential of SHG microscopy for optical assessment of the wound healing process in vivo.

  5. Analyse de "Time Lapse" optiques stéréo et d'images radar satellitaires : application à la mesure du déplacement de glaciers

    OpenAIRE

    Pham, Ha Thai

    2015-01-01

    Earth observation by image acquisition systems allows the survey of temporal evolution of natural phenomena such as earthquakes, volcanoes or gravitational movements. Various techniques exist including satellite imagery, terrestrial photogrammetry and in-situ measurements. Image time series from automatic cameras (Time Lapse) are a growing source of information since they offer an interesting compromise in terms of spatial coverage and observation frequency in order to measure surface motion ...

  6. Two-photon imaging and analysis of neural network dynamics

    Science.gov (United States)

    Lütcke, Henry; Helmchen, Fritjof

    2011-08-01

    The glow of a starry night sky, the smell of a freshly brewed cup of coffee or the sound of ocean waves breaking on the beach are representations of the physical world that have been created by the dynamic interactions of thousands of neurons in our brains. How the brain mediates perceptions, creates thoughts, stores memories and initiates actions remains one of the most profound puzzles in biology, if not all of science. A key to a mechanistic understanding of how the nervous system works is the ability to measure and analyze the dynamics of neuronal networks in the living organism in the context of sensory stimulation and behavior. Dynamic brain properties have been fairly well characterized on the microscopic level of individual neurons and on the macroscopic level of whole brain areas largely with the help of various electrophysiological techniques. However, our understanding of the mesoscopic level comprising local populations of hundreds to thousands of neurons (so-called 'microcircuits') remains comparably poor. Predominantly, this has been due to the technical difficulties involved in recording from large networks of neurons with single-cell spatial resolution and near-millisecond temporal resolution in the brain of living animals. In recent years, two-photon microscopy has emerged as a technique which meets many of these requirements and thus has become the method of choice for the interrogation of local neural circuits. Here, we review the state-of-research in the field of two-photon imaging of neuronal populations, covering the topics of microscope technology, suitable fluorescent indicator dyes, staining techniques, and in particular analysis techniques for extracting relevant information from the fluorescence data. We expect that functional analysis of neural networks using two-photon imaging will help to decipher fundamental operational principles of neural microcircuits.

  7. Two-photon imaging and analysis of neural network dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Luetcke, Henry; Helmchen, Fritjof [Brain Research Institute, University of Zurich, Winterthurerstrasse 190, CH-8057 Zurich (Switzerland)

    2011-08-15

    The glow of a starry night sky, the smell of a freshly brewed cup of coffee or the sound of ocean waves breaking on the beach are representations of the physical world that have been created by the dynamic interactions of thousands of neurons in our brains. How the brain mediates perceptions, creates thoughts, stores memories and initiates actions remains one of the most profound puzzles in biology, if not all of science. A key to a mechanistic understanding of how the nervous system works is the ability to measure and analyze the dynamics of neuronal networks in the living organism in the context of sensory stimulation and behavior. Dynamic brain properties have been fairly well characterized on the microscopic level of individual neurons and on the macroscopic level of whole brain areas largely with the help of various electrophysiological techniques. However, our understanding of the mesoscopic level comprising local populations of hundreds to thousands of neurons (so-called 'microcircuits') remains comparably poor. Predominantly, this has been due to the technical difficulties involved in recording from large networks of neurons with single-cell spatial resolution and near-millisecond temporal resolution in the brain of living animals. In recent years, two-photon microscopy has emerged as a technique which meets many of these requirements and thus has become the method of choice for the interrogation of local neural circuits. Here, we review the state-of-research in the field of two-photon imaging of neuronal populations, covering the topics of microscope technology, suitable fluorescent indicator dyes, staining techniques, and in particular analysis techniques for extracting relevant information from the fluorescence data. We expect that functional analysis of neural networks using two-photon imaging will help to decipher fundamental operational principles of neural microcircuits.

  8. Whole brain imaging with Serial Two-Photon Tomography

    Directory of Open Access Journals (Sweden)

    Stephen P Amato

    2016-03-01

    Full Text Available Imaging entire mouse brains at submicron resolution has historically been a challenging undertaking and largely confined to the province of dedicated atlasing initiatives. The has limited systematic investigations into important areas of neuroscience, such as neural circuits, brain mapping and neurodegeneration. In this paper, we describe in detail Serial Two-Photon (STP tomography, a robust, reliable method for imaging entire brains with histological detail. We provide examples of how the basic methodology can be extended to other imaging modalities, such as optical coherence tomography, in order to provide unique contrast mechanisms. Furthermore we provide a survey of the research that STP tomography has enabled in the field of neuroscience, provide examples of how this technology enables quantitative whole brain studies, and discuss the current limitations of STP tomography-based approaches

  9. High contrast two-photon imaging of fingermarks

    Science.gov (United States)

    Stoltzfus, Caleb R.; Rebane, Aleksander

    2016-04-01

    Optically-acquired fingermarks are widely used as evidence across law enforcement agencies as well as in the courts of law. A common technique for visualizing latent fingermarks on nonporous surfaces consists of cyanoacrylate fuming of the fingerprint material, followed by impregnation with a fluorescent dye, which under ultra violet (UV) illumination makes the fingermarks visible and thus accessible for digital recording. However, there exist critical circumstances, when the image quality is compromised due to high background scattering, high auto-fluorescence of the substrate material, or other detrimental photo-physical and photo-chemical effects such as light-induced damage to the sample. Here we present a novel near-infrared (NIR), two-photon induced fluorescence imaging modality, which significantly enhances the quality of the fingermark images, especially when obtained from highly reflective and/or scattering surfaces, while at the same time reducing photo-damage to sensitive forensic samples.

  10. In vivo spectral imaging of different cell types in the small intestine by two-photon excited autofluorescence

    Science.gov (United States)

    Orzekowsky-Schroeder, Regina; Klinger, Antje; Martensen, Björn; Blessenohl, Maike; Gebert, Andreas; Vogel, Alfred; Hüttmann, Gereon

    2011-11-01

    Spectrally resolved two-photon excited autofluorescence imaging is used to distinguish different cell types and functional areas during dynamic processes in the living gut. Excitation and emission spectra of mucosal tissue and tissue components are correlated to spectra of endogenous chromophores. We show that selective excitation with only two different wavelengths within the tuning range of a Ti:sapphire femtosecond laser system yields excellent discrimination between enterocytes, antigen presenting cells and lysosomes based on the excitation and emission properties of their autofluorescence. The method is employed for time-lapse microscopy over up to 8 h. Changes of the spectral signature with the onset of photodamage are demonstrated, and their origin is discussed.

  11. Time-lapse and UAV Thermal Imaging of Glacial and Periglacial Environments in the Peruvian Andes (Cordillera Blanca, Peru)

    Science.gov (United States)

    McKenzie, J. M.; Wigmore, O.; Aubry-Wake, C.; Mark, B. G.; Hellstrom, R. A.; Lautz, L.

    2015-12-01

    In the tropics, the acquisition of high-resolution geospatial data of high-mountain glacial and periglacial systems presents unique challenges due to remote site access and very high elevations. For glaciers and hydrologic systems, a key variable of interest is surface temperature as it constrains glacier melt rates, traces hydrologic processes, and is needed for the calibration of energy budget models. We present results from two studies that acquired high resolution temperature data from the Cuchillacocha Glacier, Peru (9.24°S, 77.21°W). The glacier resides on the western drainage of the Cordillera Blanca with an elevation range of 4700 to 6096 m. In the first study we use high resolution time-lapse infrared imagery (5-10 minute interval over 3 days; 0.6 m2 pixel size) to observe diel changes in the surface energy budget of the glacier and to demonstrate how radiation from bare rock adjacent to the glacier may affect melt rates. In the second study we use a newly developed, inexpensive unmanned aerial vehicle (UAV) for high resolution multispectral mapping of the glacier (2 cm resolution orthomosaic and 5 cm resolution DEM). We present results showing how the time-lapse and the high-resolution UAV imagery can be combined to further strengthen our understanding of the Cuchillacocha Glacier's energy budget and possible insights about turbulent heat fluxes. While the new instruments provide unprecedented data acquisition capabilities, there is an outstanding need for proper data correction. Spatial/thermal control points and post-processing algorithms are needed to produce quantifiable datasets. For example, our post-processed time-lapse imagery has an r2 > 0.9 after emissivity, transmissivity and offset corrections.

  12. Two-Photon Ghost Image and Interference-Diffraction

    Science.gov (United States)

    Shih, Y. H.; Sergienko, A. V.; Pittman, T. B.; Strekalov, D. V.; Klyshko, D. N.

    1996-01-01

    One of the most surprising consequences of quantum mechanics is entanglement of two or more distance particles. The two-particle entangled state was mathematically formulated by Schrodinger. Based on this unusual quantum behavior, EPR defined their 'physical reality' and then asked the question: 'Can Quantum-Mechanical Description of Physical Reality Be Considered Complete?' One may not appreciate EPR's criterion of physical reality and insist that 'no elementary quantum phenomenon is a phenomenon until it is a recorded phenomenon'. Optical spontaneous parametric down conversion (SPDC) is the most effective mechanism to generate an EPR type entangled two-photon state. In SPDC, an optical beam, called the pump, is incident on a birefringent crystal. The pump is intense enough so that nonlinear effects lead to the conversion of pump photons into pairs of photons, historically called signal and idler. Technically, the SPDC is said to be type-1 or type-2, depending on whether the signal and idler beams have parallel or orthogonal polarization. The SPDC conversion efficiency is typically on the order of 10(exp -9) to 10(exp -11), depending on the SPDC nonlinear material. The signal and idler intensities are extremely low, only single photon detection devices can register them. The quantum entanglement nature of SPDC has been demonstrated in EPR-Bohm experiments and Bell's inequality measurements. The following two experiments were recently performed in our laboratory, which are more closely related to the original 1935 EPR gedankenezperiment. The first experiment is a two-photon optical imaging type experiment, which has been named 'ghost image' by the physics community. The signal and idler beams of SPDC are sent in different directions, so that the detection of the signal and idler photons can be performed by two distant photon counting detectors. An aperture object (mask) is placed in front of the signal photon detector and illuminated by the signal beam through a

  13. Heterogeneous cell-cycle behavior in response to UVB irradiation by a population of single cancer cells visualized by time-lapse FUCCI imaging.

    Science.gov (United States)

    Miwa, Shinji; Yano, Shuya; Kimura, Hiroaki; Yamamoto, Mako; Toneri, Makoto; Murakami, Takashi; Hayashi, Katsuhiro; Yamamoto, Norio; Fujiwara, Toshiyoshi; Tsuchiya, Hiroyuki; Hoffman, Robert M

    2015-01-01

    The present study analyzed the heterogeneous cell-cycle dependence and fate of single cancer cells in a population treated with UVB using a fluorescence ubiquitination-based cell-cycle (FUCCI) imaging system. HeLa cells expressing FUCCI were irradiated by 100 or 200 J/m(2) UVB. Modulation of the cell-cycle and apoptosis were observed by time-lapse confocal microscopy imaging every 30 min for 72 h. Correlation between cell survival and factors including cell-cycle phase at the time of the irradiation of UVB, mitosis and the G1/S transition were analyzed using the Kaplan-Meier method along with the log rank test. Time-lapse FUCCI imaging of HeLa cells demonstrated that UVB irradiation induced cell-cycle arrest in S/G2/M phase in the majority of the cells. The cells irradiated by 100 or 200 J/m(2) UVB during G0/G1 phase had a higher survival rate than the cells irradiated during S/G2/M phase. A minority of cells could escape S/G2/M arrest and undergo mitosis which significantly correlated with decreased survival of the cells. In contrast, G1/S transition significantly correlated with increased survival of the cells after UVB irradiation. UVB at 200 J/m(2) resulted in a greater number of apoptotic cells.

  14. Study of time-lapse processing for dynamic hydrologic conditions. [electronic satellite image analysis console for Earth Resources Technology Satellites imagery

    Science.gov (United States)

    Serebreny, S. M.; Evans, W. E.; Wiegman, E. J.

    1974-01-01

    The usefulness of dynamic display techniques in exploiting the repetitive nature of ERTS imagery was investigated. A specially designed Electronic Satellite Image Analysis Console (ESIAC) was developed and employed to process data for seven ERTS principal investigators studying dynamic hydrological conditions for diverse applications. These applications include measurement of snowfield extent and sediment plumes from estuary discharge, Playa Lake inventory, and monitoring of phreatophyte and other vegetation changes. The ESIAC provides facilities for storing registered image sequences in a magnetic video disc memory for subsequent recall, enhancement, and animated display in monochrome or color. The most unique feature of the system is the capability to time lapse the imagery and analytic displays of the imagery. Data products included quantitative measurements of distances and areas, binary thematic maps based on monospectral or multispectral decisions, radiance profiles, and movie loops. Applications of animation for uses other than creating time-lapse sequences are identified. Input to the ESIAC can be either digital or via photographic transparencies.

  15. A Sparse Bayesian Imaging Technique for Efficient Recovery of Reservoir Channels With Time-Lapse Seismic Measurements

    KAUST Repository

    Sana, Furrukh

    2016-06-01

    Subsurface reservoir flow channels are characterized by high-permeability values and serve as preferred pathways for fluid propagation. Accurate estimation of their geophysical structures is thus of great importance for the oil industry. The ensemble Kalman filter (EnKF) is a widely used statistical technique for estimating subsurface reservoir model parameters. However, accurate reconstruction of the subsurface geological features with the EnKF is challenging because of the limited measurements available from the wells and the smoothing effects imposed by the \\\\ell _{2} -norm nature of its update step. A new EnKF scheme based on sparse domain representation was introduced by Sana et al. (2015) to incorporate useful prior structural information in the estimation process for efficient recovery of subsurface channels. In this paper, we extend this work in two ways: 1) investigate the effects of incorporating time-lapse seismic data on the channel reconstruction; and 2) explore a Bayesian sparse reconstruction algorithm with the potential ability to reduce the computational requirements. Numerical results suggest that the performance of the new sparse Bayesian based EnKF scheme is enhanced with the availability of seismic measurements, leading to further improvement in the recovery of flow channels structures. The sparse Bayesian approach further provides a computationally efficient framework for enforcing a sparse solution, especially with the possibility of using high sparsity rates through the inclusion of seismic data.

  16. A-3 Construction Time Lapse

    Science.gov (United States)

    2009-01-01

    A time lapse from start to finish of steel erection for the 235-foot tall A-3 Test Stand. Ground work for the stand was broken in August 2008 and the final structural steel beam was placed April 9, 2009.

  17. Automatic detection and analysis of cell motility in phase-contrast time-lapse images using a combination of maximally stable extremal regions and Kalman filter approaches.

    Science.gov (United States)

    Kaakinen, M; Huttunen, S; Paavolainen, L; Marjomäki, V; Heikkilä, J; Eklund, L

    2014-01-01

    Phase-contrast illumination is simple and most commonly used microscopic method to observe nonstained living cells. Automatic cell segmentation and motion analysis provide tools to analyze single cell motility in large cell populations. However, the challenge is to find a sophisticated method that is sufficiently accurate to generate reliable results, robust to function under the wide range of illumination conditions encountered in phase-contrast microscopy, and also computationally light for efficient analysis of large number of cells and image frames. To develop better automatic tools for analysis of low magnification phase-contrast images in time-lapse cell migration movies, we investigated the performance of cell segmentation method that is based on the intrinsic properties of maximally stable extremal regions (MSER). MSER was found to be reliable and effective in a wide range of experimental conditions. When compared to the commonly used segmentation approaches, MSER required negligible preoptimization steps thus dramatically reducing the computation time. To analyze cell migration characteristics in time-lapse movies, the MSER-based automatic cell detection was accompanied by a Kalman filter multiobject tracker that efficiently tracked individual cells even in confluent cell populations. This allowed quantitative cell motion analysis resulting in accurate measurements of the migration magnitude and direction of individual cells, as well as characteristics of collective migration of cell groups. Our results demonstrate that MSER accompanied by temporal data association is a powerful tool for accurate and reliable analysis of the dynamic behaviour of cells in phase-contrast image sequences. These techniques tolerate varying and nonoptimal imaging conditions and due to their relatively light computational requirements they should help to resolve problems in computationally demanding and often time-consuming large-scale dynamical analysis of cultured cells.

  18. Time-lapse imaging as a tool to investigate contractility of the epididymal duct--effects of cGMP signaling.

    Directory of Open Access Journals (Sweden)

    Andrea Mietens

    Full Text Available The well orchestrated function of epididymal smooth muscle cells ensures transit of spermatozoa through the epididymal duct during which spermatozoa acquire motility and fertilizing capacity. Relaxation of smooth muscle cells is mediated by cGMP signaling and components of this pathway are found within the male reproductive tract. Whereas contractile function of caudal parts of the rat epididymal duct can be examined in organ bath studies, caput and corpus regions are fragile and make it difficult to mount them in an organ bath. We developed an ex vivo time-lapse imaging-based approach to investigate the contractile pattern in these parts of the epididymal duct. Collagen-embedding allowed immobilization without impeding contractility or diffusion of drugs towards the duct and therefore facilitated subsequent movie analyses. The contractile pattern was made visible by placing virtual sections through the acquired image stack to track wall movements over time. By this, simultaneous evaluation of contractile activity at different positions of the observed duct segment was possible. With each contraction translating into a spike, drug-induced alterations in contraction frequency could be assessed easily. Peristaltic contractions were also detectable and throughout all regions in the proximal epididymis we found regular spontaneous contractile activity that elicited movement of intraluminal contents. Stimulating cGMP production by natriuretic peptide ANP or inhibiting degradation of cGMP by the phosphodiesterase 5 inhibitor sildenafil significantly reduced contractile frequency in isolated duct segments from caput and corpus. RT-PCR analysis after laser-capture microdissection localized the corresponding molecules to the smooth muscle layer of the duct. Our time-lapse imaging approach proved to be feasible to assess contractile function in all regions of the epididymal duct under near physiological conditions and provides a tool to evaluate acute

  19. Time lapse imaging analysis of the effect of ER stress modulators on apoptotic cell assessed by caspase3/7 activation in NG108-15 cells

    Directory of Open Access Journals (Sweden)

    Ayako Saito

    2016-03-01

    Full Text Available This paper reports the data from the long term time lapse imaging of neuronal cell line NG108-15 that were treated with apoptosis inducer or various ER stress inducers. Use of the fluorescent reporter for activated caspase3/7 in combination with the conventional light microscope allowed us to investigate the time course of apoptosis induction at the single cell level. Quantitative as well as qualitative data are presented here to show the effect of two different ER stress modulating chemical compounds on caspase3/7-dependent apoptosis in neuronal cell line NG108-15 cells. Additional results and interpretation of our data concerning ER stress and apoptosis in NG108-15 cells can be found in Suga et al. (2015 [1] and in Suga et al. (2015 [2].

  20. Two-photon fluorescence probes for imaging of mitochondria and lysosomes.

    Science.gov (United States)

    Yang, Wanggui; Chan, Pui Shan; Chan, Miu Shan; Li, King Fai; Lo, Pik Kwan; Mak, Nai Ki; Cheah, Kok Wai; Wong, Man Shing

    2013-04-28

    Novel biocompatible cyanines show not only a very large two-photon cross-section of up to 5130 GM at 910 nm in aqueous medium for high-contrast and -brightness two-photon fluorescence live cell imaging but also highly selective subcellular localization properties including localization of mitochondria and lysosomes.

  1. Imaging theory and resolution improvement of two-photon confocal microscopy

    Institute of Scientific and Technical Information of China (English)

    唐志列; 杨初平; 裴红津; 梁瑞生; 刘颂豪

    2002-01-01

    The nonlinear effect of two-photon excitation on the imaging property of two-photonconfocal microscopy has been analyzed by the two-photon fluorescence intensity transfer functionderived in this paper. The two-photon fluorescence intensity transfer function in a confocal micros-copy is given. Furthermore the three-dimensional point spread function (3D-PSF) and thethree-dimensional optical transfer function (3D-OTF) of two-photon confocal microscopy are de-rived based on the nonlinear effect of two-photon excitation. The imaging property of two-photonconfocal microscopy is discussed in detail based on 3D-OTF. Finally the spatial resolution limit oftwo-photon confocal microscopy is discussed according to the uncertainty principle.

  2. New algorithm to determine true colocalization in combination with image restoration and time-lapse confocal microscopy to MAP kinases in mitochondria.

    Directory of Open Access Journals (Sweden)

    Jorge Ignacio Villalta

    Full Text Available The subcellular localization and physiological functions of biomolecules are closely related and thus it is crucial to precisely determine the distribution of different molecules inside the intracellular structures. This is frequently accomplished by fluorescence microscopy with well-characterized markers and posterior evaluation of the signal colocalization. Rigorous study of colocalization requires statistical analysis of the data, albeit yet no single technique has been established as a standard method. Indeed, the few methods currently available are only accurate in images with particular characteristics. Here, we introduce a new algorithm to automatically obtain the true colocalization between images that is suitable for a wide variety of biological situations. To proceed, the algorithm contemplates the individual contribution of each pixel's fluorescence intensity in a pair of images to the overall Pearsońs correlation and Manders' overlap coefficients. The accuracy and reliability of the algorithm was validated on both simulated and real images that reflected the characteristics of a range of biological samples. We used this algorithm in combination with image restoration by deconvolution and time-lapse confocal microscopy to address the localization of MEK1 in the mitochondria of different cell lines. Appraising the previously described behavior of Akt1 corroborated the reliability of the combined use of these techniques. Together, the present work provides a novel statistical approach to accurately and reliably determine the colocalization in a variety of biological images.

  3. Enabling user-guided segmentation and tracking of surface-labeled cells in time-lapse image sets of living tissues.

    Science.gov (United States)

    Mashburn, David N; Lynch, Holley E; Ma, Xiaoyan; Hutson, M Shane

    2012-05-01

    To study the process of morphogenesis, one often needs to collect and segment time-lapse images of living tissues to accurately track changing cellular morphology. This task typically involves segmenting and tracking tens to hundreds of individual cells over hundreds of image frames, a scale that would certainly benefit from automated routines; however, any automated routine would need to reliably handle a large number of sporadic, and yet typical problems (e.g., illumination inconsistency, photobleaching, rapid cell motions, and drift of focus or of cells moving through the imaging plane). Here, we present a segmentation and cell tracking approach based on the premise that users know their data best-interpreting and using image features that are not accounted for in any a priori algorithm design. We have developed a program, SeedWater Segmenter, that combines a parameter-less and fast automated watershed algorithm with a suite of manual intervention tools that enables users with little to no specialized knowledge of image processing to efficiently segment images with near-perfect accuracy based on simple user interactions.

  4. A semi-Markov model for mitosis segmentation in time-lapse phase contrast microscopy image sequences of stem cell populations.

    Science.gov (United States)

    Liu, An-An; Li, Kang; Kanade, Takeo

    2012-02-01

    We propose a semi-Markov model trained in a max-margin learning framework for mitosis event segmentation in large-scale time-lapse phase contrast microscopy image sequences of stem cell populations. Our method consists of three steps. First, we apply a constrained optimization based microscopy image segmentation method that exploits phase contrast optics to extract candidate subsequences in the input image sequence that contains mitosis events. Then, we apply a max-margin hidden conditional random field (MM-HCRF) classifier learned from human-annotated mitotic and nonmitotic sequences to classify each candidate subsequence as a mitosis or not. Finally, a max-margin semi-Markov model (MM-SMM) trained on manually-segmented mitotic sequences is utilized to reinforce the mitosis classification results, and to further segment each mitosis into four predefined temporal stages. The proposed method outperforms the event-detection CRF model recently reported by Huh as well as several other competing methods in very challenging image sequences of multipolar-shaped C3H10T1/2 mesenchymal stem cells. For mitosis detection, an overall precision of 95.8% and a recall of 88.1% were achieved. For mitosis segmentation, the mean and standard deviation for the localization errors of the start and end points of all mitosis stages were well below 1 and 2 frames, respectively. In particular, an overall temporal location error of 0.73 ± 1.29 frames was achieved for locating daughter cell birth events.

  5. Fast two-photon neuronal imaging and control using a spatial light modulator and ruthenium compounds

    Science.gov (United States)

    Peterka, Darcy S.; Nikolenko, Volodymyr; Fino, Elodie; Araya, Roberto; Etchenique, Roberto; Yuste, Rafael

    2010-02-01

    We have developed a spatial light modulator (SLM) based microscope that uses diffraction to shape the incoming two-photon laser source to any arbitrary light pattern. This allows the simultaneous imaging or photostimulation of different regions of a sample with three-dimensional precision at high frame rates. Additionally, we have combined this microscope with a new class of two photon active neuromodulators with Ruthenium BiPyridine (RuBi) based cages that offer great flexibility for neuronal control.

  6. NeuronGrowth, a software for automatic quantification of neurite and filopodial dynamics from time-lapse sequences of digital images.

    Science.gov (United States)

    Fanti, Zian; Martinez-Perez, M Elena; De-Miguel, Francisco F

    2011-10-01

    We developed NeuronGrowth, a software for the automatic quantification of extension and retraction of neurites and filopodia, from time-lapse sequences of two-dimensional digital micrographs. NeuronGrowth requires a semiautomatic characterization of individual neurites in a reference frame, which is then used for automatic tracking and measurement of every neurite over the whole image sequence. Modules for sequence alignment, background subtraction, flat field correction, light normalization, and cropping have been integrated to improve the quality of the analysis. Moreover, NeuronGrowth incorporates a deconvolution filter that corrects the shadow-cast effect of differential interference contrast (DIC) images. NeuronGrowth was tested by analyzing the formation of outgrowth patterns by individual leech neurons cultured under two different conditions. Phase contrast images were obtained from neurons plated on CNS homogenates and DIC images were obtained from similar neurons plated on ganglion capsules as substrates. Filopodia were measured from fluorescent growth-cones of chick dorsal root ganglion cells. Quantitative data of neurite extension and retraction obtained by three different users applying NeuronGrowth and two other manually operated software packages were similar. However, NeuronGrowth required less user participation and had a better time performance when compared with the other software packages. NeuronGrowth may be used in general to quantify the dynamics of tubular structures such as blood vessels. NeuronGrowth is a free plug-in for the free software ImageJ and can be downloaded along with a user manual, a troubleshooting section and other information required for its use from http://www.ifc.unam.mx or http://www.ifc.unam.mx/ffm/index.html.

  7. Time-lapse imaging reveals symmetric neurogenic cell division of GFAP-expressing progenitors for expansion of postnatal dentate granule neurons.

    Directory of Open Access Journals (Sweden)

    Takashi Namba

    Full Text Available Granule cells in the hippocampus, a region critical for memory and learning, are generated mainly during the early postnatal period but neurogenesis continues in adulthood. Postnatal neuronal production is carried out by primary progenitors that express glial fibrillary acidic protein (GFAP and they are assumed to function as stem cells. A central question regarding postnatal dentate neurogenesis is how astrocyte-like progenitors produce neurons. To reveal cell division patterns and the process of neuronal differentiation of astrocyte-like neural progenitors, we performed time-lapse imaging in cultured hippocampal slices from early postnatal transgenic mice with mouse GFAP promoter-controlled enhanced green fluorescent protein (mGFAP-eGFP Tg mice in combination with a retrovirus carrying a red fluorescent protein gene. Our results showed that the majority of GFAP-eGFP+ progenitor cells that express GFAP, Sox2 and nestin divided symmetrically to produce pairs of GFAP+ cells (45% or pairs of neuron-committed cells (45%, whereas a minority divided asymmetrically to generate GFAP+ cells and neuron-committed cells (10%. The present results suggest that a substantial number of GFAP-expressing progenitors functions as transient amplifying progenitors, at least in an early postnatal dentate gyrus, although a small population appears to be stem cell-like progenitors. From the present data, we discuss possible cell division patterns of adult GFAP+ progenitors.

  8. Time-lapse imaging reveals symmetric neurogenic cell division of GFAP-expressing progenitors for expansion of postnatal dentate granule neurons.

    Science.gov (United States)

    Namba, Takashi; Mochizuki, Hideki; Suzuki, Ryusuke; Onodera, Masafumi; Yamaguchi, Masahiro; Namiki, Hideo; Shioda, Seiji; Seki, Tatsunori

    2011-01-01

    Granule cells in the hippocampus, a region critical for memory and learning, are generated mainly during the early postnatal period but neurogenesis continues in adulthood. Postnatal neuronal production is carried out by primary progenitors that express glial fibrillary acidic protein (GFAP) and they are assumed to function as stem cells. A central question regarding postnatal dentate neurogenesis is how astrocyte-like progenitors produce neurons. To reveal cell division patterns and the process of neuronal differentiation of astrocyte-like neural progenitors, we performed time-lapse imaging in cultured hippocampal slices from early postnatal transgenic mice with mouse GFAP promoter-controlled enhanced green fluorescent protein (mGFAP-eGFP Tg mice) in combination with a retrovirus carrying a red fluorescent protein gene. Our results showed that the majority of GFAP-eGFP+ progenitor cells that express GFAP, Sox2 and nestin divided symmetrically to produce pairs of GFAP+ cells (45%) or pairs of neuron-committed cells (45%), whereas a minority divided asymmetrically to generate GFAP+ cells and neuron-committed cells (10%). The present results suggest that a substantial number of GFAP-expressing progenitors functions as transient amplifying progenitors, at least in an early postnatal dentate gyrus, although a small population appears to be stem cell-like progenitors. From the present data, we discuss possible cell division patterns of adult GFAP+ progenitors.

  9. Simultaneous two-photon imaging and photo-stimulation with structured light illumination.

    Science.gov (United States)

    Dal Maschio, Marco; Difato, Francesco; Beltramo, Riccardo; Blau, Axel; Benfenati, Fabio; Fellin, Tommaso

    2010-08-30

    Holographic microscopy is increasingly recognized as a promising tool for the study of the central nervous system. Here we present a "holographic module", a simple optical path that can be combined with commercial scanheads for simultaneous imaging and uncaging with structured two-photon light. The present microscope is coupled to two independently tunable lasers and has two principal configurations: holographic imaging combined with galvo-steered uncaging and holographic uncaging combined with conventional scanning imaging. We applied this flexible system for simultaneous two-photon imaging and photostimulation of neuronal cells with complex light patterns, opening new perspectives for the study of brain function in situ and in vivo.

  10. Time-lapse electrical resistivity imaging of the thermally affected zone of a Borehole Thermal Energy Storage system near Torino (Northern Italy)

    Science.gov (United States)

    Giordano, N.; Arato, A.; Comina, C.; Mandrone, G.

    2017-05-01

    A Borehole Thermal Energy Storage living lab was built up nearby Torino (Northern Italy). This living lab aims at testing the ability of the alluvial deposits of the north-western Po Plain to store the thermal energy collected by solar thermal panels and the efficiency of energy storage systems in this climatic context. Different monitoring approaches have been tested and analyzed since the start of the thermal injection in April 2014. Underground temperature monitoring is constantly undertaken by means of several temperature sensors located along the borehole heat exchangers and within the hydraulic circuit. Nevertheless, this can provide only pointwise information about underground temperature distribution. For this reason, a geophysical approach is proposed in order to image the thermally affected zone (TAZ) caused by the heat injection: surface electrical resistivity measurements were carried out with this purpose. In the present paper, results of time-lapse acquisitions during a heating day are reported with the aim of imaging the thermal plume evolution within the subsoil. Resistivity data, calibrated on local temperature measurements, have shown their potentiality in imaging the heated plume of the system and depicting its evolution throughout the day. Different types of data processing were adopted in order to face issues mainly related to a highly urbanized environment. The use of apparent resistivity proved to be in valid agreement with the results of different inversion approaches. The inversion processes did not significantly improve the qualitative and quantitative TAZ imaging in comparison to the pseudo-sections. This suggested the usefulness of apparent resistivity data alone for a rough monitoring of TAZ in this kind of applications.

  11. Time-lapse fluorescence imaging of Arabidopsis root growth with rapid manipulation of the root environment using the RootChip.

    Science.gov (United States)

    Grossmann, Guido; Meier, Matthias; Cartwright, Heather N; Sosso, Davide; Quake, Stephen R; Ehrhardt, David W; Frommer, Wolf B

    2012-07-07

    The root functions as the physical anchor of the plant and is the organ responsible for uptake of water and mineral nutrients such as nitrogen, phosphorus, sulfate and trace elements that plants acquire from the soil. If we want to develop sustainable approaches to producing high crop yield, we need to better understand how the root develops, takes up a wide spectrum of nutrients, and interacts with symbiotic and pathogenic organisms. To accomplish these goals, we need to be able to explore roots in microscopic detail over time periods ranging from minutes to days. We developed the RootChip, a polydimethylsiloxane (PDMS)- based microfluidic device, which allows us to grow and image roots from Arabidopsis seedlings while avoiding any physical stress to roots during preparation for imaging(1) (Figure 1). The device contains a bifurcated channel structure featuring micromechanical valves to guide the fluid flow from solution inlets to each of the eight observation chambers(2). This perfusion system allows the root microenvironment to be controlled and modified with precision and speed. The volume of the chambers is approximately 400 nl, thus requiring only minimal amounts of test solution. Here we provide a detailed protocol for studying root biology on the RootChip using imaging-based approaches with real time resolution. Roots can be analyzed over several days using time lapse microscopy. Roots can be perfused with nutrient solutions or inhibitors, and up to eight seedlings can be analyzed in parallel. This system has the potential for a wide range of applications, including analysis of root growth in the presence or absence of chemicals, fluorescence-based analysis of gene expression, and the analysis of biosensors, e.g. FRET nanosensors(3).

  12. In vivo two-photon calcium imaging in the visual system.

    Science.gov (United States)

    Ohki, Kenichi; Reid, R Clay

    2014-04-01

    Two-photon imaging of calcium-sensitive dyes in vivo has become a common tool used by neuroscientists, largely because of the development of bolus loading techniques, which can label every neuron in a local circuit with calcium-sensitive dye. Like multielectrode recordings, two-photon imaging paired with bolus loading provides a method for monitoring many neurons at once, but, in addition, it provides a means for determining the precise location of every neuron. Thus, it is an ideal method for studying the fine-scale functional architecture of the cortex and guiding the experimenter to individual neurons that can be targeted for further anatomical study. Two-photon calcium imaging enables study of the fine structure of functional maps in the visual cortex in cats and rodents. In mice, it can allow the characterization of specific cell types when paired with transgenic or retrograde labeling.

  13. Two-photon calcium imaging in mice navigating a virtual reality environment.

    Science.gov (United States)

    Leinweber, Marcus; Zmarz, Pawel; Buchmann, Peter; Argast, Paul; Hübener, Mark; Bonhoeffer, Tobias; Keller, Georg B

    2014-02-20

    In recent years, two-photon imaging has become an invaluable tool in neuroscience, as it allows for chronic measurement of the activity of genetically identified cells during behavior(1-6). Here we describe methods to perform two-photon imaging in mouse cortex while the animal navigates a virtual reality environment. We focus on the aspects of the experimental procedures that are key to imaging in a behaving animal in a brightly lit virtual environment. The key problems that arise in this experimental setup that we here address are: minimizing brain motion related artifacts, minimizing light leak from the virtual reality projection system, and minimizing laser induced tissue damage. We also provide sample software to control the virtual reality environment and to do pupil tracking. With these procedures and resources it should be possible to convert a conventional two-photon microscope for use in behaving mice.

  14. Enhanced two-photon fluorescence imaging and therapy of cancer cells via Gold@bridged silsesquioxane nanoparticles.

    Science.gov (United States)

    Croissant, Jonas; Maynadier, Marie; Mongin, Olivier; Hugues, Vincent; Blanchard-Desce, Mireille; Chaix, Arnaud; Cattoën, Xavier; Wong Chi Man, Michel; Gallud, Audrey; Gary-Bobo, Magali; Garcia, Marcel; Raehm, Laurence; Durand, Jean-Olivier

    2015-01-21

    A two-photon photosensitizer with four triethoxysilyl groups is synthesized through the click reaction. This photosensitizer allows the design of bridged silsesquioxane (BS) nanoparticles through a sol-gel process; moreover, gold core BS shells or BS nanoparticles decorated with gold nanospheres are synthesized. An enhancement of the two-photon properties is noted with gold and the nanoparticles are efficient for two-photon imaging and two-photon photodynamic therapy of cancer cells.

  15. Time-lapse photogrammetry in geomorphic studies

    Science.gov (United States)

    Eltner, Anette; Kaiser, Andreas

    2017-04-01

    Image based approaches to reconstruct the earth surface (Structure from Motion - SfM) are establishing as a standard technology for high resolution topographic data. This is amongst other advantages due to the comparatively ease of use and flexibility of data generation. Furthermore, the increased spatial resolution led to its implementation at a vast range of applications from sub-mm to tens-of-km scale. Almost fully automatic calculation of referenced digital elevation models allows for a significant increase of temporal resolution, as well, potentially up to sub-second scales. Thereby, the setup of a time-lapse multi-camera system is necessary and different aspects need to be considered: The camera array has to be temporary stable or potential movements need to be compensated by temporary stable reference targets/areas. The stability of the internal camera geometry has to be considered due to a usually significantly lower amount of images of the scene, and thus redundancy for parameter estimation, compared to more common SfM applications. Depending on the speed of surface change, synchronisation has to be very accurate. Due to the usual application in the field, changing environmental conditions important for lighting and visual range are also crucial factors to keep in mind. Besides these important considerations much potential is comprised by time-lapse photogrammetry. The integration of multi-sensor systems, e.g. using thermal cameras, enables the potential detection of other processes not visible with RGB-images solely. Furthermore, the implementation of low-cost sensors allows for a significant increase of areal coverage and their setup at locations, where a loss of the system cannot be ruled out. The usage of micro-computers offers smart camera triggering, e.g. acquiring images with increased frequency controlled by a rainfall-triggered sensor. In addition these micro-computers can enable on-site data processing, e.g. recognition of increased surface

  16. Ionomycin-induced calcium influx induces neurite degeneration in mouse neuroblastoma cells: analysis of a time-lapse live cell imaging system.

    Science.gov (United States)

    Nakamura, Saki; Nakanishi, Ayumi; Takazawa, Minami; Okihiro, Shunsuke; Urano, Shiro; Fukui, Koji

    2016-01-01

    Reactive oxygen species induce neuronal cell death. However, the detailed mechanisms of cell death have not yet been elucidated. Previously, we reported neurite degeneration before the induction of cell death. Here, we attempted to elucidate the mechanisms of neurite degeneration before the induction of cell death using the neuroblastoma N1E-115 cell line and a time-lapse live cell imaging system. Treatment with the calcium ionophore ionomycin induced cell death and neurite degeneration in a concentration- and time-dependent manner. Treatment with a low concentration of ionomycin immediately produced a significant calcium influx into the intracellular region in N1E-115 cells. After 1-h incubation with ionomycin, the fluorescence emission of MitoSOX(TM) increased significantly compared to the control. Finally, analysis using a new mitochondrial specific fluorescence dye, MitoPeDPP, indicated that treatment with ionomycin significantly increased the mitochondrial lipid hydroperoxide production in N1E-115 cells. The fluorescence emissions of Fluo-4 AM and MitoPeDPP were detected in the cell soma and neurite regions in ionomycin-treated N1E-115 cells. However, the emissions of neurites were much lower than those of the cell soma. TBARS values of ionomycin-treated cells significantly increased compared to the control. These results indicate that ionomycin induces calcium influx into the intracellular region and reactive oxygen species production in N1E-115 cells. Lipid hydroperoxide production was induced in ionomycin-treated N1E-115 cells. Calcium influx into the intracellular region is a possible activator of neurite degeneration.

  17. [Intensity loss of two-photon excitation fluorescence microscopy images of mouse oocyte chromosomes].

    Science.gov (United States)

    Zhao, Feng-Ying; Wu, Hong-Xin; Chen, Die-Yan; Ma, Wan-Yun

    2014-07-01

    As an optical microscope with high resolution, two-photon excitation (TPE) fluorescence microscope is widely used in noninvasive 3D optical imaging of biological samples. Compared with confocal laser scanning microscope, TPE fluorescence microscope provides a deeper detecting depth. In spite of that, the image quality of sample always declines as the detecting depth increases when a noninvasive 3D optical imaging of thicker samples is performed. Mouse oocytes with a large diameter, which play an important role in clinical and biological fields, have obvious absorption and scattering effects. In the present paper, we performed compensation for two-photon fluorescence images of mouse oocyte chromosomes. Using volume as a parameter, the attenuation degree of these chromosomes was also studied. The result of our data suggested that there exists a severe axial intensity loss in two-photon microscopic images of mouse oocytes due to the absorption and scattering effects. It is necessary to make compensation for these images of mouse oocyte chromosomes obtained from two-photon microscopic system. It will be specially needed in studying the quantitative three-dimensional information of mouse oocytes.

  18. Non-invasive imaging of skin cancer with fluorescence lifetime imaging using two photon tomography

    Science.gov (United States)

    Patalay, Rakesh; Talbot, Clifford; Alexandrov, Yuriy; Munro, Ian; Breunig, Hans Georg; König, Karsten; Warren, Sean; Neil, Mark A. A.; French, Paul M. W.; Chu, Anthony; Stamp, Gordon W.; Dunsby, Christopher

    2011-07-01

    Multispectral fluorescence lifetime imaging (FLIM) using two photon microscopy as a non-invasive technique for the diagnosis of skin lesions is described. Skin contains fluorophores including elastin, keratin, collagen, FAD and NADH. This endogenous contrast allows tissue to be imaged without the addition of exogenous agents and allows the in vivo state of cells and tissues to be studied. A modified DermaInspect® multiphoton tomography system was used to excite autofluorescence at 760 nm in vivo and on freshly excised ex vivo tissue. This instrument simultaneously acquires fluorescence lifetime images in four spectral channels between 360-655 nm using time-correlated single photon counting and can also provide hyperspectral images. The multispectral fluorescence lifetime images were spatially segmented and binned to determine lifetimes for each cell by fitting to a double exponential lifetime model. A comparative analysis between the cellular lifetimes from different diagnoses demonstrates significant diagnostic potential.

  19. Two-photon imaging and spectroscopy of fresh human colon biopsies

    Science.gov (United States)

    Cicchi, R.; Sturiale, A.; Nesi, G.; Tonelli, F.; Pavone, F. S.

    2012-03-01

    Two-photon fluorescence (TPEF) microscopy is a powerful tool to image human tissues up to 200 microns depth without any exogenously added probe. TPEF can take advantage of the autofluorescence of molecules intrinsically contained in a biological tissue, as such NADH, elastin, collagen, and flavins. Two-photon microscopy has been already successfully used to image several types of tissues, including skin, muscles, tendons, bladder. Nevertheless, its usefulness in imaging colon tissue has not been deeply investigated yet. In this work we have used combined two-photon excited fluorescence (TPEF), second harmonic generation microscopy (SHG), fluorescence lifetime imaging microscopy (FLIM), and multispectral two-photon emission detection (MTPE) to investigate different kinds of human ex-vivo fresh biopsies of colon. Morphological and spectroscopic analyses allowed to characterize both healthy mucosa, polyp, and colon samples in a good agreement with common routine histology. Even if further analysis, as well as a more significant statistics on a large number of samples would be helpful to discriminate between low, mild, and high grade cancer, our method is a promising tool to be used as diagnostic confirmation of histological results, as well as a diagnostic tool in a multiphoton endoscope or colonoscope to be used in in-vivo imaging applications.

  20. Image registration and averaging of low laser power two-photon fluorescence images of mouse retina.

    Science.gov (United States)

    Alexander, Nathan S; Palczewska, Grazyna; Stremplewski, Patrycjusz; Wojtkowski, Maciej; Kern, Timothy S; Palczewski, Krzysztof

    2016-07-01

    Two-photon fluorescence microscopy (TPM) is now being used routinely to image live cells for extended periods deep within tissues, including the retina and other structures within the eye . However, very low laser power is a requirement to obtain TPM images of the retina safely. Unfortunately, a reduction in laser power also reduces the signal-to-noise ratio of collected images, making it difficult to visualize structural details. Here, image registration and averaging methods applied to TPM images of the eye in living animals (without the need for auxiliary hardware) demonstrate the structural information obtained with laser power down to 1 mW. Image registration provided between 1.4% and 13.0% improvement in image quality compared to averaging images without registrations when using a high-fluorescence template, and between 0.2% and 12.0% when employing the average of collected images as the template. Also, a diminishing return on image quality when more images were used to obtain the averaged image is shown. This work provides a foundation for obtaining informative TPM images with laser powers of 1 mW, compared to previous levels for imaging mice ranging between 6.3 mW [Palczewska G., Nat Med.20, 785 (2014) Sharma R., Biomed. Opt. Express4, 1285 (2013)].

  1. Visualizing heterogeneity of photosynthetic properties of plant leaves with two-photon fluorescence lifetime imaging microscopy

    NARCIS (Netherlands)

    Iermak, Ievgeniia; Vink, Jochem; Bader, Arjen N.; Wientjes, Emilie; Amerongen, van Herbert

    2016-01-01

    Two-photon fluorescence lifetime imaging microscopy (FLIM) was used to analyse the distribution and properties of Photosystem I (PSI) and Photosystem II (PSII) in palisade and spongy chloroplasts of leaves from the C3 plant Arabidopsis thaliana and the C4 plant Miscanthus x giganteus. This was ac

  2. Gold Core Mesoporous Organosilica Shell Degradable Nanoparticles for Two-Photon Imaging and Gemcitabine Monophosphate Delivery

    KAUST Repository

    Rhamani, Saher

    2017-09-12

    The synthesis of gold core degradable mesoporous organosilica shell nanoparticles is described. The nanopaticles were very efficient for two-photon luminescence imaging of cancer cells and for in vitro gemcitabine monophosphate delivery, allowing promising theranostic applications in the nanomedicine field.

  3. Applying two-photon excitation fluorescence lifetime imaging microscopy to study photosynthesis in plant leaves

    NARCIS (Netherlands)

    Broess, K.; Borst, J.W.; Amerongen, van H.

    2009-01-01

    This study investigates to which extent two-photon excitation (TPE) fluorescence lifetime imaging microscopy can be applied to study picosecond fluorescence kinetics of individual chloroplasts in leaves. Using femtosecond 860 nm excitation pulses, fluorescence lifetimes can be measured in leaves of

  4. Scanless functional imaging of hippocampal networks using patterned two-photon illumination through GRIN lenses

    KAUST Repository

    Moretti, Claudio

    2016-09-12

    Patterned illumination through the phase modulation of light is increasingly recognized as a powerful tool to investigate biological tissues in combination with two-photon excitation and light-sensitive molecules. However, to date two-photon patterned illumination has only been coupled to traditional microscope objectives, thus limiting the applicability of these methods to superficial biological structures. Here, we show that phase modulation can be used to efficiently project complex two-photon light patterns, including arrays of points and large shapes, in the focal plane of graded index (GRIN) lenses. Moreover, using this approach in combination with the genetically encoded calcium indicator GCaMP6, we validate our system performing scanless functional imaging in rodent hippocampal networks in vivo ~1.2 mm below the brain surface. Our results open the way to the application of patterned illumination approaches to deep regions of highly scattering biological tissues, such as the mammalian brain.

  5. Two-photon excited fluorescence microendoscopic imaging using a GRIN lens

    Science.gov (United States)

    Yan, Wei; Peng, Xiao; Lin, Danying; Wang, Qi; Gao, Jian; Zhou, Jie; Ye, Tong; Qu, Junle; Niu, Hanben

    2015-03-01

    With the rapid development of life sciences, there is an increasing demand for intravital fluorescence imaging of small animals. However, large dimensions and limited working distances of objective lenses in traditional fluorescence microscopes have limited the imaging applications mostly to superficial tissues. To overcome this disadvantage, researchers have developed the graded-index (GRIN) probes with small diameters for imaging internal organs of small animals in a minimally invasive fashion. Here, we present the development of a fluorescence endoscopic imaging system based on a GRIN lens using two-photon excitation. Experimental results showed that this system could perform dynamic fluorescence microendoscopic imaging and monitor the blood flow in anesthetized living mice using two-photon excitation.

  6. Mitigating thermal mechanical damage potential during two-photon dermal imaging.

    Science.gov (United States)

    Masters, Barry R; So, Peter T C; Buehler, Christof; Barry, Nicholas; Sutin, Jason D; Mantulin, William W; Gratton, Enrico

    2004-01-01

    Two-photon excitation fluorescence microscopy allows in vivo high-resolution imaging of human skin structure and biochemistry with a penetration depth over 100 microm. The major damage mechanism during two-photon skin imaging is associated with the formation of cavitation at the epidermal-dermal junction, which results in thermal mechanical damage of the tissue. In this report, we verify that this damage mechanism is of thermal origin and is associated with one-photon absorption of infrared excitation light by melanin granules present in the epidermal-dermal junction. The thermal mechanical damage threshold for selected Caucasian skin specimens from a skin bank as a function of laser pulse energy and repetition rate has been determined. The experimentally established thermal mechanical damage threshold is consistent with a simple heat diffusion model for skin under femtosecond pulse laser illumination. Minimizing thermal mechanical damage is vital for the potential use of two-photon imaging in noninvasive optical biopsy of human skin in vivo. We describe a technique to mitigate specimen thermal mechanical damage based on the use of a laser pulse picker that reduces the laser repetition rate by selecting a fraction of pulses from a laser pulse train. Since the laser pulse picker decreases laser average power while maintaining laser pulse peak power, thermal mechanical damage can be minimized while two-photon fluorescence excitation efficiency is maximized.

  7. Application of quantitative time-lapse imaging (QTLI) for evaluation of Mrp2-based drug–drug interaction induced by liver metabolites

    Energy Technology Data Exchange (ETDEWEB)

    Nakanishi, Takeo; Ikenaga, Miho; Fukuda, Hajime; Matsunaga, Norikazu; Tamai, Ikumi, E-mail: tamai@p.kanazawa-w.ac.jp

    2012-09-01

    We previously reported a quantitative time-lapse imaging (QTLI)-based analysis method to assess drug–drug interactions (DDI) at multidrug resistance-associated protein 2 (Mrp2) in rat sandwich-cultured hepatocyte (SCH) system, utilizing the fluorescent Mrp2 substrate, 5-(and 6)-carboxy-2′,7′-dichlorofluorescein (CDF). Here, we aimed to examine the feasibility of using QTLI to evaluate DDI involving drug metabolite(s) generated in hepatocytes. We used estradiol (E2) and bilirubin as model compounds; both are not substrates of MRP2, whereas their hepatic metabolites, estradiol-17β-glucuronide (E17G) or bilirubin glucuronides, are known to be its substrates as well as inhibitors. When rat SCHs were pre-exposed with E2, fluorescence of CDF accumulated in bile canaliculi decreased depending upon both the duration of pre-exposure and the concentration of extracellular E2. The decrease corresponded with the increase in intracellular concentration of E17G in hepatocytes. Furthermore, cytotoxicity of vinblastine, a substrate of MRP2, was enhanced in SCHs treated with E2. Similarly, CDF accumulated in bile canaliculi was significantly reduced in rat SCHs pre-exposed with bilirubin. In conclusion, these results suggest that phase II biotransformation of a competitor is reflected in alteration of MRP2-mediated CDF transport detected in QTLI. The QTLI might provide a convenient platform to evaluate transporter-based DDIs involving hepatic metabolites of drug candidates without the need to identify the metabolites. -- Highlights: ► Mrp2-mediated CDF transport is inhibited by E2, but not E17G in vesicle study. ► Both E2 and E17G do not compromise CDF formation from CDFDA in hepatocytes. ► CDF accumulation in bile canaliculi is inhibited by E2 or E17G in QTLI. ► Increasing exposure to E2 decreases CDF accumulation in bile canaliculi in QTLI. ► QTLI is feasible to assess Mrp2-based DDI involving drug metabolite in hepatocytes.

  8. Multiscale vision model for event detection and reconstruction in two-photon imaging data

    DEFF Research Database (Denmark)

    Brazhe, Alexey; Mathiesen, Claus; Lind, Barbara;

    2014-01-01

    on a modified multiscale vision model, an object detection framework based on the thresholding of wavelet coefficients and hierarchical trees of significant coefficients followed by nonlinear iterative partial object reconstruction, for the analysis of two-photon calcium imaging data. The framework is discussed...... of the multiscale vision model is similar in the denoising, but provides a better segmenation of the image into meaningful objects, whereas other methods need to be combined with dedicated thresholding and segmentation utilities....

  9. Acute two-photon imaging of the neurovascular unit in the cortex of active mice

    Science.gov (United States)

    Tran, Cam Ha T.; Gordon, Grant R.

    2015-01-01

    In vivo two-photon scanning fluorescence imaging is a powerful technique to observe physiological processes from the millimeter to the micron scale in the intact animal. In neuroscience research, a common approach is to install an acute cranial window and head bar to explore neocortical function under anesthesia before inflammation peaks from the surgery. However, there are few detailed acute protocols for head-restrained and fully awake animal imaging of the neurovascular unit during activity. This is because acutely performed awake experiments are typically untenable when the animal is naïve to the imaging apparatus. Here we detail a method that achieves acute, deep-tissue two-photon imaging of neocortical astrocytes and microvasculature in behaving mice. A week prior to experimentation, implantation of the head bar alone allows mice to train for head-immobilization on an easy-to-learn air-supported ball treadmill. Following just two brief familiarization sessions to the treadmill on separate days, an acute cranial window can subsequently be installed for immediate imaging. We demonstrate how running and whisking data can be captured simultaneously with two-photon fluorescence signals with acceptable movement artifacts during active motion. We also show possible applications of this technique by (1) monitoring dynamic changes to microvascular diameter and red blood cells in response to vibrissa sensory stimulation, (2) examining responses of the cerebral microcirculation to the systemic delivery of pharmacological agents using a tail artery cannula during awake imaging, and (3) measuring Ca2+ signals from synthetic and genetically encoded Ca2+ indicators in astrocytes. This method will facilitate acute two-photon fluorescence imaging in awake, active mice and help link cellular events within the neurovascular unit to behavior. PMID:25698926

  10. Acute two-photon imaging of the neurovascular unit in the cortex of active mice

    Directory of Open Access Journals (Sweden)

    Cam Ha Thai Tran

    2015-02-01

    Full Text Available In vivo two-photon scanning fluorescence imaging is a powerful technique to observe physiological processes from the millimeter to the micron scale in the intact animal. In neuroscience research, a common approach is to install an acute cranial window and head bar to explore neocortical function under anesthesia before inflammation peaks from the surgery. However, there are few detailed acute protocols for head-restrained and fully awake animal imaging of the neurovascular unit during activity. This is because acutely performed awake experiments are typically untenable when the animal is naïve to the imaging apparatus. Here we detail a protocol that achieves acute, deep-tissue two-photon imaging of neocortical astrocytes and microvasculature in behaving mice. A week prior to experimentation, implantation of the head bar alone allows mice to train for head-immobilization on an easy-to-learn air-supported ball treadmill. Following just two brief familiarization sessions to the treadmill on separate days, an acute cranial window can subsequently be installed for immediate imaging. We demonstrate how running and whisking data can be captured simultaneously with two-photon fluorescence signals with acceptable movement artifacts during active motion. We also show possible applications of this technique by 1 monitoring dynamic changes to microvascular diameter and red blood cells movements in response to vibrissa sensory stimulation, 2 examining responses of the cerebral microcirculation to the systemic delivery of pharmacological agents using a tail artery cannula during awake imaging, and 3 measuring Ca2+ signals from synthetic and genetically encoded Ca2+ indicators in astrocytes. This method will facilitate acute two-photon fluorescence imaging in awake, active mice and help link cellular events within the neurovascular unit to behaviour.

  11. In vitro imaging of thyroid tissues using two-photon excited fluorescence and second harmonic generation.

    Science.gov (United States)

    Huang, Zufang; Li, Zuanfang; Chen, Rong; Lin, Juqiang; Li, Yongzeng; Li, Chao

    2010-08-01

    To evaluate the feasibility of two-photon excited fluorescence (TPEF) and second harmonic generation (SHG) imaging to discriminate the normal, nodular goiter and papillary cancerous thyroid tissue. In total, 45 fresh thyroid specimens (normal, 15; nodular goiter, 12; and papillary cancerous, 18) from 31 subjects were directly imaged by the TPEF and SHG combination method. The microstructure of follicle and collagen structure in thyroid tissue were clearly identified, morphologic changes between normal, nodular goiter, and papillary cancerous thyroid tissue were well characterized by using two-photon excitation fluorescence. SHG imaging of the collagen matrix also revealed the differences between normal and abnormal. Our preliminary study suggests that the TPEF and SHG combination method might be a useful tool in revealing pathologic changes in thyroid tissue.

  12. Rational Design of Fluorescent Phthalazinone Derivatives for One- and Two-Photon Imaging.

    Science.gov (United States)

    Yang, Lingfei; Zhu, Yuanjun; Shui, Mengyang; Zhou, Tongliang; Cai, Yuanbo; Wang, Wei; Xu, Fengrong; Niu, Yan; Wang, Chao; Zhang, Jun-Long; Xu, Ping; Yuan, Lan; Liang, Lei

    2016-08-22

    Phthalazinone derivatives were designed as optical probes for one- and two-photon fluorescence microscopy imaging. The design strategy involves stepwise extension and modification of pyridazinone by 1) expansion of pyridazinone to phthalazinone, a larger conjugated system, as the electron acceptor, 2) coupling of electron-donating aromatic groups such as N,N-diethylaminophenyl, thienyl, naphthyl, and quinolyl to the phthalazinone, and 3) anchoring of an alkyl chain to the phthalazinone with various terminal substituents such as triphenylphosphonio, morpholino, triethylammonio, N-methylimidazolio, pyrrolidino, and piperidino. Theoretical calculations were utilized to verify the initial design. The desired fluorescent probes were synthesized by two different routes in considerable yields. Twenty-two phthalazinone derivatives were synthesized and their photophysical properties were measured. Selected compounds were applied in cell imaging, and valuable information was obtained. Furthermore, the designed compounds showed excellent performance in two-photon microscopic imaging of mouse brain slices.

  13. Imaging marine virus CroV and its host Cafeteria roenbergensis with two-photon microscopy

    Science.gov (United States)

    Cao, Bin; Chakraborty, Sayan; Sun, Wenqing; Aghvami, Seyedmohammadali; Fischer, Matthias G.; Qian, Wei; Xiao, Chuan; Li, Chunqiang

    2014-02-01

    We use two-photon microscopy to monitor the infection process of marine zooplankton, Cafeteria roenbergensis (C.roenbergensis), by Cafeteria roenbergensis virus (CroV), a giant DNA virus named after its host. Here, we image C.roenbergensis in culture by two-photon excited NADH autofluorescence at video-rate (30 frame/s), and the movement of C.roenbergensis is recorded in live videos. Moreover, CroV is stained with DNA dye SYBR gold and recorded simultaneously with this two-photon microscope. We observed the initial infection moment with this method. The result demonstrates the potential use of two-photon microscopy to investigate the fast dynamic interaction between C.roenbergensis with virus CroV. After catching this initial moment, we will freeze the sample in liquid nitrogen for cryo-electron microscopy (EM) study to resolve the virus-host interaction at molecular level. The long-term goal is to study similar fast moving pathogen-host interaction process which could lead to important medical applications.

  14. Diagnosis of basal cell carcinoma by two photon excited fluorescence combined with lifetime imaging

    Science.gov (United States)

    Fan, Shunping; Peng, Xiao; Liu, Lixin; Liu, Shaoxiong; Lu, Yuan; Qu, Junle

    2014-02-01

    Basal cell carcinoma (BCC) is the most common type of human skin cancer. The traditional diagnostic procedure of BCC is histological examination with haematoxylin and eosin staining of the tissue biopsy. In order to reduce complexity of the diagnosis procedure, a number of noninvasive optical methods have been applied in skin examination, for example, multiphoton tomography (MPT) and fluorescence lifetime imaging microscopy (FLIM). In this study, we explored two-photon optical tomography of human skin specimens using two-photon excited autofluorescence imaging and FLIM. There are a number of naturally endogenous fluorophores in skin sample, such as keratin, melanin, collagen, elastin, flavin and porphyrin. Confocal microscopy was used to obtain structures of the sample. Properties of epidermic and cancer cells were characterized by fluorescence emission spectra, as well as fluorescence lifetime imaging. Our results show that two-photon autofluorescence lifetime imaging can provide accurate optical biopsies with subcellular resolution and is potentially a quantitative optical diagnostic method in skin cancer diagnosis.

  15. Enhanced two-photon excited fluorescence from imaging agents using true thermal light

    Science.gov (United States)

    Jechow, Andreas; Seefeldt, Michael; Kurzke, Henning; Heuer, Axel; Menzel, Ralf

    2013-12-01

    Two-photon excited fluorescence (TPEF) is a standard technique in modern microscopy, but is still affected by photodamage to the probe. It has been proposed that TPEF can be enhanced using entangled photons, but this has proven challenging. Recently, it was shown that some features of entangled photons can be mimicked with thermal light, which finds application in ghost imaging, subwavelength lithography and metrology. Here, we use true thermal light from a superluminescent diode to demonstrate TPEF that is enhanced compared to coherent light, using two common fluorophores and luminescent quantum dots, which suit applications in imaging and microscopy. We find that the TPEF rate is directly proportional to the measured degree of second-order coherence, as predicted by theory. Our results show that photon bunching in thermal light can be exploited in two-photon microscopy, with the photon statistic providing a new degree of freedom.

  16. Two-photon microscopy for imaging germinal centers and T follicular helper cells.

    Science.gov (United States)

    Clatworthy, Menna R

    2015-01-01

    One of the principle features of immune cells is their dynamic nature. Lymphocytes circulate in the blood between secondary lymphoid organs and tissues in an effort to maximize the likelihood of a rapid and appropriate immune response to invading pathogens and tissue damage. Conventional experimental techniques such as histology and flow cytometry have greatly increased our understanding of immune cells, but in the last decade, two-photon microscopy has revolutionized our ability to interrogate the dynamic behavior of immune cells, a facet so critical to their function. Two-photon microscopy relies on the excitation of fluorophores by simultaneous application of two photons of longer wavelength light. This allows a greater depth of imaging with minimal photodamage. Thus, living tissues can be imaged, including immune cells in lymph nodes. This technique has been used to interrogate the events occurring in a germinal center response and the interactions between cells in the germinal center, including T follicular helper cells (Tfh), germinal center B cells, and follicular dendritic cells (FDC). Herein, a method is described by which the interactions between Tfh and B cells within a germinal center in a popliteal lymph node can be imaged in a live mouse.

  17. Intravital two-photon imaging: a versatile tool for dissecting the immune system.

    Science.gov (United States)

    Ishii, Taeko; Ishii, Masaru

    2011-03-01

    During the past decade, multi-photon or 'two-photon' excitation microscopy has launched a new era in the field of biological imaging. The near-infrared excitation laser for two-photon microscopy can penetrate thicker specimens, enabling the visualisation of living cell behaviour deep within tissues and organs without thin sectioning. The minimised photobleaching and toxicity enables the visualisation of live and intact specimens for extended periods. In this brief review, recent findings in intravital two-photon imaging for the physiology and pathology of the immune system are discussed. The immune system configures highly dynamic networks, where many cell types actively travel throughout the body and interact with each other in specific areas. Hence, real-time intravital imaging may be a powerful tool for dissecting the mechanisms of this dynamic system. The most unique characteristic of the immune system is its highly dynamic nature. A variety of cell types, such as lymphocytes, macrophages and dendritic cells (DCs), are continuously circulating throughout the body, migrating through the peripheral tissues and interacting with each other in their respective niches. Conventional methodologies in immunology, such as flow cytometry, cell or tissue culture, biochemistry and histology, have brought tremendous achievement within this field, although the dynamics of immune cells in an entire animal remain less clear. Technological progress of fluorescence microscopy has enabled us to visualise the intact biological phenomenon that has been uninvestigated. Among the advancements, the recent emergence and prevalence of two-photon, excitation-based, laser microscopy has revolutionised the research field, such that the dynamic behaviour of cells deep inside living organs can be visualised and analysed.

  18. Adaptive optics for in vivo two-photon calcium imaging of neuronal networks

    Science.gov (United States)

    Meimon, Serge; Conan, Jean-Marc; Mugnier, Laurent M.; Michau, Vincent; Cossart, Rosa; Malvache, Arnaud

    2014-03-01

    The landscape of biomedical research in neuroscience has changed dramatically in recent years as a result of spectacular progress in dynamic microscopy. However, the optical accessibility of deep brain structures or deeper regions of the surgically exposed hippocampus (a few 100 microns typically) remains limited, due to volumic aberrations created by the sample inhomogeneities. Adaptive optics can correct for these aberrations. Our goal is to realize a novel adaptive optics module dedicated to in vivo two-photon calcium imaging of the hippocampus. The key issue in adaptive optics is the ability to perform an accurate and reliable wavefront sensing. In two- photon microscopy indirect methods are required. Two families of approaches have been proposed so far, the modal sensorless technique and a method based on pupil segmentation. We present here a formal comparison of these approaches, in particular as a function of the amount of aberrations.

  19. Additive controlled synthesis of gold nanorods (GNRs) for two-photon luminescence imaging of cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhu Jing; Roy, Indrajit; Hu Rui; Ding Hong; Zhao Lingling; He, Guang S; Prasad, Paras N [Institute for Lasers, Photonics and Biophotonics, University at Buffalo, State University of New York, Buffalo, NY 14260-4200 (United States); Yong, Ken-Tye [School of Electrical and Electronic Engineering, Nanyang Technological University, Singapore 639798 (Singapore); Swihart, Mark T [Department of Chemical and Biological Engineering, University at Buffalo, State University of New York, Buffalo, NY 14260-4200 (United States); Cui Yiping, E-mail: ktyong@ntu.edu.sg, E-mail: cyp@seu.edu.cn, E-mail: pnprasad@buffalo.edu [Advanced Photonics Center, School of Electronic Science and Engineering, Southeast University, Nanjing 210096 (China)

    2010-07-16

    Gold nanorods (GNRs) with a longitudinal surface plasmon resonance peak that is tunable from 600 to 1100 nm have been fabricated in a cetyl trimethylammoniumbromide (CTAB) micellar medium using hydrochloric acid and silver nitrate as additives to control their shape and size. By manipulating the concentrations of silver nitrate and hydrochloric acid, the aspect ratio of the GNRs was reliably and reproducibly tuned from 2.5 to 8. The GNRs were first coated with polyelectrolyte multilayers and then bioconjugated to transferrin (Tf) to target pancreatic cancer cells. Two-photon imaging excited from the bioconjugated GNRs demonstrated receptor-mediated uptake of the bioconjugates into Panc-1 cells, overexpressing the transferrin receptor (TfR). The bioconjugated GNR formulation exhibited very low toxicity, suggesting that it is biocompatible and potentially suitable for targeted two-photon bioimaging.

  20. Additive controlled synthesis of gold nanorods (GNRs) for two-photon luminescence imaging of cancer cells.

    Science.gov (United States)

    Zhu, Jing; Yong, Ken-Tye; Roy, Indrajit; Hu, Rui; Ding, Hong; Zhao, Lingling; Swihart, Mark T; He, Guang S; Cui, Yiping; Prasad, Paras N

    2010-07-16

    Gold nanorods (GNRs) with a longitudinal surface plasmon resonance peak that is tunable from 600 to 1100 nm have been fabricated in a cetyl trimethylammoniumbromide (CTAB) micellar medium using hydrochloric acid and silver nitrate as additives to control their shape and size. By manipulating the concentrations of silver nitrate and hydrochloric acid, the aspect ratio of the GNRs was reliably and reproducibly tuned from 2.5 to 8. The GNRs were first coated with polyelectrolyte multilayers and then bioconjugated to transferrin (Tf) to target pancreatic cancer cells. Two-photon imaging excited from the bioconjugated GNRs demonstrated receptor-mediated uptake of the bioconjugates into Panc-1 cells, overexpressing the transferrin receptor (TfR). The bioconjugated GNR formulation exhibited very low toxicity, suggesting that it is biocompatible and potentially suitable for targeted two-photon bioimaging.

  1. Correction-free remotely scanned two-photon in vivo mouse retinal imaging

    Institute of Scientific and Technical Information of China (English)

    Adi Schejter Bar-Noam; Nairouz Farah; Shy Shoham

    2016-01-01

    Non-invasive fluorescence retinal imaging in small animals is an important requirement for an array of translational vision applications.The in vivotwo-photon imaging of the mouse retina may enable the long-term investigation of the structure and function of healthy and diseased retinal tissue.However,to date,this has only been possible using relatively complex adaptive-optics systems.Here,the optical modeling of the murine eye and of the imaging system is used to achieve correction-free two-photon microscopy through the pupil of a mouse eye to yield high-quality,optically sectioned fundus images.By remotely scanning the focus using an electronically tunable lens,high-resolution three-dimensional fluorescein angiograms and cellular-scale images are acquired,thus introducing a correction-free baseline performance level for two-photon in vivo retinal imaging.Moreover,the system enables functional calcium imaging of repeated retinal responses to light stimulation using the genetically encoded indicator,GCaMP6s.These results and the simplicity of the new add-on optics are an important step toward several structural,functional,and multimodal imaging applications that will benefit from the tight optical sectioning and the use of near-infrared light.

  2. Time-lapse microscopy using smartphone with augmented reality markers.

    Science.gov (United States)

    Baek, Dongyoub; Cho, Sungmin; Yun, Kyungwon; Youn, Keehong; Bang, Hyunwoo

    2014-04-01

    A prototype system that replaces the conventional time-lapse imaging in microscopic inspection for use with smartphones is presented. Existing time-lapse imaging requires a video data feed between a microscope and a computer that varies depending on the type of image grabber. Even with proper hardware setups, a series of tedious and repetitive tasks is still required to relocate to the region-of-interest (ROI) of the specimens. In order to simplify the system and improve the efficiency of time-lapse imaging tasks, a smartphone-based platform utilizing microscopic augmented reality (μ-AR) markers is proposed. To evaluate the feasibility and efficiency of the proposed system, a user test was designed and performed, measuring the elapse time for a trial of the task starting from the execution of the application software to the completion of restoring and imaging of an ROI saved in advance. The results of the user test showed that the average elapse time was 65.3 ± 15.2 s with 6.86 ± 3.61 μm of position error and 0.08 ± 0.40 degrees of angle error. This indicates that the time-lapse imaging task was accomplished rapidly with a high level of accuracy. Thus, simplification of both the system and the task was achieved via our proposed system. Copyright © 2014 Wiley Periodicals, Inc.

  3. A femtosecond Raman generator for long wavelength two-photon and third harmonic generation imaging

    Science.gov (United States)

    Trägârdh, J.; Schniete, J.; Parsons, M.; McConnell, G.

    2016-12-01

    We demonstrate a femtosecond single pass Raman generator based on an YVO4 crystal pumped by a high energy fiber laser at a wavelength of 1064 nm and a repetition rate of 1 MHz. The Raman generator shifts the pump wavelength to 1175 nm, in a broadband spectrum, making it suitable for multi-photon microscopy. We use the Raman generator for third harmonic generation imaging of live plant specimens as well as for two-photon fluorescence imaging of red fluorescent protein expressing HeLa cells. We demonstrate that the photo-damage to a live specimen is low.

  4. Two-photon fluorescent sensor for K+ imaging in live cells (Conference Presentation)

    Science.gov (United States)

    Sui, Binglin; Yue, Xiling; Kim, Bosung; Belfield, Kevin D.

    2016-03-01

    It is difficult to overstate the physiological importance of potassium for life as its indispensable roles in a variety of biological processes are widely known. As a result, efficient methods for determining physiological levels of potassium are of paramount importance. Despite this, relatively few K+ fluorescence sensors have been reported, with only one being commercially available. A new two-photon excited fluorescent K+ sensor is reported. The sensor is comprised of three moieties, a highly selective K+ chelator as the K+ recognition unit, a boron-dipyrromethene (BODIPY) derivative modified with phenylethynyl groups as the fluorophore, and two polyethylene glycol chains to afford water solubility. The sensor displays very high selectivity (physiological metal cations. Upon binding K+, the sensor switches from non-fluorescent to highly fluorescent, emitting red to near-IR (NIR) fluorescence. The sensor exhibited a good two-photon absorption cross section, 500 GM at 940 nm. Moreover, it is not sensitive to pH in the physiological pH range. Time-dependent cell imaging studies via both one- and two-photon fluorescence microscopy demonstrate that the sensor is suitable for dynamic K+ sensing in living cells.

  5. Two-photon fluorescence and fluorescence imaging of two styryl heterocyclic dyes combined with DNA.

    Science.gov (United States)

    Gao, Chao; Liu, Shu-yao; Zhang, Xian; Liu, Ying-kai; Qiao, Cong-de; Liu, Zhao-e

    2016-03-01

    Two new styryl heterocyclic two-photon (TP) materials, 4-[4-(N-methyl)styrene]-imidazo [4,5-f][1,10] phenanthroline-benzene iodated salt (probe-1) and 4,4-[4-(N-methyl)styrene]-benzene iodated salt (probe-2) were successfully synthesized and studied as potential fluorescent probes of DNA detection. The linear and nonlinear photophysical properties of two compounds in different solvents were investigated. The absorption, one- and two-photon fluorescent spectra of the free dye and dye-DNA complex were also examined to evaluate their photophysical properties. The binding constants of dye-DNA were obtained according to Scatchard equation with good values. The results showed that two probes could be used as fluorescent DNA probes by two-photon excitation, and TP fluorescent properties of probe-1 are superior to that of probe-2. The fluorescent method date indicated that the mechanisms of dye-DNA complex interaction may be groove binding for probe-1 and electrostatic interaction for probe-2, respectively. The MTT assay experiments showed two probes are low toxicity. Moreover, the TP fluorescence imaging of DNA detection in living cells at 800 nm indicated that the ability to locate in cell nuclei of probe-1 is better than that of probe-2.

  6. Two-photon absorption and transient photothermal imaging of pigments in tissues

    Science.gov (United States)

    Ye, Tong; Fu, Dan; Matthews, Thomas E.; Hong, Lian; Simon, John D.; Warren, Warren S.

    2008-02-01

    As a main pigment in skin tissues, melanin plays an important role in photo-protecting skin from UV radiation. However, melanogenesis may be altered due to disease or environmental factors; for example, sun exposure may cause damage and mutation of melanocytes and induce melanoma. Imaging pigmentation changes may provide invaluable information to catch the malignant transformation in its early stage and in turn improve the prognosis of patients. We have demonstrated previously that transmission mode, two-photon, one- or two-color absorption microscopy could provide remarkable contrast in imaging melanin in skin. In this report we demonstrate significantly improved sensitivity, so that we are now able to image in epi-mode (or back reflection) in two-photon absorption. This improvement makes possible for us to characterize the different types of pigmentation on the skin in vivo at virtually any location. Another finding is that we can also image transient photothermal dynamics due to the light absorption of melanin. By carefully choosing excitation and probe wavelengths, we might be able to image melanin in different structures under different micro-environments in skin, which could provide useful photochemical and photophysical insights in understanding how pigments are involved in photoprotection and photodamage of cells.

  7. An automated detection for axonal boutons in vivo two-photon imaging of mouse

    Science.gov (United States)

    Li, Weifu; Zhang, Dandan; Xie, Qiwei; Chen, Xi; Han, Hua

    2017-02-01

    Activity-dependent changes in the synaptic connections of the brain are tightly related to learning and memory. Previous studies have shown that essentially all new synaptic contacts were made by adding new partners to existing synaptic elements. To further explore synaptic dynamics in specific pathways, concurrent imaging of pre and postsynaptic structures in identified connections is required. Consequently, considerable attention has been paid for the automated detection of axonal boutons. Different from most previous methods proposed in vitro data, this paper considers a more practical case in vivo neuron images which can provide real time information and direct observation of the dynamics of a disease process in mouse. Additionally, we present an automated approach for detecting axonal boutons by starting with deconvolving the original images, then thresholding the enhanced images, and reserving the regions fulfilling a series of criteria. Experimental result in vivo two-photon imaging of mouse demonstrates the effectiveness of our proposed method.

  8. Highly Efficient and Excitation Tunable Two-Photon Luminescence Platform For Targeted Multi-Color MDRB Imaging Using Graphene Oxide

    Science.gov (United States)

    Pramanik, Avijit; Fan, Zhen; Chavva, Suhash Reddy; Sinha, Sudarson Sekhar; Ray, Paresh Chandra

    2014-08-01

    Multiple drug-resistance bacteria (MDRB) infection is one of the top three threats to human health according to the World Health Organization (WHO). Due to the large penetration depth and reduced photodamage, two-photon imaging is an highly promising technique for clinical MDRB diagnostics. Since most commercially available water-soluble organic dyes have low two-photon absorption cross-section and rapid photobleaching tendency, their applications in two-photon imaging is highly limited. Driven by the need, in this article we report extremely high two-photon absorption from aptamer conjugated graphene oxide (σ2PA = 50800 GM) which can be used for highly efficient two-photon fluorescent probe for MDRB imaging. Reported experimental data show that two-photon photoluminescence imaging color, as well as luminescence peak position can be tuned from deep blue to red, just by varying the excitation wavelength without changing its chemical composition and size. We have demonstrated that graphene oxide (GO) based two-photon fluorescence probe is capable of imaging of multiple antibiotics resistance MRSA in the first and second biological transparency windows using 760-1120 nm wavelength range.

  9. Two-photon autofluorescence lifetime and SHG imaging of healthy and diseased human corneas

    Science.gov (United States)

    Batista, Ana; Breunig, Hans Georg; Uchugonova, Aisada; Seitz, Berthold; Morgado, António Miguel; König, Karsten

    2015-03-01

    Corneal function can be drastically affected by several degenerations and dystrophies, leading to blindness. Early diagnosis of corneal disease is of major importance and it may be accomplished by monitoring changes of the metabolic state and structural organization, the first detectable pathological signs, by two-photon excitation autofluorescence lifetime and second-harmonic generation imaging. In this study, we propose to use these imaging techniques to differentiate between healthy and pathological corneas. Images were acquired using a laser-scanning microscope with a broadband sub-15 femtosecond near-infrared pulsed laser and a 16-channel photomultiplier tube detector for signal collection. This setup allows the simultaneous excitation of metabolic co-factors and to identify them based on their fluorescence spectra. We were able to discriminate between healthy and pathological corneas using two-photon excitation autofluorescence lifetime and second-harmonic generation imaging from corneal epithelium and stroma. Furthermore, differences between different pathologies were observed. Alterations in the metabolic state of corneal epithelial cells were observed using the autofluorescence lifetime of the metabolic co-factors. In the corneal stroma, we observed not only alterations in the collagen fibril structural organization but also alterations in the autofluorescence lifetime. Further tests are required as the number of pathological samples must be increased. In the future, we intend to establish a correlation between the metabolic and structural changes and the disease stage. This can be a step forward in achieving early diagnosis.

  10. Multidimensional two-photon imaging and spectroscopy of fresh human bladder biopsies

    Science.gov (United States)

    Cicchi, Riccardo; Crisci, Alfonso; Cosci, Alessandro; Nesi, Gabriella; Giancane, Saverio; Carini, Marco; Pavone, Francesco S.

    2010-02-01

    Two-photon microscopy has been successfully used to image several types of tissues, including skin, muscles, tendons. Nevertheless, its usefulness in imaging bladder tissue has not been investigated yet. In this work we used combined twophoton excited fluorescence, second-harmonic generation microscopy, fluorescence lifetime imaging microscopy, and multispectral two-photon emission detection to investigate different kinds of human ex-vivo fresh biopsies of bladder. Morphological and spectroscopic analyses allowed to characterize both healthy mucosa and carcinoma in-situ samples in a good agreement with common routine histology. Cancer cells showed different morphology with respect to the corresponding healthy cells: they appeared more elongated and with a larger nucleus to cytoplasm ratio. From the spectroscopic point of view, differences between the two tissue types in both spectral emission and fluorescence lifetime distribution were found. Even if further analysis, as well as a more significant statistics on a larger number of samples would be helpful to discriminate between low, mild, and high grade cancer, our method is a promising tool to be used as diagnostic confirmation of histological results, as well to be implemented in a multi-photon endoscope or in a spectroscopic for in in-vivo imaging applications.

  11. Time-lapse seismic within reservoir engineering

    OpenAIRE

    Oldenziel, T.

    2003-01-01

    Time-lapse 3D seismic is a fairly new technology allowing dynamic reservoir characterisation in a true volumetric sense. By investigating the differences between multiple seismic surveys, valuable information about changes in the oil/gas reservoir state can be captured. Its interpretation involves different disciplines, of which the main three are: reservoir management, rock physics, and seismics. The main challenge is expressed as "How to optimally benefit from time-lapse seismic". The chall...

  12. A novel Kalman filter based video image processing scheme for two-photon fluorescence microscopy

    Science.gov (United States)

    Sun, Wenqing; Huang, Xia; Li, Chunqiang; Xiao, Chuan; Qian, Wei

    2016-03-01

    Two-photon fluorescence microscopy (TPFM) is a perfect optical imaging equipment to monitor the interaction between fast moving viruses and hosts. However, due to strong unavoidable background noises from the culture, videos obtained by this technique are too noisy to elaborate this fast infection process without video image processing. In this study, we developed a novel scheme to eliminate background noises, recover background bacteria images and improve video qualities. In our scheme, we modified and implemented the following methods for both host and virus videos: correlation method, round identification method, tree-structured nonlinear filters, Kalman filters, and cell tracking method. After these procedures, most of noises were eliminated and host images were recovered with their moving directions and speed highlighted in the videos. From the analysis of the processed videos, 93% bacteria and 98% viruses were correctly detected in each frame on average.

  13. In-vivo two-photon imaging of the honey bee antennal lobe

    CERN Document Server

    Haase, Albrecht; Trona, Federica; Anfora, Gianfranco; Vallortigara, Giorgio; Antolini, Renzo; Vinegoni, Claudio

    2010-01-01

    Due to the honey bee's importance as a simple neural model, there is a great need for new functional imaging modalities. Herein we report on the use of two-photon microscopy for in-vivo functional and morphological imaging of the honey bee's olfactory system focusing on its primary centers, the antennal lobes (ALs). Our imaging platform allows for simultaneously obtaining both morphological measurements of the AL and in-vivo calcium recording of neural activities. By applying external odor stimuli to the bee's antennas, we were able to record the characteristic odor response maps. Compared to previous works where conventional fluorescence microscopy is used, our approach offers all the typical advantages of multi-photon imaging, providing substantial enhancement in both spatial and temporal resolutions while minimizing photo-damages and autofluorescence contribution with a four-fold improvement in the functional signal. Moreover, the multi-photon associated extended penetration depth allows for functional ima...

  14. Two-photon imaging of lymphoma cells targeted by gold nanoparticles

    Institute of Scientific and Technical Information of China (English)

    Xiaochao Qu; Jing Wang; Cuiping Yao; Zhenxi Zhang

    2008-01-01

    Gold nanoparticles (NPs) have highly efficient multi-photon-induced luminescence. In this paper, we record the two-photon images of gold NPs, lymphoma cell line Karpas 299, and Karpas 299 incubated with 30-nm-diameter gold NPs and ACT-1 antibody conjugates (Au30-ACT-1 conjugates) by using a multi-photon microscopy system. Due to the specific conjugation of ACT-1 antibody and celt membrane receptor CD25, gold NPs are only bound to the surface of cell membrane of Karpas 299. The luminescence intensity of gold NPs is higher than that of cells at 750-nm laser excitation. By comparing the images of Karpas 299 cells incubated with and without gold NPs, it is found that by means of gold NPs, we can get clear cell images with lower excitation power. Their excellent optical and chemical properties make gold NPs an attractive contrast agent for cellular imaging.

  15. A Two-Photon Ratiometric Fluorescent Probe for Imaging Carboxylesterase 2 in Living Cells and Tissues.

    Science.gov (United States)

    Jin, Qiang; Feng, Lei; Wang, Dan-Dan; Dai, Zi-Ru; Wang, Ping; Zou, Li-Wei; Liu, Zhi-Hong; Wang, Jia-Yue; Yu, Yang; Ge, Guang-Bo; Cui, Jing-Nan; Yang, Ling

    2015-12-30

    In this study, a two-photon ratiometric fluorescent probe NCEN has been designed and developed for highly selective and sensitive sensing of human carboxylesterase 2 (hCE2) based on the catalytic properties and substrate preference of hCE2. Upon addition of hCE2, the probe could be readily hydrolyzed to release 4-amino-1,8-naphthalimide (NAH), which brings remarkable red-shift in fluorescence (90 nm) spectrum. The newly developed probe exhibits good specificity, ultrahigh sensitivity, and has been successfully applied to determine the real activities of hCE2 in complex biological samples such as cell and tissue preparations. NCEN has also been used for two-photon imaging of intracellular hCE2 in living cells as well as in deep-tissues for the first time, and the results showed that the probe exhibited high ratiometric imaging resolution and deep-tissue imaging depth. All these findings suggested that this probe holds great promise for applications in bioimaging of endogenous hCE2 in living cells and in exploring the biological functions of hCE2 in complex biological systems.

  16. In situ imaging of the mouse cochlea using two-photon microscopy

    Science.gov (United States)

    Yang, Xin; Pu, Ye; Psaltis, Demetri; Stankovic, Konstantina M.

    2013-04-01

    Intracochlear imaging is of great interest clinically because cochlea is the central organ of hearing. However, intracochlear imaging is technologically challenging due to the cochlea's small size and encasement in bone. The state-of- the-art imaging techniques are not adequate for high resolution cellular imaging to establish diagnosis without destroying the cochlea. We report in situ imaging of intact mouse cochlea using endogenous two-photon excitation fluorescence (TPEF) as the contrast mechanism. TPEF eliminates the need for exogenous labeling and eradicating the staining-induced artifacts. We used a natural, membranous opening into the cochlea, the round window, as the optical access to reach the organ of Corti, requiring no additional slicing or opening. Our approach provides the maximum non-invasiveness in the imaging process. TPEF exhibits strong contrast allowing deep imaging of mouse cochlea with cellular and even subcellular resolution. Inner hair cell, outer hair cell and supporting cell are clearly identifiable in TPEF images. Distinct morphological differences are observed between healthy and noise-exposed cochleae, allowing detection of specific, noise-induced pathologic changes. The TPEF images taken through the round window are correlated with the whole mount sections, verifying their reliability. Compared with one-photon excitation fluorescence (OPEF) confocal microscope and wide-field transmission microscope images taken under the same magnification and resolution, TPEF images demonstrate clear advantages in terms of sharpness, signal to noise ratio and contrast. These capabilities provide a working foundation for microendoscopy-based clinical diagnostics of sensorineural hearing loss.

  17. Multiscale vision model for event detection and reconstruction in two-photon imaging data

    DEFF Research Database (Denmark)

    Brazhe, Alexey; Mathiesen, Claus; Lind, Barbara Lykke;

    2014-01-01

    Reliable detection of calcium waves in multiphoton imaging data is challenging because of the low signal-to-noise ratio and because of the unpredictability of the time and location of these spontaneous events. This paper describes our approach to calcium wave detection and reconstruction based...... on a modified multiscale vision model, an object detection framework based on the thresholding of wavelet coefficients and hierarchical trees of significant coefficients followed by nonlinear iterative partial object reconstruction, for the analysis of two-photon calcium imaging data. The framework is discussed...... in the context of detection and reconstruction of intercellular glial calcium waves. We extend the framework by a different decomposition algorithm and iterative reconstruction of the detected objects. Comparison with several popular state-of-the-art image denoising methods shows that performance...

  18. Nanoshells for in vivo imaging using two-photon excitation microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Gao Liang; Nammalvar, Vengadesan [Department of Bioengineering, Rice University, Houston, TX 77005 (United States); Vadakkan, Tegy J, E-mail: lg3@rice.edu, E-mail: venkyn@rice.edu [Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030 (United States)

    2011-09-07

    Gold nanoshells have been intensively investigated and applied to various biomedical fields because of their flexible optical tunability and biological compatibility. They hold great potential to serve as luminescent contrast agents excitable with near-infrared (NIR) lasers. In this paper, we describe the development of nanoshells with a peak of plasmon resonance at 800 nm and their subsequent use for in vivo blood vessel imaging using two-photon excitation microscopy at an excitation wavelength of 750 nm. We were able to image single nanoshell particles in blood vessels and generate optical contrast for blood vessel structure using luminescent signals. These results confirm the feasibility of engineering nanoshells with controlled optical properties for single-particle-based in vivo imaging.

  19. Nanoshells for in vivo imaging using two-photon excitation microscopy.

    Science.gov (United States)

    Gao, Liang; Vadakkan, Tegy J; Nammalvar, Vengadesan

    2011-09-07

    Gold nanoshells have been intensively investigated and applied to various biomedical fields because of their flexible optical tunability and biological compatibility. They hold great potential to serve as luminescent contrast agents excitable with near-infrared (NIR) lasers. In this paper, we describe the development of nanoshells with a peak of plasmon resonance at 800 nm and their subsequent use for in vivo blood vessel imaging using two-photon excitation microscopy at an excitation wavelength of 750 nm. We were able to image single nanoshell particles in blood vessels and generate optical contrast for blood vessel structure using luminescent signals. These results confirm the feasibility of engineering nanoshells with controlled optical properties for single-particle-based in vivo imaging.

  20. Two-photon fluorescence and confocal reflected light imaging of thick tissue structures

    Science.gov (United States)

    Kim, Ki H.; So, Peter T. C.; Kochevar, Irene E.; Masters, Barry R.; Gratton, Enrico

    1998-04-01

    The technology of two-photon excitation has opened a window of opportunity for developing non-invasive medical diagnostic tools capable of monitoring thick tissue biochemical states. Using cellular endogenous chromophores, (beta) -nicotinamide- adenine dinucleotide phosphate [NAD(P)H], the cellular metabolic rates in living human skin were determined. Although important functional information can be obtained from the fluorescence spectroscopy of endogenous chromophores, these chromophores are rather poor contrast enhancing agent for mapping cellular morphology. First, most endogenous chromophores are confined to the cellular cytoplasm which prevents the visualization of other cellular organelles. Second, there is significant variability in the distribution and the quantum yield of endogenous chromophores which depends on tissue biochemistry but prevents consistent comparison of cellular morphology. On the other hand, the deep tissue cellular morphology has been imaged with excellent resolution using reflected light confocal microscopy. In reflected light microscopy, the image contrast originates from the index of refraction differences of the cellular structures. The organelle boundaries with significant index differences such as the plasma membrane and the nucleus envelope can be consistently visualized. A combination of morphological and functional information is required for a thorough tissue study. This presentation describes the development of a new microscope which is capable of simultaneously collecting both two-photon fluorescence and confocal reflected light signals. Promising biomedical applications include the non-invasive diagnosis of skin cancer and the study of wound healing.

  1. Two photon fluorescence imaging of lipid membrane domains and potentials using advanced fluorescent probes

    Science.gov (United States)

    Kilin, Vasyl; Darwich, Zeinab; Richert, Ludovic; Didier, Pascal; Klymchenko, Andrey; Mély, Yves

    2013-02-01

    Biomembranes are ordered and dynamic nanoscale structures critical for cell functions. The biological functions of the membranes strongly depend on their physicochemical properties, such as electrostatics, phase state, viscosity, polarity and hydration. These properties are essential for the membrane structure and the proper folding and function of membrane proteins. To monitor these properties, fluorescence techniques and notably, two-photon microscopy appear highly suited due to their exquisite sensitivity and their capability to operate in complex biological systems, such as living cells and tissues. In this context, we have developed multiparametric environment-sensitive fluorescent probes tailored for precise location in the membrane bilayer. We notably developed probes of the 3-hydroxychromone family, characterized by an excited state intramolecular proton transfer reaction, which generates two tautomeric emissive species with well-separated emission bands. As a consequence, the response of these probes to changes in their environment could be monitored through changes in the ratios of the two bands, as well as through changes in the fluorescence lifetimes. Using two-photon ratiometric imaging and FLIM, these probes were used to monitor the surface membrane potential, and were applied to detect apoptotic cells and image membrane domains.

  2. Two-Photon Microscopy Allows Imaging and Characterization of Cochlear Microvasculature In Vivo

    Directory of Open Access Journals (Sweden)

    Friedrich Ihler

    2015-01-01

    Full Text Available Impairment of cochlear blood flow has been discussed as factor in the pathophysiology of various inner ear disorders. However, the microscopic study of cochlear microcirculation is limited due to small scale and anatomical constraints. Here, two-photon fluorescence microscopy is applied to visualize cochlear microvessels. Guinea pigs were injected with Fluorescein isothiocyanate- or Texas red-dextrane as plasma marker. Intravital microscopy was performed in four animals and explanted cochleae from four animals were studied. The vascular architecture of the cochlea was visualized up to a depth of 90.0±22.7 μm. Imaging yielded a mean contrast-to-noise ratio (CNR of 3.3±1.7. Mean diameter in vivo was 16.5±6.0 μm for arterioles and 8.0±2.4 μm for capillaries. In explanted cochleae, the diameter of radiating arterioles and capillaries was measured with 12.2±1.6 μm and 6.6±1.0 μm, respectively. The difference between capillaries and arterioles was statistically significant in both experimental setups (P<0.001 and P=0.022, two-way ANOVA. Measured vessel diameters in vivo and ex vivo were in agreement with published data. We conclude that two-photon fluorescence microscopy allows the investigation of cochlear microvessels and is potentially a valuable tool for inner ear research.

  3. Maximum imaging depth of two-photon autofluorescence microscopy in epithelial tissues.

    Science.gov (United States)

    Durr, Nicholas J; Weisspfennig, Christian T; Holfeld, Benjamin A; Ben-Yakar, Adela

    2011-02-01

    Endogenous fluorescence provides morphological, spectral, and lifetime contrast that can indicate disease states in tissues. Previous studies have demonstrated that two-photon autofluorescence microscopy (2PAM) can be used for noninvasive, three-dimensional imaging of epithelial tissues down to approximately 150 μm beneath the skin surface. We report ex-vivo 2PAM images of epithelial tissue from a human tongue biopsy down to 370 μm below the surface. At greater than 320 μm deep, the fluorescence generated outside the focal volume degrades the image contrast to below one. We demonstrate that these imaging depths can be reached with 160 mW of laser power (2-nJ per pulse) from a conventional 80-MHz repetition rate ultrafast laser oscillator. To better understand the maximum imaging depths that we can achieve in epithelial tissues, we studied image contrast as a function of depth in tissue phantoms with a range of relevant optical properties. The phantom data agree well with the estimated contrast decays from time-resolved Monte Carlo simulations and show maximum imaging depths similar to that found in human biopsy results. This work demonstrates that the low staining inhomogeneity (∼ 20) and large scattering coefficient (∼ 10 mm(-1)) associated with conventional 2PAM limit the maximum imaging depth to 3 to 5 mean free scattering lengths deep in epithelial tissue.

  4. Comparison of calcium imaging in dorsal root ganglion neurons by using laser scanning confocal and two-photon microscopy

    Science.gov (United States)

    Huang, Yimei; Yang, Hongqin; Chen, Jiangxu; Shen, Xiuqiu; Zheng, Liqin; Wang, Yuhua; Xie, Shusen

    2012-03-01

    As one of the most important second messengers, calcium in nerve cells plays a critical role in neuronal processes, including excitability, neurotransmitter release, synaptic plasticity. Modulation of the calcium concentration is an important means of regulating diverse neuronal functions. To evaluate the role of calcium, quantitative measurement of cytosolic free calcium concentrations is necessary. There are several optical techniques that are available for measurement of calcium in live cells. Laser scanning confocal microscopy and two-photon microscopy are two prevalent techniques for their advantage in spatial resolution. In this paper, calcium in dorsal root ganglion neurons was imaged by laser scanning confocal microscopy and two-photon microscopy with Fluo-3, a calcium specific fluorescence probe. Both of spatial resolution and photobleaching, two common limitations of optical image modality, were compared between laser scanning confocal microscopy and two-photon microscopy, respectively. Three dimension images showed that laser scanning confocal microscopy and two-photon microscopy had not only similar lateral resolution but also parallel vertical resolution. However, Laser scanning confocal microscopy had an advantage over the two-photon microcopy in photobleaching. These results indicated that laser scanning confocal microscopy was more suitable than two-photon microscopy to be applied in imaging calcium in dorsal root ganglion neurons with Fluo-3.

  5. Accurate, precise modeling of cell proliferation kinetics from time-lapse imaging and automated image analysis of agar yeast culture arrays

    Directory of Open Access Journals (Sweden)

    Zhao Lue

    2007-01-01

    Full Text Available Abstract Background Genome-wide mutant strain collections have increased demand for high throughput cellular phenotyping (HTCP. For example, investigators use HTCP to investigate interactions between gene deletion mutations and additional chemical or genetic perturbations by assessing differences in cell proliferation among the collection of 5000 S. cerevisiae gene deletion strains. Such studies have thus far been predominantly qualitative, using agar cell arrays to subjectively score growth differences. Quantitative systems level analysis of gene interactions would be enabled by more precise HTCP methods, such as kinetic analysis of cell proliferation in liquid culture by optical density. However, requirements for processing liquid cultures make them relatively cumbersome and low throughput compared to agar. To improve HTCP performance and advance capabilities for quantifying interactions, YeastXtract software was developed for automated analysis of cell array images. Results YeastXtract software was developed for kinetic growth curve analysis of spotted agar cultures. The accuracy and precision for image analysis of agar culture arrays was comparable to OD measurements of liquid cultures. Using YeastXtract, image intensity vs. biomass of spot cultures was linearly correlated over two orders of magnitude. Thus cell proliferation could be measured over about seven generations, including four to five generations of relatively constant exponential phase growth. Spot area normalization reduced the variation in measurements of total growth efficiency. A growth model, based on the logistic function, increased precision and accuracy of maximum specific rate measurements, compared to empirical methods. The logistic function model was also more robust against data sparseness, meaning that less data was required to obtain accurate, precise, quantitative growth phenotypes. Conclusion Microbial cultures spotted onto agar media are widely used for genotype

  6. Quantitative Imaging of Molecular Order in Lipid Membranes Using Two-Photon Fluorescence Polarimetry

    Science.gov (United States)

    Gasecka, Alicja; Han, Tsai-Jung; Favard, Cyril; Cho, Bong Rae; Brasselet, Sophie

    2009-01-01

    Abstract We present a polarimetric two-photon microscopy technique to quantitatively image the local static molecular orientational behavior in lipid and cell membranes. This approach, based on a tunable excitation polarization state complemented by a polarized readout, is easily implementable and does not require hypotheses on the molecular angular distribution such as its mean orientation, which is a main limitation in traditional fluorescence anisotropy measurements. The method is applied to the investigation of the molecular angular distribution in giant unilamellar vesicles formed by liquid-ordered and liquid-disordered micro-domains, and in COS-7 cell membranes. The highest order contrast between ordered and disordered domains is obtained for dyes locating within the membrane acyl chains. PMID:19917241

  7. Ultrabroadband ghost imaging exploiting optoelectronic amplified spontaneous emission and two-photon detection.

    Science.gov (United States)

    Hartmann, Sébastien; Molitor, Andreas; Elsäßer, Wolfgang

    2015-12-15

    Ghost imaging (GI) is one of the recent fascinating and probably counterintuitive topics of quantum optics. Here, we present an alternative classical GI scheme using spectrally ultrabroadband amplified spontaneous emission from an optoelectronic quantum dot based superluminescent diode source. This light source exhibits highly incoherent properties regarding both first- and second-order correlations with a 70 nm-wide optical spectrum as well as thermal-like photon statistics. Exploiting a two-photon-absorption detection method, we demonstrate for the first time, to the best of our knowledge, a GI experiment handling the corresponding femtosecond correlation timescales. By introducing compact broadband light sources to GI, this work contributes toward practical application of GI.

  8. Two-photon calcium imaging during fictive navigation in virtual environments

    Directory of Open Access Journals (Sweden)

    Misha Benjamin Ahrens

    2013-06-01

    Full Text Available A full understanding of nervous system function requires recording from large populations of neurons during naturalistic behaviors. Here we enable paralyzed larval zebrafish to fictively navigate two-dimensional virtual environments while we record optically from many neurons with two-photon imaging. Electrical recordings from motor nerves in the tail are decoded into intended forward swims and turns, which are used to update a virtual environment displayed underneath the fish. Several behavioral features - such as turning responses to whole-field motion and dark avoidance - are well-replicated in this virtual setting. We readily observed neuronal populations in the hindbrain with laterally selective responses that correlated with right or left optomotor behavior. We also observed neurons in the habenula, pallium, and midbrain with response properties specific to environmental features. Beyond single-cell correlations, the classification of network activity in such virtual settings promises to reveal principles of brainwide neural dynamics during behavior.

  9. Two-photon calcium imaging during fictive navigation in virtual environments.

    Science.gov (United States)

    Ahrens, Misha B; Huang, Kuo Hua; Narayan, Sujatha; Mensh, Brett D; Engert, Florian

    2013-01-01

    A full understanding of nervous system function requires recording from large populations of neurons during naturalistic behaviors. Here we enable paralyzed larval zebrafish to fictively navigate two-dimensional virtual environments while we record optically from many neurons with two-photon imaging. Electrical recordings from motor nerves in the tail are decoded into intended forward swims and turns, which are used to update a virtual environment displayed underneath the fish. Several behavioral features-such as turning responses to whole-field motion and dark avoidance-are well-replicated in this virtual setting. We readily observed neuronal populations in the hindbrain with laterally selective responses that correlated with right or left optomotor behavior. We also observed neurons in the habenula, pallium, and midbrain with response properties specific to environmental features. Beyond single-cell correlations, the classification of network activity in such virtual settings promises to reveal principles of brainwide neural dynamics during behavior.

  10. Ultra-bright and -stable red and near-infrared squaraine fluorophores for in vivo two-photon imaging.

    Science.gov (United States)

    Podgorski, Kaspar; Terpetschnig, Ewald; Klochko, Oleksii P; Obukhova, Olena M; Haas, Kurt

    2012-01-01

    Fluorescent dyes that are bright, stable, small, and biocompatible are needed for high-sensitivity two-photon imaging, but the combination of these traits has been elusive. We identified a class of squaraine derivatives with large two-photon action cross-sections (up to 10,000 GM) at near-infrared wavelengths critical for in vivo imaging. We demonstrate the biocompatibility and stability of a red-emitting squaraine-rotaxane (SeTau-647) by imaging dye-filled neurons in vivo over 5 days, and utility for sensitive subcellular imaging by synthesizing a specific peptide-conjugate label for the synaptic protein PSD-95.

  11. Ultra-bright and -stable red and near-infrared squaraine fluorophores for in vivo two-photon imaging.

    Directory of Open Access Journals (Sweden)

    Kaspar Podgorski

    Full Text Available Fluorescent dyes that are bright, stable, small, and biocompatible are needed for high-sensitivity two-photon imaging, but the combination of these traits has been elusive. We identified a class of squaraine derivatives with large two-photon action cross-sections (up to 10,000 GM at near-infrared wavelengths critical for in vivo imaging. We demonstrate the biocompatibility and stability of a red-emitting squaraine-rotaxane (SeTau-647 by imaging dye-filled neurons in vivo over 5 days, and utility for sensitive subcellular imaging by synthesizing a specific peptide-conjugate label for the synaptic protein PSD-95.

  12. Steady state anisotropy two-photon microscopy resolves multiple, spectrally similar fluorophores, enabling in vivo multilabel imaging.

    Science.gov (United States)

    Dubach, J Matthew; Vinegoni, Claudio; Weissleder, Ralph

    2014-08-01

    The use of spectrally distinguishable fluorescent dyes enables imaging of multiple targets. However, in two-photon microscopy, the number of fluorescent labels with distinct emission spectra that can be effectively excited and resolved is constrained by the confined tuning range of the excitation laser and the broad and overlapping nature of fluorophore two-photon absorption spectra. This limitation effectively reduces the number of available imaging channels. Here, we demonstrate that two-photon steady state anisotropy imaging (2PSSA) offers the capability to resolve otherwise unresolvable fluorescent tracers both in live cells and in mouse tumor models. This approach expands the number of biological targets that can be imaged simultaneously, increasing the total amount of information that can be obtained through imaging.

  13. Multispot two-photon imaging of mice heart tissue detecting calcium waves

    Science.gov (United States)

    de Mauro, C.; Cecchetti, C. A.; Alfieri, D.; Borile, G.; Mongillo, M.; Pavone, F. S.

    2012-06-01

    High rate, full field image acquisition in multiphoton imaging is achievable by parallelization of the excitation and of the detection paths. Via a Diffractive Optical Elements (DOEs) which splits a pulsed laser, and a spatial resolved descanned detection path, a new approach to microscopy has been developed. By exploiting the three operating mode, single beam, 16 beamlets or 64 beamlets, the best experimental conditions can be found by adapting the power per beamlet. This Multiphoton Multispot system (MCube) has been characterized in thick tissue samples, and subsequently used for the first time for Ca2+ imaging of acute heart slices. A test sample with fixed mice heart slices with embedded sub-resolution fluorescent beads has been used to test the capability of optical axial resolution up to ~200 microns in depth. Radial and axial resolutions of 0.6 microns and 3 microns have been respectively obtained with a 40X water immersion objective, getting close to the theoretical limit. Then images of heart slices cardiomyocites, loaded with Fluo4-AM have been acquired. The formation of Ca2+ waves during electrostimulated beating has been observed, and the possibility of easily acquire full frame images at 15 Hz (16 beamlets) has been demonstrated, towards the in vivo study of time resolved cellular dynamics and arrhythmia trigger mechanisms in particular. A very high speed two-photon Random Access system for in vivo electrophysiological studies, towards the correlation of voltage and calcium signals in arrhythmia phenomena, is now under developing at Light4tech.

  14. Time-lapse seismic within reservoir engineering

    NARCIS (Netherlands)

    Oldenziel, T.

    2003-01-01

    Time-lapse 3D seismic is a fairly new technology allowing dynamic reservoir characterisation in a true volumetric sense. By investigating the differences between multiple seismic surveys, valuable information about changes in the oil/gas reservoir state can be captured. Its interpretation involves d

  15. Time-lapse seismic within reservoir engineering

    NARCIS (Netherlands)

    Oldenziel, T.

    2003-01-01

    Time-lapse 3D seismic is a fairly new technology allowing dynamic reservoir characterisation in a true volumetric sense. By investigating the differences between multiple seismic surveys, valuable information about changes in the oil/gas reservoir state can be captured. Its interpretation involves d

  16. Time-lapse seismic within reservoir engineering

    NARCIS (Netherlands)

    Oldenziel, T.

    2003-01-01

    Time-lapse 3D seismic is a fairly new technology allowing dynamic reservoir characterisation in a true volumetric sense. By investigating the differences between multiple seismic surveys, valuable information about changes in the oil/gas reservoir state can be captured. Its interpretation involves

  17. Ultra-thin rigid endoscope: Two-photon imaging through a graded-index multi-mode fiber

    CERN Document Server

    Sivankutty, Siddharth; Cossart, Rosa; Bouwmans, Géraud; Monneret, Serge; Rigneault, Hervé

    2015-01-01

    Rigid endoscopes like graded-index (GRIN) lenses are known tools in biological imaging, but it is conceptually difficult to miniaturize them. In this letter, we demonstrate an ultra-thin rigid endoscope with a diameter of only 125 microns. In addition, we identify a domain where two-photon endoscopic imaging with fs-pulse excitation is possible. We validate the ultra-thin rigid endoscope consisting of a few cm of graded-index multi-mode fiber by using it to acquire optically sectioned two-photon fluorescence endoscopic images of three-dimensional samples.

  18. Advances in two photon scanning and scanless microscopy technologies for functional neural circuit imaging.

    Science.gov (United States)

    Schultz, Simon R; Copeland, Caroline S; Foust, Amanda J; Quicke, Peter; Schuck, Renaud

    2017-01-01

    Recent years have seen substantial developments in technology for imaging neural circuits, raising the prospect of large scale imaging studies of neural populations involved in information processing, with the potential to lead to step changes in our understanding of brain function and dysfunction. In this article we will review some key recent advances: improved fluorophores for single cell resolution functional neuroimaging using a two photon microscope; improved approaches to the problem of scanning active circuits; and the prospect of scanless microscopes which overcome some of the bandwidth limitations of current imaging techniques. These advances in technology for experimental neuroscience have in themselves led to technical challenges, such as the need for the development of novel signal processing and data analysis tools in order to make the most of the new experimental tools. We review recent work in some active topics, such as region of interest segmentation algorithms capable of demixing overlapping signals, and new highly accurate algorithms for calcium transient detection. These advances motivate the development of new data analysis tools capable of dealing with spatial or spatiotemporal patterns of neural activity, that scale well with pattern size.

  19. Imaging-guided two-photon excitation-emission-matrix measurements of human skin tissues

    Science.gov (United States)

    Yu, Yingqiu; Lee, Anthony M. D.; Wang, Hequn; Tang, Shuo; Zhao, Jianhua; Lui, Harvey; Zeng, Haishan

    2012-07-01

    There are increased interests on using multiphoton imaging and spectroscopy for skin tissue characterization and diagnosis. However, most studies have been done with just a few excitation wavelengths. Our objective is to perform a systematic study of the two-photon fluorescence (TPF) properties of skin fluorophores, normal skin, and diseased skin tissues. A nonlinear excitation-emission-matrix (EEM) spectroscopy system with multiphoton imaging guidance was constructed. A tunable femtosecond laser was used to vary excitation wavelengths from 730 to 920 nm for EEM data acquisition. EEM measurements were performed on excised fresh normal skin tissues, seborrheic keratosis tissue samples, and skin fluorophores including: NADH, FAD, keratin, melanin, collagen, and elastin. We found that in the stratum corneum and upper epidermis of normal skin, the cells have large sizes and the TPF originates from keratin. In the lower epidermis, cells are smaller and TPF is dominated by NADH contributions. In the dermis, TPF is dominated by elastin components. The depth resolved EEM measurements also demonstrated that keratin structure has intruded into the middle sublayers of the epidermal part of the seborrheic keratosis lesion. These results suggest that the imaging guided TPF EEM spectroscopy provides useful information for the development of multiphoton clinical devices for skin disease diagnosis.

  20. Measurement of drug-target engagement in live cells by two-photon fluorescence anisotropy imaging.

    Science.gov (United States)

    Vinegoni, Claudio; Fumene Feruglio, Paolo; Brand, Christian; Lee, Sungon; Nibbs, Antoinette E; Stapleton, Shawn; Shah, Sunil; Gryczynski, Ignacy; Reiner, Thomas; Mazitschek, Ralph; Weissleder, Ralph

    2017-07-01

    The ability to directly image and quantify drug-target engagement and drug distribution with subcellular resolution in live cells and whole organisms is a prerequisite to establishing accurate models of the kinetics and dynamics of drug action. Such methods would thus have far-reaching applications in drug development and molecular pharmacology. We recently presented one such technique based on fluorescence anisotropy, a spectroscopic method based on polarization light analysis and capable of measuring the binding interaction between molecules. Our technique allows the direct characterization of target engagement of fluorescently labeled drugs, using fluorophores with a fluorescence lifetime larger than the rotational correlation of the bound complex. Here we describe an optimized protocol for simultaneous dual-channel two-photon fluorescence anisotropy microscopy acquisition to perform drug-target measurements. We also provide the necessary software to implement stream processing to visualize images and to calculate quantitative parameters. The assembly and characterization part of the protocol can be implemented in 1 d. Sample preparation, characterization and imaging of drug binding can be completed in 2 d. Although currently adapted to an Olympus FV1000MPE microscope, the protocol can be extended to other commercial or custom-built microscopes.

  1. A two-photon fluorescent probe with a large turn-on signal for imaging hydrogen sulfide in living tissues

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Kaibo [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082 (China); Lin, Weiying, E-mail: weiyinglin2013@163.com [Institute of Fluorescent Probes for Biological Imaging, University of Jinan, Jinan, Shandong 250022 (China); State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082 (China); Tan, Li; Cheng, Dan [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082 (China)

    2015-01-01

    Highlights: • A two-photon fluorescent probe for sensing H{sub 2}S was developed. • The probe shows a large turn on signal (120-fold enhancement). • The probe is suitable for fluorescence imaging of H{sub 2}S in living cells and tissues. • The probe was capable of detecting H{sub 2}S up to 170 μm depth in live tissues. - Abstract: A two-photon fluorescence turn-on H{sub 2}S probe GCTPOC–H{sub 2}S based on a two-photon platform with a large cross-section, GCTPOC, and a sensitive H{sub 2}S recognition site, dinitrophenyl ether was constructed. The probe GCTPOC–H{sub 2}S exhibits desirable properties such as high sensitivity, high selectivity, functioning well at physiological pH and low cytotoxicity. In particular, the probe shows a 120-fold enhancement in the presence of Na{sub 2}S (500 μM), which is larger than the reported two-photon fluorescent H{sub 2}S probes. The large fluorescence enhancement of the two-photon probe GCTPOC–H{sub 2}S renders it attractive for imaging H{sub 2}S in living tissues with deep tissue penetration. Significantly, we have demonstrated that the probe GCTPOC–H{sub 2}S is suitable for fluorescence imaging of H{sub 2}S in living tissues with deep penetration by using two-photon microscopy. The further application of the two-photon probe for the investigation of biological functions and pathological roles of H{sub 2}S in living systems is under progress.

  2. Enhanced multi-spectral imaging of live breast cancer cells using immunotargeted gold nanoshells and two-photon excitation microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Bickford, Lissett; Sun Jiantang; Fu, Kun; Lewinski, Nastassja; Nammalvar, Vengadesan; Chang, Joseph; Drezek, Rebekah [Department of Bioengineering, Rice University, Houston, TX 77005 (United States)], E-mail: drezek@rice.edu

    2008-08-06

    We demonstrate the capability of using immunotargeted gold nanoshells as contrast agents for in vitro two-photon microscopy. The two-photon luminescence properties of different-sized gold nanoshells are first validated using near-infrared excitation at 780 nm. The utility of two-photon microscopy as a tool for imaging live HER2-overexpressing breast cancer cells labeled with anti-HER2-conjugated nanoshells is then explored and imaging results are compared to normal breast cells. Five different imaging channels are simultaneously examined within the emission wavelength range of 451-644 nm. Our results indicate that under near-infrared excitation, superior contrast of SK-BR-3 cancer cells labeled with immunotargeted nanoshells occurs at an emission wavelength ranging from 590 to 644 nm. Luminescence from labeled normal breast cells and autofluorescence from unlabeled cancer and normal cells remain imperceptible under the same conditions.

  3. Enhanced multi-spectral imaging of live breast cancer cells using immunotargeted gold nanoshells and two-photon excitation microscopy

    Science.gov (United States)

    Bickford, Lissett; Sun, Jiantang; Fu, Kun; Lewinski, Nastassja; Nammalvar, Vengadesan; Chang, Joseph; Drezek, Rebekah

    2008-08-01

    We demonstrate the capability of using immunotargeted gold nanoshells as contrast agents for in vitro two-photon microscopy. The two-photon luminescence properties of different-sized gold nanoshells are first validated using near-infrared excitation at 780 nm. The utility of two-photon microscopy as a tool for imaging live HER2-overexpressing breast cancer cells labeled with anti-HER2-conjugated nanoshells is then explored and imaging results are compared to normal breast cells. Five different imaging channels are simultaneously examined within the emission wavelength range of 451-644 nm. Our results indicate that under near-infrared excitation, superior contrast of SK-BR-3 cancer cells labeled with immunotargeted nanoshells occurs at an emission wavelength ranging from 590 to 644 nm. Luminescence from labeled normal breast cells and autofluorescence from unlabeled cancer and normal cells remain imperceptible under the same conditions.

  4. Visualizing heterogeneity of photosynthetic properties of plant leaves with two-photon fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Iermak, Ievgeniia; Vink, Jochem; Bader, Arjen N; Wientjes, Emilie; van Amerongen, Herbert

    2016-09-01

    Two-photon fluorescence lifetime imaging microscopy (FLIM) was used to analyse the distribution and properties of Photosystem I (PSI) and Photosystem II (PSII) in palisade and spongy chloroplasts of leaves from the C3 plant Arabidopsis thaliana and the C4 plant Miscanthus x giganteus. This was achieved by separating the time-resolved fluorescence of PSI and PSII in the leaf. It is found that the PSII antenna size is larger on the abaxial side of A. thaliana leaves, presumably because chloroplasts in the spongy mesophyll are "shaded" by the palisade cells. The number of chlorophylls in PSI on the adaxial side of the A. thaliana leaf is slightly higher. The C4 plant M. x giganteus contains both mesophyll and bundle sheath cells, which have a different PSI/PSII ratio. It is shown that the time-resolved fluorescence of bundle sheath and mesophyll cells can be analysed separately. The relative number of chlorophylls, which belong to PSI (as compared to PSII) in the bundle sheath cells is at least 2.5 times higher than in mesophyll cells. FLIM is thus demonstrated to be a useful technique to study the PSI/PSII ratio and PSII antenna size in well-defined regions of plant leaves without having to isolate pigment-protein complexes.

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

    Science.gov (United States)

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

    2013-03-01

    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 (λex=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.

  6. Two-photon absorption-induced photoacoustic imaging of Rhodamine B dyed polyethylene spheres using a femtosecond laser.

    Science.gov (United States)

    Langer, Gregor; Bouchal, Klaus-Dieter; Grün, Hubert; Burgholzer, Peter; Berer, Thomas

    2013-09-23

    In the present paper we demonstrate the possibility to image dyed solids, i.e. Rhodamine B dyed polyethylene spheres, by means of two-photon absorption-induced photoacoustic scanning microscopy. A two-photon luminescence image is recorded simultaneously with the photoacoustic image and we show that location and size of the photoacoustic and luminescence image match. In the experiments photoacoustic signals and luminescence signals are generated by pulses from a femtosecond laser. Photoacoustic signals are acquired with a hydrophone; luminescence signals with a spectrometer or an avalanche photo diode. In addition we derive the expected dependencies between excitation intensity and photoacoustic signal for single-photon absorption, two-photon absorption and for the combination of both. In order to verify our setup and evaluation method the theoretical predictions are compared with experimental results for liquid and solid specimens, i.e. a carbon fiber, Rhodamine B solution, silicon, and Rhodamine B dyed microspheres. The results suggest that the photoacoustic signals from the Rhodamine B dyed microspheres do indeed stem from two-photon absorption.

  7. Second harmonic imaging of plants tissues and cell implosion using two-photon process in ZnO nanoparticles.

    Science.gov (United States)

    Urban, Ben E; Neogi, Purnima B; Butler, Sween J; Fujita, Yasuhisa; Neogi, Arup

    2012-03-01

    The optical properties of colloidal ZnO nanoparticle (NP) solutions, with size ranging from several nm to around 200 nm, have been tailored to have high optical nonlinearity for bioimaging with no auto-fluorescence above 750 nm and minimal auto-fluorescence below 750 nm. The high second harmonic conversion efficiency enables selective tissue imaging and cell tracking using tunable near-infrared femtosecond laser source ranging from 750-980 nm. For laser energies exceeding the two-photon energy of the bandgap of ZnO (half of 3.34 eV), the SHG signal greatly decreases and the two-photon emission becomes the dominant signal. The heat generated due to two-photon absorption within the ZnO NPs enable selective cell or localized tissue destruction using excitation wavelength ranging from 710-750 nm. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Time Lapse Camera Images for Observation and Visualization of Snow Lines on two Benchmark Glaciers in High Asia - Zhadang and Halji

    Science.gov (United States)

    Schröter, B.; Buchroithner, M. F.; Pieczonka, T.

    2015-12-01

    Glaciers are characteristic elements of high mountain environments and represent key indicators for the ongoing climate change. The covering snowpack considerably affects the glacier-ice surface temperature and thus the meltdown of the glaciers which in recent decades has been accelerating worldwide. Therefore, the detailed investigation of the transient snow is of high importance. Zhadang Glacier is located in the Nyainqentanglha Mountain Range on the central part of the Tibetan Plateau (30°28.24' N, 90°38.69' E). The glacier is debris-free and flows from 6,090 to 5,515 m a.s.l. Recent measurements have shown that the whole glacier is below the ELA and experiences significant glacier volume loss. In May 2010 two terrestrial cameras had been installed there and were continuously operating until September 2012 generating 6,225 images of the glacier with a frequency of three resp. six images per day. In order to use this dataset for snow line mapping all images had to be georeferenced and orthorectified. The biggest challenge was the problem of shifting camera positions due to deformations of the ground and hence the offset in the image coordinates. This was resolved by combining the manual orthorectification of one image per week with a subsequent spline interpolation to determine the changed image coordinates. The actual orthorectification was finally realized by applying a fully automated batch processing of all images. The most favorable image of each day was chosen for the manual snow line mapping process. The final aim was the calculation of the mean elevation of the snow line for every day of the data collecting period materialized by intersecting the mapped snow lines with resampled SRTM 3 data. Considering the fact that there were several weeks either with full snow cover or without any snow this aim could be achieved. The findings have been used for the evaluation of a glacier mass balance model developed at RWTH Aachen, Germany, showing a high level of

  9. Multicolor excitation two-photon microscopy: in vivo imaging of cells and tissues

    Science.gov (United States)

    Li, Dong; Zheng, Wei; Qu, Jianan Y.

    2010-02-01

    Two-photon microscopy based on endogenous fluorescence provides non-invasive imaging of living biological system. Reduced nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD), keratin, collagen and elastin are the endogenous fluorophores widely used as the contrast agents for imaging metabolism and morphology of living cells and tissue. The fluorescence of tryptophan, a kind of essential amino acid, conveys the information on cellular protein content, structure and microenvironment. However, it can't be effectively excited by the commonly used Ti:sapphire femtosecond laser. Because each endogenous fluorophore provides limited information, it is desirable to simultaneously excite fluorescence from as many fluorophores as possible to obtain accurate biochemical and morphological information on biomedical samples. In this study, we demonstrate that the supercontinuum generation from a photonic crystal fiber (PCF) excited by an ultrafast source can be used to excite multiple endogenous nonlinear optical signals simultaneously. By employing the spectral lifetime detection capability, this technology provides a unique approach to sense the fine structure, protein distribution and cellular metabolism of cells and tissues in vivo. In particular, with application of acetic acid, a safe contrast agent used for detection cervical cancer for many years, the tryptophan signals reveal cellular morphology and even cell-cell junctions clearly. Moreover, it was found that the pH value dependent lifetime of tryptophan fluorescence could provide the qualitative information on the gradient of pH value in epithelial tissue. Finally, we will demonstrate the potential of our multi-color TPEF microscopy to investigate the early development of cancer in epithelial tissue.

  10. Two-photon fluorescence and second-harmonic generation imaging of collagen in human tissue based on multiphoton microscopy.

    Science.gov (United States)

    Jiang, Xingshan; Zhong, Jiazhao; Liu, Yuchun; Yu, Haibo; Zhuo, Shuangmu; Chen, Jianxin

    2011-01-01

    Multiphoton microscopic imaging of collagen plays an important role in noninvasive diagnoses of human tissue. In this study, two-photon fluorescence and second-harmonic generation (SHG) imaging of collagen in human skin dermis and submucosa of colon and stomach tissues were investigated based on multiphoton microscopy (MPM). Our results show that multiphoton microscopic image of collagen bundles exhibits apparently different pattern in human tissues. The collagen bundles can simultaneously reveal its SHG and two-photon excited fluorescence images in the submucosa of colon and stomach, whereas it solely emit SHG signal in skin dermis. The intensity spectral information from tissues further demonstrated the above results. This indicates that collagen bundles have completely different space arrangement in these tissues. Our experimental results bring more detailed information of collagen for the application of MPM in human noninvasive imaging. Copyright © 2011 Wiley Periodicals, Inc.

  11. Cellular imaging of deep organ using two-photon Bessel light-sheet nonlinear structured illumination microscopy

    Science.gov (United States)

    Zhao, Ming; Zhang, Han; Li, Yu; Ashok, Amit; Liang, Rongguang; Zhou, Weibin; Peng, Leilei

    2014-01-01

    In vivo fluorescent cellular imaging of deep internal organs is highly challenging, because the excitation needs to penetrate through strong scattering tissue and the emission signal is degraded significantly by photon diffusion induced by tissue-scattering. We report that by combining two-photon Bessel light-sheet microscopy with nonlinear structured illumination microscopy (SIM), live samples up to 600 microns wide can be imaged by light-sheet microscopy with 500 microns penetration depth, and diffused background in deep tissue light-sheet imaging can be reduced to obtain clear images at cellular resolution in depth beyond 200 microns. We demonstrate in vivo two-color imaging of pronephric glomeruli and vasculature of zebrafish kidney, whose cellular structures located at the center of the fish body are revealed in high clarity by two-color two-photon Bessel light-sheet SIM. PMID:24876996

  12. Cellular imaging of deep organ using two-photon Bessel light-sheet nonlinear structured illumination microscopy.

    Science.gov (United States)

    Zhao, Ming; Zhang, Han; Li, Yu; Ashok, Amit; Liang, Rongguang; Zhou, Weibin; Peng, Leilei

    2014-05-01

    In vivo fluorescent cellular imaging of deep internal organs is highly challenging, because the excitation needs to penetrate through strong scattering tissue and the emission signal is degraded significantly by photon diffusion induced by tissue-scattering. We report that by combining two-photon Bessel light-sheet microscopy with nonlinear structured illumination microscopy (SIM), live samples up to 600 microns wide can be imaged by light-sheet microscopy with 500 microns penetration depth, and diffused background in deep tissue light-sheet imaging can be reduced to obtain clear images at cellular resolution in depth beyond 200 microns. We demonstrate in vivo two-color imaging of pronephric glomeruli and vasculature of zebrafish kidney, whose cellular structures located at the center of the fish body are revealed in high clarity by two-color two-photon Bessel light-sheet SIM.

  13. 时差成像系统在IVF-ET的临床应用进展%Progress on clinical application of time-lapse imaging in IVF-ET

    Institute of Scientific and Technical Information of China (English)

    武丽红; 韩伟; 黄国宁

    2015-01-01

    时差成像系统(time-lapse imaging,TLI)作为一种新的胚胎选择体系,越来越受到国内、外辅助生殖领域临床医生与胚胎学家的青睐.近年来,TLI应用于体外受精(in vitro fertilization,IVF)中的临床研究也逐渐增多.胚胎学家应用这一系统后不需将胚胎从培养箱中移出即可通过视频图像连续观察其整个动态发育过程,以及获得预测胚胎发育潜能的动力学参数.应用TLI提供的动力学参数结合形态学选择移植胚胎,可以改善胚胎着床率和临床妊娠率.但TLI技术无论从硬件还是软件方面均面临着巨大挑战;另外,TLI系统的安全性亦有待进一步评估.

  14. Label-free imaging immune cells and collagen in atherosclerosis with two-photon and second harmonic generation microscopy

    Directory of Open Access Journals (Sweden)

    Chunqiang Li

    2016-01-01

    Full Text Available Atherosclerosis has been recognized as a chronic inflammation disease, in which many types of cells participate in this process, including lymphocytes, macrophages, dendritic cells (DCs, mast cells, vascular smooth muscle cells (SMCs. Developments in imaging technology provide the capability to observe cellular and tissue components and their interactions. The knowledge of the functions of immune cells and their interactions with other cell and tissue components will facilitate our discovery of biomarkers in atherosclerosis and prediction of the risk factor of rupture-prone plaques. Nonlinear optical microscopy based on two-photon excited autofluorescence and second harmonic generation (SHG were developed to image mast cells, SMCs and collagen in plaque ex vivo using endogenous optical signals. Mast cells were imaged with two-photon tryptophan autofluorescence, SMCs were imaged with two-photon NADH autofluorescence, and collagen were imaged with SHG. This development paves the way for further study of mast cell degranulation, and the effects of mast cell derived mediators such as induced synthesis and activation of matrix metalloproteinases (MMPs which participate in the degradation of collagen.

  15. Light-induced damage and its diagnosis in two-photon excited autofluorescence imaging of retinal pigment epithelium cells

    Science.gov (United States)

    Chen, Danni; Qu, Junle; Xu, Gaixia; Zhao, Lingling; Niu, Hanben

    2007-05-01

    In this paper, a novel method for the differentiation of the retinal pigment epithelium (RPE) cells after light-induced damage by two-photon excitation is presented. Fresh samples of RPE cells of pig eyes are obtained from local slaughterhouse. Light-induced damage is produced by the output from Ti: sapphire laser which is focused onto the RPE layer. We study the change of the autofluorescence properties of RPE after two-photon excitation with the same wavelength. Preliminary results show that after two-photon excitation, there are two clear changes in the emission spectrum. The first change is the blue-shift of the emission peak. The emission peak of the intact RPE is located at 592nm, and after excitation, it shifts to 540nm. It is supposed that the excitation has led to the increased autofluorescence of flavin whose emission peak is located at 540nm. The second change is the increased intensity of the emission peak, which might be caused by the accelerated aging because the autofluorescence of RPE would increase during aging process. Experimental results indicate that two-photon excitation could not only lead to the damage of the RPE cells in multiphoton RPE imaging, but also provide an evaluation of the light-induced damage.

  16. Time-lapse microscopy of lung endothelial cells under hypoxia

    Science.gov (United States)

    Mehrvar, Shima; Ghanian, Zahra; Kondouri, Ganesh; Camara, Amadou S.; Ranji, Mahsa

    2017-02-01

    Objective: This study utilizes fluorescence microscopy to assess the effect of the oxygen tension on the production of reactive oxygen species (ROS) in mitochondria of fetal pulmonary artery endothelial cells (FPAECs). Introduction: Hypoxia is a severe oxygen stress, which mostly causes irreversible injury in lung cells. However, in some studies, it is reported that hypoxia decreases the severity of injuries. In this study, ROS production level was examined in hypoxic FPAECs treated with pentachlorophenol (PCP, uncoupler). This work was accomplished by monitoring and quantifying the changes in the level of the produced ROS in hypoxic cells before and after PCP treatment. Materials and methods: The dynamic of the mitochondrial ROS production in two groups of FPAECs was measured over time using time-lapse microscopy. For the first group, cells were incubated in 3% hypoxic condition for 2 hours and then continuously were exposed to hypoxic condition for imaging as well. For the second group, cells were incubated in normal oxygen condition. Time lapse images of the cells loaded with Mito-SOX (ROS indicator) were acquired, and the red fluorescence intensity profile of the cells was calculated. Changes in the level of the fluorescence intensity profile while they are treated with PCP indicates the dynamics of the ROS level. Results: The intensity profiles of the PCP-treated cells in the first group showed 47% lower ROS production rate than the PCP-treated cells in the second group. Conclusion: Time lapse microscopy revealed that hypoxic cells have lower ROS generation while treated with PCP. Therefore, this result suggests that hypoxia decreased electron transport chain activity in uncoupled chain.

  17. Identification of biogeochemical hot spots using time-lapse hydrogeophysics

    Science.gov (United States)

    Franz, T. E.; Loecke, T.; Burgin, A.

    2016-12-01

    The identification and monitoring of biogeochemical hot spots and hot moments is difficult using point based sampling techniques and sensors. Without proper monitoring and accounting of water, energy, and trace gas fluxes it is difficult to assess the environmental footprint of land management practices. One key limitation is optimal placement of sensors/chambers that adequately capture the point scale fluxes and thus a reasonable integration to landscape scale flux. In this work we present time-lapse hydrogeophysical imaging at an old agricultural field converted into a wetland mitigation bank near Dayton, Ohio. While the wetland was previously instrumented with a network of soil sensors and surface chambers to capture a suite of state variables and fluxes, we hypothesize that time-lapse hydrogeophysical imaging is an underutilized and critical reconnaissance tool for effective network design and landscape scaling. Here we combine the time-lapse hydrogeophysical imagery with the multivariate statistical technique of Empirical Orthogonal Functions (EOF) in order to isolate the spatial and temporal components of the imagery. Comparisons of soil core information (e.g. soil texture, soil carbon) from around the study site and organized within like spatial zones reveal statistically different mean values of soil properties. Moreover, the like spatial zones can be used to identify a finite number of future sampling locations, evaluation of the placement of existing sensors/chambers, upscale/downscale observations, all of which are desirable techniques for commercial use in precision agriculture. Finally, we note that combining the EOF analysis with continuous monitoring from point sensors or remote sensing products may provide a robust statistical framework for scaling observations through time as well as provide appropriate datasets for use in landscape biogeochemical models.

  18. Imaging of Fluoride Ion in Living Cells and Tissues with a Two-Photon Ratiometric Fluorescence Probe

    Directory of Open Access Journals (Sweden)

    Xinyue Zhu

    2015-01-01

    Full Text Available A reaction-based two-photon (TP ratiometric fluorescence probe Z2 has been developed and successfully applied to detect and image fluoride ion in living cells and tissues. The Z2 probe was designed designed to utilize an ICT mechanism between n-butylnaphthalimide as a fluorophore and tert-butyldiphenylsilane (TBDPS as a response group. Upon addition of fluoride ion, the Si-O bond in the Z2 would be cleaved, and then a stronger electron-donating group was released. The fluorescent changes at 450 and 540 nm, respectively, made it possible to achieve ratiometric fluorescence detection. The results indicated that the Z2 could ratiometrically detect and image fluoride ion in living cells and tissues in a depth of 250 μm by two-photon microscopy (TPM.

  19. Mouse tail vertebrae adapt to cyclic mechanical loading by increasing bone formation rate and decreasing bone resorption rate as shown by time-lapsed in vivo imaging of dynamic bone morphometry.

    Science.gov (United States)

    Lambers, Floor M; Schulte, Friederike A; Kuhn, Gisela; Webster, Duncan J; Müller, Ralph

    2011-12-01

    It is known that mechanical loading leads to an increase in bone mass through a positive shift in the balance between bone formation and bone resorption. How the remodeling sites change over time as an effect of loading remains, however, to be clarified. The purpose of this paper was to investigate how bone formation and resorption sites are modulated by mechanical loading over time by using a new imaging technique that extracts three dimensional formation and resorption parameters from time-lapsed in vivo micro-computed tomography images. To induce load adaptation, the sixth caudal vertebra of C57BL/6 mice was cyclically loaded through pins in the adjacent vertebrae at either 8 N or 0 N (control) three times a week for 5 min (3000 cycles) over a total of 4 weeks. The results showed that mechanical loading significantly increased trabecular bone volume fraction by 20% (pbone formation rate was on average 23% greater (pbone resorption rate was on average 25% smaller (pbone formation rate for the 8 N group was mostly an effect of a significantly increased surface of bone formation sites (on average 16%, pbone formation packages was less affected (on average 5% greater, pbone resorption sites was significantly reduced (on average 15%, pbone increased significantly by 24% (pbone decreased significantly by 24% (ptail vertebrae adapt to mechanical loading by increasing the surface of formation sites and decreasing the surface of resorption sites, leading to an overall increase in bone strength. This new imaging technique will provide opportunities to investigate in vivo bone remodeling in the context of disease and treatment options, with the added value that both bone formation and bone resorption parameters can be nondestructively calculated over time.

  20. A new endoplasmic reticulum-targeted two-photon fluorescent probe for imaging of superoxide anion in diabetic mice.

    Science.gov (United States)

    Xiao, Haibin; Liu, Xiao; Wu, Chuanchen; Wu, Yaohuan; Li, Ping; Guo, Xiaomeng; Tang, Bo

    2017-05-15

    Excessive or unfolded proteins accumulation in endoplasmic reticulum (ER) will cause ER stress, which has evolved to involve in various metabolic diseases. In particular, ER stress plays an important role in the pathogenesis of diabetes. Both ER stress and course of diabetes accompany oxidative stress and production of reactive oxygen species (ROS), among which superoxide anion (O2(•-)) is the first produced ROS and has been recognized as cell signaling mediator involved in the physiological and pathological process of diabetes. Hence, the development of effective monitoring methods of O2(•-) in live cells and in vivo is of great importance for ascertaining the onset and progress of related diseases. Herein, a new endoplasmic reticulum-targeted two-photon fluorescent probe termed ER-BZT is designed and synthesized for imaging of O2(•-). The probe ER-BZT shows high sensitivity, selectivity, stability, and low cytotoxicity. Based on these superior properties, the rise of O2(•-) levels in endoplasmic reticulum induced with different stimuli is visualized by one- and two-photon fluorescence imaging. Most importantly, by utilizing ER-BZT, the two-photon fluorescence imaging results demonstrate that the endogenous O2(•-) concentration in abdominal or hepatic tissue of diabetic mice is higher than that in normal mice. Meanwhile, after treated with metformin, a broad-spectrum antidiabetic drug, the diabetic mice exhibit depressed O2(•-) level. The proposed two-photon probe, ER-BZT might serve as perfect tool to image the O2(•-) fluctuations and study the relevance between O2(•-) and various diseases in live cells and in vivo.

  1. In Vivo Non Linear Optical (NLO) Imaging in Live Rabbit Eyes Using the Heidelberg Two-Photon Laser Ophthalmoscope

    Science.gov (United States)

    Hao, Ming; Flynn, Kevin; Nien-Shy, Chyong; Jester, Bryan E.; Winkler, Moritz; Brown, Donald J.; La Schiazza, Olivier; Bille, Josef; Jester, James V.

    2010-01-01

    Imaging of non-linear optical (NLO) signals generated from the eye using ultrafast pulsed lasers has been limited to the study of ex vivo tissues because of the use of conventional microscopes with slow scan speeds. The purpose of this study was to evaluate the ability of a novel, high scan rate ophthalmoscope to generate NLO signals using an attached femtosecond laser. NLO signals were generated and imaged in live, anesthetized albino rabbits using a newly designed Heidelberg Two-Photon Laser Ophthalmoscope with attached 25 mW femtosecond laser having a central wavelength of 780 nm, pulsewidth of 75 fs, and a repetition rate of 50 MHz. To assess two-photon excited fluorescent (TPEF) signal generation, cultured rabbit corneal fibroblasts (RCF) were first labeled by Blue-green fluorescent FluoSpheres (1 μm diameter) and then cells were micro-injected into the central cornea. Clumps of RCF cells could be detected by both reflectance and TPEF imaging at 6 hours after injection. By 6 days, RCF containing fluorescent microspheres confirmed by TPEF showed a more spread morphology and had migrated from the original injection site. Overall, this study demonstrates the potential of using NLO microscopy to sequentially detect TPEF signals from live, intact corneas. We conclude that further refinement of the Two-photon laser Ophthalmoscope should lead to the development of an important, new clinical instrument capable of detecting NLO signals from patient corneas. PMID:20558159

  2. Two-photon cryomicroscope

    Science.gov (United States)

    Breunig, H. G.; Köhler, C.; König, K.

    2012-03-01

    We report on a new two-photon cryomicroscope which consist of a compact laser-scanning microscope combined with a motorized heating and freezing stage. Samples can be cooled down to -196 °C (77 K) and heated up to 600 °C (873 K) with adjustable heating/freezing rates between 0.01 K / min and 150 K / min. Two-photon imaging is realized by near infrared femtosecond-laser pulse excitation. The abilities of the two-photon cryomicroscope are illustrated in several measurements: imaging of fluorescent microspheres inside a piece of ice illustrates the feasibility of deep-microscopic imaging inside frozen sample. The temperature-dependent structural integrity of collagen is monitored by detection of second harmonic generation signals from porcine cornea. The measurements reveal also the dependence of the collagendenaturation temperature on hydration state of the cornea collagen. Furthermore, the potential of the two-photon cryomicroscope for optimization of freezing and thawing procedures as well as to evaluate the viability of frozen cells and tissue is discussed.

  3. Estimating the hydrogeological parameters of an unconfined aquifer with the time-lapse resistivity-imaging method during pumping tests: Case studies at the Pengtsuo and Dajou sites, Taiwan

    Science.gov (United States)

    Chang, Ping-Yu; Chang, Lian-Cheng; Hsu, Shao-Yiu; Tsai, Jui-Pin; Chen, Wen-Fu

    2017-09-01

    We conducted time-lapse resistivity imaging during pumping tests at the Pengtsuo and Dajou test sites in Taiwan in order to examine the feasibility of estimating hydrogeological parameters with resistivity variations. Core logs reveal that the subsurface consists mainly of at least 100-m-thick gravel and sand at the two test sites. The resistivity differences between the pumping stages and pre-pumping background are well correlated to water level changes that are due to the dewatering of pumping activity. Therefore, it is possible to use the geometry of resistivity anomalies to estimate the hydraulic conductivity of the unconfined aquifer using the distance-drawdown equation for pumping tests in unsaturated aquifers. For each site, we used the contours of resistivity variations and recorded water levels in the pumping well to depict the bottom of the drawdown cone. The estimated hydraulic conductivity and specific yield, respectively, are 1.33 × 10- 4 m/s and 0.12 at the Pengtsuo site, and are 2.50 × 10- 4 m/s and 0.22 at the Dajou site. These values are consistent with the parameters that engineers from Taiwan Sugar Company calculated previously regarding groundwater-level variations in multiple wells (9.65 × 10- 5 m/s and 0.13 at Pengtsuo, and 1.00 × 10- 3 m/s and 0.19 at Dajou). This consistency suggests that resistivity imaging can perhaps serve as an alternative way to yield information about hydrogeological parameters.

  4. Label-free near-infrared reflectance microscopy as a complimentary tool for two-photon fluorescence brain imaging.

    Science.gov (United States)

    Allegra Mascaro, Anna Letizia; Costantini, Irene; Margoni, Emilia; Iannello, Giulio; Bria, Alessandro; Sacconi, Leonardo; Pavone, Francesco S

    2015-11-01

    In vivo two-photon imaging combined with targeted fluorescent indicators is currently extensively used for attaining critical insights into brain functionality and structural plasticity. Additional information might be gained from back-scattered photons from the near-infrared (NIR) laser without introducing any exogenous labelling. Here, we describe a complimentary and versatile approach that, by collecting the reflected NIR light, provides structural details on axons and blood vessels in the brain, both in fixed samples and in live animals under a cranial window. Indeed, by combining NIR reflectance and two-photon imaging of a slice of hippocampus from a Thy1-GFPm mouse, we show the presence of randomly oriented axons intermingled with sparsely fluorescent neuronal processes. The back-scattered photons guide the contextualization of the fluorescence structure within brain atlas thanks to the recognition of characteristic hippocampal structures. Interestingly, NIR reflectance microscopy allowed the label-free detection of axonal elongations over the superficial layers of mouse cortex under a cranial window in vivo. Finally, blood flow can be measured in live preparations, thus validating label free NIR reflectance as a tool for monitoring hemodynamic fluctuations. The prospective versatility of this label-free technique complimentary to two-photon fluorescence microscopy is demonstrated in a mouse model of photothrombotic stroke in which the axonal degeneration and blood flow remodeling can be investigated.

  5. NIR-to-NIR Two-Photon Scanning Laser Microscopy Imaging of Single Nanoparticles Doped by Yb(III) Complexes.

    Science.gov (United States)

    Bourdolle, Adrien; D'Aléo, Anthony; Philippot, Cécile; Baldeck, Patrice L; Guyot, Yannick; Dubois, Fabien; Ibanez, Alain; Andraud, Chantal; Brasselet, Sophie; Maury, Olivier

    2016-01-04

    The photophysical and nonlinear optical properties of water-soluble chromophore-functionalised tris-dipicolinate complexes [LnL3](3-) (Ln=Yb and Nd) are thoroughly studied, revealing that only the Yb(III) luminescence can be sensitized by a two-photon excitation process. The stability of the complex in water is strongly enhanced by embedding in dispersible organosilicate nanoparticles (NPs). Finally, the spectroscopic properties of [NBu4]3 [YbL3] are studied in solution and in the solid state. The high brightness of the NPs allows imaging them as single objects using a modified two-photon microscopy setup in a NIR-to-NIR configuration.

  6. Novel Bis-β-diketone-type Ligand and Its Copper and Zinc Complexes for Two-photon Biological Imaging

    Institute of Scientific and Technical Information of China (English)

    ZHOU Shuang-sheng; XUE Xuan; WEI Dong; JIANG Bo; WANG Jia-feng; LU Cheng-hua

    2012-01-01

    A curcumin derivative ligand,1,7-bis(3-methoxyl-4-oxyethylacetate)phenyl-1,6-heptadiene-3,5-diketone (diethyl acetatecurcumin,abbreviated as HL),and its Cu(Ⅱ) and Zn(Ⅱ) complexes have been synthesized and characterized by elemental analyses,infrared(IR),1H NMR and molar conductivity.The experimental results show that the resulting complexes bear strong two-photon excited fluorescence(TPEF) in N,N-dimethyformamide solvent,which has been proven to be potentially useful for two-photon microscopy imaging in living cells.In addition,cytotoxicity tests show that the low-micromolar concentrations of metal-ligand complex(ML2) did not cause significant reduction in cell viability over a pcriod of,at least,24 h and should be safe for further biological studies.

  7. Two-photon excitation with pico-second fluorescence lifetime imaging to detect nuclear association of flavanols.

    Science.gov (United States)

    Mueller-Harvey, Irene; Feucht, Walter; Polster, Juergen; Trnková, Lucie; Burgos, Pierre; Parker, Anthony W; Botchway, Stanley W

    2012-03-16

    Two-photon excitation enabled for the first time the observation and measurement of excited state fluorescence lifetimes from three flavanols in solution, which were ~1.0 ns for catechin and epicatechin, but <45 ps for epigallocatechin gallate (EGCG). The shorter lifetime for EGCG is in line with a lower fluorescence quantum yield of 0.003 compared to catechin (0.015) and epicatechin (0.018). In vivo experiments with onion cells demonstrated that tryptophan and quercetin, which tend to be major contributors of background fluorescence in plant cells, have sufficiently low cross sections for two-photon excitation at 630 nm and therefore do not interfere with detection of externally added or endogenous flavanols in Allium cepa or Taxus baccata cells. Applying two-photon excitation to flavanols enabled 3-D fluorescence lifetime imaging microscopy and showed that added EGCG penetrated the whole nucleus of onion cells. Interestingly, EGCG and catechin showed different lifetime behaviour when bound to the nucleus: EGCG lifetime increased from <45 to 200 ps, whilst catechin lifetime decreased from 1.0 ns to 500 ps. Semi-quantitative measurements revealed that the relative ratios of EGCG concentrations in nucleoli associated vesicles: nucleus: cytoplasm were ca. 100:10:1. Solution experiments with catechin, epicatechin and histone proteins provided preliminary evidence, via the appearance of a second lifetime (τ(2)=1.9-3.1 ns), that both flavanols may be interacting with histone proteins. We conclude that there is significant nuclear absorption of flavanols. This advanced imaging using two-photon excitation and biophysical techniques described here will prove valuable for probing the intracellular trafficking and functions of flavanols, such as EGCG, which is the major flavanol of green tea.

  8. Semi-automated discrimination of retinal pigmented epithelial cells in two-photon fluorescence images of mouse retinas

    Science.gov (United States)

    Alexander, Nathan S.; Palczewska, Grazyna; Palczewski, Krzysztof

    2015-01-01

    Automated image segmentation is a critical step toward achieving a quantitative evaluation of disease states with imaging techniques. Two-photon fluorescence microscopy (TPM) has been employed to visualize the retinal pigmented epithelium (RPE) and provide images indicating the health of the retina. However, segmentation of RPE cells within TPM images is difficult due to small differences in fluorescence intensity between cell borders and cell bodies. Here we present a semi-automated method for segmenting RPE cells that relies upon multiple weak features that differentiate cell borders from the remaining image. These features were scored by a search optimization procedure that built up the cell border in segments around a nucleus of interest. With six images used as a test, our method correctly identified cell borders for 69% of nuclei on average. Performance was strongly dependent upon increasing retinosome content in the RPE. TPM image analysis has the potential of providing improved early quantitative assessments of diseases affecting the RPE. PMID:26309765

  9. Glucagon-Secreting Alpha Cell Selective Two-Photon Fluorescent Probe TP-α: For Live Pancreatic Islet Imaging.

    Science.gov (United States)

    Agrawalla, Bikram Keshari; Chandran, Yogeswari; Phue, Wut-Hmone; Lee, Sung-Chan; Jeong, Yun-Mi; Wan, Si Yan Diana; Kang, Nam-Young; Chang, Young-Tae

    2015-04-29

    Two-photon (TP) microscopy has an advantage for live tissue imaging which allows a deeper tissue penetration up to 1 mm comparing to one-photon (OP) microscopy. While there are several OP fluorescence probes in use for pancreatic islet imaging, TP imaging of selective cells in live islet still remains a challenge. Herein, we report the discovery of first TP live pancreatic islet imaging probe; TP-α (Two Photon-alpha) which can selectively stain glucagon secreting alpha cells. Through fluorescent image based screening using three pancreatic cell lines, we discovered TP-α from a TP fluorescent dye library TPG (TP-Green). In vitro fluorescence test showed that TP-α have direct interaction and appear glucagon with a significant fluorescence increase, but not with insulin or other hormones/analytes. Finally, TP-α was successfully applied for 3D imaging of live islets by staining alpha cell directly. The newly developed TP-α can be a practical tool to evaluate and identify live alpha cells in terms of localization, distribution and availability in the intact islets.

  10. Deep-red polymer dots with bright two-photon fluorescence and high biocompatibility for in vivo mouse brain imaging

    Science.gov (United States)

    Alifu, Nuernisha; Sun, Zezhou; Zebibula, Abudureheman; Zhu, Zhenggang; Zhao, Xinyuan; Wu, Changfeng; Wang, Yalun; Qian, Jun

    2017-09-01

    With high contrast and deep penetration, two-photon fluorescence (2PF) imaging has become one of the most promising in vivo fluorescence imaging techniques. To obtain good imaging contrast, fluorescent nanoprobes with good 2PF properties are highly needed. In this work, bright 2PF polymer dots (P dots) were applied for in vivo mouse brain imaging. Deep-red emissive P dots with PFBT as the donor and PFDBT5 as the acceptor were synthesized and used as a contrast agent. P dots were further encapsulated by poly(styrene-co-maleic anhydride) (PSMA) and grafted with poly(ethylene glycol) (PEG). The P dots-PEG exhibit large two-photon absorption (2PA) cross-sections (δ≥8500 g), good water dispersibility, and high biocompatibility. P dots-PEG was further utilized first time for in vivo vascular imaging of mouse ear and brain, under 690-900 nm femtosecond (fs) laser excitation. Due to the large 2PA cross-section and deep-red emission, a large imaging depth ( 720 μm) was achieved.

  11. Choreography of cell motility and interaction dynamics imaged by two-photon microscopy in lymphoid organs.

    Science.gov (United States)

    Cahalan, Michael D; Parker, Ian

    2008-01-01

    The immune system is the most diffuse cellular system in the body. Accordingly, long-range migration of cells and short-range communication by local chemical signaling and by cell-cell contacts are vital to the control of an immune response. Cellular homing and migration within lymphoid organs, antigen recognition, and cell signaling and activation are clearly vital during an immune response, but these events had not been directly observed in vivo until recently. Introduced to the field of immunology in 2002, two-photon microscopy is the method of choice for visualizing living cells deep within native tissue environments, and it is now revealing an elegant cellular choreography that underlies the adaptive immune response to antigen challenge. We review cellular dynamics and molecular factors that contribute to basal motility of lymphocytes in the lymph node and cellular interactions leading to antigen capture and recognition, T cell activation, B cell activation, cytolytic effector function, and antibody production.

  12. Two-photon fluorescence imaging and femtosecond laser microsurgery to study drosophila dorsal closure

    Science.gov (United States)

    Thayil K. N., Anisha; Pereira, Andrea; Mathew, Manoj; Artigas, David; Martín Blanco, Enrique; Loza-Alvarez, Pablo

    2008-02-01

    Dorsal closure is a key morphogenic process that occurs at the last stages of Drosophila melanogaster embryogenesis. It involves a well coordinated rearrangement and movement of tissues that resemble epithelial wound healing in mammals. The cell dynamics and intracellular signaling pathways that accompany hole closure are expected to be similar during would healing providing a model system to study epithelial healing. Here we demonstrate the use of two-photon fluorescence microscope together with femtosecond laser ablation to examine the epithelial wound healing during embryonic dorsal closure. By using tightly focused NIR femtosecond pulses of subnanojoule energy we are able to produce highly confined microsurgery on the epithelial cells of a developing embryo. We observed that drosophila epidermis heals from the laser wounds with increased activity of actin near the wound edges.

  13. Two-photon instant structured illumination microscopy improves the depth penetration of super-resolution imaging in thick scattering samples.

    Science.gov (United States)

    Winter, Peter W; York, Andrew G; Nogare, Damian Dalle; Ingaramo, Maria; Christensen, Ryan; Chitnis, Ajay; Patterson, George H; Shroff, Hari

    2014-09-20

    Fluorescence imaging methods that achieve spatial resolution beyond the diffraction limit (super-resolution) are of great interest in biology. We describe a super-resolution method that combines two-photon excitation with structured illumination microscopy (SIM), enabling three-dimensional interrogation of live organisms with ~150 nm lateral and ~400 nm axial resolution, at frame rates of ~1 Hz. By performing optical rather than digital processing operations to improve resolution, our microscope permits super-resolution imaging with no additional cost in acquisition time or phototoxicity relative to the point-scanning two-photon microscope upon which it is based. Our method provides better depth penetration and inherent optical sectioning than all previously reported super-resolution SIM implementations, enabling super-resolution imaging at depths exceeding 100 μm from the coverslip surface. The capability of our system for interrogating thick live specimens at high resolution is demonstrated by imaging whole nematode embryos and larvae, and tissues and organs inside zebrafish embryos.

  14. Extended two-photon microscopy in live samples with Bessel beams: steadier focus, faster volume scans, and simpler stereoscopic imaging

    Science.gov (United States)

    Thériault, Gabrielle; Cottet, Martin; Castonguay, Annie; McCarthy, Nathalie; De Koninck, Yves

    2014-01-01

    Two-photon microscopy has revolutionized functional cellular imaging in tissue, but although the highly confined depth of field (DOF) of standard set-ups yields great optical sectioning, it also limits imaging speed in volume samples and ease of use. For this reason, we recently presented a simple and retrofittable modification to the two-photon laser-scanning microscope which extends the DOF through the use of an axicon (conical lens). Here we demonstrate three significant benefits of this technique using biological samples commonly employed in the field of neuroscience. First, we use a sample of neurons grown in culture and move it along the z-axis, showing that a more stable focus is achieved without compromise on transverse resolution. Second, we monitor 3D population dynamics in an acute slice of live mouse cortex, demonstrating that faster volumetric scans can be conducted. Third, we acquire a stereoscopic image of neurons and their dendrites in a fixed sample of mouse cortex, using only two scans instead of the complete stack and calculations required by standard systems. Taken together, these advantages, combined with the ease of integration into pre-existing systems, make the extended depth-of-field imaging based on Bessel beams a strong asset for the field of microscopy and life sciences in general. PMID:24904284

  15. Multi-beam two-photon imaging of fast Ca2+ signals in the Langendorff mouse heart.

    Science.gov (United States)

    Hammer, Karin; Lipp, Peter; Kaestner, Lars

    2014-11-03

    Although the role of calcium (Ca(2+)) in excitation-contraction coupling in the heart can be comprehensively studied at the cellular level, propagation of Ca(2+) signals intercellularly requires tissue-based investigations. To access cells below the epicardium, an optical-sectioning technique is necessary. Multi-photon microscopy allows reliable imaging for penetration to depths of up to 0.5 mm. Here, we provide a protocol that uses multibeam two-photon microscopy for measuring Ca(2+) signals in a Langendorff-perfused mouse heart.

  16. In vivo reactive neural plasticity investigation by means of correlative two photon: electron microscopy

    Science.gov (United States)

    Allegra Mascaro, A. L.; Cesare, P.; Sacconi, L.; Grasselli, G.; Mandolesi, G.; Maco, B.; Knott, G.; Huang, L.; De Paola, V.; Strata, P.; Pavone, F. S.

    2013-02-01

    In the adult nervous system, different populations of neurons correspond to different regenerative behavior. Although previous works showed that olivocerebellar fibers are capable of axonal regeneration in a suitable environment as a response to injury1, we have hitherto no details about the real dynamics of fiber regeneration. We set up a model of singularly axotomized climbing fibers (CF) to investigate their reparative properties in the adult central nervous system (CNS) in vivo. Time lapse two-photon imaging has been combined to laser nanosurgery2, 3 to define a temporal pattern of the degenerative event and to follow the structural rearrangement after injury. To characterize the damage and to elucidate the possible formation of new synaptic contacts on the sprouted branches of the lesioned CF, we combined two-photon in vivo imaging with block face scanning electron microscopy (FIB-SEM). Here we describe the approach followed to characterize the reactive plasticity after injury.

  17. Two-photon excitation with pico-second fluorescence lifetime imaging to detect nuclear association of flavanols

    Energy Technology Data Exchange (ETDEWEB)

    Mueller-Harvey, Irene, E-mail: i.mueller-harvey@reading.ac.uk [Chemistry and Biochemistry Laboratory, Food Production and Quality Research Division, School of Agriculture, Policy and Development, University of Reading, P O Box 236, Reading RG6 6AT (United Kingdom); Feucht, Walter, E-mail: walter.feucht@gmail.com [Department of Plant Sciences, Technical University of Munich (TUM), Wissenschaftszentrum Weihenstephan (WZW), D-85354 Freising (Germany); Polster, Juergen, E-mail: j.polster@wzw.tum.de [Department of Physical Biochemistry, Technical University of Munich (TUM), Wissenschaftszentrum Weihenstephan (WZW), D-85354 Freising (Germany); Trnkova, Lucie, E-mail: lucie.trnkova@uhk.cz [University of Hradec Kralove, Faculty of Science, Department of Chemistry, Rokitanskeho 62, 50003 Hradec Kralove (Czech Republic); Burgos, Pierre, E-mail: pierre.burgos@stfc.ac.uk [Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell-Oxford, Didcot, Oxfordshire, OX11 0QX (United Kingdom); Parker, Anthony W., E-mail: tony.parker@stfc.ac.uk [Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell-Oxford, Didcot, Oxfordshire, OX11 0QX (United Kingdom); Botchway, Stanley W., E-mail: stan.botchway@stfc.ac.uk [Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell-Oxford, Didcot, Oxfordshire, OX11 0QX (United Kingdom)

    2012-03-16

    Highlights: Black-Right-Pointing-Pointer This fluorescence lifetime imaging microscopy (FLIM) technique for flavanols overcomes autofluorescence interference in cells. Black-Right-Pointing-Pointer Plant flavanols differed in their lifetimes. Black-Right-Pointing-Pointer Dissolved and bound flavanols revealed contrasting lifetime changes. Black-Right-Pointing-Pointer This technique will allow studying of flavanol trafficking in live cells. - Abstract: Two-photon excitation enabled for the first time the observation and measurement of excited state fluorescence lifetimes from three flavanols in solution, which were {approx}1.0 ns for catechin and epicatechin, but <45 ps for epigallocatechin gallate (EGCG). The shorter lifetime for EGCG is in line with a lower fluorescence quantum yield of 0.003 compared to catechin (0.015) and epicatechin (0.018). In vivo experiments with onion cells demonstrated that tryptophan and quercetin, which tend to be major contributors of background fluorescence in plant cells, have sufficiently low cross sections for two-photon excitation at 630 nm and therefore do not interfere with detection of externally added or endogenous flavanols in Allium cepa or Taxus baccata cells. Applying two-photon excitation to flavanols enabled 3-D fluorescence lifetime imaging microscopy and showed that added EGCG penetrated the whole nucleus of onion cells. Interestingly, EGCG and catechin showed different lifetime behaviour when bound to the nucleus: EGCG lifetime increased from <45 to 200 ps, whilst catechin lifetime decreased from 1.0 ns to 500 ps. Semi-quantitative measurements revealed that the relative ratios of EGCG concentrations in nucleoli associated vesicles: nucleus: cytoplasm were ca. 100:10:1. Solution experiments with catechin, epicatechin and histone proteins provided preliminary evidence, via the appearance of a second lifetime ({tau}{sub 2} = 1.9-3.1 ns), that both flavanols may be interacting with histone proteins. We conclude that there

  18. Noninvasive two-photon imaging reveals retinyl ester storage structures in the eye.

    Science.gov (United States)

    Imanishi, Yoshikazu; Batten, Matthew L; Piston, David W; Baehr, Wolfgang; Palczewski, Krzysztof

    2004-02-01

    Visual sensation in vertebrates is triggered when light strikes retinal photoreceptor cells causing photoisomerization of the rhodopsin chromophore 11-cis-retinal to all-trans-retinal. The regeneration of preillumination conditions of the photoreceptor cells requires formation of 11-cis-retinal in the adjacent retinal pigment epithelium (RPE). Using the intrinsic fluorescence of all-trans-retinyl esters, noninvasive two-photon microscopy revealed previously uncharacterized structures (6.9 +/- 1.1 microm in length and 0.8 +/- 0.2 microm in diameter) distinct from other cellular organelles, termed the retinyl ester storage particles (RESTs), or retinosomes. These structures form autonomous all-trans-retinyl ester-rich intracellular compartments distinct from other organelles and colocalize with adipose differentiation-related protein. As demonstrated by in vivo experiments using wild-type mice, the RESTs participate in 11-cis-retinal formation. RESTs accumulate in Rpe65-/- mice incapable of carrying out the enzymatic isomerization, and correspondingly, are absent in the eyes of Lrat-/- mice deficient in retinyl ester synthesis. These results indicate that RESTs located close to the RPE plasma membrane are essential components in 11-cis-retinal production.

  19. FocusStack and StimServer: A new open source MATLAB toolchain for visual stimulation and analysis of two-photon calcium neuronal imaging data

    Directory of Open Access Journals (Sweden)

    Dylan Richard Muir

    2015-01-01

    Full Text Available Two-photon calcium imaging of neuronal responses is an increasingly accessible technology for probing population responses in cortex at single cell resolution, and with reasonable and improving temporal resolution. However, analysis of two-photon data is usually performed using ad-hoc solutions. To date, no publicly available software exists for straightforward analysis of stimulus-triggered two-photon imaging experiments. In addition, the increasing data rates of two-photon acquisition systems imply increasing cost of computing hardware required for in-memory analysis. Here we present a Matlab toolbox, FocusStack, for simple and efficient analysis of two-photon calcium imaging stacks on consumer-level hardware, with minimal memory footprint. We also present a Matlab toolbox, StimServer, for generation and sequencing of visual stimuli, designed to be triggered over a network link from a two-photon acquisition system. FocusStack is compatible out of the box with several existing two-photon acquisition systems, and is simple to adapt to arbitrary binary file formats. Analysis tools such as stack alignment for movement correction, automated cell detection and peri-stimulus time histograms are already provided, and further tools can be easily incorporated. Both packages are available as publicly-accessible source-code repositories.

  20. FocusStack and StimServer: a new open source MATLAB toolchain for visual stimulation and analysis of two-photon calcium neuronal imaging data.

    Science.gov (United States)

    Muir, Dylan R; Kampa, Björn M

    2014-01-01

    Two-photon calcium imaging of neuronal responses is an increasingly accessible technology for probing population responses in cortex at single cell resolution, and with reasonable and improving temporal resolution. However, analysis of two-photon data is usually performed using ad-hoc solutions. To date, no publicly available software exists for straightforward analysis of stimulus-triggered two-photon imaging experiments. In addition, the increasing data rates of two-photon acquisition systems imply increasing cost of computing hardware required for in-memory analysis. Here we present a Matlab toolbox, FocusStack, for simple and efficient analysis of two-photon calcium imaging stacks on consumer-level hardware, with minimal memory footprint. We also present a Matlab toolbox, StimServer, for generation and sequencing of visual stimuli, designed to be triggered over a network link from a two-photon acquisition system. FocusStack is compatible out of the box with several existing two-photon acquisition systems, and is simple to adapt to arbitrary binary file formats. Analysis tools such as stack alignment for movement correction, automated cell detection and peri-stimulus time histograms are already provided, and further tools can be easily incorporated. Both packages are available as publicly-accessible source-code repositories.

  1. Choosing the best embryo by time lapse versus standard morphology.

    Science.gov (United States)

    Kirkegaard, Kirstine; Ahlström, Aishling; Ingerslev, Hans Jakob; Hardarson, Thorir

    2015-02-01

    Within the past few years the morphological evaluation of in vitro fertilized embryos has been extended to include continuous surveillance, enabled by the introduction of time-lapse incubators developed specifically for IVF treatment. As a result time-lapse monitoring has been implemented in many clinics worldwide. The proposed benefits compared with culture in a standard incubator and fixed time-point evaluation are uninterrupted culture, a flexible workflow in the laboratory, and improved embryo selection. The latter is based on the reasonable assumption that more frequent observations will provide substantially more information on the relationship between development, timing, and embryo viability. Several retrospective studies have confirmed a relationship between time-lapse parameters and embryo viability evaluated by developmental competence, aneuploidy, and clinical pregnancy. Furthermore a much anticipated randomized study has shown improved pregnancy rates (PRs) after culture in a time-lapse incubator combined with selection using a hierarchical time-lapse selection model. At present this is the only randomized study on possible benefits of time lapse in human embryology. Strict evidence may still seem too weak to introduce time lapse in routine clinical setting. This aim of this review is therefore to perform a balanced discussion of the evidence for time-lapse monitoring. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  2. MULTIPHOTON MICROSCOPIC IMAGING OF MOUSE INTESTINAL MUCOSA BASED ON TWO-PHOTON EXCITED FLUORESCENCE AND SECOND HARMONIC GENERATION

    Directory of Open Access Journals (Sweden)

    REN'AN XU

    2013-01-01

    Full Text Available Multiphoton microscopy (MPM, based on two-photon excited fluorescence and second harmonic generation, enables direct noninvasive visualization of tissue architecture and cell morphology in live tissues without the administration of exogenous contrast agents. In this paper, we used MPM to image the microstructures of the mucosa in fresh, unfixed, and unstained intestinal tissue of mouse. The morphology and distribution of the main components in mucosa layer such as columnar cells, goblet cells, intestinal glands, and a little collagen fibers were clearly observed in MPM images, and then compared with standard H&E images from paired specimens. Our results indicate that MPM combined with endoscopy and miniaturization probes has the potential application in the clinical diagnosis and in vivo monitoring of early intestinal cancer.

  3. Noninvasive two-photon microscopy imaging of mouse retina and retinal pigment epithelium through the pupil of the eye.

    Science.gov (United States)

    Palczewska, Grazyna; Dong, Zhiqian; Golczak, Marcin; Hunter, Jennifer J; Williams, David R; Alexander, Nathan S; Palczewski, Krzysztof

    2014-07-01

    Two-photon excitation microscopy can image retinal molecular processes in vivo. Intrinsically fluorescent retinyl esters in subcellular structures called retinosomes are an integral part of the visual chromophore regeneration pathway. Fluorescent condensation products of all-trans-retinal accumulate in the eye with age and are also associated with age-related macular degeneration (AMD). Here, we report repetitive, dynamic imaging of these compounds in live mice through the pupil of the eye. By leveraging advanced adaptive optics, we developed a data acquisition algorithm that permitted the identification of retinosomes and condensation products in the retinal pigment epithelium by their characteristic localization, spectral properties and absence in genetically modified or drug-treated mice. This imaging approach has the potential to detect early molecular changes in retinoid metabolism that trigger light- and AMD-induced retinal defects and to assess the effectiveness of treatments for these conditions.

  4. Noninvasive two-photon fluorescence microscopy imaging of mouse retina and RPE through the pupil of the eye

    Science.gov (United States)

    Palczewska, Grazyna; Dong, Zhiqian; Golczak, Marcin; Hunter, Jennifer J.; Williams, David R.; Alexander, Nathan S.; Palczewski, Krzysztof

    2014-01-01

    Two-photon excitation microscopy (TPM) can image retinal molecular processes in vivo. Intrinsically fluorescent retinyl esters in sub-cellular structures called retinosomes are an integral part of the visual chromophore regeneration pathway. Fluorescent condensation products of all–trans–retinal accumulate in the eye with age and are also associated with age-related macular degeneration (AMD). Here we report repetitive, dynamic imaging of these compounds in live mice, through the pupil of the eye. Leveraging advanced adaptive optics we developed a data acquisition algorithm that permitted the identification of retinosomes and condensation products in the retinal pigment epithelium (RPE) by their characteristic localization, spectral properties, and absence in genetically modified or drug-treated mice. This imaging approach has the potential to detect early molecular changes in retinoid metabolism that trigger light and AMD-induced retinal defects and to assess the effectiveness of treatments for these conditions. PMID:24952647

  5. Simultaneous Two-photon in Vivo Imaging of Synaptic Inputs and Postsynaptic Targets in the Mouse Retrosplenial Cortex.

    Science.gov (United States)

    Łukasiewicz, Kacper; Robacha, Magdalena; Bożycki, Łukasz; Radwanska, Kasia; Czajkowski, Rafał

    2016-01-01

    This video shows the craniotomy procedure that allows chronic imaging of neurons in the mouse retrosplenial cortex (RSC) using in vivo two-photon microscopy in Thy1-GFP transgenic mouse line. This approach creates a possibility to investigate the correlation of behavioural manipulations with changes in neuronal morphology in vivo. The cranial window implantation procedure was considered to be limited only to the easily accessible cortex regions such as the barrel field. Our approach allows visualization of neurons in the highly vascularized RSC. RSC is an important element of the brain circuit responsible for spatial memory, previously deemed to be problematic for in vivo two-photon imaging. The cranial window implantation over the RSC is combined with an injection of mCherry-expressing recombinant adeno-associated virus (rAAV(mCherry)) into the dorsal hippocampus. The expressed mCherry spreads out to axonal projections from the hippocampus to RSC, enabling the visualization of changes in both presynaptic axonal boutons and postsynaptic dendritic spines in the cortex. This technique allows long-term monitoring of experience-dependent structural plasticity in RSC.

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

    Science.gov (United States)

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

    2012-03-01

    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 (λex=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.

  7. Lighting the Way to See Inside Two-Photon Absorption Materials: Structure-Property Relationship and Biological Imaging.

    Science.gov (United States)

    Zhang, Qiong; Tian, Xiaohe; Zhou, Hongping; Wu, Jieying; Tian, Yupeng

    2017-02-23

    ions, including transition metals and lanthanides, can serve as an important part of the structure to control the intramolecular charge-transfer process that drives the 2PA process. As templates, transition metal ions can assemble simple to more sophisticated ligands in a variety of multipolar arrangements resulting in interesting and tailorable electronic and optical properties, depending on the nature of the metal center and the energetics of the metal-ligand interactions, such as intraligand charge-transfer (ILCT) and metal-ligand charge-transfer (MLCT) processes. Lanthanide complexes are attractive for a number of reasons: (i) their visible emissions are quite long-lived; (ii) their absorption and emission can be tuned with the aid of appropriate photoactive ligands; (iii) the accessible energy-transfer path between the photo-active ligands and the lanthanide ion can facilitate efficient lanthanide-based 2PA properties. Thus, the above materials with excellent 2PA properties should be applied in two-photon applications, especially two-photon fluorescence microscopy (TPFM) and related emission-based applications. Furthermore, the progress of research into the use of those new 2PA materials with moderate 2PA cross section in the near-infrared region, good Materials 2017, 10, 223 2 of 37 biocompatibility, and enhanced two-photon excited fluorescence for two-photon bio-imaging is summarized. In addition, several possible future directions in this field are also discussed (146 references).

  8. Lighting the Way to See Inside Two-Photon Absorption Materials: Structure–Property Relationship and Biological Imaging

    Science.gov (United States)

    Zhang, Qiong; Tian, Xiaohe; Zhou, Hongping; Wu, Jieying; Tian, Yupeng

    2017-01-01

    ions, including transition metals and lanthanides, can serve as an important part of the structure to control the intramolecular charge-transfer process that drives the 2PA process. As templates, transition metal ions can assemble simple to more sophisticated ligands in a variety of multipolar arrangements resulting in interesting and tailorable electronic and optical properties, depending on the nature of the metal center and the energetics of the metal-ligand interactions, such as intraligand charge-transfer (ILCT) and metal-ligand charge-transfer (MLCT) processes. Lanthanide complexes are attractive for a number of reasons: (i) their visible emissions are quite long-lived; (ii) their absorption and emission can be tuned with the aid of appropriate photoactive ligands; (iii) the accessible energy-transfer path between the photo-active ligands and the lanthanide ion can facilitate efficient lanthanide-based 2PA properties. Thus, the above materials with excellent 2PA properties should be applied in two-photon applications, especially two-photon fluorescence microscopy (TPFM) and related emission-based applications. Furthermore, the progress of research into the use of those new 2PA materials with moderate 2PA cross section in the near-infrared region, good biocompatibility, and enhanced two-photon excited fluorescence for two-photon bio-imaging is summarized. In addition, several possible future directions in this field are also discussed (146 references). PMID:28772584

  9. Lighting the Way to See Inside Two-Photon Absorption Materials: Structure–Property Relationship and Biological Imaging

    Directory of Open Access Journals (Sweden)

    Qiong Zhang

    2017-02-01

    properties. The metal ions, including transition metals and lanthanides, can serve as an important part of the structure to control the intramolecular charge-transfer process that drives the 2PA process. As templates, transition metal ions can assemble simple to more sophisticated ligands in a variety of multipolar arrangements resulting in interesting and tailorable electronic and optical properties, depending on the nature of the metal center and the energetics of the metal-ligand interactions, such as intraligand charge-transfer (ILCT and metal-ligand charge-transfer (MLCT processes. Lanthanide complexes are attractive for a number of reasons: (i their visible emissions are quite long-lived; (ii their absorption and emission can be tuned with the aid of appropriate photoactive ligands; (iii the accessible energy-transfer path between the photo-active ligands and the lanthanide ion can facilitate efficient lanthanide-based 2PA properties. Thus, the above materials with excellent 2PA properties should be applied in two-photon applications, especially two-photon fluorescence microscopy (TPFM and related emission-based applications. Furthermore, the progress of research into the use of those new 2PA materials with moderate 2PA cross section in the near-infrared region, good Materials 2017, 10, 223 2 of 37 biocompatibility, and enhanced two-photon excited fluorescence for two-photon bio-imaging is summarized. In addition, several possible future directions in this field are also discussed (146 references.

  10. Synthesis, crystals of centrosymmetric triphenylamine chromophores bearing prodigious two-photon absorption cross-section and biological imaging

    Science.gov (United States)

    Wang, Shichao; Xu, Shasha; Wang, Yiming; Tian, Xiaohe; Zhang, Yujin; Wang, Chuankui; Wu, Jieying; Yang, Jiaxiang; Tian, Yupeng

    2017-02-01

    Two centrosymmetric D-π-D type triphenylamine chromophores with long π-conjugated bridge and strong electron-donating moiety were designed, synthesized and fully characterized. The crystal analysis revealed that multiple Csbnd H ⋯ π interactions existed in two chromophores, which played a crucial role in generating molecular 1D chains and 2D layers structures. Linear and nonlinear optical properties of the chromophores were systematically investigated with the aid of theoretical calculations. Two chromophores both exhibited intense and wide-dispersed one-photon/two-photon excited fluorescence, bear prodigious 2PA cross section (δ). Especially for Dye2, with ethyoxyl groups, displayed the strong 2PA activity, large cross-sections (δmax > 16,000 GM) and high NLO efficiency (δmax/MW > 16 GM/(g·mol)) in the range of 680-830 nm in DMF. In addition, one- and two-photon fluorescence microscopy images of HepG2 cells incubated with Dye2 were obtained and found that Dye2 could effectively uptake toward living cells and display a uniformly localized in cytosolic space.

  11. Automated image analysis for diameters and branching points of cerebral penetrating arteries and veins captured with two-photon microscopy.

    Science.gov (United States)

    Sugashi, Takuma; Yoshihara, Kouichi; Kawaguchi, Hiroshi; Takuwa, Hiroyuki; Ito, Hiroshi; Kanno, Iwao; Yamada, Yukio; Masamoto, Kazuto

    2014-01-01

    The present study was aimed to characterize 3-dimensional (3D) morphology of the cortical microvasculature (e.g., penetrating artery and emerging vein), using two-photon microscopy and automated analysis for their cross-sectional diameters and branching positions in the mouse cortex. We observed that both artery and vein had variable cross-sectional diameters across cortical depths. The mean diameter was similar for both artery (17 ± 5 μm) and vein (15 ± 5 μm), and there were no detectable differences over depths of 50-400 μm. On the other hand, the number of branches was slightly increased up to 400-μm depth for both the artery and vein. The mean number of branches per 0.1 mm vessel length was 1.7 ± 1.2 and 3.8 ± 1.6 for the artery and vein, respectively. This method allows for quantification of the large volume data of microvascular images captured with two-photon microscopy. This will contribute to the morphometric analysis of the cortical microvasculature in functioning brains.

  12. Energy transfer in aminonaphthalimide-boron-dipyrromethene (BODIPY) dyads upon one- and two-photon excitation: applications for cellular imaging.

    Science.gov (United States)

    Collado, Daniel; Remón, Patricia; Vida, Yolanda; Najera, Francisco; Sen, Pratik; Pischel, Uwe; Perez-Inestrosa, Ezequiel

    2014-03-01

    Aminonaphthalimide-BODIPY energy transfer cassettes were found to show very fast (kEET ≈ 10(10)-10(11) s(-1) and efficient BODIPY fluorescence sensitization. This was observed upon one- and two-photon excitation, which extends the application range of the investigated bichromophoric dyads in terms of accessible excitation wavelengths. In comparison with the direct excitation of the BODIPY chromophore, the two-photon absorption cross-section δ of the dyads is significantly incremented by the presence of the aminonaphthalimide donor [δ ≈ 10 GM for the BODIPY versus 19-26 GM in the dyad at λ(exc)=840 nm; 1 GM (Goeppert-Mayer unit)=10(-50) cm(4) smolecule(-1) photon-(1)]. The electronic decoupling of the donor and acceptor, which is a precondition for the energy transfercassette concept, was demonstrated by time-dependent density functional theory calculations. The applicability of the new probes in the one- and twophoton excitation mode was demonstrated in a proof-of-principle approach in the fluorescence imaging of HeLa cells. To the best of our knowledge, this is the first demonstration of the merging of multiphoton excitation with the energy transfer cassette concept for a BODIPY-containing dyad.

  13. Two-photon imaging of neural activity and structural plasticity in the rodent spinal cord

    OpenAIRE

    Johannssen, H

    2011-01-01

    In my PhD thesis, I used two‐photon imaging to investigate neuronal circuits and glia cells in the spinal cord of living mice. To achieve this, a major effort first was to establish a mouse spinal cord preparation suitable for stable and long‐lasting imaging experiments. Without adequate stabilisation, the spinal cord was prone to large‐scale movement artefacts clearly hampering high‐resolution imaging in vivo. To overcome these limitations, I employed strategies to optimise th...

  14. Multiphoton microscopic imaging of adipose tissue based on second-harmonic generation and two-photon excited fluorescence.

    Science.gov (United States)

    Huang, Zufang; Zhuo, Shuangmu; Chen, Jianxin; Chen, Rong; Jiang, Xingshan

    2008-01-01

    The fresh adipose tissue was investigated by the use of multiphoton microscopy (MPM) based on two-photon excited fluorescence and second-harmonic generation (SHG). Microstructure of collagen and adipose cells in the adipose tissue is clearly imaged at a subcellular level with the excitation light wavelengths of 850 and 730 nm, respectively. The emission spectrum of collagen SHG signal and NADH and FAD fluorescence signal can also be obtained, which can be used to quantify the content of collagen and adipose cells and reflect the degree of pathological changes when comparing normal tissue with abnormal adipose tissue in the same condition. The results indicate that MPM has the potential to be applied to investigate the adipose tissue and can be used in the research field of lipid and connective tissues.

  15. Age-related structural abnormalities in the human retina-choroid complex revealed by two-photon excited autofluorescence imaging.

    Science.gov (United States)

    Han, Meng; Giese, Guenter; Schmitz-Valckenberg, Steffen; Bindewald-Wittich, Almut; Holz, Frank G; Yu, Jiayi; Bille, Josef F; Niemz, Markolf H

    2007-01-01

    The intensive metabolism of photoreceptors is delicately maintained by the retinal pigment epithelium (RPE) and the choroid. Dysfunction of either the RPE or choroid may lead to severe damage to the retina. Two-photon excited autofluorescence (TPEF) from endogenous fluorophores in the human retina provides a novel opportunity to reveal age-related structural abnormalities in the retina-choroid complex prior to apparent pathological manifestations of age-related retinal diseases. In the photoreceptor layer, the regularity of the macular photoreceptor mosaic is preserved during aging. In the RPE, enlarged lipofuscin granules demonstrate significantly blue-shifted autofluorescence, which coincides with the depletion of melanin pigments. Prominent fibrillar structures in elderly Bruch's membrane and choriocapillaries represent choroidal structure and permeability alterations. Requiring neither slicing nor labeling, TPEF imaging is an elegant and highly efficient tool to delineate the thick, fragile, and opaque retina-choroid complex, and may provide clues to the trigger events of age-related macular degeneration.

  16. Microscopic imaging of glyceraldehyde-induced tissue glycation with intrinsic second harmonic generation and two-photon fluorescence contrasts

    Science.gov (United States)

    Hwang, Yu Jer; Granelli, Joseph; Tirumalasetty, Manasa; Lyubovitsky, Julia

    2013-02-01

    The bioinspired approaches to tissue strengthening and preservation rely on non-toxic cross-linking agents one of which is glyceraldehyde. In this study we used multiphoton microscopy that employs second harmonic generation (SHG) contrast to evaluate collagen microstructures and two-photon fluorescence (TPF) contrast to monitor progression of cross-linking upon treatment of tissues with glyceraldehyde. We examined collagen hydrogels assembled at 37 °C and 27 °C, bovine scleral and corneal tissues, skin as well as rat tail tendons. The results show a different effect of glyceraldehyde on collagen microstructures within the above tissues. This effect depends on the original microstructural assembly of collagen within a specific tissue. Our data suggests that epidermis (in skin and cornea) will protect collagen from cross-linking with glyceraldehyde. The work highlights benefits of monitoring progression of collagen cross-linking and effects of cross-linking on fiber microstructures as imaged with SHG and TPF signals.

  17. Two-photon excitation fluorescence imaging of the living juxtaglomerular apparatus.

    Science.gov (United States)

    Peti-Peterdi, János; Morishima, Shigeru; Bell, P Darwin; Okada, Yasunobu

    2002-07-01

    Recently, multiphoton excitation fluorescence microscopy has been developed that offers important advantages over confocal imaging, particularly for in vivo visualization of thick tissue samples. We used this state-of-the-art technique to capture high-quality images and study the function of otherwise inaccessible cell types and complex cell structures of the juxtaglomerular apparatus (JGA) in living preparations of the kidney. This structure has multiple cell types that exhibit a complex array of functions, which regulate the process of filtrate formation and renal hemodynamics. We report, for the first time, on high-resolution three-dimensional morphology and Z-sectioning through isolated, perfused kidney glomeruli, tubules, and JGA. Time-series images show how alterations in tubular fluid composition cause striking changes in single-cell volume of the unique macula densa tubular epithelium in situ and how they also affect glomerular filtration through alterations in associated structures within the JGA. In addition, calcium imaging of the glomerulus and JGA demonstrates the utility of this system in capturing the complexity of events and effects that are exerted by the specific hypertensive autacoid angiotensin II. This imaging approach to the study of isolated, perfused live tissue with multiphoton microscopy may be applied to other biological systems in which multiple cell types form a functionally integrated syncytium.

  18. Quantifying distortions in two-photon remote focussing images using a volumetric calibration specimen

    Directory of Open Access Journals (Sweden)

    Alexander David Corbett

    2014-10-01

    Full Text Available Remote focussing microscopy allows sharp, in-focus images to be acquired at speed from outside of the focal plane of an objective lens without any agitation of the specimen. However, without careful optical alignment, the advantages of remote focussing microscopy could be compromised by the introduction of depth-dependent scaling artefacts. To achieve an ideal alignment in a point-scanning remote focussing microscope, the lateral (XY scan mirror pair must be imaged onto the back focal plane of both the reference and imaging objectives, in a telecentric arrangement. However, for many commercial objective lenses, it can be difficult to accurately locate the position of the back focal plane. This paper investigates the impact of this limitation on the fidelity of three-dimensional data sets of living cardiac tissue, specifically the introduction of distortions. These distortions limit the accuracy of sarcomere measurements taken directly from raw volumetric data. The origin of the distortion is first identified through simulation of a remote focussing microscope. Using a novel three-dimensional calibration specimen it was then possible to quantify experimentally the size of the distortion as a function of objective misalignment. Finally, by first approximating and then compensating the distortion in imaging data from whole heart rodent studies, the variance of sarcomere length measurements was reduced by almost 50%.

  19. A NEW INVERSION METHOD OF TIME-LAPSE SEISMIC

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Time-Lapse Seismic improves oil recovery ratio by dynamic reservoir monitoring. Because of the large number of seismic explorations in the process of time-lapse seismic inversion, traditional methods need plenty of inversion calculations which cost high computational works. The method is therefore inefficient. In this paper, in order to reduce the repeating computations in traditional, a new time-lapse seismic inversion method is put forward. Firstly a homotopy-regularization method is proposed for the first time inversion. Secondly, with the first time inversion results as the initial value of following model, a model of the second time inversion is rebuilt by analyzing the characters of time-lapse seismic and localized inversion method is designed by using the model. Finally, through simulation, the comparison between traditional method and the new scheme is given. Our simulation results show that the new scheme could save the algorithm computations greatly.

  20. Organic nanostructure-based probes for two-photon imaging of mitochondria and microbes with emission between 430 nm and 640 nm.

    Science.gov (United States)

    Yang, Xinglong; Wang, Nuoxin; Zhang, Lingmin; Dai, Luru; Shao, Huawu; Jiang, Xingyu

    2017-04-06

    Multi-photon excitation and versatile fluorescent probes are in high need for biological imaging, since one probe can satisfy many needs as a biosensor. Herein we synthesize a series of two-photon excited probes based on tetraphenylethene (TPE) structures (TPE-Acr, TPE-Py, and TPE-Quino), which can image both mammalian cells and bacteria based on aggregation-induced emission (AIE) without washing them. Because of cationic moieties, the fluorescent molecules can aggregate into nanoscale fluorescent organic nanoscale dots to image mitochondria and bacteria with tunable emissions using both one-photon and two-photon excitation. Our research demonstrates that these AIE-dots expand the functions of luminescent organic dots to construct efficient fluorescent sensors applicable to both one-photon and two-photon excitation for bio-imaging of bacteria and mammalian cells.

  1. Photolytic-interference-free, femtosecond, two-photon laser-induced fluorescence imaging of atomic oxygen in flames

    Science.gov (United States)

    Kulatilaka, Waruna D.; Roy, Sukesh; Jiang, Naibo; Gord, James R.

    2016-02-01

    Ultrashort-pulse lasers are well suited for nonlinear diagnostic techniques such as two-photon laser-induced fluorescence (TPLIF) because the signals generated scale as the laser intensity squared. Furthermore, the broad spectral bandwidths associated with nearly Fourier-transform-limited ultrashort pulses effectively contribute to efficient nonlinear excitation by coupling through a large number of in-phase photon pairs, thereby producing strong fluorescence signals. Additionally, femtosecond (fs)-duration amplified laser systems typically operate at 1-10 kHz repetition rates, enabling high-repetition-rate imaging in dynamic environments. In previous experiments, we have demonstrated utilization of fs pulses for kilohertz (kHz)-rate, interference-free imaging of atomic hydrogen (H) in flames. In the present study, we investigate the utilization of fs-duration pulses to photolytic-interference-free TPLIF imaging of atomic oxygen (O). In TPLIF of O, photodissociation of vibrationally excited carbon dioxide (CO2) is known to be the prominent interference that produces additional O atoms in the medium. We have found that through the use of fs excitation, such interferences can be virtually eliminated in premixed laminar methane flames, which paves the way for two-dimensional imaging of O at kHz data rates. Such measurements can provide critical data for validating complex, multidimensional turbulent-combustion models as well as for investigating flame dynamics in practical combustion devices.

  2. Single Cell Assay for Molecular Diagnostics and Medicine: Monitoring Intracellular Concentrations of Macromolecules by Two-photon Fluorescence Lifetime Imaging.

    Science.gov (United States)

    Pliss, Artem; Peng, Xiao; Liu, Lixin; Kuzmin, Andrey; Wang, Yan; Qu, Junle; Li, Yuee; Prasad, Paras N

    2015-01-01

    Molecular organization of a cell is dynamically transformed along the course of cellular physiological processes, pathologic developments or derived from interactions with drugs. The capability to measure and monitor concentrations of macromolecules in a single cell would greatly enhance studies of cellular processes in heterogeneous populations. In this communication, we introduce and experimentally validate a bio-analytical single-cell assay, wherein the overall concentration of macromolecules is estimated in specific subcellular domains, such as structure-function compartments of the cell nucleus as well as in nucleoplasm. We describe quantitative mapping of local biomolecular concentrations, either intrinsic relating to the functional and physiological state of a cell, or altered by a therapeutic drug action, using two-photon excited fluorescence lifetime imaging (FLIM). The proposed assay utilizes a correlation between the fluorescence lifetime of fluorophore and the refractive index of its microenvironment varying due to changes in the concentrations of macromolecules, mainly proteins. Two-photon excitation in Near-Infra Red biological transparency window reduced the photo-toxicity in live cells, as compared with a conventional single-photon approach. Using this new assay, we estimated average concentrations of proteins in the compartments of nuclear speckles and in the nucleoplasm at ~150 mg/ml, and in the nucleolus at ~284 mg/ml. Furthermore, we show a profound influence of pharmaceutical inhibitors of RNA synthesis on intracellular protein density. The approach proposed here will significantly advance theranostics, and studies of drug-cell interactions at the single-cell level, aiding development of personal molecular medicine.

  3. Dynamic characterization of hydrophobic and hydrophilic solutes in oleic-acid enhanced transdermal delivery using two-photon fluorescence microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Tseng, Te-Yu; Yang, Chiu-Sheng; Chen, Yang-Fang [Department of Physics, National Taiwan University, Taipei, Taiwan (China); Tsai, Tsung-Hua [Department of Dermatology, Far Eastern Memorial Hospital, New Taipei City, Taiwan (China); Dong, Chen-Yuan, E-mail: cydong@phys.ntu.edu.tw [Department of Physics, National Taiwan University, Taipei, Taiwan (China); Center for Quantum Science and Engineering, National Taiwan University, Taipei, Taiwan (China); Center for Optoelectronic Biomedicine, National Taiwan University, Taipei, Taiwan (China)

    2014-10-20

    In this letter, we propose an efficient methodology of investigating dynamic properties of sulforhodamine B and rhodamine B hexyl ester molecules transporting across ex-vivo human stratum corneum with and without oleic acid enhancement. Three-dimensional, time-lapse fluorescence images of the stratum corneum can be obtained using two-photon fluorescence microscopy. Furthermore, temporal quantifications of transport enhancements in diffusion parameters can be achieved with the use of Fick's second law. Dynamic characterization of solutes transporting across the stratum corneum is an effective method for understanding transient phenomena in transdermal delivery of probe molecules, leading to improved delivery strategies of molecular species for therapeutic purposes.

  4. Two-dimensional imaging of molecular hydrogen in H2-air diffusion flames using two-photon laser-induced fluorescence

    Science.gov (United States)

    Lempert, W.; Kumar, V.; Glesk, I.; Miles, R.; Diskin, G.

    1991-01-01

    The use of a tunable ArF laser at 193.26 nm to record simultaneous single-laser-shot, planar images of molecular hydrogen and hot oxygen in a turbulent H2-air diffusion flame. Excitation spectra of fuel and oxidant-rich flame zones confirm a partial overlap of the two-photon H2 and single-photon O2 Schumann-Runge absorption bands. UV Rayleigh scattering images of flame structure and estimated detection limits for the H2 two-photon imaging are also presented.

  5. Upconverting nanoparticles: a versatile platform for wide-field two-photon microscopy and multi-modal in vivo imaging.

    Science.gov (United States)

    Park, Yong Il; Lee, Kang Taek; Suh, Yung Doug; Hyeon, Taeghwan

    2015-03-21

    Lanthanide-doped upconverting nanoparticles (UCNPs) have recently attracted enormous attention in the field of biological imaging owing to their unique optical properties: (1) efficient upconversion photoluminescence, which is intense enough to be detected at the single-particle level with a (nonscanning) wide-field microscope setup equipped with a continuous wave (CW) near-infrared (NIR) laser (980 nm), and (2) resistance to photoblinking and photobleaching. Moreover, the use of NIR excitation minimizes adverse photoinduced effects such as cellular photodamage and the autofluorescence background. Finally, the cytotoxicity of UCNPs is much lower than that of other nanoparticle systems. All these advantages can be exploited simultaneously without any conflicts, which enables the establishment of a novel UCNP-based platform for wide-field two-photon microscopy. UCNPs are also useful for multimodal in vivo imaging because simple variations in the composition of the lattice atoms and dopant ions integrated into the particles can be easily implemented, yielding various distinct biomedical activities relevant to magnetic resonance imaging (MRI), computed tomography (CT), and positron emission tomography (PET). These multiple functions embedded in a single type of UCNPs play a crucial role in precise disease diagnosis. The application of UCNPs is extended to therapeutic fields such as photodynamic and photothermal cancer therapies through advanced surface conjugation schemes.

  6. Cytosolic NADH-NAD+ Redox Visualized in Brain Slices by Two-Photon Fluorescence Lifetime Biosensor Imaging

    Science.gov (United States)

    Mongeon, Rebecca; Venkatachalam, Veena

    2016-01-01

    Abstract Aim: Cytosolic NADH-NAD+ redox state is central to cellular metabolism and a valuable indicator of glucose and lactate metabolism in living cells. Here we sought to quantitatively determine NADH-NAD+ redox in live cells and brain tissue using a fluorescence lifetime imaging of the genetically-encoded single-fluorophore biosensor Peredox. Results: We show that Peredox exhibits a substantial change in its fluorescence lifetime over its sensing range of NADH-NAD+ ratio. This allows changes in cytosolic NADH redox to be visualized in living cells using a two-photon scanning microscope with fluorescence lifetime imaging capabilities (2p-FLIM), using time-correlated single photon counting. Innovation: Because the lifetime readout is absolutely calibrated (in nanoseconds) and is independent of sensor concentration, we demonstrate that quantitative assessment of NADH redox is possible using a single fluorophore biosensor. Conclusion: Imaging of the sensor in mouse hippocampal brain slices reveals that astrocytes are typically much more reduced (with higher NADH:NAD+ ratio) than neurons under basal conditions, consistent with the hypothesis that astrocytes are more glycolytic than neurons. Antioxid. Redox Signal. 25, 553–563. PMID:26857245

  7. Curious case of gravitational lensing by binary black holes: A tale of two photon spheres, new relativistic images, and caustics

    Science.gov (United States)

    Patil, Mandar; Mishra, Priti; Narasimha, D.

    2017-01-01

    Binary black holes have been in the limelight of late due to the detection of gravitational waves from coalescing compact binaries in the events GW150914 and GW151226. In this paper we study gravitational lensing by the binary black holes modeled as an equal mass Majumdar-Papapetrou dihole metric and show that this system displays features that are quite unprecedented and absent in any other lensing configuration investigated so far in the literature. We restrict our attention to the light rays which move on the plane midway between the two identical black holes, which allows us to employ various techniques developed for the equatorial lensing in the spherically symmetric spacetimes. If distance between the two black holes is below a certain threshold value, then the system admits two photon spheres. As in the case of a single black hole, infinitely many relativistic images are formed due to the light rays which turn back from the region outside the outer (unstable) photon sphere, all of which lie beyond a critical angular radius with respect to the lens. However, in the presence of the inner (stable) photon sphere, the effective potential after admitting minimum turns upwards and blows up for the smaller values of radii and the light rays that enter the outer photon sphere can turn back, leading to the formation of a new set of infinitely many relativistic images, all of which lie below the critical radius from the lens mentioned above. As the distance between the two black holes is increased, two photon spheres approach one another, merge and eventually disappear. In the absence of the photon sphere, apart from the formation of a finite number of discrete relativistic images, the system remarkably admits a radial caustic, which has never been observed in the context of relativistic lensing before. Thus the system of the binary black hole admits novel features both in the presence and absence of photon spheres. We discuss possible observational signatures and

  8. Using Student-Produced Time-Lapse Plant Movies to Communicate Concepts in Plant Biology

    Directory of Open Access Journals (Sweden)

    Marcia Harrison-Pitaniello

    2013-01-01

    Full Text Available Why do students think plants are “boring”?  One factor may be that they do not see plant movement in real (i.e., their time.  This attitude may negatively impact their understanding of plant biology.  Time-lapse movies of plants allow students to see the sophistication of movements involved in both organ development and orientation.  The objective of this project was to develop simple methods to capture image sequences for lab analysis and for converting into movies.  The technology for making time-lapse movies is now easily attainable and fairly inexpensive, allowing its use for skill levels from grade school through college undergraduates.  Presented are example time-lapse movie exercises from both an undergraduate plant physiology course and outreach activities.  The time-lapse plant exercises are adaptable to explore numerous topics that incorporate science standards core concepts, competencies, and disciplinary practices as well as to integrate higher order thinking skills and build skills in hypothesis development and communicating results to various audiences.

  9. Determination of Optimum Frame Rates for Observation of Construction Operations from Time-Lapse Movies

    Directory of Open Access Journals (Sweden)

    Y. M. Ibrahim

    2012-07-01

    Full Text Available Construction professionals have been using time-lapse movies in monitoring construction operations. However, some amount of detail is always lost in the interval between two consecutive frames in a time-lapse movie. This poses the question: By how much can the frame rate be lowered from the standard 30fps (frames per second to allow for the accurate observation of construction operations from a time-lapse movie? This paper addresses the problem by establishing the optimum frame rates for observation of activities related to mortar mixing and block handling. The activities were first recorded at the standard rate of 30fps. Using the Adobe Premier Pro video editing software, the records were then segregated into still images from which 15 different time-lapse movies of various time intervals were generated. The movies were then shown to 25 Construction Managers. A structured questionnaire was employed to capture the level of accuracy with which Construction Managers could interpret the job site situation from each movie. The results suggest that 1fpm (frame per minute is sufficient for the accurate tracking of labourers involved in mortar mixing while 1 frame in every 20 seconds is sufficient for accurate identification of number of cement bags used. However, for tracking number of blocks off-loaded, and those damaged, 1 frame in every 2 seconds is required.

  10. LFP-guided targeting of a cortical barrel column for in vivo two-photon calcium imaging.

    Science.gov (United States)

    Lee, Joon-Hyuk; Shin, Hee-Sup; Lee, Kwang-Hyung; Chung, Sooyoung

    2015-10-29

    Two-photon microscopy of bulk-loaded functional dyes is an outstanding physiological technique that enables simultaneous functional mapping of hundreds of brain cells in vivo at single-cell resolution. However, precise targeting of a specific cortical location is not easy due to its fine dimensionality. To enable precise targeting, intrinsic-signal optical imaging is often additionally performed. However, the intrinsic-signal optical imaging is not only time-consuming but also ineffective in ensuring precision. Here, we propose an alternative method for precise targeting based on local field potential (LFP) recording, a conventional electrophysiological method. The heart of this method lies in use of the same glass pipette to record LFPs and to eject calcium dye. After confirming the target area by LFP using a glass pipette, the calcium dye is ejected from the same pipette without a time delay or spatial adjustment. As a result, the calcium dye is loaded into the same ensemble of brain cells from which the LFP was obtained. As a validation of the proposed LFP-based method, we targeted and successfully loaded calcium dye into layer 2/3 of a mouse barrel column.

  11. Long-Term Two-Photon Calcium Imaging of Neuronal Populations with Subcellular Resolution in Adult Non-human Primates.

    Science.gov (United States)

    Sadakane, Osamu; Masamizu, Yoshito; Watakabe, Akiya; Terada, Shin-Ichiro; Ohtsuka, Masanari; Takaji, Masafumi; Mizukami, Hiroaki; Ozawa, Keiya; Kawasaki, Hiroshi; Matsuzaki, Masanori; Yamamori, Tetsuo

    2015-12-01

    Two-photon imaging with genetically encoded calcium indicators (GECIs) enables long-term observation of neuronal activity in vivo. However, there are very few studies of GECIs in primates. Here, we report a method for long-term imaging of a GECI, GCaMP6f, expressed from adeno-associated virus vectors in cortical neurons of the adult common marmoset (Callithrix jacchus), a small New World primate. We used a tetracycline-inducible expression system to robustly amplify neuronal GCaMP6f expression and up- and downregulate it for more than 100 days. We succeeded in monitoring spontaneous activity not only from hundreds of neurons three-dimensionally distributed in layers 2 and 3 but also from single dendrites and axons in layer 1. Furthermore, we detected selective activities from somata, dendrites, and axons in the somatosensory cortex responding to specific tactile stimuli. Our results provide a way to investigate the organization and plasticity of cortical microcircuits at subcellular resolution in non-human primates.

  12. Multimodal second harmonic generation and two photon fluorescence imaging of microdomain calcium contraction coupling in single cardiomyocytes

    Science.gov (United States)

    Chan, James; Awasthi, Samir; Izu, Leighton; Mao, Ziliang; Jian, Zhong; Landas, Trevor; Lerner, Aaron; Shimkunas, Rafael; Woldeyesus, Rahwa; Bossuyt, Julie; Wood, Brittani; Chen, Yi-Je; Matthews, Dennis; Lieu, Deborah; Chiamvimonvat, Nipavan; Lam, Kit; Chen-Izu, Ye

    2016-11-01

    The objective of this study was to develop a method for simultaneously measuring the calcium and contraction dynamics of single, live cardiomyocytes at high spatial resolutions. Such measurements are important to investigate local calcium release and the mechanical response at the sarcomere level (i.e. the basic unit of contraction), which have important implications in cardiac dysfunction and arrhythmias in conditions such as hypertension, atrial fibrillation, and myocardial infarction. Here, we describe a multimodal second harmonic generation (SHG) and two photon fluorescence (2PF) microscopy technique that is used to simultaneously measure subsarcomere calcium and contraction events at high spatial and temporal resolutions. The method takes advantage of the label-free nature of SHG for imaging the sarcomeres and the high spatial colocalization of the SHG signal and the fluorescence signal excited from calcium indicators. This microscope was used to measure calcium sparks and waves and associated contractions in subcellular microdomains, leading to the generation of subcellular strain. We anticipate this new imaging tool will play an important role in studying mechanical stress-induced heart disease.

  13. Volumetric label-free imaging and 3D reconstruction of mammalian cochlea based on two-photon excitation fluorescence microscopy

    Science.gov (United States)

    Zhang, Xianzeng; Geng, Yang; Ye, Qing; Zhan, Zhenlin; Xie, Shusen

    2013-11-01

    The visualization of the delicate structure and spatial relationship of intracochlear sensory cells has relied on the laborious procedures of tissue excision, fixation, sectioning and staining for light and electron microscopy. Confocal microscopy is advantageous for its high resolution and deep penetration depth, yet disadvantageous due to the necessity of exogenous labeling. In this study, we present the volumetric imaging of rat cochlea without exogenous dyes using a near-infrared femtosecond laser as the excitation mechanism and endogenous two-photon excitation fluorescence (TPEF) as the contrast mechanism. We find that TPEF exhibits strong contrast, allowing cellular and even subcellular resolution imaging of the cochlea, differentiating cell types, visualizing delicate structures and the radial nerve fiber. Our results further demonstrate that 3D reconstruction rendered with z-stacks of optical sections enables better revealment of fine structures and spatial relationships, and easily performed morphometric analysis. The TPEF-based optical biopsy technique provides great potential for new and sensitive diagnostic tools for hearing loss or hearing disorders, especially when combined with fiber-based microendoscopy.

  14. NEAR-IR TWO PHOTON MICROSCOPY IMAGING OF SILICA NANOPARTICLES FUNCTIONALIZED WITH ISOLATED SENSITIZED Yb(III) CENTERS

    Energy Technology Data Exchange (ETDEWEB)

    Lapadula, Giuseppe; Bourdolle, Adrien; Allouche, Florian; Conley, Matthew P.; Maron, Laurent; Lukens, Wayne W.; Guyot, Yannick; Andraud, Chantal; Brasselet, Sophie; Copé; ret, Christophe; Maury, Olivier; Andersen, Richard A.

    2013-01-12

    Bright nano objects emitting in the near infrared with a maximal cross section of 41.4 x 103 GM (Goppert Mayer), were prepared by implanting ca. 180 4,4 diethylaminostyryl 2,2 bipyridine (DEAS) Yb(III) complexes on the surface of 12 nm silica nanoparticles. The surface complexes ([DEAS Ln SiO2], Ln =Y,Yb) were characterized using IR, solid state NMR, UV Vis, EXAFS spectroscopies in combination with the preparation and characterization of similar molecular analogues by analytical techniques (IR, solution NMR, UV Vis, X ray crystallography) as well as DFT calculations. Starting from the partial dehydroxylation of the silica at 700 C on high vacuum having 0.8 OH.nm 2, the grafting of Ln(N(SiMe3)2)3 generate ≤SiO Ln(N(SiMe3)2)2, which upon thermal step and coordination of the DEAS chromophore yields (≤SiO)3Ln(DEAS). Surface and molecular analogues display similar properties, in terms of DEAS binding constants absorption maxima and luminescence properties (intense emission band assigned to a ligand centered CT fluorescence and life time) in the solid state, consistent with the molecular nature of the surface species. The densely functionalized nanoparticles can be dispersed via ultra-sonication in small ca. 15-20 nm aggregates (1 to 6 elementary particles) that were detected using two photon microscopy imaging at 720 nm excitation, making them promising nano objects for bio imaging.

  15. Calcium rubies: a family of red-emitting functionalizable indicators suitable for two-photon Ca2+ imaging.

    Science.gov (United States)

    Collot, Mayeul; Loukou, Christina; Yakovlev, Aleksey V; Wilms, Christian D; Li, Dongdong; Evrard, Alexis; Zamaleeva, Alsu; Bourdieu, Laurent; Léger, Jean-François; Ropert, Nicole; Eilers, Jens; Oheim, Martin; Feltz, Anne; Mallet, Jean-Maurice

    2012-09-12

    We designed Calcium Rubies, a family of functionalizable BAPTA-based red-fluorescent calcium (Ca(2+)) indicators as new tools for biological Ca(2+) imaging. The specificity of this Ca(2+)-indicator family is its side arm, attached on the ethylene glycol bridge that allows coupling the indicator to various groups while leaving open the possibility of aromatic substitutions on the BAPTA core for tuning the Ca(2+)-binding affinity. Using this possibility we now synthesize and characterize three different CaRubies with affinities between 3 and 22 μM. Their long excitation and emission wavelengths (peaks at 586/604 nm) allow their use in otherwise challenging multicolor experiments, e.g., when combining Ca(2+) uncaging or optogenetic stimulation with Ca(2+) imaging in cells expressing fluorescent proteins. We illustrate this capacity by the detection of Ca(2+) transients evoked by blue light in cultured astrocytes expressing CatCh, a light-sensitive Ca(2+)-translocating channelrhodopsin linked to yellow fluorescent protein. Using time-correlated single-photon counting, we measured fluorescence lifetimes for all CaRubies and demonstrate a 10-fold increase in the average lifetime upon Ca(2+) chelation. Since only the fluorescence quantum yield but not the absorbance of the CaRubies is Ca(2+)-dependent, calibrated two-photon fluorescence excitation measurements of absolute Ca(2+) concentrations are feasible.

  16. Endogenous two-photon fluorescence imaging elucidates metabolic changes related to enhanced glycolysis and glutamine consumption in precancerous epithelial tissues.

    Science.gov (United States)

    Varone, Antonio; Xylas, Joanna; Quinn, Kyle P; Pouli, Dimitra; Sridharan, Gautham; McLaughlin-Drubin, Margaret E; Alonzo, Carlo; Lee, Kyongbum; Münger, Karl; Georgakoudi, Irene

    2014-06-01

    Alterations in the balance between different metabolic pathways used to meet cellular bioenergetic and biosynthetic demands are considered hallmarks of cancer. Optical imaging relying on endogenous fluorescence has been used as a noninvasive approach to assess tissue metabolic changes during cancer development. However, quantitative correlations of optical assessments with variations in the concentration of relevant metabolites or in the specific metabolic pathways that are involved have been lacking. In this study, we use high-resolution, depth-resolved imaging, relying entirely on endogenous two-photon excited fluorescence in combination with invasive biochemical assays and mass spectrometry to demonstrate the sensitivity and quantitative nature of optical redox ratio tissue assessments. We identify significant differences in the optical redox ratio of live, engineered normal and precancerous squamous epithelial tissues. We establish that while decreases in the optical redox ratio are associated with enhanced levels of glycolysis relative to oxidative phosphorylation, increases in glutamine consumption to support energy production are associated with increased optical redox ratio values. Such mechanistic insights in the origins of optical metabolic assessments are critical for exploiting fully the potential of such noninvasive approaches to monitor and understand important metabolic changes that occur in live tissues at the onset of cancer or in response to treatment.

  17. Functional double-shelled silicon nanocrystals for two-photon fluorescence cell imaging: spectral evolution and tuning

    Science.gov (United States)

    Chandra, Sourov; Ghosh, Batu; Beaune, Grégory; Nagarajan, Usharani; Yasui, Takao; Nakamura, Jin; Tsuruoka, Tohru; Baba, Yoshinobu; Shirahata, Naoto; Winnik, Françoise M.

    2016-04-01

    Functional near-IR (NIR) emitting nanoparticles (NPs) adapted for two-photon excitation fluorescence cell imaging were obtained starting from octadecyl-terminated silicon nanocrystals (ncSi-OD) of narrow photoluminescence (PL) spectra having no long emission tails, continuously tunable over the 700-1000 nm window, PL quantum yields exceeding 30%, and PL lifetimes of 300 μs or longer. These NPs, consisting of a Pluronic F127 shell and a core made up of assembled ncSi-OD kept apart by an octadecyl (OD) layer, were readily internalized into the cytosol, but not the nucleus, of NIH3T3 cells and were non-toxic. Asymmetrical field-flow fractionation (AF4) analysis was carried out to determine the size of the NPs in water. HiLyte Fluor 750 amine was linked via an amide link to NPs prepared with Pluronic-F127-COOH, as a first demonstration of functional NIR-emitting water dispersible ncSi-based nanoparticles.Functional near-IR (NIR) emitting nanoparticles (NPs) adapted for two-photon excitation fluorescence cell imaging were obtained starting from octadecyl-terminated silicon nanocrystals (ncSi-OD) of narrow photoluminescence (PL) spectra having no long emission tails, continuously tunable over the 700-1000 nm window, PL quantum yields exceeding 30%, and PL lifetimes of 300 μs or longer. These NPs, consisting of a Pluronic F127 shell and a core made up of assembled ncSi-OD kept apart by an octadecyl (OD) layer, were readily internalized into the cytosol, but not the nucleus, of NIH3T3 cells and were non-toxic. Asymmetrical field-flow fractionation (AF4) analysis was carried out to determine the size of the NPs in water. HiLyte Fluor 750 amine was linked via an amide link to NPs prepared with Pluronic-F127-COOH, as a first demonstration of functional NIR-emitting water dispersible ncSi-based nanoparticles. Electronic supplementary information (ESI) available. See DOI: 10.1039/c6nr01437b

  18. Fluorescent detection and imaging of Hg{sup 2+} using a novel phenanthroline derivative based single- and two-photon excitation

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xian, E-mail: zhangx@qlu.edu.cn; Li, Long-long; Liu, Ying-kai

    2016-02-01

    A novel phenanthroline derivative, 4-[4-(N-methyl)styrene]-imidazo[4,5-f][1,10]phenanthroline-benzene iodated salt (MSIPBI), was synthesized, and the linear absorption and fluorescent spectra of MSIPBI in different solvents were investigated. The photophysical properties in unbound and in ligand–metal complexes were evaluated by UV absorption and one- and two-photon fluorescent spectra, and the quantum yields, two-photon active cross-sections and the binding constant of dye–metal were calculated. The results indicated that MSIPBI has a large Stokes shift (more than 167 nm), and the dye was selective and sensitive for the detection of Hg{sup 2+} with a two-photon active cross-section of 55.5 GM in tris–HCl buffer solution at 800 nm. Furthermore, the results of the fluorescence microscopy imaging indicated that MSIPBI is an efficient fluorescent probe for the detection of Hg{sup 2+} in living cells by one- and two-photon excitation. Moreover, the experiments of determination Hg{sup 2+} in river water and tap water were finished. - Highlights: • A novel phenanthroline derivative (MSIPBI) has been synthesized. • The dye of MSIPBI was selective and sensitive to detect Hg{sup 2+}. • MSIPBI has a large Stokes shift (≥ 167 nm). • Hg{sup 2+} in living cells was successfully imaged by one- and two-photon excitation.

  19. Time Lapse Photography From Arctic Buoys

    Science.gov (United States)

    Valentic, T. A.; Matrai, P.; Woods, J. E.

    2013-12-01

    We have equipped a number of buoys with cameras that have been deployed throughout the Arctic. These systems need to be simple, reliable and low power. The images are transmitted over an Iridium satellite link and assembled into long running movies. We have captured a number of interesting events, observed the ice dynamics through the year and visits by local wildlife. Each of the systems have been deployed for periods of up to a year, with images every hour. The cameras have proved to be a great outreach tool and are routinely watched by number of people on our websites. This talk will present the techniques used in developing these camera systems, the methods used for reliably transmitting the images and the process for generating the movies.

  20. Multifunctional superparamagnetic nanoshells: combining two-photon luminescence imaging, surface-enhanced Raman scattering and magnetic separation

    Science.gov (United States)

    Jin, Xiulong; Li, Haiyan; Wang, Shanshan; Kong, Ni; Xu, Hong; Fu, Qihua; Gu, Hongchen; Ye, Jian

    2014-11-01

    With the increasing need for multi-purpose analysis in the biomedical field, traditional single diagnosis methods cannot meet the requirements. Therefore new multifunctional technologies and materials for the integration of sample collection, sensing and imaging are in great demand. Core-shell nanoparticles offer a unique platform to combine multifunctions in a single particle. In this work, we have constructed a novel type of core-shell superparamagnetic nanoshell (Fe3O4@SiO2@Au), composed of a Fe3O4 cluster core, a thin Au shell and a SiO2 layer in between. The obtained multifunctional nanoparticles combine the magnetic properties and plasmonic optical properties effectively, which were well investigated by a number of experimental characterization methods and theoretical simulations. We have demonstrated that Fe3O4@SiO2@Au nanoparticles can be utilized for two-photon luminescence (TPL) imaging, near-infrared surface-enhanced Raman scattering (NIR SERS) and cell collection by magnetic separation. The TPL intensity could be further greatly enhanced through the plasmon coupling effect in the self-assembled nanoparticle chains, which were triggered by an external magnetic field. In addition, Fe3O4@SiO2@Au nanoparticles may have great potential applications such as enhanced magnetic resonance imaging (MRI) and photo-thermotherapy. Successful combination of multifunctions including magnetic response, biosensing and bioimaging in single nanoparticles allows further manipulation, real-time tracking, and intracellular molecule analysis of live cells at a single-cell level.With the increasing need for multi-purpose analysis in the biomedical field, traditional single diagnosis methods cannot meet the requirements. Therefore new multifunctional technologies and materials for the integration of sample collection, sensing and imaging are in great demand. Core-shell nanoparticles offer a unique platform to combine multifunctions in a single particle. In this work, we have

  1. Two-photon physics

    Energy Technology Data Exchange (ETDEWEB)

    Bardeen, W.A.

    1981-10-01

    A new experimental frontier has recently been opened to the study of two photon processes. The first results of many aspects of these reactions are being presented at this conference. In contrast, the theoretical development of research ito two photon processes has a much longer history. This talk reviews the many different theoretical ideas which provide a detailed framework for our understanding of two photon processes.

  2. Curious case of gravitational lensing by binary black holes: a tale of two photon spheres, new relativistic images and caustics

    CERN Document Server

    Patil, Mandar; Narasimha, D

    2016-01-01

    Binary black holes have been in limelight off late due to the detection of gravitational waves from coalescing compact binaries in the events GW150914 and GW151226. In this paper we study gravitational lensing by the binary black holes modeled as equal mass Majumdar-Papapetrou dihole metric and show that this system displays features that are quite unprecedented and absent in any other lensing configuration investigated so far. We restrict our attention to the light rays which move on the plane midway between the two identical black holes, which allows us to employ techniques developed for the equatorial lensing in spherically symmetric spacetimes. If distance between the two black holes is below a certain threshold value, the system admits two photon spheres. As in the case of single black hole, infinitely many relativistic images are formed due to the light rays which turn back from the region outside the outer (unstable) photon sphere, all of which lie beyond a critical angular radius with respect to the l...

  3. Indole-based cyanine as a nuclear RNA-selective two-photon fluorescent probe for live cell imaging.

    Science.gov (United States)

    Guo, Lei; Chan, Miu Shan; Xu, Di; Tam, Dick Yan; Bolze, Frédéric; Lo, Pik Kwan; Wong, Man Shing

    2015-05-15

    We have demonstrated that the subcellular targeting properties of the indole-based cyanines can be tuned by the functional substituent attached onto the indole moiety in which the first example of a highly RNA-selective and two-photon active fluorescent light-up probe for high contrast and brightness TPEF images of rRNA in the nucleolus of live cells has been developed. It is important to find that this cyanine binds much stronger toward RNA than DNA in a buffer solution as well as selectively stains and targets to rRNA in the nucleolus. Remarkably, the TPEF brightness (Φσmax) is dramatically increased with 11-fold enhancement in the presence of rRNA, leading to the record high Φσmax of 228 GM for RNA. This probe not only shows good biocompatibility and superior photostability but also offers general applicability to various live cell lines including HeLa, HepG2, MCF-7, and KB cells and excellent counterstaining compatibility with commercially available DNA or protein trackers.

  4. Two-Photon Processor and SeNeCA: a freely available software package to process data from two-photon calcium imaging at speeds down to several milliseconds per frame.

    Science.gov (United States)

    Tomek, Jakub; Novak, Ondrej; Syka, Josef

    2013-07-01

    Two-Photon Processor (TPP) is a versatile, ready-to-use, and freely available software package in MATLAB to process data from in vivo two-photon calcium imaging. TPP includes routines to search for cell bodies in full-frame (Search for Neural Cells Accelerated; SeNeCA) and line-scan acquisition, routines for calcium signal calculations, filtering, spike-mining, and routines to construct parametric fields. Searching for somata in artificial in vivo data, our algorithm achieved better performance than human annotators. SeNeCA copes well with uneven background brightness and in-plane motion artifacts, the major problems in simple segmentation methods. In the fast mode, artificial in vivo images with a resolution of 256 × 256 pixels containing ≈ 100 neurons can be processed at a rate up to 175 frames per second (tested on Intel i7, 8 threads, magnetic hard disk drive). This speed of a segmentation algorithm could bring new possibilities into the field of in vivo optophysiology. With such a short latency (down to 5-6 ms on an ordinary personal computer) and using some contemporary optogenetic tools, it will allow experiments in which a control program can continuously evaluate the occurrence of a particular spatial pattern of activity (a possible correlate of memory or cognition) and subsequently inhibit/stimulate the entire area of the circuit or inhibit/stimulate a different part of the neuronal system. TPP will be freely available on our public web site. Similar all-in-one and freely available software has not yet been published.

  5. Two-photon excited surface plasmon enhanced energy transfer between DAPI and gold nanoparticles: Opportunities in intra-cellular imaging and sensing

    Science.gov (United States)

    Zhang, Yinan; Birch, David J. S.; Chen, Yu

    2011-09-01

    We have demonstrated energy transfer between 4'-6-Diamidino-2-phenylindole (DAPI), a commonly used DNA label, and gold nanoparticles under two-photon excitation in solution using fluorescence lifetime imaging microscopy (FLIM). With comparable size and concentration, gold nanorods (GNRs) are shown to provide more efficient energy transfer than gold nanospheres (GNSs). We attribute this transfer enhancement effect to the longitudinal surface plasmon mode of GNRs overlapping with the excitation wavelength. Energy transfer under two-photon excitation between GNRs and DAPI has also been observed in cell culture and found to be in accord with the solution phase results.

  6. Direct Vpr-Vpr Interaction in Cells monitored by two Photon Fluorescence Correlation Spectroscopy and Fluorescence Lifetime Imaging

    Directory of Open Access Journals (Sweden)

    Mély Yves

    2008-09-01

    Full Text Available Abstract Background The human immunodeficiency virus type 1 (HIV-1 encodes several regulatory proteins, notably Vpr which influences the survival of the infected cells by causing a G2/M arrest and apoptosis. Such an important role of Vpr in HIV-1 disease progression has fuelled a large number of studies, from its 3D structure to the characterization of specific cellular partners. However, no direct imaging and quantification of Vpr-Vpr interaction in living cells has yet been reported. To address this issue, eGFP- and mCherry proteins were tagged by Vpr, expressed in HeLa cells and their interaction was studied by two photon fluorescence lifetime imaging microscopy and fluorescence correlation spectroscopy. Results Results show that Vpr forms homo-oligomers at or close to the nuclear envelope. Moreover, Vpr dimers and trimers were found in the cytoplasm and in the nucleus. Point mutations in the three α helices of Vpr drastically impaired Vpr oligomerization and localization at the nuclear envelope while point mutations outside the helical regions had no effect. Theoretical structures of Vpr mutants reveal that mutations within the α-helices could perturb the leucine zipper like motifs. The ΔQ44 mutation has the most drastic effect since it likely disrupts the second helix. Finally, all Vpr point mutants caused cell apoptosis suggesting that Vpr-mediated apoptosis functions independently from Vpr oligomerization. Conclusion We report that Vpr oligomerization in HeLa cells relies on the hydrophobic core formed by the three α helices. This oligomerization is required for Vpr localization at the nuclear envelope but not for Vpr-mediated apoptosis.

  7. Development of image-guided targeted two-photon PDT for the treatment of head and neck cancers

    Science.gov (United States)

    Spangler, Charles W.; Starkey, Jean R.; Liang, Bo; Fedorka, Sara; Yang, Hao; Jiang, Huabei

    2014-03-01

    There has been significant effort over the past two decades in the treatment of malignancies of epithelial origin, including some of the most devastating of cancers, such as colorectal cancer (CRC), squamous call carcinoma of the head and neck (HNSCC), and carcinomas of the pancreas, lungs, (both Small Cell and Non-Small Cell), renal cell, prostate, bladder and breast. Recurring, refractory HNSCC is a particularly difficult cancer to treat once the tumors recur due to mutations that are resistant to repeat chemotherapy and radiation. In addition, repeat surgery is often difficult due to the requirement of significant surgical margins that may not be possible due to the attending potential functional deficits (e.g., salivary glands, nerves and major blood vessels in confined areas). In this study FaDu HNSCC xenograft tumors in SCID mice were imaged, and "optical", as opposed to "surgical" margins defined for the tumor being treated. The subsequent two-photon treatment irradiation was computer-controlled to carry out the tumor treatment by rastering the laser beam throughout the tumor volume plus the defined optical margins simultaneously. In our initial studies, up to 85% regression in tumor volume was observed in 5 days post PDT, with complete tumor regression in 18 days. No re-growth was observed up to 41 days post-PDT, with little or no scarring and complete hair re-growth. However, competition between imaging and PDT moieties was also observed in some mouse models, possibly favoring tumor re-growth. Strategies to selectively optimize the PDT effect will be discussed.

  8. Nonlinear spectral imaging of human hypertrophic scar based on two-photon excited fluorescence and second-harmonic generation.

    Science.gov (United States)

    Chen, G; Chen, J; Zhuo, S; Xiong, S; Zeng, H; Jiang, X; Chen, R; Xie, S

    2009-07-01

    A noninvasive method using microscopy and spectroscopy for analysing the morphology of collagen and elastin and their biochemical variations in skin tissue will enable better understanding of the pathophysiology of hypertrophic scars and facilitate improved clinical management and treatment of this disease. To obtain simultaneously microscopic images and spectra of collagen and elastin fibres in ex vivo skin tissues (normal skin and hypertrophic scar) using a nonlinear spectral imaging method, and to compare the morphological structure and spectral characteristics of collagen and elastin fibres in hypertrophic scar tissues with those of normal skin, to determine whether this approach has potential for in vivo assessment of the pathophysiology of human hypertrophic scars and for monitoring treatment responses as well as for tracking the process of development of hypertrophic scars in clinic. Ex vivo human skin specimens obtained from six patients aged from 10 to 50 years old who were undergoing skin plastic surgery were examined. Five patients had hypertrophic scar lesions and one patient had no scar lesion before we obtained his skin specimen. A total of 30 tissue section samples of 30 mum thickness were analysed by the use of a nonlinear spectral imaging system consisting of a femtosecond excitation light source, a high-throughput scanning inverted microscope, and a spectral imaging detection system. The high-contrast and high-resolution second harmonic generation (SHG) images of collagen and two-photon excited fluorescence (TPEF) images of elastin fibres in hypertrophic scar tissues and normal skin were acquired using the extracting channel tool of the system. The emission spectra were analysed using the image-guided spectral analysis method. The depth-dependent decay constant of the SHG signal and the image texture characteristics of hypertrophic scar tissue and normal skin were used to quantitatively assess the amount, distribution and orientation of their

  9. Time-Lapse Acoustic Impedance Inversion in CO2 Sequestration Study (Weyburn Field, Canada)

    Science.gov (United States)

    Wang, Y.; Morozov, I. B.

    2016-12-01

    Acoustic-impedance (AI) pseudo-logs are useful for characterising subtle variations of fluid content during seismic monitoring of reservoirs undergoing enhanced oil recovery and/or geologic CO2 sequestration. However, highly accurate AI images are required for time-lapse analysis, which may be difficult to achieve with conventional inversion approaches. In this study, two enhancements of time-lapse AI analysis are proposed. First, a well-known uncertainty of AI inversion is caused by the lack of low-frequency signal in reflection seismic data. To resolve this difficulty, we utilize an integrated AI inversion approach combining seismic data, acoustic well logs and seismic-processing velocities. The use of well logs helps stabilizing the recursive AI inverse, and seismic-processing velocities are used to complement the low-frequency information in seismic records. To derive the low-frequency AI from seismic-processing velocity data, an empirical relation is determined by using the available acoustic logs. This method is simple and does not require subjective choices of parameters and regularization schemes as in the more sophisticated joint inversion methods. The second improvement to accurate time-lapse AI imaging consists in time-variant calibration of reflectivity. Calibration corrections consist of time shifts, amplitude corrections, spectral shaping and phase rotations. Following the calibration, average and differential reflection amplitudes are calculated, from which the average and differential AI are obtained. The approaches are applied to a time-lapse 3-D 3-C dataset from Weyburn CO2 sequestration project in southern Saskatchewan, Canada. High quality time-lapse AI volumes are obtained. Comparisons with traditional recursive and colored AI inversions (obtained without using seismic-processing velocities) show that the new method gives a better representation of spatial AI variations. Although only early stages of monitoring seismic data are available, time-lapse

  10. Two-photon microscopy with diffractive optical elements and spatial light modulators

    Directory of Open Access Journals (Sweden)

    Brendon O Watson

    2010-09-01

    Full Text Available Two-photon microscopy is often performed at slow frame rates, due to the need to serially scan all points in a field of view with a single laser beam. To overcome this problem, we have developed two optical methods that split and multiplex a laser beam across the sample. In the first method a diffractive optical element (DOE generates a fixed number of beamlets that are scanned in parallel, resulting in a corresponding increase in speed, or in signal-to-noise ratio, in time-lapse measurements. The second method uses a computer-controlled spatial light modulator (SLM, to generate any arbitrary spatio-temporal light pattern. With an SLM one can image or photostimulate any predefined region of the image, such as neurons or dendritic spines. In addition, SLMs can be used to mimic a large number of optical transfer functions, including light path corrections or as adaptive optical devices.

  11. Selected time-lapse movies of the east rift zone eruption of KĪlauea Volcano, 2004–2008

    Science.gov (United States)

    Orr, Tim R.

    2011-01-01

    Since 2004, the U.S. Geological Survey's Hawaiian Volcano Observatory has used mass-market digital time-lapse cameras and network-enabled Webcams for visual monitoring and research. The 26 time-lapse movies in this report were selected from the vast collection of images acquired by these camera systems during 2004–2008. Chosen for their content and broad aesthetic appeal, these image sequences document a variety of flow-field and vent processes from Kīlauea's east rift zone eruption, which began in 1983 and is still (as of 2011) ongoing.

  12. Asante Calcium Green and Asante Calcium Red--novel calcium indicators for two-photon fluorescence lifetime imaging.

    Science.gov (United States)

    Jahn, Karolina; Hille, Carsten

    2014-01-01

    For a comprehensive understanding of cellular processes and potential dysfunctions therein, an analysis of the ubiquitous intracellular second messenger calcium is of particular interest. This study examined the suitability of the novel Ca2+-sensitive fluorescent dyes Asante Calcium Red (ACR) and Asante Calcium Green (ACG) for two-photon (2P)-excited time-resolved fluorescence measurements. Both dyes displayed sufficient 2P fluorescence excitation in a range of 720-900 nm. In vitro, ACR and ACG exhibited a biexponential fluorescence decay behavior and the two decay time components in the ns-range could be attributed to the Ca(2+)-free and Ca(2+)-bound dye species. The amplitude-weighted average fluorescence decay time changed in a Ca(2+)-dependent way, unraveling in vitro dissociation constants K(D) of 114 nM and 15 nM for ACR and ACG, respectively. In the presence of bovine serum albumin, the absorption and steady-state fluorescence behavior of ACR was altered and its biexponential fluorescence decay showed about 5-times longer decay time components indicating dye-protein interactions. Since no ester derivative of ACG was commercially available, only ACR was evaluated for 2P-excited fluorescence lifetime imaging microscopy (2P-FLIM) in living cells of American cockroach salivary glands. In living cells, ACR also exhibited a biexponential fluorescence decay with clearly resolvable short (0.56 ns) and long (2.44 ns) decay time components attributable to the Ca(2+)-free and Ca(2+)-bound ACR species. From the amplitude-weighted average fluorescence decay times, an in situ K(D) of 180 nM was determined. Thus, quantitative [Ca(2+)]i recordings were realized, unraveling a reversible dopamine-induced [Ca(2+)]i elevation from 21 nM to 590 nM in salivary duct cells. It was concluded that ACR is a promising new Ca(2+) indicator dye for 2P-FLIM recordings applicable in diverse biological systems.

  13. Asante Calcium Green and Asante Calcium Red--novel calcium indicators for two-photon fluorescence lifetime imaging.

    Directory of Open Access Journals (Sweden)

    Karolina Jahn

    Full Text Available For a comprehensive understanding of cellular processes and potential dysfunctions therein, an analysis of the ubiquitous intracellular second messenger calcium is of particular interest. This study examined the suitability of the novel Ca2+-sensitive fluorescent dyes Asante Calcium Red (ACR and Asante Calcium Green (ACG for two-photon (2P-excited time-resolved fluorescence measurements. Both dyes displayed sufficient 2P fluorescence excitation in a range of 720-900 nm. In vitro, ACR and ACG exhibited a biexponential fluorescence decay behavior and the two decay time components in the ns-range could be attributed to the Ca(2+-free and Ca(2+-bound dye species. The amplitude-weighted average fluorescence decay time changed in a Ca(2+-dependent way, unraveling in vitro dissociation constants K(D of 114 nM and 15 nM for ACR and ACG, respectively. In the presence of bovine serum albumin, the absorption and steady-state fluorescence behavior of ACR was altered and its biexponential fluorescence decay showed about 5-times longer decay time components indicating dye-protein interactions. Since no ester derivative of ACG was commercially available, only ACR was evaluated for 2P-excited fluorescence lifetime imaging microscopy (2P-FLIM in living cells of American cockroach salivary glands. In living cells, ACR also exhibited a biexponential fluorescence decay with clearly resolvable short (0.56 ns and long (2.44 ns decay time components attributable to the Ca(2+-free and Ca(2+-bound ACR species. From the amplitude-weighted average fluorescence decay times, an in situ K(D of 180 nM was determined. Thus, quantitative [Ca(2+]i recordings were realized, unraveling a reversible dopamine-induced [Ca(2+]i elevation from 21 nM to 590 nM in salivary duct cells. It was concluded that ACR is a promising new Ca(2+ indicator dye for 2P-FLIM recordings applicable in diverse biological systems.

  14. Two-photon excited endogenous fluorescence for label-free in vivo imaging ingestion of disease-causing bacteria by human leukocytes

    Science.gov (United States)

    Zeng, Yan; Yan, Bo; Sun, Qiqi; Teh, Seng Khoon; Zhang, Wei; Wen, Zilong; Qu, Jianan Y.

    2013-02-01

    Real time and in vivo monitoring leukocyte behavior provides unique information to understand the physiological and pathological process of infection. In this study, we demonstrate that two-photon excited reduced nicotinamide adenine dinucleotide (NADH) fluorescence provides imaging contrast to distinguish granulocyte and agranulocyte. By using spectral and time-resolved NADH fluorescence, we study the immune response of human neutrophils against bacterial infection (Escherichia coli). The two-photon excited NADH fluorescence images clearly review the morphological changes from resting neutrophils (round shape) to activated neutrophils (ruffle shape) during phagocytosis. The free-tobound NADH ratio of neutrophils decreases after ingesting disease-causing pathogen: Escherichia coli. This finding may provide a new optical tool to investigate inflammatory processes by using NADH fluorescence in vivo.

  15. A two-photon fluorescent probe for exogenous and endogenous superoxide anion imaging in vitro and in vivo.

    Science.gov (United States)

    Li, Run-Qing; Mao, Zhi-Qiang; Rong, Lei; Wu, Nian; Lei, Qi; Zhu, Jing-Yi; Zhuang, Lin; Zhang, Xian-Zheng; Liu, Zhi-Hong

    2017-01-15

    Herein, we report a novel quinoline derivative-based two-photon fluorescent probe 6-(dimethylamino)quinoline-2-benzothiazoline (HQ), which is capable of tracking superoxide anion in organisms with specific "turn-on" fluorescence response based on extension of π-conjugations and moderate ICT process. The probe exhibited favorable photophysical properties, a broad linear range and high photostability. It can specifically detect superoxide anion with a significant fluorescence enhancement and great linearity from 0 to 500μM in PBS buffer. Furthermore, HQ shows low cytotoxicity and excellent photostability toward living cells and organisms, which was able to monitor endogenous superoxide anion fluxes in living cells and in vivo. For the first time, endogenous superoxide anion in lung inflammation was visualized successfully by using HQ through two-photon microscopy, and the probe HQ shows great potential for fast in-situ detecting of inflammatory response in live organisms.

  16. The application of time-lapse photography for the observation of snow processes in mountainous catchments

    Science.gov (United States)

    Garvelmann, J.; Pohl, S.; Weiler, M.

    2012-04-01

    For the forecast of snowmelt flood events in mountainous catchments it is very important to know the spatial distribution and temporal evolution of the snowcover. Topography and vegetation have the most important influence on the spatio-temporal variability of the snowcover. In order to accomplish a continuous observation of the quantity and the status of the snowcover, an extensive measurement network consisting of numerous standalone snow and meteorological sensors and time-lapse photography was established in three catchments in the Black Forest, a typical mid latitude medium elevation mountain range. Catchments with different topographic characteristic and areal extent were specifically chosen for this study. Within the catchments, a stratified sampling design was used to cover a wide range of altitudes and exposures. In order to investigate the influence of a vegetation cover on the snow processes beneath sensors and cameras have been installed under the forest canopy and on adjacent open field sites, respectively. In the presented study the application of spatially distributed time-lapse cameras for the observation of snow processes and snowcover properties at the catchment scale will be discussed. Image analysis software was applied to extract information about snowdepth, snow albedo and canopy interception from the digital images. A measurement scale with a black/white board was installed in the focus of every camera to allow a determination of the snowdepth at every camera location while the black/white board was used to provide a white balance for the albedo estimation. The albedo provides important information about the status of the snowcover and its temporal evolution is a crucial factor for the snowmelt energy balance. Furthermore the time-lapse images provided a continuous observation of the forest canopy allowing the estimation of the interception efficiency and the temporal evolution of the snow interception for different topographic situations

  17. Probabilistic 3-D time-lapse inversion of magnetotelluric data: Application to an enhanced geothermal system

    Science.gov (United States)

    Rosas-Carbajal, Marina; Linde, Nicolas; Peacock, Jared R.; Zyserman, F. I.; Kalscheuer, Thomas; Thiel, Stephan

    2015-01-01

    Surface-based monitoring of mass transfer caused by injections and extractions in deep boreholes is crucial to maximize oil, gas and geothermal production. Inductive electromagnetic methods, such as magnetotellurics, are appealing for these applications due to their large penetration depths and sensitivity to changes in fluid conductivity and fracture connectivity. In this work, we propose a 3-D Markov chain Monte Carlo inversion of time-lapse magnetotelluric data to image mass transfer following a saline fluid injection. The inversion estimates the posterior probability density function of the resulting plume, and thereby quantifies model uncertainty. To decrease computation times, we base the parametrization on a reduced Legendre moment decomposition of the plume. A synthetic test shows that our methodology is effective when the electrical resistivity structure prior to the injection is well known. The centre of mass and spread of the plume are well retrieved.We then apply our inversion strategy to an injection experiment in an enhanced geothermal system at Paralana, South Australia, and compare it to a 3-D deterministic time-lapse inversion. The latter retrieves resistivity changes that are more shallow than the actual injection interval, whereas the probabilistic inversion retrieves plumes that are located at the correct depths and oriented in a preferential north-south direction. To explain the time-lapse data, the inversion requires unrealistically large resistivity changes with respect to the base model. We suggest that this is partly explained by unaccounted subsurface heterogeneities in the base model from which time-lapse changes are inferred.

  18. Calibrating vadose zone models with time-lapse gravity data

    DEFF Research Database (Denmark)

    Christiansen, Lars; Binning, Philip John; Rosbjerg, Dan

    2011-01-01

    The vadose zone plays an important role in the hydrologic cycle. Various geophysical methods can determine soil water content variations in time and space in volumes ranging from a few cubic centimeters to several cubic meters. In contrast to the established methods, time-lapse gravity measurements...... of changes in soil water content do not rely on a petrophysical relationship between the measured quantity and the water content but give a direct measure of the mass change in the soil. Only recently has the vadose zone been systematically incorporated when ground-based gravity data are used to infer...... hydrologic information. In this study, changes in the soil water content gave rise to a measurable signal in a forced infiltration experiment on a 107-m2 grassland area. Time-lapse gravity data were able to constrain the van Genuchten soil hydraulic parameters in both a synthetic example and a field...

  19. N-doped carbon dots derived from bovine serum albumin and formic acid with one- and two-photon fluorescence for live cell nuclear imaging.

    Science.gov (United States)

    Tan, Mingqian; Li, Xintong; Wu, Hao; Wang, Beibei; Wu, Jing

    2015-12-01

    Carbon dots with both one- and two-photon fluorescence have drawn great attention for biomedical imaging. Herein, nitrogen-doped carbon dots were facilely developed by one-pot hydrothermal method using bovine serum albumin and formic acid as carbon sources. They are highly water-soluble with strong fluorescence when excited with ultraviolet or near infrared light. The carbon dots have a diameter of ~8.32 nm and can emit strong two-photon induced fluorescence upon excitation at 750 nm with a femtosecond laser. X-ray photoelectron spectrometer analysis revealed that the carbon dots contained three components, C, N and O, corresponding to the peak at 285, 398 and 532 eV, respectively. The Fourier-transform infrared spectroscopy analysis revealed that there are carboxyl and carboxylic groups on the surface, which allowed further linking of functional molecules. pH stability study demonstrated that the carbon dots are able to be used in a wide range of pH values. The fluorescence mechanism is also discussed in this study. Importantly, these carbon dots are biocompatible and highly photostable, which can be directly applied for both one- and two-photon living cell imaging. After proper surface functionalization with TAT peptide, they can be used as fluorescent probes for live cell nuclear-targeted imaging.

  20. Deep two-photon microscopic imaging through brain tissue using the second singlet state from fluorescent agent chlorophyll α in spinach leaf.

    Science.gov (United States)

    Shi, Lingyan; Rodríguez-Contreras, Adrián; Budansky, Yury; Pu, Yang; Nguyen, Thien An; Alfano, Robert R

    2014-06-01

    Two-photon (2P) excitation of the second singlet (S₂) state was studied to achieve deep optical microscopic imaging in brain tissue when both the excitation (800 nm) and emission (685 nm) wavelengths lie in the "tissue optical window" (650 to 950 nm). S₂ state technique was used to investigate chlorophyll α (Chl α) fluorescence inside a spinach leaf under a thick layer of freshly sliced rat brain tissue in combination with 2P microscopic imaging. Strong emission at the peak wavelength of 685 nm under the 2P S₂ state of Chl α enabled the imaging depth up to 450 μm through rat brain tissue.

  1. Time-lapse 3D VSP monitoring of a carbon dioxide injection project at Delhi Field, Louisiana

    Science.gov (United States)

    Lubis, Muhammad Husni Mubarak

    Delhi Field is a producing oil field located in northeastern Louisiana. The estimated original oil in place (OOIP) is 357 mmbo and approximately 54% of OOIP has been produced through the primary production and water-flooding. A CO2-EOR program has been implemented since November 2009 to recover an additional 17% of OOIP. Reservoir surveillance using time-lapse 3D seismic data has been conducted to monitor the CO2 sweep efficiency. The goal of this study is to monitor the CO2 flow-path in the area around the injector using time-lapse 3D VSP data. For this purpose, two 3D VSPs acquired in June 2010 and again in August 2011 were processed together. Fluid substitution and VSP modeling were performed to understand the influence of pore-fluid saturation change on VSP records. A cross-equalization was performed to improve the similarity of the datasets. This step is important to reduce the ambiguity in time-lapse observation. The splice of a 3D VSP image into the surface seismic data becomes the key point in determining the reflector of the reservoir. By integrating the observation from the modeling and the splice of 3D VSP image to surface seismic, the CO2 flow-path from injector 164-3 can be identified from 3D time-lapse VSP data. The CO2 was not radially distributed around the injector, but moved toward southwest direction. This finding is also consistent with the flow-path interpreted from surface seismic. This consistency implies that time-lapse 3D VSP surveys at Delhi Field confirm and augment the time-lapse interpretation from surface seismic data.

  2. Label-free NIR reflectance imaging as a complimentary tool for two-photon fluorescence microscopy: multimodal investigation of stroke (Conference Presentation)

    Science.gov (United States)

    Allegra Mascaro, Anna Letizia; Costantini, Irene; Margoni, Emilia; Iannello, Giulio; Bria, Alessandro; Sacconi, Leonardo; Pavone, Francesco S.

    2016-03-01

    Two-photon imaging combined with targeted fluorescent indicators is extensively used for visualizing critical features of brain functionality and structural plasticity. Back-scattered photons from the NIR laser provide complimentary information without introducing any exogenous labelling. Here, we describe a versatile approach that, by collecting the reflected NIR light, provides structural details on the myelinated axons and blood vessels in the brain, both in fixed samples and in live animals. Indeed, by combining NIR reflectance and two-photon imaging of a slice of hippocampus from Thy1-GFPm mice, we show the presence of randomly oriented axons intermingled with sparsely fluorescent neuronal processes. The back-scattered photons guide the contextualization of the fluorescence structure within brain atlas thanks to the recognition of characteristic hippocampal structures. Label-free detection of axonal elongations over the layer 2/3 of mouse cortex under a cranial window was also possible in live brain. Finally, blood flow could be measured in vivo, thus validating label free NIR reflectance as a tool for monitoring hemodynamic fluctuations. The prospective versatility of this label-free technique complimentary to two-photon fluorescence microscopy is demonstrated in a mouse model of photothrombotic stroke in which the axonal degeneration and blood flow remodeling can be investigated simultaneously.

  3. Multifunctional biocompatible graphene oxide quantum dots decorated magnetic nanoplatform for efficient capture and two-photon imaging of rare tumor cells.

    Science.gov (United States)

    Shi, Yongliang; Pramanik, Avijit; Tchounwou, Christine; Pedraza, Francisco; Crouch, Rebecca A; Chavva, Suhash Reddy; Vangara, Aruna; Sinha, Sudarson Sekhar; Jones, Stacy; Sardar, Dhiraj; Hawker, Craig; Ray, Paresh Chandra

    2015-05-27

    Circulating tumor cells (CTCs) are extremely rare cells in blood containing billions of other cells. The selective capture and identification of rare cells with sufficient sensitivity is a real challenge. Driven by this need, this manuscript reports the development of a multifunctional biocompatible graphene oxide quantum dots (GOQDs) coated, high-luminescence magnetic nanoplatform for the selective separation and diagnosis of Glypican-3 (GPC3)-expressed Hep G2 liver cancer tumor CTCs from infected blood. Experimental data show that an anti-GPC3-antibody-attached multifunctional nanoplatform can be used for selective Hep G2 hepatocellular carcinoma tumor cell separation from infected blood containing 10 tumor cells/mL of blood in a 15 mL sample. Reported data indicate that, because of an extremely high two-photon absorption cross section (40530 GM), an anti-GPC3-antibody-attached GOQDs-coated magnetic nanoplatform can be used as a two-photon luminescence platform for selective and very bright imaging of a Hep G2 tumor cell in a biological transparency window using 960 nm light. Experimental results with nontargeted GPC3(-) and SK-BR-3 breast cancer cells show that multifunctional-nanoplatform-based cell separation, followed by two-photon imaging, is highly selective for Hep G2 hepatocellular carcinoma tumor cells.

  4. Cell and brain tissue imaging of the flavonoid fisetin using label-free two-photon microscopy.

    Science.gov (United States)

    Krasieva, Tatiana B; Ehren, Jennifer; O'Sullivan, Thomas; Tromberg, Bruce J; Maher, Pamela

    2015-10-01

    Over the last few years, we have identified an orally active, novel neuroprotective and cognition-enhancing molecule, the flavonoid fisetin. Fisetin not only has direct antioxidant activity but it can also increase the intracellular levels of glutathione, the major intracellular antioxidant. Fisetin can also activate key neurotrophic factor signaling pathways. In addition, it has anti-inflammatory activity against microglia and astrocytes and inhibits the activity of lipoxygenases, thereby reducing the production of pro-inflammatory eicosanoids and their by-products. However, key questions about its targets and brain penetration remain. In this study, we used label-free two-photon microscopy of intrinsic fisetin fluorescence to examine the localization of fisetin in living nerve cells and the brains of living mice. In cells, fisetin but not structurally related flavonols with different numbers of hydroxyl groups, localized to the nucleoli suggesting that key targets of fisetin may reside in this organelle. In the mouse brain, following intraperitoneal injection and oral administration, fisetin rapidly distributed to the blood vessels of the brain followed by a slower dispersion into the brain parenchyma. Thus, these results provide further support for the effects of fisetin on brain function. In addition, they suggest that label-free two-photon microscopy may prove useful for studying the intracellular and tissue distribution of other intrinsically-fluorescent flavonoids. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Two-photon spectral fluorescence lifetime and second-harmonic generation imaging of the porcine cornea with a 12-femtosecond laser microscope

    Science.gov (United States)

    Batista, Ana; Breunig, Hans Georg; Uchugonova, Aisada; Morgado, António Miguel; König, Karsten

    2016-03-01

    Five dimensional microscopy with a 12-fs laser scanning microscope based on spectrally resolved two-photon autofluorescence lifetime and second-harmonic generation (SHG) imaging was used to characterize all layers of the porcine cornea. This setup allowed the simultaneous excitation of both metabolic cofactors, NAD(P)H and flavins, and their discrimination based on their spectral emission properties and fluorescence decay characteristics. Furthermore, the architecture of the stromal collagen fibrils was assessed by SHG imaging in both forward and backward directions. Information on the metabolic state and the tissue architecture of the porcine cornea were obtained with subcellular resolution, and high temporal and spectral resolutions.

  6. Combined nonlinear laser imaging (two-photon excitation fluorescence, second and third-harmonic generation, and fluorescence lifetime imaging microscopies) in ovarian tumors

    Science.gov (United States)

    Adur, J.; Pelegati, V. B.; de Thomaz, A. A.; Bottcher-Luiz, F.; Andrade, L. A. L. A.; Almeida, D. B.; Carvalho, H. F.; Cesar, C. L.

    2012-03-01

    We applied Two-photon Excited Fluorescence (TPEF), Second/Third Harmonic Generation (SHG and THG) and Fluorescence Lifetime Imaging (FLIM) Non Linear Optics (NLO) Laser-Scanning Microscopy within the same imaging platform to evaluate their use as a diagnostic tool in ovarian tumors. We assess of applicability of this multimodal approach to perform a pathological evaluation of serous and mucinous tumors in human samples. The combination of TPEF-SHG-THG imaging provided complementary information about the interface epithelium/stromal, such as the transformation of epithelium surface (THG) and the overall fibrillar tissue architecture (SHG). The fact that H&E staining is the standard method used in clinical pathology and that the stored samples are usually fixed makes it important a re-evaluation of these samples with NLO microscopy to compare new results with a library of already existing samples. FLIM, however, depends on the chemical environment around the fluorophors that was completely changed after fixation; therefore it only makes sense in unstained samples. Our FLIM results in unstained samples demonstrate that it is possible to discriminate healthy epithelia from serous or mucinous epithelia. Qualitative and quantitative analysis of the different imaging modalities used showed that multimodal nonlinear microscopy has the potential to differentiate between cancerous and healthy ovarian tissue.

  7. Time-lapse seismic waveform modelling and attribute analysis using hydromechanical models for a deep reservoir undergoing depletion

    Science.gov (United States)

    He, Y.-X.; Angus, D. A.; Blanchard, T. D.; Wang, G.-L.; Yuan, S.-Y.; Garcia, A.

    2016-04-01

    Extraction of fluids from subsurface reservoirs induces changes in pore pressure, leading not only to geomechanical changes, but also perturbations in seismic velocities and hence observable seismic attributes. Time-lapse seismic analysis can be used to estimate changes in subsurface hydromechanical properties and thus act as a monitoring tool for geological reservoirs. The ability to observe and quantify changes in fluid, stress and strain using seismic techniques has important implications for monitoring risk not only for petroleum applications but also for geological storage of CO2 and nuclear waste scenarios. In this paper, we integrate hydromechanical simulation results with rock physics models and full-waveform seismic modelling to assess time-lapse seismic attribute resolution for dynamic reservoir characterization and hydromechanical model calibration. The time-lapse seismic simulations use a dynamic elastic reservoir model based on a North Sea deep reservoir undergoing large pressure changes. The time-lapse seismic traveltime shifts and time strains calculated from the modelled and processed synthetic data sets (i.e. pre-stack and post-stack data) are in a reasonable agreement with the true earth models, indicating the feasibility of using 1-D strain rock physics transform and time-lapse seismic processing methodology. Estimated vertical traveltime shifts for the overburden and the majority of the reservoir are within ±1 ms of the true earth model values, indicating that the time-lapse technique is sufficiently accurate for predicting overburden velocity changes and hence geomechanical effects. Characterization of deeper structure below the overburden becomes less accurate, where more advanced time-lapse seismic processing and migration is needed to handle the complex geometry and strong lateral induced velocity changes. Nevertheless, both migrated full-offset pre-stack and near-offset post-stack data image the general features of both the overburden and

  8. Time-Lapse Video Microscopy for Assessment of EYFP-Parkin Aggregation as a Marker for Cellular Mitophagy.

    Science.gov (United States)

    Di Sante, Gabriele; Casimiro, Mathew C; Pestell, Timothy G; Pestell, Richard G

    2016-05-04

    Time-lapse video microscopy can be defined as the real time imaging of living cells. This technique relies on the collection of images at different time points. Time intervals can be set through a computer interface that controls the microscope-integrated camera. This kind of microscopy requires both the ability to acquire very rapid events and the signal generated by the observed cellular structure during these events. After the images have been collected, a movie of the entire experiment is assembled to show the dynamic of the molecular events of interest. Time-lapse video microscopy has a broad range of applications in the biomedical research field and is a powerful and unique tool for following the dynamics of the cellular events in real time. Through this technique, we can assess cellular events such as migration, division, signal transduction, growth, and death. Moreover, using fluorescent molecular probes we are able to mark specific molecules, such as DNA, RNA or proteins and follow them through their molecular pathways and functions. Time-lapse video microscopy has multiple advantages, the major one being the ability to collect data at the single-cell level, that make it a unique technology for investigation in the field of cell biology. However, time-lapse video microscopy has limitations that can interfere with the acquisition of high quality images. Images can be compromised by both external factors; temperature fluctuations, vibrations, humidity and internal factors; pH, cell motility. Herein, we describe a protocol for the dynamic acquisition of a specific protein, Parkin, fused with the enhanced yellow fluorescent protein (EYFP) in order to track the selective removal of damaged mitochondria, using a time-lapse video microscopy approach.

  9. Interferometric full-waveform inversion of time-lapse data

    KAUST Repository

    Sinha, Mrinal

    2017-08-17

    One of the key challenges associated with time-lapse surveys is ensuring the repeatability between the baseline and monitor surveys. Non-repeatability between the surveys is caused by varying environmental conditions over the course of different surveys. To overcome this challenge, we propose the use of interferometric full waveform inversion (IFWI) for inverting the velocity model from data recorded by baseline and monitor surveys. A known reflector is used as the reference reflector for IFWI, and the data are naturally redatumed to this reference reflector using natural reflections as the redatuming operator. This natural redatuming mitigates the artifacts introduced by the repeatability errors that originate above the reference reflector.

  10. Inter and intra-observer variability of time-lapse annotations

    DEFF Research Database (Denmark)

    Sundvall Germeys, Linda Karin M; Ingerslev, Hans Jakob; Knudsen, Ulla Breth;

    already: The commonly employed method used to assess embryos in IVF treatments is based on static evaluation of morphology in a microscope, but this is limited by substantial intra- and inter-observer variation. Time-lapse imaging has been proposed as a method to refine embryo selection by adding new......-lapse recordings. One observer performed the assessment twice. Twenty-five parameters were annotated and the inter- and intra-observer agreement was assessed by calculating Intra-class correlation coefficients (ICC). Main results and the role of chance: Extremely close agreement (ICC 0.99) was found for dynamic...

  11. A software solution for recording circadian oscillator features in time-lapse live cell microscopy

    Directory of Open Access Journals (Sweden)

    Salmon Patrick

    2010-07-01

    Full Text Available Abstract Background Fluorescent and bioluminescent time-lapse microscopy approaches have been successfully used to investigate molecular mechanisms underlying the mammalian circadian oscillator at the single cell level. However, most of the available software and common methods based on intensity-threshold segmentation and frame-to-frame tracking are not applicable in these experiments. This is due to cell movement and dramatic changes in the fluorescent/bioluminescent reporter protein during the circadian cycle, with the lowest expression level very close to the background intensity. At present, the standard approach to analyze data sets obtained from time lapse microscopy is either manual tracking or application of generic image-processing software/dedicated tracking software. To our knowledge, these existing software solutions for manual and automatic tracking have strong limitations in tracking individual cells if their plane shifts. Results In an attempt to improve existing methodology of time-lapse tracking of a large number of moving cells, we have developed a semi-automatic software package. It extracts the trajectory of the cells by tracking theirs displacements, makes the delineation of cell nucleus or whole cell, and finally yields measurements of various features, like reporter protein expression level or cell displacement. As an example, we present here single cell circadian pattern and motility analysis of NIH3T3 mouse fibroblasts expressing a fluorescent circadian reporter protein. Using Circadian Gene Express plugin, we performed fast and nonbiased analysis of large fluorescent time lapse microscopy datasets. Conclusions Our software solution, Circadian Gene Express (CGE, is easy to use and allows precise and semi-automatic tracking of moving cells over longer period of time. In spite of significant circadian variations in protein expression with extremely low expression levels at the valley phase, CGE allows accurate and

  12. 3-D time-lapse electrical resistivity monitoring of injected CO2 in a shallow aquifer

    DEFF Research Database (Denmark)

    Doetsch, Joseph A. J.A.; Auken, Esben; Christiansen, Anders Vest C A.V.C.;

    2013-01-01

    Contamination of potable groundwater by leaking CO2 is a potential risk of carbon sequestration. With the help of a field experiment, we investigate if surface electrical resistivity tomography (ERT) can detect dissolved CO2 in a shallow aquifer. For this purpose, we injected CO2 at a depth of 5...... inversion reveals the geology at the site consisting of aeolian sands near the surface and glacial sands below 5 m depth. Time-lapse inversions clearly image the dissolved CO2 plume with decreased electrical resistivity values. We can follow the CO2 plume as it spreads and moves with the groundwater...

  13. Time-lapse ultrashort pulse microscopy of infection in three-dimensional versus two-dimensional culture environments reveals enhanced extra-chromosomal virus replication compartment formation

    Science.gov (United States)

    Gibbs, Holly C.; Sing, Garwin; Armas, Juan Carlos González; Campbell, Colin J.; Ghazal, Peter; Yeh, Alvin T.

    2013-03-01

    The mechanisms that enable viruses to harness cellular machinery for their own survival are primarily studied in cell lines cultured in two-dimensional (2-D) environments. However, there are increasing reports of biological differences between cells cultured in 2-D versus three-dimensional (3-D) environments. Here we report differences in host-virus interactions based on differences in culture environment. Using ultrashort pulse microscopy (UPM), a form of two-photon microscopy that utilizes sub-10-fs pulses to efficiently excite fluorophores, we have shown that de novo development of extra-chromosomal virus replication compartments (VRCs) upon murine cytomegalovirus (mCMV) infection is markedly enhanced when host cells are cultured in 3-D collagen gels versus 2-D monolayers. In addition, time-lapse imaging revealed that mCMV-induced VRCs have the capacity to grow by coalescence. This work supports the future potential of 3-D culture as a useful bridge between traditional monolayer cultures and animal models to study host-virus interactions in a more physiologically relevant environment for the development of effective anti-viral therapeutics. These advances will require broader adoption of modalities, such as UPM, to image deep within scattering tissues.

  14. Very-High-Resolution Time-Lapse Photography for Plant and Ecosystems Research

    Directory of Open Access Journals (Sweden)

    Mary H. Nichols

    2013-09-01

    Full Text Available Premise of the study: Traditional photography is a compromise between image detail and area covered. We report a new method for creating time-lapse sequences of very-high-resolution photographs to produce zoomable images that facilitate observation across a range of spatial and temporal scales. Methods and Results: A robotic camera mount and software were used to capture images of the growth and movement in Brassica rapa every 15 s in the laboratory. The resultant time-lapse sequence (http://timemachine.gigapan.org/wiki/Plant_Growth captures growth detail such as circumnutation. A modified, solar-powered system was deployed at a remote field site in southern Arizona. Images were collected every 2 h over a 3-mo period to capture the response of vegetation to monsoon season rainfall (http://timemachine.gigapan.org/wiki/Arizona_Grasslands. Conclusions: A technique for observing time sequences of both individual plant and ecosystem response at a range of spatial scales is available for use in the laboratory and in the field.

  15. Time-lapse 3D ground-penetrating radar during plot-scale infiltration experiments

    Science.gov (United States)

    Allroggen, Niklas; Jackisch, Conrad; Tronicke, Jens

    2016-04-01

    In electrical resistive soils, surface-based ground-penetrating radar (GPR) is known as the geophysical tool providing the highest spatial resolution. Thus, 2D and 3D GPR surveys are commonly used for imaging subsurface structures or estimating soil moisture content. Due to its sensitivity to soil moisture and its non-invasive character, GPR provides a large potential to monitor soil moisture variation at high temporal and spatial resolution. As shown in previous experiments, the acquisition of time-lapse GPR data under field conditions requires a high data quality in terms of repeatability as well as spatial and temporal resolution. We present hydrogeophysical field experiments at the plot scale (1m x 1m), during which we record time-lapse 3D GPR. For GPR data acquisition, we use a pulseEKKO PRO GPR system equipped with a pair of 500 MHz antennas in combination with a specially designed metal-free measuring platform. Additionally, we collect tracer and soil moisture data, which are used to improve the interpretation of the GPR data with special focus on preferential flow paths and their structured advective flow field. After an accurate time-lapse GPR data processing, we compare 3D reflection events before and after infiltration and quantitatively interpret their relative time-shift in terms of soil moisture variations. Thereby, we are able to account for basically all of the infiltrated water. The first experiments demonstrate the general applicability of our experimental approach but are limited by the number of acquired time steps and measurement during the sprinkling period (the time of the highest temporal dynamics) are not possible at all. Based on this experience we redesign our experimental setup to continuously collect GPR data during irrigation and infiltration. Thereby, we strongly increase the temporal resolution of our measurements, improve the interpretability of the GPR data, and monitor the temporal and spatial dynamics of shallow subsurface

  16. Two-photon deep imaging through skin and skull of Zebra finches: preliminary studies for in-vivo brain metabolism monitoring

    Science.gov (United States)

    Abi-Haidar, D.; Olivier, T.; Mottin, S.; Vignal, C.; Mathevon, N.

    2007-02-01

    Zebra Finches are songbirds which constitute a model for neuro-ethologists to study the neuro-mechanisms of vocal recognition. For this purpose, in vivo and non invasive monitoring of brain activity is required during acoustical stimulation. MRI (Magnetic Resonance Imaging) or NIRS (Near InfraRed Spectroscopy) are suitable methods for these measurements, even though MRI is difficult to link quantitatively with neural activity and NIRS suffers from a poor resolution. In the particular case of songbirds (whose skin is thin and quite transparent and whose skull structure is hollow), two-photon microscopy enables a quite deep penetration in tissues and could be an alternative. We present here preliminary studies on the feasability of two-photon microscopy in these conditions. To do so, we chose to image hollow fibers, filled with Rhodamine B, through the skin of Zebra finches in order to evaluate the spatial resolution we may expect in future in vivo experiments. Moreover, we used the reflectance-mode confocal configuration to evaluate the exponential decrease of backreflected light in skin and in skull samples. Following this procedure recently proposed by S.L. Jacques and co-workers, we planned to determine the scattering coefficient μ s and the anisotropy g of these tissues and make a comparison between fixed and fresh skin and skull samples for future Monte Carlo simulations of the scattering in our particular multi-layered structure.

  17. Rock Slope Monitoring from 4D Time-Lapse Structure from Motion Analysis

    Science.gov (United States)

    Kromer, Ryan; Abellan, Antonio; Chyz, Alex; Hutchinson, Jean

    2017-04-01

    more accurate 3D models using time-lapse photos taken from an array of cameras than photos taken from a single camera from multiple positions. For this survey setup, it was possible to detect mm to cm level of changes, which is of sufficient accuracy to detect the pre-failure stage of rockfalls, as well as small rockfall events. Additionally, cameras mounted in a static array can be operated remotely and automatically. Time-lapse SfM photogrammetry can be a cost effective alternative to terrestrial laser scanning for rockfall prone areas and facilitates the study of surface processes with high spatial and temporal detail. We gratefully acknowledge support from the NSERC collaborative research and development grant. References Eltner, A., Kaiser, A., Castillo, C.; Rock, G., Neugirg, F., Abellán, A. Image-based surface reconstruction in geomorphometry—Merits, limits and developments. Earth Surf. Dyn. 2016, 4, 359-389. Kromer, R. A., Abellán, A., Hutchinson, D. J., Lato, M., Edwards, T., & Jaboyedoff, M. A 4D filtering and calibration technique for small-scale point cloud change detection with a terrestrial laser scanner. Remote Sensing 2015, 7(10), 13029-13052.

  18. Combined time-lapse cinematography and immuno-electron microscopy.

    Science.gov (United States)

    Balfour, B M; Goscicka, T; MacKenzie, J L; Gautam, A; Tate, M; Clark, J

    1990-04-01

    A method was developed to record interactions between mobile non-adherent immunocytes by time-lapse cinematography and then to study the same cells by immuno-electron microscopy, using monoclonal antibodies against surface components. For this purpose a modified stage was designed to fit an inverted microscope. The attachment included a device to cool the culture chamber with N2 gas, a micro-injector for monoclonal antibody and immuno-gold treatment, and two pairs of washing needles to change the medium without disturbance. The technique was first employed to study the formation of aggregates around the antigen-presenting cells in cultures containing cells from hyper-immunized animals. Recently peripheral blood cells from normal subjects and patients with immune deficiency syndromes were stimulated with pokeweed mitogen, cluster formation was recorded, and the cells were processed for immuno-electron microscopy.

  19. A Prototype System for Time-Lapse Electrical Resistivity Tomographies

    Directory of Open Access Journals (Sweden)

    Raffaele Luongo

    2012-01-01

    Full Text Available A prototype system for time-lapse acquisition of 2D electrical resistivity tomography (ERT and time domain reflectometry (TDR measurements was installed in a test site affected by a landslide in Basilicata region (southern Italy. The aim of the system is to monitor in real-time the rainwater infiltration into the soil and obtain information about the variation of the water content in the first layers of the subsoil and the possible influence of this variation on landslide activity. A rain gauge placed in the test site gives information on the rainfall intensity and frequency and suggests the acquisition time interval. The installed system and the preliminary results are presented in this paper.

  20. Time-lapse microscopy patent upheld in Europe.

    Science.gov (United States)

    Pearce, David

    2017-02-01

    A case for revoking Stanford University's European patent 2430454 on time-lapse microscopy was set out in Reproductive BioMedicine Online by Sterckx et al. in 2014, on the grounds that the patent claimed a method of diagnosis that was excluded under a provision of the European Patent Convention. An opposition at the European Patent Office in which this ground was raised has recently concluded with a decision that the patent is not excluded from patentability under European patent law and is to be upheld. An appeal from this decision has been filed, but the possibility of the decision being overturned is, in this author's opinion, very limited. Copyright © 2016 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  1. Label-free imaging of brain and brain tumor specimens with combined two-photon excited fluorescence and second harmonic generation microscopy

    Science.gov (United States)

    Jiang, Liwei; Wang, Xingfu; Wu, Zanyi; Du, Huiping; Wang, Shu; Li, Lianhuang; Fang, Na; Lin, Peihua; Chen, Jianxin; Kang, Dezhi; Zhuo, Shuangmu

    2017-10-01

    Label-free imaging techniques are gaining acceptance within the medical imaging field, including brain imaging, because they have the potential to be applied to intraoperative in situ identifications of pathological conditions. In this paper, we describe the use of two-photon excited fluorescence (TPEF) and second harmonic generation (SHG) microscopy in combination for the label-free detection of brain and brain tumor specimens; gliomas. Two independently detecting channels were chosen to subsequently collect TPEF/SHG signals from the specimen to increase TPEF/SHG image contrasts. Our results indicate that the combined TPEF/SHG microscopic techniques can provide similar rat brain structural information and produce a similar resolution like conventional H&E staining in neuropathology; including meninges, cerebral cortex, white-matter structure corpus callosum, choroid plexus, hippocampus, striatum, and cerebellar cortex. It can simultaneously detect infiltrating human brain tumor cells, the extracellular matrix collagen fiber of connective stroma within brain vessels and collagen depostion in tumor microenvironments. The nuclear-to-cytoplasmic ratio and collagen content can be extracted as quantitative indicators for differentiating brain gliomas from healthy brain tissues. With the development of two-photon fiberscopes and microendoscope probes and their clinical applications, the combined TPEF and SHG microcopy may become an important multimodal, nonlinear optical imaging approach for real-time intraoperative histological diagnostics of residual brain tumors. These occur in various brain regions during ongoing surgeries through the method of simultaneously identifying tumor cells, and the change of tumor microenvironments, without the need for the removal biopsies and without the need for tissue labelling or fluorescent markers.

  2. Monitoring channel head erosion processes in response to an artificially induced abrupt base level change using time-lapse photography

    Science.gov (United States)

    Nichols, M. H.; Nearing, M.; Hernandez, M.; Polyakov, V. O.

    2016-07-01

    Gullies that terminate at a vertical-wall are ubiquitous throughout arid and semiarid regions. Multi-year assessments of gully evolution and headcut advance are typically accomplished using traditional ground surveys and aerial photographs, with much recent research focused on integrating data collected at very high spatial resolutions using new techniques such as aerial surveys with blimps or kites and ground surveys with LiDar scanners. However, knowledge of specific processes that drive headcut advance is limited due to inadequate observation and documentation of flash floods and subsequent erosion that can occur at temporal resolutions not captured through repeat surveys. This paper presents a method for using very-high temporal resolution ground-based time-lapse photography to capture short-duration flash floods and gully head evolution in response. In 2004, a base level controlling concrete weir was removed from the outlet of a 1.29 ha semiarid headwater drainage on the Walnut Gulch Experimental Watershed in southeastern Arizona, USA. During the ten year period from 2004 to 2014 the headcut migrated upchannel a total of 14.5 m reducing the contributing area at the headwall by 9.5%. Beginning in July 2012, time-lapse photography was employed to observe event scale channel evolution dynamics. The most frequent erosion processes observed during three seasons of time-lapse photography were plunge pool erosion and mass wasting through sidewall or channel headwall slumping that occurred during summer months. Geomorphic change during the ten year period was dominated by a single piping event in August 2014 that advanced the channel head 7.4 m (51% of the overall advance) and removed 11.3 m3 of sediment. High temporal resolution time-lapse photography was critical for identifying subsurface erosion processes, in the absence of time-lapse images piping would not have been identified as an erosion mechanism responsible for advancing the gully headwall at this site.

  3. Possible mechanism of polyspermy block in human oocytes observed by time-lapse cinematography.

    Science.gov (United States)

    Mio, Yasuyuki; Iwata, Kyoko; Yumoto, Keitaro; Kai, Yoshiteru; Sargant, Haruka C; Mizoguchi, Chizuru; Ueda, Minako; Tsuchie, Yuka; Imajo, Akifumi; Iba, Yumiko; Nishikori, Kyoko

    2012-09-01

    To analyze the fertilization process related to polyspermy block in human oocytes using an in vitro culturing system for time-lapse cinematography. We had 122 oocytes donated for this study from couples that provided informed consent. We recorded human oocytes at 2,000 to 2,800 frames every 10 s during the fertilization process and thereafter every 2 min using a new in vitro culture system originally developed by the authors for time-lapse cinematography. We displayed 30 frames per second for analysis of the polyspermy block during fertilization. Three oocytes showed the leading and following sperm within the zona pellucida in the same microscopic field. The dynamic images obtained during the fertilization process using this new system revealed that once a leading sperm penetrated the zona pellucida and attached to the oocyte membrane, a following sperm was arrested from further penetration into the zona pellucida within 10 s. The present results strongly suggest the existence of a novel mechanism of polyspermy block that takes place at the zona pellucida immediately after fertilization. These findings are clearly different from previous mechanisms describing polyspermy block as the oocyte membrane block to sperm penetration and the zona reaction. The finding presented herein thus represents a novel discovery about the highly complicated polyspermy block mechanism occurring in human oocytes.

  4. Time-lapse cinematography in living Drosophila tissues: preparation of material.

    Science.gov (United States)

    Davis, Ilan; Parton, Richard M

    2006-11-01

    The fruit fly, Drosophila melanogaster, has been an extraordinarily successful model organism for studying the genetic basis of development and evolution. It is arguably the best-understood complex multicellular model system, owing its success to many factors. Recent developments in imaging techniques, in particular sophisticated fluorescence microscopy methods and equipment, now allow cellular events to be studied at high resolution in living material. This ability has enabled the study of features that tend to be lost or damaged by fixation, such as transient or dynamic events. Although many of the techniques of live cell imaging in Drosophila are shared with the greater community of cell biologists working on other model systems, studying living fly tissues presents unique difficulties in keeping the cells alive, introducing fluorescent probes, and imaging through thick hazy cytoplasm. This protocol outlines the preparation of major tissue types amenable to study by time-lapse cinematography and different methods for keeping them alive.

  5. Femtosecond, two-photon-absorption, laser-induced-fluorescence (fs-TALIF) imaging of atomic hydrogen and oxygen in non-equilibrium plasmas

    Science.gov (United States)

    Schmidt, Jacob B.; Roy, Sukesh; Kulatilaka, Waruna D.; Shkurenkov, Ivan; Adamovich, Igor V.; Lempert, Walter R.; Gord, James R.

    2017-01-01

    Femtosecond, two-photon-absorption laser-induced fluorescence (fs-TALIF) is employed to measure space- and time-resolved distributions of atomic hydrogen and oxygen in moderate-pressure, non-equilibrium, nanosecond-duration pulsed-discharge plasmas. Temporally and spatially resolved hydrogen and oxygen TALIF images are obtained over a range of low-temperature plasmas in mixtures of helium and argon at 100 Torr total pressure. The high-peak-intensity, low-average-energy fs pulses combined with the increased spectral bandwidth compared to traditional ns-duration laser pulses provide a large number of photon pairs that are responsible for the two-photon excitation, which results in an enhanced TALIF signal. Krypton and xenon TALIF are used for quantitative calibration of the hydrogen and oxygen concentrations, respectively, with similar excitation schemes being employed. This enables 2D collection of atomic-hydrogen and -oxygen TALIF signals with absolute number densities ranging from 2  ×  1012 cm-3 to 6  ×  1015 cm-3 and 1  ×  1013 cm-3 to 3  ×  1016 cm-3, respectively. These 2D images are the first application of TALIF imaging in moderate-pressure plasma discharges. 1D self-consistent modeling predictions show agreement with experimental results within the estimated experimental error of 25%. The present results can be used to further the development of higher fidelity kinetic models while quantifying plasma-source characteristics.

  6. Nonlinear spectral imaging of human normal skin, basal cell carcinoma and squamous cell carcinoma based on two-photon excited fluorescence and second-harmonic generation

    Science.gov (United States)

    Xiong, S. Y.; Yang, J. G.; Zhuang, J.

    2011-10-01

    In this work, we use nonlinear spectral imaging based on two-photon excited fluorescence (TPEF) and second harmonic generation (SHG) for analyzing the morphology of collagen and elastin and their biochemical variations in basal cell carcinoma (BCC), squamous cell carcinoma (SCC) and normal skin tissue. It was found in this work that there existed apparent differences among BCC, SCC and normal skin in terms of their thickness of the keratin and epithelial layers, their size of elastic fibers, as well as their distribution and spectral characteristics of collagen. These differences can potentially be used to distinguish BCC and SCC from normal skin, and to discriminate between BCC and SCC, as well as to evaluate treatment responses.

  7. 4D (x-y-z-t) imaging of thick biological samples by means of Two-Photon inverted Selective Plane Illumination Microscopy (2PE-iSPIM).

    Science.gov (United States)

    Lavagnino, Zeno; Sancataldo, Giuseppe; d'Amora, Marta; Follert, Philipp; De Pietri Tonelli, Davide; Diaspro, Alberto; Cella Zanacchi, Francesca

    2016-04-01

    In the last decade light sheet fluorescence microscopy techniques, such as selective plane illumination microscopy (SPIM), has become a well established method for developmental biology. However, conventional SPIM architectures hardly permit imaging of certain tissues since the common sample mounting procedure, based on gel embedding, could interfere with the sample morphology. In this work we propose an inverted selective plane microscopy system (iSPIM), based on non-linear excitation, suitable for 3D tissue imaging. First, the iSPIM architecture provides flexibility on the sample mounting, getting rid of the gel-based mounting typical of conventional SPIM, permitting 3D imaging of hippocampal slices from mouse brain. Moreover, all the advantages brought by two photon excitation (2PE) in terms of reduction of scattering effects and contrast improvement are exploited, demonstrating an improved image quality and contrast compared to single photon excitation. The system proposed represents an optimal platform for tissue imaging and it smooths the way to the applicability of light sheet microscopy to a wider range of samples including those that have to be mounted on non-transparent surfaces.

  8. Iceberg Calving and Flow Dynamics at Helheim Glacier, East Greenland, from Time-Lapse Photography

    Science.gov (United States)

    Hamilton, G. S.; Khan, S. A.; Schild, K. M.; Stearns, L. A.; Nettles, M.; Ahlstrøm, A. P.; Andersen, M. L.; Davis, J. L.; Ekström, G.; Elósegui, P.; Forsberg, R.; de Juan, J.; Larsen, T. B.; Stenseng, L.

    2008-12-01

    Helheim Glacier in East Greenland is the focus of coordinated studies aimed at understanding tidewater outlet-glacier dynamics and kinematics, and their link to glacial earthquakes. As part of this effort, we installed three time-lapse cameras overlooking the calving terminus of the glacier during the Arctic summer of 2008. Images were captured every five minutes during the mostly unattended period of operation. Several interesting aspects of the glacier's behavior are observed in the image sequences, including vertical displacement of the glacier terminus by ocean tides, and very large calving events. These observations, in combination with simultaneous measurements of ice flow, ocean tides (including tsunamis) and seismic activity, contribute to our understanding of the dynamics of Helheim Glacier and the source mechanism of glacial earthquakes.

  9. Network Analysis of Time-Lapse Microscopy Recordings

    Directory of Open Access Journals (Sweden)

    Erik eSmedler

    2014-09-01

    Full Text Available Multicellular organisms rely on intercellular communication to regulate important cellular processes critical to life. To further our understanding of those processes there is a need to scrutinize dynamical signaling events and their functions in both cells and organisms. Here, we report a method and provide MATLAB code that analyzes time-lapse microscopy recordings to identify and characterize network structures within large cell populations, such as interconnected neurons. The approach is demonstrated using intracellular calcium (Ca2+ recordings in neural progenitors and cardiac myocytes, but could be applied to a wide variety of biosensors employed in diverse cell types and organisms. In this method, network structures are analyzed by applying cross-correlation signal processing and graph theory to single-cell recordings. The goal of the analysis is to determine if the single cell activity constitutes a network of interconnected cells and to decipher the properties of this network. The method can be applied in many fields of biology in which biosensors are used to monitor signaling events in living cells. Analyzing intercellular communication in cell ensembles can reveal essential network structures that provide important biological insights.

  10. Automated filtering of intrinsic movement artifacts during two-photon intravital microscopy.

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    Denis Soulet

    Full Text Available In vivo imaging using two-photon microscopy is an essential tool to explore the dynamic of physiological events deep within biological tissues for short or extended periods of time. The new capabilities offered by this technology (e.g. high tissue penetrance, low toxicity have opened a whole new era of investigations in modern biomedical research. However, the potential of using this promising technique in tissues of living animals is greatly limited by the intrinsic irregular movements that are caused by cardiac and respiratory cycles and muscular and vascular tone. Here, we show real-time imaging of the brain, spinal cord, sciatic nerve and myenteric plexus of living mice using a new automated program, named Intravital_Microscopy_Toolbox, that removes frames corrupted with motion artifacts from time-lapse videos. Our approach involves generating a dissimilarity score against precalculated reference frames in a specific reference channel, thus allowing the gating of distorted, out-of-focus or translated frames. Since the algorithm detects the uneven peaks of image distortion caused by irregular animal movements, the macro allows a fast and efficient filtering of the image sequence. In addition, extra features have been implemented in the macro, such as XY registration, channel subtraction, extended field of view with maximum intensity projection, noise reduction with average intensity projections, and automated timestamp and scale bar overlay. Thus, the Intravital_Microscopy_Toolbox macro for ImageJ provides convenient tools for biologists who are performing in vivo two-photon imaging in tissues prone to motion artifacts.

  11. Intravital two-photon microscopy of immune cell dynamics in corneal lymphatic vessels.

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    Philipp Steven

    Full Text Available BACKGROUND: The role of lymphatic vessels in tissue and organ transplantation as well as in tumor growth and metastasis has drawn great attention in recent years. METHODOLOGY/PRINCIPAL FINDINGS: We now developed a novel method using non-invasive two-photon microscopy to simultaneously visualize and track specifically stained lymphatic vessels and autofluorescent adjacent tissues such as collagen fibrils, blood vessels and immune cells in the mouse model of corneal neovascularization in vivo. The mouse cornea serves as an ideal tissue for this technique due to its easy accessibility and its inducible and modifiable state of pathological hem- and lymphvascularization. Neovascularization was induced by suture placement in corneas of Balb/C mice. Two weeks after treatment, lymphatic vessels were stained intravital by intrastromal injection of a fluorescently labeled LYVE-1 antibody and the corneas were evaluated in vivo by two-photon microscopy (TPM. Intravital TPM was performed at 710 nm and 826 nm excitation wavelengths to detect immunofluorescence and tissue autofluorescence using a custom made animal holder. Corneas were then harvested, fixed and analyzed by histology. Time lapse imaging demonstrated the first in vivo evidence of immune cell migration into lymphatic vessels and luminal transport of individual cells. Cells immigrated within 1-5.5 min into the vessel lumen. Mean velocities of intrastromal corneal immune cells were around 9 µm/min and therefore comparable to those of T-cells and macrophages in other mucosal surfaces. CONCLUSIONS: To our knowledge we here demonstrate for the first time the intravital real-time transmigration of immune cells into lymphatic vessels. Overall this study demonstrates the valuable use of intravital autofluorescence two-photon microscopy in the model of suture-induced corneal vascularizations to study interactions of immune and subsequently tumor cells with lymphatic vessels under close as possible

  12. Low-cost motility tracking system (LOCOMOTIS for time-lapse microscopy applications and cell visualisation.

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    Adam E Lynch

    Full Text Available Direct visualisation of cells for the purpose of studying their motility has typically required expensive microscopy equipment. However, recent advances in digital sensors mean that it is now possible to image cells for a fraction of the price of a standard microscope. Along with low-cost imaging there has also been a large increase in the availability of high quality, open-source analysis programs. In this study we describe the development and performance of an expandable cell motility system employing inexpensive, commercially available digital USB microscopes to image various cell types using time-lapse and perform tracking assays in proof-of-concept experiments. With this system we were able to measure and record three separate assays simultaneously on one personal computer using identical microscopes, and obtained tracking results comparable in quality to those from other studies that used standard, more expensive, equipment. The microscopes used in our system were capable of a maximum magnification of 413.6×. Although resolution was lower than that of a standard inverted microscope we found this difference to be indistinguishable at the magnification chosen for cell tracking experiments (206.8×. In preliminary cell culture experiments using our system, velocities (mean µm/min ± SE of 0.81 ± 0.01 (Biomphalaria glabrata hemocytes on uncoated plates, 1.17 ± 0.004 (MDA-MB-231 breast cancer cells, 1.24 ± 0.006 (SC5 mouse Sertoli cells and 2.21 ± 0.01 (B. glabrata hemocytes on Poly-L-Lysine coated plates, were measured and are consistent with previous reports. We believe that this system, coupled with open-source analysis software, demonstrates that higher throughput time-lapse imaging of cells for the purpose of studying motility can be an affordable option for all researchers.

  13. Low-cost motility tracking system (LOCOMOTIS) for time-lapse microscopy applications and cell visualisation.

    Science.gov (United States)

    Lynch, Adam E; Triajianto, Junian; Routledge, Edwin

    2014-01-01

    Direct visualisation of cells for the purpose of studying their motility has typically required expensive microscopy equipment. However, recent advances in digital sensors mean that it is now possible to image cells for a fraction of the price of a standard microscope. Along with low-cost imaging there has also been a large increase in the availability of high quality, open-source analysis programs. In this study we describe the development and performance of an expandable cell motility system employing inexpensive, commercially available digital USB microscopes to image various cell types using time-lapse and perform tracking assays in proof-of-concept experiments. With this system we were able to measure and record three separate assays simultaneously on one personal computer using identical microscopes, and obtained tracking results comparable in quality to those from other studies that used standard, more expensive, equipment. The microscopes used in our system were capable of a maximum magnification of 413.6×. Although resolution was lower than that of a standard inverted microscope we found this difference to be indistinguishable at the magnification chosen for cell tracking experiments (206.8×). In preliminary cell culture experiments using our system, velocities (mean µm/min ± SE) of 0.81 ± 0.01 (Biomphalaria glabrata hemocytes on uncoated plates), 1.17 ± 0.004 (MDA-MB-231 breast cancer cells), 1.24 ± 0.006 (SC5 mouse Sertoli cells) and 2.21 ± 0.01 (B. glabrata hemocytes on Poly-L-Lysine coated plates), were measured and are consistent with previous reports. We believe that this system, coupled with open-source analysis software, demonstrates that higher throughput time-lapse imaging of cells for the purpose of studying motility can be an affordable option for all researchers.

  14. Two-photon-excited fluorescence (TPEF) and fluorescence lifetime imaging (FLIM) with sub-nanosecond pulses and a high analog bandwidth signal detection

    Science.gov (United States)

    Eibl, Matthias; Karpf, Sebastian; Hakert, Hubertus; Weng, Daniel; Huber, Robert

    2017-02-01

    Two-photon excited fluorescence (TPEF) microscopy and fluorescence lifetime imaging (FLIM) are powerful imaging techniques in bio-molecular science. The need for elaborate light sources for TPEF and speed limitations for FLIM, however, hinder an even wider application. We present a way to overcome this limitations by combining a robust and inexpensive fiber laser for nonlinear excitation with a fast analog digitization method for rapid FLIM imaging. The applied sub nanosecond pulsed laser source is synchronized to a high analog bandwidth signal detection for single shot TPEF- and single shot FLIM imaging. The actively modulated pulses at 1064nm from the fiber laser are adjustable from 50ps to 5ns with kW of peak power. At a typically applied pulse lengths and repetition rates, the duty cycle is comparable to typically used femtosecond pulses and thus the peak power is also comparable at same cw-power. Hence, both types of excitation should yield the same number of fluorescence photons per time on average when used for TPEF imaging. However, in the 100ps configuration, a thousand times more fluorescence photons are generated per pulse. In this paper, we now show that the higher number of fluorescence photons per pulse combined with a high analog bandwidth detection makes it possible to not only use a single pulse per pixel for TPEF imaging but also to resolve the exponential time decay for FLIM. To evaluate the performance of our system, we acquired FLIM images of a Convallaria sample with pixel rates of 1 MHz where the lifetime information is directly measured with a fast real time digitizer. With the presented results, we show that longer pulses in the many-10ps to nanosecond regime can be readily applied for TPEF imaging and enable new imaging modalities like single pulse FLIM.

  15. Correlating two-photon excited fluorescence imaging of breast cancer cellular redox state with seahorse flux analysis of normalized cellular oxygen consumption

    Science.gov (United States)

    Hou, Jue; Wright, Heather J.; Chan, Nicole; Tran, Richard; Razorenova, Olga V.; Potma, Eric O.; Tromberg, Bruce J.

    2016-06-01

    Two-photon excited fluorescence (TPEF) imaging of the cellular cofactors nicotinamide adenine dinucleotide and oxidized flavin adenine dinucleotide is widely used to measure cellular metabolism, both in normal and pathological cells and tissues. When dual-wavelength excitation is used, ratiometric TPEF imaging of the intrinsic cofactor fluorescence provides a metabolic index of cells-the "optical redox ratio" (ORR). With increased interest in understanding and controlling cellular metabolism in cancer, there is a need to evaluate the performance of ORR in malignant cells. We compare TPEF metabolic imaging with seahorse flux analysis of cellular oxygen consumption in two different breast cancer cell lines (MCF-7 and MDA-MB-231). We monitor metabolic index in living cells under both normal culture conditions and, for MCF-7, in response to cell respiration inhibitors and uncouplers. We observe a significant correlation between the TPEF-derived ORR and the flux analyzer measurements (R=0.7901, p<0.001). Our results confirm that the ORR is a valid dynamic index of cell metabolism under a range of oxygen consumption conditions relevant for cancer imaging.

  16. Opposite reactivity of meningeal versus cortical microvessels to the nitric oxide donor glyceryl trinitrate evaluated in vivo with two-photon imaging.

    Directory of Open Access Journals (Sweden)

    Evgeny Pryazhnikov

    Full Text Available Vascular changes underlying headache in migraine patients induced by Glyceryl trinitrate (GTN were previously studied with various imaging techniques. Despite the long history of medical and experimental use of GTN, its effects on the brain vasculature are still poorly understood presumably due to low spatial resolution of the imaging modalities used so far. We took advantage of the micrometer-scale vertical resolution of two-photon microscopy to differentiate between the vasodynamic effects of GTN on meningeal versus cortical vessels imaged simultaneously in anesthetized rats through either thinned skull or glass-sealed cranial window. Intermediate and small calibre vessels were visualized in vivo by imaging intravascular fluorescent dextran, and detection of blood flow direction allowed identification of individual arterioles and venules. We found that i.p.-injected GTN induced a transient constriction of meningeal arterioles, while their cortical counterparts were, in contrast, dilated. These opposing effects of GTN were restricted to arterioles, whereas the effects on venules were insignificant. Interestingly, the NO synthase inhibitor L-NAME did not affect the diameter of meningeal vessels but induced a constriction of cortical vessels. The different cellular environment in cortex versus meninges as well as distinct vessel wall anatomical features probably play crucial role in the observed phenomena. These findings highlight differential region- and vessel-type-specific effects of GTN on cranial vessels, and may implicate new vascular mechanisms of NO-mediated primary headaches.

  17. Evaluation of the oxidative stress of psoriatic fibroblasts based on spectral two-photon fluorescence lifetime imaging

    Science.gov (United States)

    Kapsokalyvas, Dimitrios; Barygina, Victoria; Cicchi, Riccardo; Fiorillo, Claudia; Pavone, Francesco S.

    2013-02-01

    Psoriasis is an autoimmune disease of the skin characterized by hyperkeratosis, hyperproliferation of the epidermis, inflammatory cell accumulation and increased dilatation of dermal papillary blood vessels. Metabolic activity is increased in the epidermis and the dermis. Oxidative stress is high mainly due to reactive oxygen species (ROS) originating from the skin environment and cellular metabolism. We employed a custom multiphoton microscope coupled with a FLIM setup to image primary culture fibroblast cells from perilesional and lesional psoriatic skin in-vitro. Twophoton excited fluorescence images revealed the morphological differences between healthy and psoriatic fibroblasts. Based on the spectral analysis of the NADH and FAD components the oxidative stress was assessed and found to be higher in psoriatic cells. Furthermore the fluorescence lifetime properties were investigated with a TCSPC FLIM module. Mean fluorescence lifetime was found to be longer in psoriatic lesional cells. Analysis of the fast (τ1) and slow (τ2) decay lifetimes revealed a decrease of the ratio of the contribution of the fast (α1) parameter to the contribution of the slow (α2) parameter. The fluorescence in the examined part of the spectrum is attributed mainly to NADH. The decrease of the ratio (α1)/ (α2) is believed to correlate strongly with the anti-oxidant properties of NADH which can lead to the variation of its population in high ROS environment. This methodology could serve as an index of the oxidative status in cells and furthermore could be used to probe the oxidative stress of tissues in-vivo.

  18. The power of single and multibeam two-photon microscopy for high-resolution and high-speed deep tissue and intravital imaging.

    Science.gov (United States)

    Niesner, Raluca; Andresen, Volker; Neumann, Jens; Spiecker, Heinrich; Gunzer, Matthias

    2007-10-01

    Two-photon microscopy is indispensable for deep tissue and intravital imaging. However, current technology based on single-beam point scanning has reached sensitivity and speed limits because higher performance requires higher laser power leading to sample degradation. We utilize a multifocal scanhead splitting a laser beam into a line of 64 foci, allowing sample illumination in real time at full laser power. This technology requires charge-coupled device field detection in contrast to conventional detection by photomultipliers. A comparison of the optical performance of both setups shows functional equivalence in every measurable parameter down to penetration depths of 200 microm, where most actual experiments are executed. The advantage of photomultiplier detection materializes at imaging depths >300 microm because of their better signal/noise ratio, whereas only charge-coupled devices allow real-time detection of rapid processes (here blood flow). We also find that the point-spread function of both devices strongly depends on tissue constitution and penetration depth. However, employment of a depth-corrected point-spread function allows three-dimensional deconvolution of deep-tissue data up to an image quality resembling surface detection.

  19. A novel technique for the in vivo imaging of autoimmune diabetes development in the pancreas by two-photon microscopy.

    Directory of Open Access Journals (Sweden)

    Ken Coppieters

    Full Text Available Type 1 diabetes (T1D is characterized by the immune-mediated destruction of beta cells in the pancreas. Little is known about the in vivo dynamic interactions between T cells and beta cells or the kinetic behavior of other immune cell subsets in the pancreatic islets. Utilizing multiphoton microscopy we have designed a technique that allows for the real-time visualization of diabetogenic T cells and dendritic cells in pancreatic islets in a live animal, including their interplay with beta cells and the vasculature. Using a custom designed stage, the pancreas was surgically exposed under live conditions so that imaging of islets under intact blood pressure and oxygen supply became possible. We demonstrate here that this approach allows for the tracking of diabetogenic leukocytes as well as vascularization phenotype of islets and accumulation of dendritic cells in islets during diabetes pathogenesis. This technique should be useful in mapping crucial kinetic events in T1D pathogenesis and in testing the impact of immune based interventions on T cell migration, extravasation and islet destruction.

  20. Exploring Time-Lapse Photography as a Means for Qualitative Data Collection

    Science.gov (United States)

    Persohn, Lindsay

    2015-01-01

    Collecting information via time-lapse photography is nothing new. Scientists and artists have been using this kind of data since the late 1800s. However, my research and experiments with time-lapse have shown that great potential may lie in its application to educational and social scientific research methods. This article is part history, part…

  1. Exploring Time-Lapse Photography as a Means for Qualitative Data Collection

    Science.gov (United States)

    Persohn, Lindsay

    2015-01-01

    Collecting information via time-lapse photography is nothing new. Scientists and artists have been using this kind of data since the late 1800s. However, my research and experiments with time-lapse have shown that great potential may lie in its application to educational and social scientific research methods. This article is part history, part…

  2. Automatic cloud top height determination using a cost-effective time-lapse camera system

    Directory of Open Access Journals (Sweden)

    H. M. Schulz

    2014-03-01

    Full Text Available A new method for the determination of cloud top heights from the footage of a time-lapse camera is presented. Contact points between cloud tops and underlying terrain are automatically detected in the camera image based on differences in the brightness, texture and movement of cloudy and non-cloudy areas. The height of the detected cloud top positions is determined by comparison with a digital elevation model projected to the view of the camera. The technique has been validated using data about the cloud immersion of a second camera as well as via visual assessment. The validation shows a high detection quality, especially regarding the requirements for the validation of satellite cloud top retrieval algorithms.

  3. Electron microscopic time-lapse visualization of surface pore filtration on particulate matter trapping process.

    Science.gov (United States)

    Sanui, Ryoko; Hanamura, Katsunori

    2016-09-01

    A scanning electron microscope (SEM) was used to dynamically visualize the particulate matter (PM) trapping process on diesel particulate filter (DPF) walls at a micro scale as 'time-lapse' images corresponding to the increase in pressure drop simultaneously measured through the DPF. This visualization and pressure drop measurement led to the conclusion that the PM trapping in surface pores was driven by PM bridging and stacking at constricted areas in porous channels. This caused a drastic increase in the pressure drop during PM accumulation at the beginning of the PM trapping process. The relationship between the porous structure of the DPF and the depth of the surface pore was investigated in terms of the porosity distribution and PM penetration depth near the wall surface with respect to depth. The pressure drop calculated with an assumed surface pore depth showed a good correspondence to the measured pressure drop.

  4. Quantifying Effusion Rates at Active Volcanoes through Integrated Time-Lapse Laser Scanning and Photography

    Directory of Open Access Journals (Sweden)

    Neil Slatcher

    2015-11-01

    Full Text Available During volcanic eruptions, measurements of the rate at which magma is erupted underpin hazard assessments. For eruptions dominated by the effusion of lava, estimates are often made using satellite data; here, in a case study at Mount Etna (Sicily, we make the first measurements based on terrestrial laser scanning (TLS, and we also include explosive products. During the study period (17–21 July 2012, regular Strombolian explosions were occurring within the Bocca Nuova crater, producing a ~50 m-high scoria cone and a small lava flow field. TLS surveys over multi-day intervals determined a mean cone growth rate (effusive and explosive products of ~0.24 m3·s−1. Differences between 0.3-m resolution DEMs acquired at 10-minute intervals captured the evolution of a breakout lava flow lobe advancing at 0.01–0.03 m3·s−1. Partial occlusion within the crater prevented similar measurement of the main flow, but integrating TLS data with time-lapse imagery enabled lava viscosity (7.4 × 105 Pa·s to be derived from surface velocities and, hence, a flux of 0.11 m3·s−1 to be calculated. Total dense rock equivalent magma discharge estimates are ~0.1–0.2 m3·s−1 over the measurement period and suggest that simultaneous estimates from satellite data are somewhat overestimated. Our results support the use of integrated TLS and time-lapse photography for ground-truthing space-based measurements and highlight the value of interactive image analysis when automated approaches, such as particle image velocimetry (PIV, fail.

  5. Gliding motility of Babesia bovis merozoites visualized by time-lapse video microscopy.

    Directory of Open Access Journals (Sweden)

    Masahito Asada

    Full Text Available BACKGROUND: Babesia bovis is an apicomplexan intraerythrocytic protozoan parasite that induces babesiosis in cattle after transmission by ticks. During specific stages of the apicomplexan parasite lifecycle, such as the sporozoites of Plasmodium falciparum and tachyzoites of Toxoplasma gondii, host cells are targeted for invasion using a unique, active process termed "gliding motility". However, it is not thoroughly understood how the merozoites of B. bovis target and invade host red blood cells (RBCs, and gliding motility has so far not been observed in the parasite. METHODOLOGY/PRINCIPAL FINDINGS: Gliding motility of B. bovis merozoites was revealed by time-lapse video microscopy. The recorded images revealed that the process included egress of the merozoites from the infected RBC, gliding motility, and subsequent invasion into new RBCs. The gliding motility of B. bovis merozoites was similar to the helical gliding of Toxoplasma tachyzoites. The trails left by the merozoites were detected by indirect immunofluorescence assay using antiserum against B. bovis merozoite surface antigen 1. Inhibition of gliding motility by actin filament polymerization or depolymerization indicated that the gliding motility was driven by actomyosin dependent process. In addition, we revealed the timing of breakdown of the parasitophorous vacuole. Time-lapse image analysis of membrane-stained bovine RBCs showed formation and breakdown of the parasitophorous vacuole within ten minutes of invasion. CONCLUSIONS/SIGNIFICANCE: This is the first report of the gliding motility of B. bovis. Since merozoites of Plasmodium parasites do not glide on a substrate, the gliding motility of B. bovis merozoites is a notable finding.

  6. Active and passive electrical and seismic time-lapse monitoring of earthen embankments

    Science.gov (United States)

    Rittgers, Justin Bradley

    In this dissertation, I present research involving the application of active and passive geophysical data collection, data assimilation, and inverse modeling for the purpose of earthen embankment infrastructure assessment. Throughout the dissertation, I identify several data characteristics, and several challenges intrinsic to characterization and imaging of earthen embankments and anomalous seepage phenomena, from both a static and time-lapse geophysical monitoring perspective. I begin with the presentation of a field study conducted on a seeping earthen dam, involving static and independent inversions of active tomography data sets, and self-potential modeling of fluid flow within a confined aquifer. Additionally, I present results of active and passive time-lapse geophysical monitoring conducted during two meso-scale laboratory experiments involving the failure and self-healing of embankment filter materials via induced vertical cracking. Identified data signatures and trends, as well as 4D inversion results, are discussed as an underlying motivation for conducting subsequent research. Next, I present a new 4D acoustic emissions source localization algorithm that is applied to passive seismic monitoring data collected during a full-scale embankment failure test. Acoustic emissions localization results are then used to help spatially constrain 4D inversion of collocated self-potential monitoring data. I then turn to time-lapse joint inversion of active tomographic data sets applied to the characterization and monitoring of earthen embankments. Here, I develop a new technique for applying spatiotemporally varying structural joint inversion constraints. The new technique, referred to as Automatic Joint Constraints (AJC), is first demonstrated on a synthetic 2D joint model space, and is then applied to real geophysical monitoring data sets collected during a full-scale earthen embankment piping-failure test. Finally, I discuss some non-technical issues related to

  7. In Vivo Time-Course Imaging of Tumor Angiogenesis in Colorectal Liver Metastases in the Same Living Mice Using Two-Photon Laser Scanning Microscopy

    Directory of Open Access Journals (Sweden)

    Koji Tanaka

    2012-01-01

    Full Text Available In vivo real-time visualization of the process of angiogenesis in secondary tumors in the same living animals presents a major challenge in metastasis research. We developed a technique for intravital imaging of colorectal liver metastasis development in live mice using two-photon laser scanning microscopy (TPLSM. We also developed time-series TPLSM in which intravital TPLSM procedures were performed several times over periods of days to months. Red fluorescent protein-expressing colorectal cancer cells were inoculated into the spleens of green fluorescent protein-expressing mice. First- and second-round intravital TPLSM allowed visualization of viable cancer cells (red in hepatic sinusoids or the space of Disse. Third-round intravital TPLSM demonstrated liver metastatic colonies consisting of viable cancer cells and surrounding stroma with tumor vessels (green. In vivo time-course imaging of tumor angiogenesis in the same living mice using time-series TPLSM could be an ideal tool for antiangiogenic drug evaluation, reducing the effects of interindividual variation.

  8. Time-lapse three-dimensional inversion of complex conductivity data using an active time constrained (ATC) approach

    Science.gov (United States)

    Karaoulis, M.; Revil, A.; Werkema, D.D.; Minsley, B.J.; Woodruff, W.F.; Kemna, A.

    2011-01-01

    Induced polarization (more precisely the magnitude and phase of impedance of the subsurface) is measured using a network of electrodes located at the ground surface or in boreholes. This method yields important information related to the distribution of permeability and contaminants in the shallow subsurface. We propose a new time-lapse 3-D modelling and inversion algorithm to image the evolution of complex conductivity over time. We discretize the subsurface using hexahedron cells. Each cell is assigned a complex resistivity or conductivity value. Using the finite-element approach, we model the in-phase and out-of-phase (quadrature) electrical potentials on the 3-D grid, which are then transformed into apparent complex resistivity. Inhomogeneous Dirichlet boundary conditions are used at the boundary of the domain. The calculation of the Jacobian matrix is based on the principles of reciprocity. The goal of time-lapse inversion is to determine the change in the complex resistivity of each cell of the spatial grid as a function of time. Each model along the time axis is called a 'reference space model'. This approach can be simplified into an inverse problem looking for the optimum of several reference space models using the approximation that the material properties vary linearly in time between two subsequent reference models. Regularizations in both space domain and time domain reduce inversion artefacts and improve the stability of the inversion problem. In addition, the use of the time-lapse equations allows the simultaneous inversion of data obtained at different times in just one inversion step (4-D inversion). The advantages of this new inversion algorithm are demonstrated on synthetic time-lapse data resulting from the simulation of a salt tracer test in a heterogeneous random material described by an anisotropic semi-variogram. ?? 2011 The Authors Geophysical Journal International ?? 2011 RAS.

  9. Two photon physics. Personal recollection

    CERN Document Server

    Ginzburg, Ilya F

    2015-01-01

    The term two--photon processes is used for the reactions in which some system of particles is produced in collision of two photons, either real or virtual. In the study of these processes our main goal was to suggest approach, allowing to extract from the data information on proper two--photon process separating it from mechanism which responsible for the production of photons. Here I present my view for history of two--photon physics. I don't try to give complete review, concentrating mainly on works of our team (which cover essential part of the topic) and some colleagues. My citation is strongly incomplete. I cite here only papers which were essential in our understanding of the problems. The choice of presented details is the result of my discussions with Gleb Kotkin and Valery Serbo. 1. Prehistory. 2. Two photon processes at e^+e^- colliders. 3. Photon colliders. 4. Notes on physical program.

  10. Quantitative imaging of fibrotic and morphological changes in liver of non-alcoholic steatohepatitis (NASH) model mice by second harmonic generation (SHG) and auto-fluorescence (AF) imaging using two-photon excitation microscopy (TPEM).

    Science.gov (United States)

    Yamamoto, Shin; Oshima, Yusuke; Saitou, Takashi; Watanabe, Takao; Miyake, Teruki; Yoshida, Osamu; Tokumoto, Yoshio; Abe, Masanori; Matsuura, Bunzo; Hiasa, Yoichi; Imamura, Takeshi

    2016-12-01

    Non-alcoholic steatohepatitis (NASH) is a common liver disorder caused by fatty liver. Because NASH is associated with fibrotic and morphological changes in liver tissue, a direct imaging technique is required for accurate staging of liver tissue. For this purpose, in this study we took advantage of two label-free optical imaging techniques, second harmonic generation (SHG) and auto-fluorescence (AF), using two-photon excitation microscopy (TPEM). Three-dimensional ex vivo imaging of tissues from NASH model mice, followed by image processing, revealed that SHG and AF are sufficient to quantitatively characterize the hepatic capsule at an early stage and parenchymal morphologies associated with liver disease progression, respectively.

  11. Spectral-element simulations of carbon dioxide (CO2) sequestration time-lapse monitoring

    Science.gov (United States)

    Morency, C.; Luo, Y.; Tromp, J.

    2009-12-01

    Geologic sequestration of CO2, a green house gas, represents an effort to reduce the large amount of CO2 generated as a by-product of fossil fuels combustion and emitted into the atmosphere. This process of sequestration involves CO2 storage deep underground. There are three main storage options: injection into hydrocarbon reservoirs, injection into methane-bearing coal beds, or injection into deep saline aquifers, that is, highly permeable porous media. The key issues involve accurate monitoring of the CO2, from the injection stage to the prediction & verification of CO2 movement over time for environmental considerations. A natural non-intrusive monitoring technique is referred to as ``4D seismics'', which involves 3D time-lapse seismic surveys. The success of monitoring the CO2 movement is subject to a proper description of the physics of the problem. We propose to realize time-lapse migrations comparing acoustic, elastic, and poroelastic simulations of 4D seismic imaging to characterize the storage zone. This approach highlights the influence of using different physical theories on interpreting seismic data, and, more importantly, on extracting the CO2 signature from the seismic wave field. Our simulations are performed using a spectral-element method, which allows for highly accurate results. Biot's equations are implemented to account for poroelastic effects. Attenuation associated with the anelasticity of the rock frame and frequency-dependent viscous resistance of the pore fluid are accommodated based upon a memory variable approach. The sensitivity of observables to the model parameters is quantified based upon finite-frequency sensitivity kernels calculated using an adjoint method.

  12. In vivo two-photon imaging of axonal dieback, blood flow, and calcium influx with methylprednisolone therapy after spinal cord injury.

    Science.gov (United States)

    Tang, Peifu; Zhang, Yiling; Chen, Chao; Ji, Xinran; Ju, Furong; Liu, Xingyu; Gan, Wen-Biao; He, Zhigang; Zhang, Shengxiang; Li, Wei; Zhang, Lihai

    2015-05-19

    Severe spinal cord injury (SCI) can cause neurological dysfunction and paralysis. However, the early dynamic changes of neurons and their surrounding environment after SCI are poorly understood. Although methylprednisolone (MP) is currently the standard therapeutic agent for treating SCI, its efficacy remains controversial. The purpose of this project was to investigate the early dynamic changes and MP's efficacy on axonal damage, blood flow, and calcium influx into axons in a mouse SCI model. YFP H-line and Thy1-GCaMP transgenic mice were used in this study. Two-photon microscopy was used for imaging of axonal dieback, blood flow, and calcium influx post-injury. We found that MP treatment attenuated progressive damage of axons, increased blood flow, and reduced calcium influx post-injury. Furthermore, microglia/macrophages accumulated in the lesion site after SCI and expressed the proinflammatory mediators iNOS, MCP-1 and IL-1β. MP treatment markedly inhibited the accumulation of microglia/macrophages and reduced the expression of the proinflammatory mediators. MP treatment also improved the recovery of behavioral function post-injury. These findings suggest that MP exerts a neuroprotective effect on SCI treatment by attenuating progressive damage of axons, increasing blood flow, reducing calcium influx, and inhibiting the accumulation of microglia/macrophages after SCI.

  13. Coupled hydrogeophysical inversion using time-lapse magnetic resonance sounding and time-lapse gravity data for hydraulic aquifer testing: Will it work in practice?

    DEFF Research Database (Denmark)

    Herckenrath, Daan; Auken, Esben; Christiansen, Lars

    2012-01-01

    Temporal changes in water content can be directly related to the time-lapse signals retrieved using magnetic resonance sounding (TL-MRS) and relative gravimetry (TL-RG). Previous studies suggest that TL-RG measurements can potentially provide accurate estimates of aquifer characteristics in an aq......Temporal changes in water content can be directly related to the time-lapse signals retrieved using magnetic resonance sounding (TL-MRS) and relative gravimetry (TL-RG). Previous studies suggest that TL-RG measurements can potentially provide accurate estimates of aquifer characteristics...

  14. Two-photon microscopy imaging of thy1GFP-M transgenic mice: a novel animal model to investigate brain dendritic cell subsets in vivo.

    Directory of Open Access Journals (Sweden)

    Claudia Laperchia

    Full Text Available Transgenic mice expressing fluorescent proteins in specific cell populations are widely used for in vivo brain studies with two-photon fluorescence (TPF microscopy. Mice of the thy1GFP-M line have been engineered for selective expression of green fluorescent protein (GFP in neuronal populations. Here, we report that TPF microscopy reveals, at the brain surface of these mice, also motile non-neuronal GFP+ cells. We have analyzed the behavior of these cells in vivo and characterized in brain sections their immunophenotype.With TPF imaging, motile GFP+ cells were found in the meninges, subarachnoid space and upper cortical layers. The striking feature of these cells was their ability to move across the brain parenchyma, exhibiting evident shape changes during their scanning-like motion. In brain sections, GFP+ cells were immunonegative to antigens recognizing motile cells such as migratory neuroblasts, neuronal and glial precursors, mast cells, and fibroblasts. GFP+ non-neuronal cells exhibited instead the characteristic features and immunophenotype (CD11c and major histocompatibility complex molecule class II immunopositivity of dendritic cells (DCs, and were immunonegative to the microglial marker Iba-1. GFP+ cells were also identified in lymph nodes and blood of thy1GFP-M mice, supporting their identity as DCs. Thus, TPF microscopy has here allowed the visualization for the first time of the motile behavior of brain DCs in situ. The results indicate that the thy1GFP-M mouse line provides a novel animal model for the study of subsets of these professional antigen-presenting cells in the brain. Information on brain DCs is still very limited and imaging in thy1GFP-M mice has a great potential for analyses of DC-neuron interaction in normal and pathological conditions.

  15. Two-photon super bunching of thermal light via multiple two-photon-path interference

    CERN Document Server

    Hong, Peilong; Zhang, Guoquan

    2012-01-01

    We propose a novel scheme to achieve two-photon super bunching of thermal light through multiple two-photon-path interference, in which two mutually first-order incoherent optical channels are introduced by inserting a modified Michelson interferometer into a traditional two-photon HBT interferometer, and the bunching peak-to-background ratio can reach 3 theoretically. Experimentally, the super bunching peak-to-background ratio was measured to be 2.4, much larger than the ratio 1.7 measured with the same thermal source in a traditional HBT interferometer. The peak-to-background ratio of two-photon super bunching of thermal light can be increased up to $2\\times1.5^n$ by inserting cascadingly $n$ pairs of mutually first-order incoherent optical channels into the traditional two-photon HBT interferometer. The two-photon super bunching of thermal light should be of great significance in improving the visibility of classical ghost imaging.

  16. Informing hydrological models with ground-based time-lapse relative gravimetry: potential and limitations

    DEFF Research Database (Denmark)

    Bauer-Gottwein, Peter; Christiansen, Lars; Rosbjerg, Dan

    2011-01-01

    Coupled hydrogeophysical inversion emerges as an attractive option to improve the calibration and predictive capability of hydrological models. Recently, ground-based time-lapse relative gravity (TLRG) measurements have attracted increasing interest because there is a direct relationship between ...

  17. Discrimination between phase and amplitude attributes in time-lapse seismic streamer data

    NARCIS (Netherlands)

    Spetzler, J.; Kvam, O.

    2006-01-01

    Time-lapse seismic experiments aim to obtain information about production-related effects in hydrocarbon reservoirs to increase the recovery percentage. However, nonrepeatability problems such as acquisition differences, overburden effects, and noise are often significantly stronger than the imprint

  18. History matching of time-lapse crosswell data using ensemble kalman filtering

    KAUST Repository

    de Matos Ravanelli, Fabio Miguel

    2015-09-03

    Data from crosswell seismic surveys is processed to provide crosswell time-lapse data to map fluid changes in a reservoir where time-lapse or 4D seismic data is unavailable or unreliable, such as in onshore reservoirs. The resultant processing results provide quantitative information for history matching purposes using a probabilistic approach to take in account uncertainties in the geological model and reduce uncertainties in reservoir production forecasts.

  19. Time-lapse crosswell seismic and VSP monitoring of injected CO2 ina brine aquifer

    Energy Technology Data Exchange (ETDEWEB)

    Daley, Thomas M.; Myer, Larry R.; Peterson, J.E.; Majer, E.L.; Hoversten,G.M.

    2006-05-30

    Seismic surveys successfully imaged a small scale C02injection (1,600 tons) conducted in a brine aquifer of the Frio Formationnear Houston, Texas. These time-lapse bore-hole seismic surveys,crosswell and vertical seismic profile (VSP), were acquired to monitorthe C02 distribution using two boreholes (the new injection well and apre-existing well used for monitoring) which are 30 m apart at a depth of1500 m. The crosswell survey provided a high-resolution image of the C02distribution between the wells via tomographic imaging of the P-wavevelocity decrease (up to 500 mls). The simultaneously acquired S-wavetomography showed little change in S-wave velocity, as expected for fluidsubstitution. A rock physics model was used to estimate C02 saturationsof 10-20 percent from the P-wave velocity change. The VSP survey resolveda large (-70 percent) change in reflection amplitude for the Friohorizon. This C02 induced reflection amplitude change allowed estimationof the C02 extent beyond the monitor well and on 3 azimuths. The VSPresult is compared with numerical modeling of C02 saturations and isseismically modeled using the velocity change estimated in the crosswellsurvey.

  20. [Embryo selection in IVF/ICSI cycles using time-lapse microscopy and the clinical outcomes].

    Science.gov (United States)

    Chen, Minghao; Huang, Jun; Zhong, Ying; Quan, Song

    2015-12-01

    To compare the clinical outcomes of embryos selected using time-lapse microscopy and traditional morphological method in IVF/ICSI cycles and evaluate the clinical value of time-lapse microscopy in early embryo monitoring and selection. e retrospectively analyzed the clinical data of 139 IVF/ICSI cycles with embryo selection based on time-lapse monitoring (TLM group, n=68) and traditional morphological method (control group, n=71). The βHCG-positive rate, clinical pregnancy rate and embryo implantation rate were compared between the 2 groups. Subgroup analysis was performed in view of female patients age and the fertilization type. The βHCG-positive rate, clinical pregnancy rate and implantation rate were 66.2%, 61.8% and 47.1% in TLM group, significantly higher than those in the control group (47.9%, 43.7% and 30.3%, respectively; P<0.05). Compared with patients below 30 years of age, patients aged between 31 and 35 years benefited more from time-lapse monitoring with improved clinical outcomes. time-lapse monitoring significantly increased the βHCG-positive rate, clinical pregnancy rate and implantation rate for patients undergoing IVF cycles, but not for those undergoing ICSI or TESA cycles. Compared with those selected using traditional morphological method, the embryos selected with time-lapse microscopy have better clinical outcomes, especially in older patients (31-35 years of age) and in IVF cycles.

  1. Two-photon microscopy for chemical neuroscience.

    Science.gov (United States)

    Ellis-Davies, Graham C R

    2011-04-20

    Microscopes using non-linear excitation of chromophores with pulsed near-IR light can generate highly localized foci of molecules in the electronic singlet state that are concentrated in volumes of less than one femtoliter. The three-dimensional confinement of excitation arises from the simultaneous absorption of two IR photons of approximately half the energy required for linear excitation. Two-photon microscopy is especially useful for two types of interrogation of neural processes. First, uncaging of signaling molecules such as glutamate, as stimulation is so refined it can be used to mimic normal unitary synaptic levels. In addition, uncaging allows complete control of the timing and position of stimulation, so the two-photon light beam provides the chemical neuroscientist with an "optical conductor's baton" which can command synaptic activity at will. A second powerful feature of two-photon microscopy is that when used for fluorescence imaging it enables the visualization of cellular structure and function in living animals at depths far beyond that possible with normal confocal microscopes. In this review I provide a survey of the many important applications of two-photon microscopy in these two fields of neuroscience, and suggest some areas for future technical development.

  2. Variogram-based inversion of time-lapse electrical resistivity data: development and application to a thermal tracing experiment

    Science.gov (United States)

    Hermans, Thomas; Nguyen, Frédéric

    2015-04-01

    Electrical resistivity tomography (ERT) has become a popular imaging methodology in a broad range of applications given its large sensitivity to subsurface parameters and its relative simplicity to implement. More particularly, time-lapse ERT is now increasingly used for monitoring purposes in many contexts such as water content, permafrost, landslide, seawater intrusion, solute transport or heat transport experiments. Specific inversion schemes have been developed for time-lapse data sets. However, in contrast with static inversions for which many techniques including geostatistical, minimum support or structural inversion are commonly applied, most of the methodologies for time-lapse inversion still rely on non-physically based spatial and/or temporal smoothing of the parameters or parameter changes. In this work, we propose a time-lapse ERT inversion scheme based on the difference inversion scheme. We replace the standard smoothness-constraint regularization operator by the parameter change covariance matrix. The objective function can be expressed as ψdiff(Δm ) = ||Wd [d - d0 + f(m0)- f(m )]||2 + λ ||||C-Δ0m.5Δm ||||2 where Wd is the data weighting matrix, d and d0 are the data sets corresponding to the considered time-step and to the background, f() is the forward operator, m and m0 are the models corresponding to the considered time-step and to the background, Δm is the parameter change (resistivity), CΔm is the parameter change covariance matrix and λ the regularization parameter. This operator takes into account the correlation between changes in resistivity at different locations through a variogram computed using independent data (e.g., electromagnetic logs). It may vary for subsequent time-steps if the correlation length is time-dependent. The methodology is first validated and compared to the standard smoothness-constraint inversion using a synthetic benchmark simulating the injection of a conductive tracer into a homogeneous aquifer inducing

  3. Live time-lapse dataset of in vitro wound healing experiments.

    Science.gov (United States)

    Zaritsky, Assaf; Natan, Sari; Kaplan, Doron; Ben-Jacob, Eshel; Tsarfaty, Ilan

    2015-01-01

    The wound healing assay is the common method to study collective cell migration in vitro. Computational analyses of live imaging exploit the rich temporal information and significantly improve understanding of complex phenomena that emerge during this mode of collective motility. Publicly available experimental data can allow application of new analyses to promote new discoveries, and assess algorithms' capabilities to distinguish between different experimental conditions. A freely-available dataset of 31 time-lapse in vitro wound healing experiments of two cell lines is presented. It consists of six different experimental conditions with 4-6 replicates each, gathered to study the effects of a growth factor on collective cell migration. The raw data is available at 'The Cell: an Image Library' repository. This Data Note provides detailed description of the data, intermediately processed data, scripts and experimental validations that have not been reported before and are currently available at GigaDB. This is the first publicly available repository of live collective cell migration data that includes independent replicates for each set of conditions. This dataset has the potential for extensive reuse. Some aspects in the data remain unexplored and can be exploited extensively to reveal new insight. The dataset could also be used to assess the performance of available and new quantification methods by demonstrating phenotypic discriminatory capabilities between the different experimental conditions. It may allow faster and more elaborated, reproducible and effective analyses, which will likely lead to new biological and biophysical discoveries.

  4. Time-lapse analysis of methane quantity in Mary Lee group of coal seams using filter-based multiple-point geostatistical simulation

    Science.gov (United States)

    Karacan, C. Özgen; Olea, Ricardo A.

    2013-01-01

    Coal seam degasification and its success are important for controlling methane, and thus for the health and safety of coal miners. During the course of degasification, properties of coal seams change. Thus, the changes in coal reservoir conditions and in-place gas content as well as methane emission potential into mines should be evaluated by examining time-dependent changes and the presence of major heterogeneities and geological discontinuities in the field. In this work, time-lapsed reservoir and fluid storage properties of the New Castle coal seam, Mary Lee/Blue Creek seam, and Jagger seam of Black Warrior Basin, Alabama, were determined from gas and water production history matching and production forecasting of vertical degasification wellbores. These properties were combined with isotherm and other important data to compute gas-in-place (GIP) and its change with time at borehole locations. Time-lapsed training images (TIs) of GIP and GIP difference corresponding to each coal and date were generated by using these point-wise data and Voronoi decomposition on the TI grid, which included faults as discontinuities for expansion of Voronoi regions. Filter-based multiple-point geostatistical simulations, which were preferred in this study due to anisotropies and discontinuities in the area, were used to predict time-lapsed GIP distributions within the study area. Performed simulations were used for mapping spatial time-lapsed methane quantities as well as their uncertainties within the study area.

  5. Higgs Decay to Two Photons

    OpenAIRE

    Marciano, William J.; Zhang, Cen; Willenbrock, Scott

    2011-01-01

    The amplitude for Higgs decay to two photons is calculated in renormalizable and unitary gauges using dimensional regularization at intermediate steps. The result is finite, gauge independent, and in agreement with previously published results. The large Higgs mass limit is examined using the Goldstone-boson equivalence theorem as a check on the use of dimensional regularization and to explain the absence of decoupling.

  6. Time-lapse microscopy studies of bystander effects induced by photosensitization

    Science.gov (United States)

    Chen, Yin-Chu; Redmond, Robert W.

    2006-02-01

    Reactive oxygen species (ROS) are involved in the pathogenesis of many critical diseases and are also utilized as cytotoxic agents in a variety of treatments for eradication of diseased tissue, including cancer. Oxidative stress ensues when the level of ROS in a system exceeds the antioxidant capacity. Oxidative stress can have local (direct) and long-range (bystander) effects in cells and tissue and this research was carried out to determine the spatial and temporal nature of the photosensitized bystander effect using time-lapse fluorescence microscopy. By initiating photosensitization in only a portion of the microscopic imaging field it was possible to differentiate direct from bystander effects in EMT-6 murine breast cancer cells in 6-well plates. Elevated ROS levels are seen immediately following photodynamic treatment in direct cells with a delayed increase in oxidative stress observed in bystander cells. Cytotoxicity is also seen at earlier times in direct cells and occurs in bystander cells in a delayed fashion. These studies confirm the existence of a bystander effect following photosensitization and implicate mediators capable of diffusing in an intercellular manner from directly photosensitized cells to bystander cells and also implicate increased oxidative stress as a mechanistic factor in generating damage in bystander cells.

  7. Technical Note: Semi-automated effective width extraction from time-lapse RGB imagery of a remote, braided Greenlandic river

    Science.gov (United States)

    Gleason, C. J.; Smith, L. C.; Finnegan, D. C.; LeWinter, A. L.; Pitcher, L. H.; Chu, V. W.

    2015-06-01

    River systems in remote environments are often challenging to monitor and understand where traditional gauging apparatus are difficult to install or where safety concerns prohibit field measurements. In such cases, remote sensing, especially terrestrial time-lapse imaging platforms, offer a means to better understand these fluvial systems. One such environment is found at the proglacial Isortoq River in southwestern Greenland, a river with a constantly shifting floodplain and remote Arctic location that make gauging and in situ measurements all but impossible. In order to derive relevant hydraulic parameters for this river, two true color (RGB) cameras were installed in July 2011, and these cameras collected over 10 000 half hourly time-lapse images of the river by September of 2012. Existing approaches for extracting hydraulic parameters from RGB imagery require manual or supervised classification of images into water and non-water areas, a task that was impractical for the volume of data in this study. As such, automated image filters were developed that removed images with environmental obstacles (e.g., shadows, sun glint, snow) from the processing stream. Further image filtering was accomplished via a novel automated histogram similarity filtering process. This similarity filtering allowed successful (mean accuracy 79.6 %) supervised classification of filtered images from training data collected from just 10 % of those images. Effective width, a hydraulic parameter highly correlated with discharge in braided rivers, was extracted from these classified images, producing a hydrograph proxy for the Isortoq River between 2011 and 2012. This hydrograph proxy shows agreement with historic flooding observed in other parts of Greenland in July 2012 and offers promise that the imaging platform and processing methodology presented here will be useful for future monitoring studies of remote rivers.

  8. The measurement of corneal thickness from center to limbus in vivo in C57BL/6 and BALB/c mice using two-photon imaging.

    Science.gov (United States)

    Zhang, Hongmin; Wang, Liya; Xie, Yanting; Liu, Susu; Deng, Xianming; He, Siyu; Chen, Guoming; Liu, Hui; Yang, Biao; Zhang, Junjie; Sun, Shengtao; Li, Xiaohua; Li, Zhijie

    2013-10-01

    The mouse corneal thickness is very important for research into the fields of eye disease. However, the in vivo corneal thickness for the entire cornea from the pupil to the limbus was not determined. We measured in vivo corneal layer thicknesses in different corneal areas, from the central cornea to the limbus, in the widely used inbred C57BL/6 and BALB/c mouse strains using two-photon (2 PH) imaging. Eight corneas of the C57BL/6 or BALB/c were scanned using a 2 PH laser scanning fluorescence microscopy system. A total of 14 thicknesses of the different corneal layers, from different corneal regions, were measured using image processing software. In both C57BL/6 and BALB/c mice, the thickness of the corneal layers was inhomogeneous in different areas of the cornea, and all of the layers had their minimum thickness at the limbus. In C57BL/6 mice, the thickness of the corneal layers gradually increased from the central to the paracentral cornea, peaked at the fifth measurement point in the paracentral area, and decreased from this point to the limbus. In BALB/c mice, the thickness of the entire cornea and corneal epithelium had its maximum at the central cornea and gradually decreased from the central cornea to the peripheral cornea and to the limbus. The thickness of the corneal stroma and endothelium had its maximum at the fourth measurement point in the paracentral cornea and gradually decreased from the paracentral cornea to the limbus. The ratio of epithelial thickness to the total corneal thickness gradually decreased from the central cornea to the limbus in both C57BL/6 and BALB/c mice. The minimum ratio was observed at the fourteenth measurement point in both C57BL/6 and BALB/c mice. The ratio of stromal and endothelial to the total corneal thickness gradually increased from the central cornea to the limbus in both C57BL/6 and BALB/c mice. The maximum ratio was observed at the fourteenth measurement point in C57BL/6 mice. The ratio at the first eight

  9. Optimal experiment design for time-lapse traveltime tomography

    Energy Technology Data Exchange (ETDEWEB)

    Ajo-Franklin, J.B.

    2009-10-01

    Geophysical monitoring techniques offer the only noninvasive approach capable of assessing both the spatial and temporal dynamics of subsurface fluid processes. Increasingly, permanent sensor arrays in boreholes and on the ocean floor are being deployed to improve the repeatability and increase the temporal sampling of monitoring surveys. Because permanent arrays require a large up-front capital investment and are difficult (or impossible) to re-configure once installed, a premium is placed on selecting a geometry capable of imaging the desired target at minimum cost. We present a simple approach to optimizing downhole sensor configurations for monitoring experiments making use of differential seismic traveltimes. In our case, we use a design quality metric based on the accuracy of tomographic reconstructions for a suite of imaging targets. By not requiring an explicit singular value decomposition of the forward operator, evaluation of this objective function scales to problems with a large number of unknowns. We also restrict the design problem by recasting the array geometry into a low dimensional form more suitable for optimization at a reasonable computational cost. We test two search algorithms on the design problem: the Nelder-Mead downhill simplex method and the Multilevel Coordinate Search algorithm. The algorithm is tested for four crosswell acquisition scenarios relevant to continuous seismic monitoring, a two parameter array optimization, several scenarios involving four parameter length/offset optimizations, and a comparison of optimal multi-source designs. In the last case, we also examine trade-offs between source sparsity and the quality of tomographic reconstructions. One general observation is that asymmetric array lengths improve localized image quality in crosswell experiments with a small number of sources and a large number of receivers. Preliminary results also suggest that high-quality differential images can be generated using only a small

  10. Globally referenced real time monitoring of mass movements using monoscopic time-lapse photography.

    Science.gov (United States)

    Kenner, Robert; Phillips, Marcia; Buchroithner, Manfred

    2016-04-01

    The creep movement of a rock glacier was monitored in daily resolution using images of an automatic in-situ time-lapse camera (AC). Displacements were calculated between the images in 2D image coordinates using the imaging velocimetry algorithm of Roesgen and Totaro, 1995. To georeference and scale these displacements, a creep velocity field captured once by a terrestrial laser scan (TLS) repeat measurement was used. The laser scan point cloud and the creep velocity vector field were projected in image coordinates of the AC to obtain a georeferencing mask, a scale mask and an azimuth mask for the 2D displacements calculated between two images. The scale mask was obtained by comparing the TLS derived displacement vectors with those of the AC, referring to a common measurement period. The automatic procedure includes the following work steps: 1. Offsets between two images are identified and corrected based on image parts representing unchanged terrain. 2. 2D displacements are calculated between all chronological image sequences. 3. Faulty displacement vectors are eliminated based on a predefined threshold for spatial direction differences. 4. The remaining displacements are georeferenced, scaled and attributed with individual displacement directions (azimuths) in global coordinates. 5. In addition to the displacement values, displacement velocities and accelerations are calculated using the date of the images. 6. For chronologically successive displacement vector fields, the spatial mean of the relative velocity is defined and expressed as a percentage of the first displacement velocity in the series. The time series of the relative velocities is expressed in chart form. 7. The spatial resolution of all georeferenced output data sets is homogenized, as they were influenced by the central projection of the photos. The described procedure proved to be a reliable, low cost method to monitor mass wasting processes. Even under difficult conditions, like thin snow coverage

  11. Towards monitoring transport of single cargos across individual nuclear pore complexes by time-lapse atomic force microscopy.

    Science.gov (United States)

    Huang, Ning-Ping; Stubenrauch, Mike; Köser, Joachim; Taschner, Nicole; Aebi, Ueli; Stolz, Martin

    2010-08-01

    A new preparation procedure was developed for the stable adsorption of either the cytoplasmic or the nuclear face of native (i.e. in physiological buffer without detergent extraction and in the absence of chemical fixatives) Xenopus oocyte nuclear envelopes (NEs) onto silicon (Si) surfaces. This yields optimal structural preservation of the nuclear pore complexes (NPCs) without compromising their functional properties. The functional viability of thus prepared NPCs was documented by time-lapse atomic force microscopy (AFM) of the reversible calcium-mediated opening (i.e. +Ca(2+)) and closing (i.e. -Ca(2+)) of the iris diaphragm-like distal ring topping the NPCs' nuclear baskets. Moreover, site-specific single colloidal gold particle detection was documented by AFM imaging one and the same NPC before and after immuno-gold labeling the sample with a nucleoporin-specific antibody. With this new preparation protocol at hand, we should eventually be able to follow by time-lapse AFM transport of single gold-conjugated cargos across individual NPCs.

  12. A surge of the glaciers Skobreen–Paulabreen, Svalbard, observed by time-lapse photographs and remote sensing data

    Directory of Open Access Journals (Sweden)

    Lene Kristensen

    2012-11-01

    Full Text Available We present observations of a surge of the glaciers Skobreen–Paulabreen, Svalbard, during 2003–05, including a time-lapse movie of the frontal advance during 2005, Advanced Spaceborne Thermal Emission (ASTER imagery and oblique aerial photographs. The surge initiated in Skobreen, and then propagated downglacier into the lower parts of Paulabreen. ASTER satellite images from different stages of the surge are used to evaluate the surge progression. Features on the glacier surface advanced 2800 m over 2.4 yr, averaging 3.2 m/day, while the front advanced less (ca. 1300 m due to contemporaneous calving. The surge resulted in a lateral displacement of the medial moraines of Paulabreen of ca. 600 m at the glacier front. The time-lapse movie captured the advance of the frontal part of the glacier, and dramatically illustrates glacier dynamic processes in an accessible way. The movie documents a range of processes such as a plug-like flow of the glacier, proglacial thrusting, incorporation of old, dead ice at the margin, and calving into the fjord. The movie provides a useful resource for researchers, educators seeking to teach and inspire students, and those wishing to communicate the fascination of glacier science to a wider public.

  13. Large-scale time-lapse microscopy of Oct4 expression in human embryonic stem cell colonies.

    Science.gov (United States)

    Bhadriraju, Kiran; Halter, Michael; Amelot, Julien; Bajcsy, Peter; Chalfoun, Joe; Vandecreme, Antoine; Mallon, Barbara S; Park, Kye-Yoon; Sista, Subhash; Elliott, John T; Plant, Anne L

    2016-07-01

    Identification and quantification of the characteristics of stem cell preparations is critical for understanding stem cell biology and for the development and manufacturing of stem cell based therapies. We have developed image analysis and visualization software that allows effective use of time-lapse microscopy to provide spatial and dynamic information from large numbers of human embryonic stem cell colonies. To achieve statistically relevant sampling, we examined >680 colonies from 3 different preparations of cells over 5days each, generating a total experimental dataset of 0.9 terabyte (TB). The 0.5 Giga-pixel images at each time point were represented by multi-resolution pyramids and visualized using the Deep Zoom Javascript library extended to support viewing Giga-pixel images over time and extracting data on individual colonies. We present a methodology that enables quantification of variations in nominally-identical preparations and between colonies, correlation of colony characteristics with Oct4 expression, and identification of rare events.

  14. Assessing spatial and temporal snowpack evolution and melt with time-lapse photography

    Science.gov (United States)

    Bush, C. E.; Ewers, B. E.; Beverly, D.; Speckman, H. N.; Hyde, K.; Ohara, N.

    2015-12-01

    Snowpack supplies and stores water for many ecosystems of the greater Rocky Mountain region. In Wyoming the snowpack supplies water to 18 states east and west of the Continental Divide. The spatial variability in physical and biological processes creates a heterogeneous pattern of snow evolution. Understanding these processes within individual plots and throughout the entire watershed increases the predictive power of snow distribution, melt rates and contribution to streamflow. However, on site sampling of snow can be an expensive and arduous process. The objective of this experiment was to quantify spatial and temporal patterns of snowpack evolution and melt rates while minimizing perturbations to snowpack through the use of time-lapse photography via trail cameras. Field cameras were assessed as a method to quantify snow depths throughout the 120 ha No Name watershed at approximately 3000 m elevation in central Wyoming. RGB trail cameras were installed at three systematically chosen sites within the watershed to correlate physical and biological drivers of snow distribution. Five stakes were placed in each site in heterogeneous spots that remained in the frame of the camera. Stakes were divided into five centimeter increments, alternating black and white bars, with red bars denoting each half meter. Images were then taken at two-hour intervals over a period of three-months and analyzed with the ImageJ program. Snowpack distributions, as well as melt rates, were variable at both the plot and watershed scales. Meteorological and physical drivers, primarily topography and radiation, accounted for the greatest variability when comparing among plot across the watershed; however, LAI and soil and air temperature were the most significant drivers within plots. Snow-melt rate increased as soils and course woody debris became exposed increasing ground and soil temperature. These data will improve process model predictions of streamflow from the watershed.

  15. Full waveform inversion of repeating seismic events to estimate time-lapse velocity changes

    Science.gov (United States)

    Kamei, R.; Lumley, D.

    2017-05-01

    Seismic monitoring provides valuable information regarding the time-varying changes in subsurface physical properties caused by natural or man-made processes. However, the resulting changes in the earth's subsurface properties are often small both in terms of magnitude and spatial extent, leading to minimal time-lapse differences in seismic amplitude or traveltime. In order to better extract information from the time-lapse data, we show that exploiting the full seismic waveform information can be critical. In this study, we develop and test methods of full waveform inversion that estimate an optimal subsurface model of time-varying elastic properties in order to fit the observed time-lapse seismic data with predicted waveforms based on numerical solutions of the wave equation. Time-lapse full waveform inversion is nonlinear and non-unique, and depends on the knowledge of the baseline velocity model before a change, and (non-)repeatability of earthquake source and sensor parameters, and of ambient and cultural noise. We propose to use repeating earthquake data sets acquired with permanent arrays of seismic sensors to enhance the repeatability of source and sensor parameters. We further develop and test time-lapse parallel, double-difference and bootstrapping inversion strategies to mitigate the dependence on the baseline velocity model. The parallel approach uses a time-invariant full waveform inversion method to estimate velocity models independently of the different source event times. The double-difference approach directly estimates velocity changes from time-lapse waveform differences, requiring excellent repeatability. The bootstrapping approach inverts for velocity models sequentially in time, implicitly constraining the time-lapse inversions, while relaxing an explicit requirement for high data repeatability. We assume that prior to the time-lapse inversion, we can estimate the true source locations and the origin time of the events, and also we can also

  16. Automated Ground-based Time-lapse Camera Monitoring of West Greenland ice sheet outlet Glaciers: Challenges and Solutions

    Science.gov (United States)

    Ahn, Y.; Box, J. E.; Balog, J.; Lewinter, A.

    2008-12-01

    Monitoring Greenland outlet glaciers using remotely sensed data has drawn a great attention in earth science communities for decades and time series analysis of sensory data has provided important variability information of glacier flow by detecting speed and thickness changes, tracking features and acquiring model input. Thanks to advancements of commercial digital camera technology and increased solid state storage, we activated automatic ground-based time-lapse camera stations with high spatial/temporal resolution in west Greenland outlet and collected one-hour interval data continuous for more than one year at some but not all sites. We believe that important information of ice dynamics are contained in these data and that terrestrial mono-/stereo-photogrammetry can provide theoretical/practical fundamentals in data processing along with digital image processing techniques. Time-lapse images over periods in west Greenland indicate various phenomenon. Problematic is rain, snow, fog, shadows, freezing of water on camera enclosure window, image over-exposure, camera motion, sensor platform drift, and fox chewing of instrument cables, and the pecking of plastic window by ravens. Other problems include: feature identification, camera orientation, image registration, feature matching in image pairs, and feature tracking. Another obstacle is that non-metric digital camera contains large distortion to be compensated for precise photogrammetric use. Further, a massive number of images need to be processed in a way that is sufficiently computationally efficient. We meet these challenges by 1) identifying problems in possible photogrammetric processes, 2) categorizing them based on feasibility, and 3) clarifying limitation and alternatives, while emphasizing displacement computation and analyzing regional/temporal variability. We experiment with mono and stereo photogrammetric techniques in the aide of automatic correlation matching for efficiently handling the enormous

  17. Construction of an 8-mm time-lapse camera for biological research

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — This report covers the construction of an 8mm camera for biological research. A time-lapse camera for use in biological research can be constructed from a super 8-mm...

  18. Exploiting the airwave for time-lapse reservoir monitoring with CSEM on land

    NARCIS (Netherlands)

    Wirianto, M.; Mulder, W.A.; Slob, E.C.

    2011-01-01

    In the application of controlled source electromagnetics for reservoir monitoring on land, repeatability errors in the source will mask the time-lapse signal due to hydrocarbon production when recording surface data close to the source. We demonstrate that at larger distances, the airwave will still

  19. Feasibility of monitoring gas hydrate production with time-lapse VSP

    Energy Technology Data Exchange (ETDEWEB)

    Kowalsky, M.B.; Nakagawa, S.; Moridis, G.J.

    2009-11-01

    In this work we begin to examine the feasibility of using time-lapse seismic methods-specifically the vertical seismic profiling (VSP) method-for monitoring changes in hydrate accumulations that are predicted to occur during production of natural gas.

  20. Time-lapse motion picture technique applied to the study of geological processes

    Science.gov (United States)

    Miller, R.D.; Crandell, D.R.

    1959-01-01

    Light-weight, battery-operated timers were built and coupled to 16-mm motion-picture cameras having apertures controlled by photoelectric cells. The cameras were placed adjacent to Emmons Glacier on Mount Rainier. The film obtained confirms the view that exterior time-lapse photography can be applied to the study of slow-acting geologic processes.

  1. Time lapse inversion of 2D ERT data for monitoring river water infiltration

    Science.gov (United States)

    Wallin, E. L.; Johnson, T. C.; Greenwood, W. J.

    2011-12-01

    Uranium transport in the 300 area is driven by both the chemical and physical effects of stage fluctuations in the Columbia River and resulting river water, ground water interaction. Because river water is less conductive than groundwater, it serves as a natural tracer that can be imaged using surface ERT. We've monitored 4 lines for 4 months over the high stage spring runoff interval to identify preferred flowpaths for river water intrusion. The four lines overlay former waste disposal sites including the processing ponds and sanitary leach trenches. We have used this ERT data set to investigate two methods of time lapse inversion, sequential and all-at-once. Each technique is a model difference approach as opposed to a data difference inversion approach. Both use a regularized inversion with model constraints that regularize spatially and temporally. For the sequential inversion, the starting and reference models are taken from the previous inversion. Both starting and reference models are taken from the first inversion for the all-at-once method. In either case an inversion is triggered if data misfit from the starting model exceeds the chi-squared convergence criteria. It was found that starting with a relatively smooth model provided better visualization of temporal conductivity changes when inverting all data sets with the same initial model, while an initial model exhibiting smaller data misfit may be used successfully as the starting point for sequential inversion. Inland conductivity changes within model cells were found to be highly correlated with river stage, and when paired with the characterization model, provide evidence of waste trenches, the processing pond, as well as the existence of a paleo-channel incised into the Ringold Formation and dipping structures on the Hanford-Ringold contact that provide preferred pathways for river water intrusion.

  2. Full waveform inversion of repeating seismic events to estimate time-lapse velocity changes

    Science.gov (United States)

    Kamei, R.; Lumley, D.

    2017-02-01

    Seismic monitoring provides valuable information regarding the time-varying changes in subsurface physical properties caused by natural or man-made processes. However, the resulting changes in the earth's subsurface properties are often small both in terms of magnitude and spatial extent, leading to minimal time-lapse differences in seismic amplitude or traveltime. In order to better extract information from the time-lapse data, we show that exploiting the full seismic waveform information can be critical. In this study, we develop and test methods of full waveform inversion that estimate an optimal subsurface model of time-varying elastic properties in order to fit the observed time-lapse seismic data with predicted waveforms based on numerical solutions of the wave equation. Time-lapse full waveform inversion is non-linear and non-unique, and depends on the knowledge of the baseline velocity model before a change, and (non-)repeatability of earthquake source and sensor parameters, and of ambient and cultural noise. We propose to use repeating earthquake data sets acquired with permanent arrays of seismic sensors to enhance the repeatability of source and sensor parameters. We further develop and test time-lapse parallel, double-difference and bootstrapping inversion strategies to mitigate the dependence on the baseline velocity model. The parallel approach uses a time-invariant full waveform inversion method to estimate velocity models independently of the different source event times. The double-difference approach directly estimates velocity changes from time-lapse waveform differences, requiring excellent repeatability. The bootstrapping approach inverts for velocity models sequentially in time, implicitly constraining the time-lapse inversions, while relaxing an explicit requirement for high data repeatability. We assume that prior to the time-lapse inversion, we can estimate the true source locations and the origin time of the events, and also we can also

  3. Overland flow dynamics through visual observation using time-lapse photographs

    Science.gov (United States)

    Silasari, Rasmiaditya; Blöschl, Günter

    2016-04-01

    Overland flow process on agricultural land is important to be investigated as it affects the stream discharge and water quality assessment. During rainfall events the formation of overland flow may happen through different processes (i.e. Hortonian or saturation excess overland flow) based on the governing soil hydraulic parameters (i.e. soil infiltration rate, soil water capacity). The dynamics of the soil water state and the processes will affect the surface runoff response which can be analyzed visually by observing the saturation patterns with a camera. Although visual observation was proven useful in laboratory experiments, the technique is not yet assessed for natural rainfall events. The aim of this work is to explore the use of time-lapse photographs of naturally occurring-saturation patterns in understanding the threshold processes of overland flow generation. The image processing produces orthographic projection of the saturation patterns which will be used to assess the dynamics of overland flow formation in relation with soil moisture state and rainfall magnitude. The camera observation was performed at Hydrological Open Air Laboratory (HOAL) catchment at Petzenkirchen, Lower Austria. The catchment covers an area of 66 ha dominated with agricultural land (87%). The mean annual precipitation and mean annual flow at catchment outlet are 750 mm and 4 l/s, respectively. The camera was set to observe the overland flow along a thalweg on an arable field which was drained in 1950s and has advantages of: (1) representing agricultural land as the dominant part of the catchment, (2) adjacent to the stream with clear visibility (no obstructing objects, such as trees), (3) drained area provides extra cases in understanding the response of tile drain outflow to overland flow formation and vice versa, and (4) in the vicinity of TDT soil moisture stations. The camera takes a picture with 1280 x 720 pixels resolution every minute and sends it directly in a PC via fiber

  4. Two-Photon Flow Cytometry

    Science.gov (United States)

    Zhog, Cheng Frank; Ye, Jing Yong; Norris, Theodore B.; Myc, Andrzej; Cao, Zhengyl; Bielinska, Anna; Thomas, Thommey; Baker, James R., Jr.

    2004-01-01

    Flow cytometry is a powerful technique for obtaining quantitative information from fluorescence in cells. Quantitation is achieved by assuring a high degree of uniformity in the optical excitation and detection, generally by using a highly controlled flow such as is obtained via hydrodynamic focusing. In this work, we demonstrate a two-beam, two- channel detection and two-photon excitation flow cytometry (T(sup 3)FC) system that enables multi-dye analysis to be performed very simply, with greatly relaxed requirements on the fluid flow. Two-photon excitation using a femtosecond near-infrared (NIR) laser has the advantages that it enables simultaneous excitation of multiple dyes and achieves very high signal-to-noise ratio through simplified filtering and fluorescence background reduction. By matching the excitation volume to the size of a cell, single-cell detection is ensured. Labeling of cells by targeted nanoparticles with multiple fluorophores enables normalization of the fluorescence signal and thus ratiometric measurements under nonuniform excitation. Quantitative size measurements can also be done even under conditions of nonuniform flow via a two-beam layout. This innovative detection scheme not only considerably simplifies the fluid flow system and the excitation and collection optics, it opens the way to quantitative cytometry in simple and compact microfluidics systems, or in vivo. Real-time detection of fluorescent microbeads in the vasculature of mouse ear demonstrates the ability to do flow cytometry in vivo. The conditions required to perform quantitative in vivo cytometry on labeled cells will be presented.

  5. In situ monitoring of corrosion mechanisms and phosphate inhibitor surface deposition during corrosion of zinc-magnesium-aluminium (ZMA) alloys using novel time-lapse microscopy.

    Science.gov (United States)

    Sullivan, James; Cooze, Nathan; Gallagher, Callum; Lewis, Tom; Prosek, Tomas; Thierry, Dominique

    2015-01-01

    In situ time-lapse optical microscopy was used to examine the microstructural corrosion mechanisms in three zinc-magnesium-aluminium (ZMA) alloy coated steels immersed in 1% NaCl pH 7. Preferential corrosion of MgZn(2) lamellae within the eutectic phases was observed in all the ZMA alloys followed by subsequent dissolution of Zn rich phases. The total extent and rate of corrosion, measured using time-lapse image analysis and scanning vibrating electrode technique (SVET) estimated mass loss, decreased as Mg and Al alloying additions were increased up to a level of 3 wt% Mg and 3.7 wt% Al. This was probably due to the increased presence of MgO and Al(2)O(3) at the alloy surface retarding the kinetics of cathodic oxygen reduction. The addition of 1 × 10(-2) mol dm(-3) Na(3)PO(4) to 1% NaCl pH 7 had a dramatic influence on the corrosion mechanism for a ZMA with passivation of anodic sites through phosphate precipitation observed using time-lapse image analysis. Intriguing rapid precipitation of filamentous phosphate was also observed and it is postulated that these filaments nucleate and grow due to super saturation effects. Polarisation experiments showed that the addition of 1 × 10(-2) mol dm(-3) Na(3)PO(4) to the 1% NaCl electrolyte promoted an anodic shift of 50 mV in open circuit potential for the ZMA alloy with a reduction in anodic current of 2.5 orders of magnitude suggesting that it was acting primarily as an anodic inhibitor supporting the inferences from the time-lapse investigations. These phosphate additions resulted in a 98% reduction in estimated mass loss as measured by SVET demonstrating the effectiveness of phosphate inhibitors for this alloy system.

  6. Using Digital Time-Lapse Videos to Teach Geomorphic Processes to Undergraduates

    Science.gov (United States)

    Clark, D. H.; Linneman, S. R.; Fuller, J.

    2004-12-01

    We demonstrate the use of relatively low-cost, computer-based digital imagery to create time-lapse videos of two distinct geomorphic processes in order to help students grasp the significance of the rates, styles, and temporal dependence of geologic phenomena. Student interviews indicate that such videos help them to understand the relationship between processes and landform development. Time-lapse videos have been used extensively in some sciences (e.g., biology - http://sbcf.iu.edu/goodpract/hangarter.html, meteorology - http://www.apple.com/education/hed/aua0101s/meteor/, chemistry - http://www.chem.yorku.ca/profs/hempsted/chemed/home.html) to demonstrate gradual processes that are difficult for many students to visualize. Most geologic processes are slower still, and are consequently even more difficult for students to grasp, yet time-lapse videos are rarely used in earth science classrooms. The advent of inexpensive web-cams and computers provides a new means to explore the temporal dimension of earth surface processes. To test the use of time-lapse videos in geoscience education, we are developing time-lapse movies that record the evolution of two landforms: a stream-table delta and a large, natural, active landslide. The former involves well-known processes in a controlled, repeatable laboratory experiment, whereas the latter tracks the developing dynamics of an otherwise poorly understood slope failure. The stream-table delta is small and grows in ca. 2 days; we capture a frame on an overhead web-cam every 3 minutes. Before seeing the video, students are asked to hypothesize how the delta will grow through time. The final time-lapse video, ca. 20-80 MB, elegantly shows channel migration, progradation rates, and formation of major geomorphic elements (topset, foreset, bottomset beds). The web-cam can also be "zoomed-in" to show smaller-scale processes, such as bedload transfer, and foreset slumping. Post-lab tests and interviews with students indicate that

  7. Determining changes in CO2 saturation from time-lapse measurements using ghost reflections retrieved by seismic interferometry

    NARCIS (Netherlands)

    Draganov, D.S.; Ghose, R.; Angelov, P.; Arts, R.

    2013-01-01

    One of the targets of time-lapse seismic monitoring of CO2 sequestration is to determine the parts of the reservoir reached by the injected CO2 and to estimate their saturation. Such information could be extracted from the time-lapse measurement using AVO or impedance information. But

  8. Determining changes in CO2 saturation from time-lapse measurements using ghost reflections retrieved by seismic interferometry

    NARCIS (Netherlands)

    Draganov, D.S.; Ghose, R.; Angelov, P.; Arts, R.

    2013-01-01

    One of the targets of time-lapse seismic monitoring of CO2 sequestration is to determine the parts of the reservoir reached by the injected CO2 and to estimate their saturation. Such information could be extracted from the time-lapse measurement using AVO or impedance information. But non-repeatabil

  9. A randomized clinical trial comparing embryo culture in a conventional incubator with a time-lapse incubator

    DEFF Research Database (Denmark)

    Kirkegaard, Kirstine; Hindkjaer, Johnny Juhl; Grøndahl, Marie Louise;

    2012-01-01

    Time-lapse monitoring allows for a flexible embryo evaluation and potentially provides new dynamic markers of embryo competence. Before introducing time-lapse monitoring in a clinical setting, the safety of the instrument must be properly documented. Accordingly, the aim of this study...

  10. Reversible Disruption of Neuronal Mitochondria by Ischemic and Traumatic Injury Revealed by Quantitative Two-Photon Imaging in the Neocortex of Anesthetized Mice.

    Science.gov (United States)

    Kislin, Mikhail; Sword, Jeremy; Fomitcheva, Ioulia V; Croom, Deborah; Pryazhnikov, Evgeny; Lihavainen, Eero; Toptunov, Dmytro; Rauvala, Heikki; Ribeiro, Andre S; Khiroug, Leonard; Kirov, Sergei A

    2017-01-11

    Mitochondria play a variety of functional roles in cortical neurons, from metabolic support and neuroprotection to the release of cytokines that trigger apoptosis. In dendrites, mitochondrial structure is closely linked to their function, and fragmentation (fission) of the normally elongated mitochondria indicates loss of their function under pathological conditions, such as stroke and brain trauma. Using in vivo two-photon microscopy in mouse brain, we quantified mitochondrial fragmentation in a full spectrum of cortical injuries, ranging from severe to mild. Severe global ischemic injury was induced by bilateral common carotid artery occlusion, whereas severe focal stroke injury was induced by Rose Bengal photosensitization. The moderate and mild traumatic injury was inflicted by focal laser lesion and by mild photo-damage, respectively. Dendritic and mitochondrial structural changes were tracked longitudinally using transgenic mice expressing fluorescent proteins localized either in cytosol or in mitochondrial matrix. In response to severe injury, mitochondrial fragmentation developed in parallel with dendritic damage signified by dendritic beading. Reconstruction from serial section electron microscopy confirmed mitochondrial fragmentation. Unlike dendritic beading, fragmentation spread beyond the injury core in focal stroke and focal laser lesion models. In moderate and mild injury, mitochondrial fragmentation was reversible with full recovery of structural integrity after 1-2 weeks. The transient fragmentation observed in the mild photo-damage model was associated with changes in dendritic spine density without any signs of dendritic damage. Our findings indicate that alterations in neuronal mitochondria structure are very sensitive to the tissue damage and can be reversible in ischemic and traumatic injuries. During ischemic stroke or brain trauma, mitochondria can either protect neurons by supplying ATP and adsorbing excessive Ca(2+), or kill neurons by

  11. Conjugates of folic acids with BSA-coated quantum dots for cancer cell targeting and imaging by single-photon and two-photon excitation.

    Science.gov (United States)

    Meng, He; Chen, Ji-Yao; Mi, Lan; Wang, Pei-Nan; Ge, Mei-Ying; Yue, Yang; Dai, Ning

    2011-01-01

    Bovine serum albumin (BSA)-coated CdTe/ZnS quantum dots (BSA-QDs) were selected to conjugate with folic acid (FA), forming FA-BSA-QDs. This study aims to develop these small FA-BSA-QDs (less than 10 nm) for the diagnosis of cancers in which the FA receptor (FR) is overexpressed. The enhancement of cellular uptake in FR-positive human nasopharyngeal carcinoma cells (KB cells) for FA-BSA-QDs was found by means of confocal fluorescence microscopy under single-photon and two-photon excitation. The uptake enhancement for FA-BSA-QDs was further evaluated by flow-cytometric analysis in 10(4) KB cells, and was about 3 times higher than for BSA-QDs on average. The uptake enhancement was suppressed when KB cells had been pretreated with excess FA, reflecting that the enhancement was mediated by the association of FR at cell membranes with FA-BSA-QDs. When human embryonic kidney cells (293T) (FR-negative cells) and KB cells, respectively, were incubated with FA-BSA-QDs (1 μM) for 40 min, the FA-BSA-QD uptake by 293T cells was much weaker than that by KB cells, demonstrating that FA-BSA-QDs could undergo preferential binding on FR-positive cancer cells. These characteristics suggest that FA-BSA-QDs are potential candidates for cancer diagnosis.

  12. Feeding behavior of lactating dairy cows as measured by time-lapse photography.

    Science.gov (United States)

    Vasilatos, R; Wangsness, P J

    1980-03-01

    Evaluation of feeding behavior of ad libitum-fed lactating dairy cows by time-lapse photography revealed 68% of the total feeding activity occurred between the daylight hours of 0600 and 1800. Cows consumed an average of 12.1 meals/day, each 20.9 min in duration. Only 58% of the total defined meal time actually was spent eating, or 253.6 min/cow per day. Estimated meal size and rate of eating, as well as total daily time spent eating, were greater for cows as compared to animals with lower energy demand. Certain feeding characteristics, such as meal frequency and duration, were variable among animals, suggesting that these behaviors may be characteristics of individual cows. Results by time-lapse photography compared well with direct measurement by weigh-cell apparatus.

  13. Time-lapse microgravity study of the Strengbach catchment (Vosges mountains, France)

    Science.gov (United States)

    Masson, Frédéric; Viville, Daniel; Pierret, Marie-Claire; Mouyen, Maxime; Hecker, Louis; Chabaux, François

    2012-06-01

    Time-lapse microgravity measurements can be used to monitor underground water storage changes. For the first time, this method has been applied to a relatively steeply sloped and forested watershed in a temperate climate. Spatial and temporal measurements were performed on the small granitic Strengbach catchment (Vosges Mountains, France) during the unusually dry spring of 2011. The survey consisted of 11 relative gravimeter measurements for 13 gravity stations from February to June 2011. Temporal variations are significantly different from one station to the other. Nevertheless, the variations have a clearly observable spatial consistency, which could be related to the climatology, the characteristics of the bedrocks and/or the topography. This preliminary result highlights the potential capacity of time-lapse microgravimetry in understanding and constraining the water mass movement within a complex watershed.

  14. Time-Lapse Monitoring of Subsurface Fluid Flow using Parsimonious Seismic Interferometry

    KAUST Repository

    Hanafy, Sherif

    2017-04-21

    A typical small-scale seismic survey (such as 240 shot gathers) takes at least 16 working hours to be completed, which is a major obstacle in case of time-lapse monitoring experiments. This is especially true if the subject that needs to be monitored is rapidly changing. In this work, we will discuss how to decrease the recording time from 16 working hours to less than one hour of recording. Here, the virtual data has the same accuracy as the conventional data. We validate the efficacy of parsimonious seismic interferometry with the time-lapse mentoring idea with field examples, where we were able to record 30 different data sets within a 2-hour period. The recorded data are then processed to generate 30 snapshots that shows the spread of water from the ground surface down to a few meters.

  15. Rapid antimicrobial susceptibility testing of clinical isolates by digital time-lapse microscopy

    DEFF Research Database (Denmark)

    Fredborg, M; Rosenvinge, F S; Spillum, E

    2015-01-01

    Rapid antimicrobial susceptibility testing (AST) is essential for early and appropriate therapy. Methods with short detection time enabling same-day treatment optimisation are highly favourable. In this study, we evaluated the potential of a digital time-lapse microscope system, the oCelloScope s......Rapid antimicrobial susceptibility testing (AST) is essential for early and appropriate therapy. Methods with short detection time enabling same-day treatment optimisation are highly favourable. In this study, we evaluated the potential of a digital time-lapse microscope system, the o...... (168 antimicrobial agent-organism combinations) demonstrated 3.6 % minor, no major and 1.2 % very major errors of the oCelloScope system compared to conventional susceptibility testing, as well as a rapid and correct phenotypic detection of strains with methicillin-resistant Staphylococcus aureus (MRSA...

  16. Calixarenes and cations: a time-lapse photography of the big-bang.

    Science.gov (United States)

    Casnati, Alessandro

    2013-08-07

    The outstanding cation complexation properties emerging from the pioneering studies on calixarene ligands during a five-year period in the early 1980s triggered a big-bang burst of publications on such macrocycles that is still lasting at a distance of more than 30 years. A time-lapse photography of this timeframe is proposed which allows the readers to pinpoint the contributions of the different research groups.

  17. Analysis of the repeatability of time-lapse 3d vsp multicomponent surveys, delhi field

    Science.gov (United States)

    Carvalho, Mariana Fernandes de

    Delhi Field is a producing oil field located in northeastern Louisiana. In order to monitor the CO2 sweep efficiency, time-lapse 3D seismic data have been acquired in this area. Time-lapse studies are increasingly used to evaluate changes in the seismic response induced by the production of hydrocarbons or the injection of water, CO2 or steam into a reservoir. A 4D seismic signal is generated by a combination of production and injection effects within the reservoir as well as non-repeatability effects. In order to get reliable results from time-lapse seismic methods, it is important to distinguish the production and injection effects from the non-repeatability effects in the 4D seismic signal. Repeatability of 4D land seismic data is affected by several factors. The most significant of them are: source and receiver geometry inaccuracies, differences in seismic sources signatures, variations in the immediate near surface and ambient non-repeatable noise. In this project, two 3D multicomponent VSP surveys acquired in Delhi Field were used to quantify the relative contribution of each factor that can affect the repeatability in land seismic data. The factors analyzed in this study were: source and receiver geometry inaccura- cies, variations in the immediate near surface and ambient non-repeatable noise. This study showed that all these factors had a significant impact on the repeatability of the successive multicomponent VSP surveys in Delhi Field. This project also shows the advantages and disadvantages in the use of different repeata- bility metrics, normalized-root-mean-square (NRMS) difference and signal-to-distortion ratio (SDR) attribute, to evaluate the level of seismic repeatability between successive time-lapse seismic surveys. It is observed that NRMS difference is greatly influenced by time-shifts and that SDR attribute combined with the time-shift may give more distinct and representative repeatability information than the NRMS difference.

  18. Unscented Kalman filter assimilation of time-lapse self-potential data for monitoring solute transport

    Science.gov (United States)

    Cui, Yi-an; Liu, Lanbo; Zhu, Xiaoxiong

    2017-08-01

    Monitoring the extent and evolution of contaminant plumes in local and regional groundwater systems from existing landfills is critical in contamination control and remediation. The self-potential survey is an efficient and economical nondestructive geophysical technique that can be used to investigate underground contaminant plumes. Based on the unscented transform, we have built a Kalman filtering cycle to conduct time-lapse data assimilation for monitoring the transport of solute based on the solute transport experiment using a bench-scale physical model. The data assimilation was formed by modeling the evolution based on the random walk model and observation correcting based on the self-potential forward. Thus, monitoring self-potential data can be inverted by the data assimilation technique. As a result, we can reconstruct the dynamic process of the contaminant plume instead of using traditional frame-to-frame static inversion, which may cause inversion artifacts. The data assimilation inversion algorithm was evaluated through noise-added synthetic time-lapse self-potential data. The result of the numerical experiment shows validity, accuracy and tolerance to the noise of the dynamic inversion. To validate the proposed algorithm, we conducted a scaled-down sandbox self-potential observation experiment to generate time-lapse data that closely mimics the real-world contaminant monitoring setup. The results of physical experiments support the idea that the data assimilation method is a potentially useful approach for characterizing the transport of contamination plumes using the unscented Kalman filter (UKF) data assimilation technique applied to field time-lapse self-potential data.

  19. Time-lapse AVO fluid inversion for dynamic reservoir characterization in Delhi Field, Louisiana

    Science.gov (United States)

    Putri, Indah Hermansyah

    In the development stage, CO2 injection is becoming more widely used in enhanced oil recovery (EOR). Delhi Oil Field is part of Phases XIII and XIV of the Reservoir Characterization Project (RCP) Colorado School of Mines. The focus of these phases is to monitor the effectiveness of the CO 2 injection in Delhi Field by using multicomponent time-lapse seismic data. In this study, I analyze the amplitude versus offset (AVO) response of the time-lapse P-wave seismic data in order to quantify the fluid probability in the field. RCP acquired four square miles of multicomponent time-lapse seismic in Delhi Field to characterize the field dynamically. RCP's two surveys, monitor 1 and monitor 2, were shot in 2010 and 2011 after the start of CO2 injection in November 2009. Time-lapse AVO modeling was performed. The modeling results show that both the top Tuscaloosa and Paluxy Formations are class III AVO, and change toward class IV AVO by increasing the CO2 saturation in the reservoir. In addition, the Paluxy Formation shows a consistent result between the synthetic and real data, however, the Tuscaloosa Formation is not consistent as it is affected by tuning. AVO fluid inversion (AFI) was performed on both the Tuscaloosa and Paluxy Formations in order to quantify the fluid probability in these formations. The inversion results are confirmed by the pseudo gamma ray model, the porosity model, the permeability model, the pressure model, and the production data. In the Tuscaloosa and Paluxy Formations, oil and CO2 are located in the good quality, high porosity, and high permeability sandstones. The presence of CO2 is also confirmed by the pressure interpretation. Furthermore, production data from both Tuscaloosa and Paluxy Formations confirm the fluid presence in the reservoir.

  20. Efficiency of time-lapse intervals and simple baits for camera surveys of wild pigs

    Science.gov (United States)

    Williams, B.L.; Holtfreter, R.W.; Ditchkoff, S.S.; Grand, J.B.

    2011-01-01

    Growing concerns surrounding established and expanding populations of wild pigs (Sus scrofa) have created the need for rapid and accurate surveys of these populations. We conducted surveys of a portion of the wild pig population on Fort Benning, Georgia, to determine if a longer time-lapse interval than had been previously used in surveys of wild pigs would generate similar detection results. We concurrently examined whether use of soured corn at camera sites affected the time necessary for pigs to locate a new camera site or the time pigs remained at a site. Our results suggest that a 9-min time-lapse interval generated dependable detection results for pigs and that soured corn neither attracted pigs to a site any quicker than plain, dry, whole-kernel corn, nor held them at a site longer. Maximization of time-lapse interval should decrease data and processing loads, and use of a simple, available bait should decrease cost and effort associated with more complicated baits; combination of these concepts should increase efficiency of wild pig surveys. ?? 2011 The Wildlife Society.

  1. Measuring fast gene dynamics in single cells with time-lapse luminescence microscopy

    Science.gov (United States)

    Mazo-Vargas, Anyimilehidi; Park, Heungwon; Aydin, Mert; Buchler, Nicolas E.

    2014-01-01

    Time-lapse fluorescence microscopy is an important tool for measuring in vivo gene dynamics in single cells. However, fluorescent proteins are limited by slow chromophore maturation times and the cellular autofluorescence or phototoxicity that arises from light excitation. An alternative is luciferase, an enzyme that emits photons and is active upon folding. The photon flux per luciferase is significantly lower than that for fluorescent proteins. Thus time-lapse luminescence microscopy has been successfully used to track gene dynamics only in larger organisms and for slower processes, for which more total photons can be collected in one exposure. Here we tested green, yellow, and red beetle luciferases and optimized substrate conditions for in vivo luminescence. By combining time-lapse luminescence microscopy with a microfluidic device, we tracked the dynamics of cell cycle genes in single yeast with subminute exposure times over many generations. Our method was faster and in cells with much smaller volumes than previous work. Fluorescence of an optimized reporter (Venus) lagged luminescence by 15–20 min, which is consistent with its known rate of chromophore maturation in yeast. Our work demonstrates that luciferases are better than fluorescent proteins at faithfully tracking the underlying gene expression. PMID:25232010

  2. Holographic Two-Photon Induced Photopolymerization

    Data.gov (United States)

    Federal Laboratory Consortium — Holographic two-photon-induced photopolymerization (HTPIP) offers distinct advantages over conventional one-photon-induced photopolymerization and current techniques...

  3. Confocal and Two-Photon Microscopy: Foundations, Applications and Advances

    Science.gov (United States)

    Diaspro, Alberto

    2001-11-01

    Confocal and Two-Photon Microscopy Foundations, Applications, and Advances Edited by Alberto Diaspro Confocal and two-photon fluorescence microscopy has provided researchers with unique possibilities of three-dimensional imaging of biological cells and tissues and of other structures such as semiconductor integrated circuits. Confocal and Two-Photon Microscopy: Foundations, Applications, and Advances provides clear, comprehensive coverage of basic foundations, modern applications, and groundbreaking new research developments made in this important area of microscopy. Opening with a foreword by G. J. Brakenhoff, this reference gathers the work of an international group of renowned experts in chapters that are logically divided into balanced sections covering theory, techniques, applications, and advances, featuring: In-depth discussion of applications for biology, medicine, physics, engineering, and chemistry, including industrial applications Guidance on new and emerging imaging technology, developmental trends, and fluorescent molecules Uniform organization and review-style presentation of chapters, with an introduction, historical overview, methodology, practical tips, applications, future directions, chapter summary, and bibliographical references Companion FTP site with full-color photographs The significant experience of pioneers, leaders, and emerging scientists in the field of confocal and two-photon excitation microscopy Confocal and Two-Photon Microscopy: Foundations, Applications, and Advances is invaluable to researchers in the biological sciences, tissue and cellular engineering, biophysics, bioengineering, physics of matter, and medicine, who use these techniques or are involved in developing new commercial instruments.

  4. A state-space Bayesian framework for estimating biogeochemical transformations using time-lapse geophysical data

    Science.gov (United States)

    Chen, Jinsong; Hubbard, Susan S.; Williams, Kenneth H.; Pride, Steve; Li, Li; Steefel, Carl; Slater, Lee

    2009-08-01

    We develop a state-space Bayesian framework to combine time-lapse geophysical data with other types of information for quantitative estimation of biogeochemical parameters during bioremediation. We consider characteristics of end products of biogeochemical transformations as state vectors, which evolve under constraints of local environments through evolution equations, and consider time-lapse geophysical data as available observations, which could be linked to the state vectors through petrophysical models. We estimate the state vectors and their associated unknown parameters over time using Markov chain Monte Carlo sampling methods. To demonstrate the use of the state-space approach, we apply it to complex resistivity data collected during laboratory column biostimulation experiments that were poised to precipitate iron and zinc sulfides during sulfate reduction. We develop a petrophysical model based on sphere-shaped cells to link the sulfide precipitate properties to the time-lapse geophysical attributes and estimate volume fraction of the sulfide precipitates, fraction of the dispersed, sulfide-encrusted cells, mean radius of the aggregated clusters, and permeability over the course of the experiments. Results of the case study suggest that the developed state-space approach permits the use of geophysical data sets for providing quantitative estimates of end-product characteristics and hydrological feedbacks associated with biogeochemical transformations. Although tested here on laboratory column experiment data sets, the developed framework provides the foundation needed for quantitative field-scale estimation of biogeochemical parameters over space and time using direct, but often sparse wellbore data with indirect, but more spatially extensive geophysical data sets.

  5. Time-lapse camera studies of sea-disposed chemical munitions in Hawaii

    Science.gov (United States)

    Edwards, Margo H.; Fornari, Daniel J.; Rognstad, Mark R.; Kelley, Christopher D.; Mah, Christopher L.; Davis, Logan K.; Flores, Kyle R. M.; Main, Erin L.; Bruso, Natalie L.

    2016-06-01

    The interactions between fauna and sea-disposed munitions provide important evidence regarding whether munitions constituents affect the health of the ocean environment and its inhabitants. To date few studies of these interactions have been conducted at deep-water disposal sites; typically observations of fauna in the vicinity of sea-disposed munitions are limited to the few minutes or hours required to collect physical samples at a specific location. During the 2012 Hawaii Undersea Military Munitions Assessment (HUMMA) field program we deployed two deep-sea time-lapse camera systems with the objectives of cataloging the diversity of fauna visiting sea-disposed chemical munitions and observing faunal behavior and physiology. Over the 1- and 3-day deployments we recorded 28 different species of fishes, crustaceans, mollusks, cnidarians, and echinoderms at the two sites. Both cameras captured the previously undocumented behavior of brisingid sea stars repositioning themselves along chemical munitions casings. Despite the fact that brisingid sea stars are able to move, for the duration of both time-lapse experiments they remained on chemical munitions casings. We interpret this result to indicate that the advantages of residing on a hard substrate slightly elevated above the seafloor outweigh the effects of chemical munitions constituents for brisingid sea stars. One type of physiological anomaly observed on several arms of the brisingid sea stars at the time-lapse sites led to the collection and examination of six specimens. As reported by Mah (2015. Deep Sea Res. II, 2015, XX-XX), these physiological features are the result of parasitic crustaceans and are not caused by chemical munitions constituents.

  6. Time-Lapse Electrical Geophysical Monitoring of Amendment-Based Biostimulation

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, Timothy C.; Versteeg, Roelof; Day-Lewis, Frederick D.; Major, William; Lane, John W.

    2015-12-02

    Biostimulation is increasingly used to accelerate microbial remediation of recalcitrant groundwater contaminants. Effective application of biostimulation requires successful emplacement of amendment in the contaminant target zone. Verification of remediation performance requires postemplacement assessment and contaminant monitoring. Sampling based approaches are expensive and provide low-density spatial and temporal information. Time-lapse electrical resistivity tomography (ERT) is an effective geophysical method for determining temporal changes in subsurface electrical conductivity. Because remedial amendments and biostimulation-related biogeochemical processes often change subsurface electrical conductivity, ERT can complement and enhance sampling-based approaches for assessing emplacement and monitoring biostimulation-based remediation. Field studies demonstrating the ability of time-lapse ERT to monitor amendment emplacement and behavior were performed during a biostimulation remediation effort conducted at the Department of Defense Reutilization and Marketing Office (DRMO) Yard, in Brandywine, Maryland, United States. Geochemical fluid sampling was used to calibrate a petrophysical relation in order to predict groundwater indicators of amendment distribution. The petrophysical relations were field validated by comparing predictions to sequestered fluid sample results, thus demonstrating the potential of electrical geophysics for quantitative assessment of amendment-related geochemical properties. Crosshole radar zero-offset profile and borehole geophysical logging were also performed to augment the data set and validate interpretation. In addition to delineating amendment transport in the first 10 months after emplacement, the time-lapse ERT results show later changes in bulk electrical properties interpreted as mineral precipitation. Results support the use of more cost-effective surfacebased ERT in conjunction with limited field sampling to improve spatial

  7. Time-lapse refraction seismic tomography for the detection of ground ice degradation

    Directory of Open Access Journals (Sweden)

    C. Hilbich

    2010-07-01

    Full Text Available The ice content of the subsurface is a major factor controlling the natural hazard potential of permafrost degradation in alpine terrain. Monitoring of changes in ice content is therefore similarly important as temperature monitoring in mountain permafrost. Although electrical resistivity tomography monitoring (ERTM proved to be a valuable tool for the observation of ice degradation, results are often ambiguous or contaminated by inversion artefacts. In theory, the sensitivity of P-wave velocity of seismic waves to phase changes between unfrozen water and ice is similar to the sensitivity of electric resistivity. Provided that the general conditions (lithology, stratigraphy, state of weathering, pore space remain unchanged over the observation period, temporal changes in the observed travel times of repeated seismic measurements should indicate changes in the ice and water content within the pores and fractures of the subsurface material. In this paper, a time-lapse refraction seismic tomography (TLST approach is applied as an independent method to ERTM at two test sites in the Swiss Alps. The approach was tested and validated based on a the comparison of time-lapse seismograms and analysis of reproducibility of the seismic signal, b the analysis of time-lapse travel time curves with respect to shifts in travel times and changes in P-wave velocities, and c the comparison of inverted tomograms including the quantification of velocity changes. Results show a high potential of the TLST approach concerning the detection of altered subsurface conditions caused by freezing and thawing processes. For velocity changes on the order of 3000 m/s even an unambiguous identification of significant ice loss is possible.

  8. Epikarstic storage and doline structural characterization with time-lapse geophysics (seismic refraction & electrical resistivity)

    Science.gov (United States)

    Valois, R.; Galibert, P.; Guérin, R.; Mendes, M.; Plagnes, V.

    2011-12-01

    Karst formations are one of the most challenging environments in terms of groundwater, engineering and environmental issues. Geophysical methods can provide useful subsurface information in karst regions concerning groundwater vulnerability assessment, exploitation or hazard estimation. First, dolines are studied as preferential pathways for the protection of karstic aquifer in south France. Geophysics helps to characterize lateral and underground morphologies of such objects and is able to detect doline hidden by the soil cover too. Electrical resistivity and seismic refraction tomographies provide information about dolines filling and could help to propose a genesis scenario. Time-lapse resistivity measurements show that the studied doline is more vulnerable to infiltration on its sides than at its centre. The epikarst could be defined as a perched aquifer above the massive carbonate rocks; it constitutes a highly fractured zone, which water stock capacities. So, the epikarst was investigated with 3D seismic refraction and results show an important velocity anisotropy linked to the fracturing and weathering of the dolostone. The 3D model presents also some large heterogeneities: a corridor with highly weathered dolostone and an unweathered pinnacle. The corridor is probably situated on vertical joints, which have conducted aggressive water. The associated weathering with residual weathered-rock keeping its initial volume could create a "ghost-rock" corridor. So, the epikarst in the dolostones of the Causse du Larzac (France) seems to be composed by "ghost-rock" developed around a specific direction of fractures. Time-lapse electrical resistivity and seismic refraction velocity were carried out on this epikarst to observe the influence of water saturation on the measurements. The results show important variations for both seismic and electrical methods and are localized in the first 6 m: in the weathered zone. So, time-lapse measurements could more easily identify

  9. Two-Photon Absorption of Metal-Assisted Chromophores.

    Science.gov (United States)

    Li, Xin; Rinkevicius, Zilvinas; Ågren, Hans

    2014-12-09

    Aiming to understand the effect of a metal surface on nonlinear optical properties and the combined effects of surface and solvent environments on such properties, we present a multiscale response theory study, integrated with dynamics of the two-photon absorption of 4-nitro-4'-amino-trans-stilbene physisorbed on noble metal surfaces, considering two such surfaces, Ag(111) and Au(111), and two solvents, cyclohexane and water, as cases for demonstration. A few conclusions of general character could be drawn: While the geometrical change of the chromophore induced by the environment was found to notably alter (diminish) the two-photon absorption cross section in the polar medium, the effects of the metal surface and solvent on the electronic structure of the chromophore surpasses the geometrical effects and leads to a considerably enhanced two-photon absorption cross section in the polar solvent. This enhancement of two-photon absorption arises essentially from the metal charge image induced enlargement of the difference between the dipole moment of the excited state and the ground state. The orientation-dependence of the two-photon absorption is found to connect with the lateral rotation of the chromophore, where the two-photon absorption reaches its maximum when the polarization of the incident light coincides with the long-axis of the chromophore. Our results demonstrate a distinct enhancement of the two-photon absorption by a metal surface and a polar medium and envisage the employment of metal-chromophore composite materials for future development of nonlinear optical materials with desirable properties.

  10. Quantification of erosion and sedimentation using time-lapse gravimetry and Lidar in southern Taiwan

    Science.gov (United States)

    Mouyen, Maxime; Steer, Philippe; Croissant, Thomas; Le Moigne, Nicolas; Hwang, Cheinway; Cheng, Ching-Chung; Masson, Frédéric; Davy, Philippe; Lague, Dimitri; Longuevergne, Laurent

    2016-04-01

    After the 2009 Morakot typhoon, which triggered numerous large landslides in Taiwan, Mouyen et al. (2013) showed for the first time the potential of time-lapse gravity survey to infer the mass of sediments transferring by landsliding or through rivers. By providing an integrated measurement of masses, gravimetry might thus be complementary to common methods used to assess the sediments discharge of rivers. But the masses of rocks displaced by Morakot were exceptionally large as a result of the record-breaking rainfalls brought by this typhoon and one might wonder to what extent time-lapse gravimetry could record such sediment transfers. In order to better assess the capabilities of this method, we set a time-lapse gravity network dedicated to the monitoring of such sediments transfers in Paolai village (south-central Taiwan). Paolai is located near the large Laonong river where temporary alluvial deposits of sediments exist and face steep mountain slopes likely to experience landslides. Both features are considered as potential source of mass transfers, and in turn of temporal gravity changes. The first base gravity measurements were done in November 2015, using absolute and relative gravimeters, and will be repeated every year, before and after the typhoon season. In the same time, we also use a terrestrial lidar to scan the geometry of both the river and the mountain slopes, hence providing a detailed topographical survey of the studied area. Adding Lidar measurements is an efficient strategy to solve for the non-uniqueness of gravity solutions. Meanwhile, we use the Eros morphodynamic model, that combine landsliding and flooding models, to investigate various scenarios of landsliding and subsequent sediment transport and compute the gravity changes on a virtual network of gravimeters. This gives us insights on the expected order of magnitudes for these surface sediment transfers, which are useful to unravel the induced gravity signal from others sources such as

  11. Time-lapse and slow-motion tracking of temperature changes: response time of a thermometer

    Science.gov (United States)

    Moggio, L.; Onorato, P.; Gratton, L. M.; Oss, S.

    2017-03-01

    We propose the use of a smartphone based time-lapse and slow-motion video techniques together with tracking analysis as valuable tools for investigating thermal processes such as the response time of a thermometer. The two simple experimental activities presented here, suitable also for high school and undergraduate students, allow one to measure in a simple yet rigorous way the response time of an alcohol thermometer and show its critical dependence on the properties of the surrounding environment giving insight into instrument characteristics, heat transfer and thermal equilibrium concepts.

  12. Soil Contamination, Advanced integrated characterisation and time-lapse Monitoring, SoilCAM project highlights

    Science.gov (United States)

    French, H. K.; Van Der Zee, S. E.; Wehrer, M.; Godio, A.; Pedersen, L. B.; Tsocano, G.

    2013-12-01

    The SoilCAM project (2008- 2012, EU-FP7-212663) aimed at improving methods for monitoring subsurace contaminant distribution and biodegradation. Two test sites were chosen, Oslo airport Gardermoen, Norway where de-icing agents infiltrate the soil during snowmelt and the Trecate site in Italy where an inland crude oil spill occurred in 1994. A number of geophysical investigation techniques were combined with soil and water sampling techniques. Data obtained from time-lapse measurements were further analysed by numerical modelling of flow and transport at different scales in order to characterise transport processes in the unsaturated and saturated zones. Laboratory experiments provided physical and biogeochemical data for model parameterisation and to select remediation methods. The geophysical techniques were used to map geological heterogeneities and to conduct time-lapse measurements of processes in the unsaturated zone. Both cross borehole and surface electrodes were used for electrical resistivity and induced polarisation surveys. Results showed clear indications of areas highly affected by de-icing chemicals along the runway at Oslo airport. The time lapse measurements along the runway at the airport showed infiltration patterns during snowmelt and were used to validate 2D unsaturated flow and transport simulations using SUTRA. The simulations illustrate the effect of layering geological structures and membranes, buried parallel to the runway, on the flow pattern. Complex interaction between bio-geo-chemical processes in a 1D vertical profile along the runway were described with the ORCHESTRA model. Smaller scale field site measurements revealed increase of iron and manganese during degradation of de-icing chemicals. At the Trecate site a combination of georadar, electrical resistivity and radio magneto telluric provided a broad outline of the geology down to 50 m. Anomalies in the Induced polarisation and electrical resistivity data from the cross borehole

  13. Live-streaming: Time-lapse video evidence of novel streamer formation mechanism and varying viscosity.

    Science.gov (United States)

    Parvinzadeh Gashti, Mazeyar; Bellavance, Julien; Kroukamp, Otini; Wolfaardt, Gideon; Taghavi, Seyed Mohammad; Greener, Jesse

    2015-07-01

    Time-lapse videos of growing biofilms were analyzed using a background subtraction method, which removed camouflaging effects from the heterogeneous field of view to reveal evidence of streamer formation from optically dense biofilm segments. In addition, quantitative measurements of biofilm velocity and optical density, combined with mathematical modeling, demonstrated that streamer formation occurred from mature, high-viscosity biofilms. We propose a streamer formation mechanism by sudden partial detachment, as opposed to continuous elongation as observed in other microfluidic studies. Additionally, streamer formation occurred in straight microchannels, as opposed to serpentine or pseudo-porous channels, as previously reported.

  14. Non-linear imaging and characterization of atherosclerotic arterial tissue using combined two photon fluorescence, second-harmonic generation and CARS microscopy

    Science.gov (United States)

    Cicchi, Riccardo; Matthäus, Christian; Meyer, Tobias; Lattermann, Annika; Dietzek, Benjamin; Brehm, Bernhard R.; Popp, Jürgen; Pavone, Francesco S.

    2014-02-01

    Atherosclerosis is among the most widespread cardiovascular diseases and one of the leading cause of death in the Western World. Characterization of arterial tissue in atherosclerotic condition is extremely interesting from the diagnostic point of view. Routinely used diagnostic methods, such as histopathological examination, are limited to morphological analysis of the examined tissues, whereas an exhaustive characterization requires a morpho-functional approach. Multimodal non-linear microscopy has the potential to bridge this gap by providing morpho-functional information on the examined tissues in a label-free way. Here we employed multiple non-linear microscopy techniques, including CARS, TPF, and SHG to provide intrinsic optical contrast from various tissue components in both arterial wall and atherosclerotic plaques. CARS and TPF microscopy were used to respectively image lipid depositions within plaques and elastin in the arterial wall. Cholesterol deposition in the lumen and collagen in the arterial wall were selectively imaged by SHG microscopy and distinguished by forward-backward SHG ratio. Image pattern analysis allowed characterizing collagen organization in different tissue regions. Different values of fiber mean size, distribution and anisotropy are calculated for lumen and media prospectively allowing for automated classification of atherosclerotic lesions. The presented method represents a promising diagnostic tool for evaluating atherosclerotic tissue and has the potential to find a stable place in clinical setting as well as to be applied in vivo in the near future.

  15. Nonlinear quantitative photoacoustic tomography with two-photon absorption

    CERN Document Server

    Ren, Kui

    2016-01-01

    Two-photon photoacoustic tomography (TP-PAT) is a non-invasive optical molecular imaging modality that aims at inferring two-photon absorption property of heterogeneous media from photoacoustic measurements. In this work, we analyze an inverse problem in quantitative TP-PAT where we intend to reconstruct optical coefficients in a semilinear elliptic PDE, the mathematical model for the propagation of near infra-red photons in tissue-like optical media with two-photon absorption, from the internal absorbed energy data. We derive uniqueness and stability results on the reconstructions of single and multiple optical coefficients, and present some numerical reconstruction results based on synthetic data to complement the theoretical analysis.

  16. Enhanced two-photon absorption using true thermal light

    CERN Document Server

    Jechow, Andreas; Kurzke, Henning; Heuer, Axel; Menzel, Ralf

    2013-01-01

    Two-photon excited fluorescence (TPEF) is a standard technique in modern microscopy but still affected by photo-damage of the probe. It was proposed that TPEF can be enhanced by using entangled photons, but has proven to be challenging. Recently it was shown that some features of entangled photons can be mimicked with thermal light, which finds application in ghost imaging, sub-wavelength lithography and metrology. Here, we utilize true thermal light from a super-luminescence diode to demonstrate enhanced TPEF compared to coherent light using two common fluorophores and luminescent quantum dots. We find that the two-photon absorption rate is directly proportional to the measured degree of second-order coherence, as predicted by theory. Our results show that photon bunching can be exploited in two-photon microscopy with the photon statistic providing a new degree of freedom.

  17. The Extreme Ice Survey: Capturing and Conveying Glacial Processes Through Time-Lapse Imagery and Narration

    Science.gov (United States)

    Balog, J. D.; Box, J. E.; Pfeffer, W. T.; Hood, E. W.; Fagre, D. B.; Anker, C.; O'Neel, S.

    2010-12-01

    The Extreme Ice Survey (EIS) uses time-lapse photography, conventional photography, and video to document rapid change in the Earth's glacial ice. The EIS team currently has 38 time-lapse cameras at sites in Greenland, Iceland, Alaska, the Rocky Mountains and Nepal. EIS supplements this ongoing record with annual repeat photography in British Columbia, Iceland, the Alps, and Bolivia. EIS imagery supplies basic knowledge in glacier dynamics to the science community, as well as compelling, engaging narratives to the general public about the immediacy of the Anthropocene and climate change. Visual materials from EIS have impacted more than 150 million people, ranging from White House staff, the U. S. Congress and government agency officials to globally influential corporate officers and all age strata of the general public. Media products include a National Geographic/NOVA special, two National Geographic magazine articles, a feature in Parade magazine (circulation 71 million), and numerous presentations on CNN, NBC, BBC and National Public Radio. Columbia Glacier, Alaska, June 2006, May 2007, June 2008 terminus indicated.

  18. The time-lapse AVO difference inversion for changes in reservoir parameters

    Science.gov (United States)

    Longxiao, Zhi; Hanming, Gu; Yan, Li

    2016-12-01

    The result of conventional time-lapse seismic processing is the difference between the amplitude and the post-stack seismic data. Although stack processing can improve the signal-to-noise ratio (SNR) of seismic data, it also causes a considerable loss of important information about the amplitude changes and only gives the qualitative interpretation. To predict the changes in reservoir fluid more precisely and accurately, we also need the quantitative information of the reservoir. To achieve this aim, we develop the method of time-lapse AVO (amplitude versus offset) difference inversion. For the inversion of reservoir changes in elastic parameters, we apply the Gardner equation as the constraint and convert the three-parameter inversion of elastic parameter changes into a two-parameter inversion to make the inversion more stable. For the inversion of variations in the reservoir parameters, we infer the relation between the difference of the reflection coefficient and variations in the reservoir parameters, and then invert reservoir parameter changes directly. The results of the theoretical modeling computation and practical application show that our method can estimate the relative variations in reservoir density, P-wave and S-wave velocity, calculate reservoir changes in water saturation and effective pressure accurately, and then provide reference for the rational exploitation of the reservoir.

  19. Time lapse seismic observations and effects of reservoir compressibility at Teal South oil field

    Science.gov (United States)

    Islam, Nayyer

    One of the original ocean-bottom time-lapse seismic studies was performed at the Teal South oil field in the Gulf of Mexico during the late 1990's. This work reexamines some aspects of previous work using modern analysis techniques to provide improved quantitative interpretations. Using three-dimensional volume visualization of legacy data and the two phases of post-production time-lapse data, I provide additional insight into the fluid migration pathways and the pressure communication between different reservoirs, separated by faults. This work supports a conclusion from previous studies that production from one reservoir caused regional pressure decline that in turn resulted in liberation of gas from multiple surrounding unproduced reservoirs. I also provide an explanation for unusual time-lapse changes in amplitude-versus-offset (AVO) data related to the compaction of the producing reservoir which, in turn, changed an isotropic medium to an anisotropic medium. In the first part of this work, I examine regional changes in seismic response due to the production of oil and gas from one reservoir. The previous studies primarily used two post-production ocean-bottom surveys (Phase I and Phase II), and not the legacy streamer data, due to the unavailability of legacy prestack data and very different acquisition parameters. In order to incorporate the legacy data in the present study, all three post-stack data sets were cross-equalized and examined using instantaneous amplitude and energy volumes. This approach appears quite effective and helps to suppress changes unrelated to production while emphasizing those large-amplitude changes that are related to production in this noisy (by current standards) suite of data. I examine the multiple data sets first by using the instantaneous amplitude and energy attributes, and then also examine specific apparent time-lapse changes through direct comparisons of seismic traces. In so doing, I identify time-delays that, when

  20. Analysis of compaction initiation in human embryos by using time-lapse cinematography.

    Science.gov (United States)

    Iwata, Kyoko; Yumoto, Keitaro; Sugishima, Minako; Mizoguchi, Chizuru; Kai, Yoshiteru; Iba, Yumiko; Mio, Yasuyuki

    2014-04-01

    To analyze the initiation of compaction in human embryos in vitro by using time-lapse cinematography (TLC), with the goal of determining the precise timing of compaction and clarifying the morphological changes underlying the compaction process. One hundred and fifteen embryos donated by couples with no further need for embryo-transfer were used in this study. Donated embryos were thawed and processed, and then their morphological behavior during the initiation of compaction was dynamically observed via time-lapse cinematography (TLC) for 5 days. Although the initiation of compaction occurred throughout the period from the 4-cell to 16-cell stage, 99 (86.1 %) embryos initiated compaction at the 8-cell stage or later, with initiation at the 8-cell stage being most frequent (22.6 %). Of these 99 embryos, 49.5 % developed into good-quality blastocysts. In contrast, of the 16 (13.9 %) embryos that initiated compaction prior to the 8-cell stage, only 18.8 % developed into good-quality blastocysts. Embryos that initiated compaction before the 8-cell stage showed significantly higher numbers of multinucleated blastomeres, due to asynchronism in nuclear division at the third mitotic division resulting from cytokinetic failure. The initiation of compaction primarily occurs at the third mitotic division or later in human embryos. Embryos that initiate compaction before the 8-cell stage are usually associated with aberrant embryonic development (i.e., cytokinetic failure accompanied by karyokinesis).

  1. Using a time lapse microgravity model for mapping seawater intrusion around Semarang

    Science.gov (United States)

    Supriyadi, Khumaedi, Yusuf, M.; Agung, W.

    2016-03-01

    A modeling of time-lapse microgravity anomaly due to sea water intrusion has been conducted. It used field data of aquifer cross section, aquifer thickness and lithology of research area. Those data were then processed using Grav3D and Surfer. Modeling results indicated that the intrusion of sea water resulting in a time-lapse microgravity anomalies of 0.12 to 0.18 mGal, at soil layer density of 0.15 g/cm3 to 0.3 g/cm3 and at depth of 30 to 100 m. These imply that the areas experiencing seawater intrusion were Tanjung Mas, SPBE Bandarharjo, Brass, Old Market Boom and Johar as the microgravity measured there were in the range of 0.12 to 0.18 mGal and the density contrast were at 0.15 g/cm3 to 0.28 g/cm3. Areas that experienced fluid reduction were Puri Anjasmoro, Kenconowungu and Puspowarno with microgravity changes from -0.06 mGal to -0.18 mGal.

  2. Time-lapse 3-D measurements of a glucose biosensor in multicellular spheroids by light sheet fluorescence microscopy in commercial 96-well plates

    Science.gov (United States)

    Maioli, Vincent; Chennell, George; Sparks, Hugh; Lana, Tobia; Kumar, Sunil; Carling, David; Sardini, Alessandro; Dunsby, Chris

    2016-11-01

    Light sheet fluorescence microscopy has previously been demonstrated on a commercially available inverted fluorescence microscope frame using the method of oblique plane microscopy (OPM). In this paper, OPM is adapted to allow time-lapse 3-D imaging of 3-D biological cultures in commercially available glass-bottomed 96-well plates using a stage-scanning OPM approach (ssOPM). Time-lapse 3-D imaging of multicellular spheroids expressing a glucose Förster resonance energy transfer (FRET) biosensor is demonstrated in 16 fields of view with image acquisition at 10 minute intervals. As a proof-of-principle, the ssOPM system is also used to acquire a dose response curve with the concentration of glucose in the culture medium being varied across 42 wells of a 96-well plate with the whole acquisition taking 9 min. The 3-D image data enable the FRET ratio to be measured as a function of distance from the surface of the spheroid. Overall, the results demonstrate the capability of the OPM system to measure spatio-temporal changes in FRET ratio in 3-D in multicellular spheroids over time in a multi-well plate format.

  3. Three-dimensional microfabrication using two-photon polymerization

    Science.gov (United States)

    Cumpston, Brian H.; Ehrlich, Jeffrey E.; Kuebler, Stephen M.; Lipson, Matthew; Marder, Seth R.; McCord-Maughon, D.; Perry, Joseph W.; Roeckel, Harold; Rumi, Maria Cristina

    1998-09-01

    Photopolymerization initiated by the simultaneous absorption of two photons is unique in its ability to produce complex three-dimensional (3D) structures from a single, thick photopolymer film. Strong 3D confinement of the polymerization process is not possible in other polymer microfabrication techniques such as LIGA, rapid prototyping, and conventional photoresist technology. Two-photon polymerization also permits the fabrication of 3D structures and the definition of lithographic features on non-planar surfaces. We have developed a wide array of chromophores which hold great promise for 3D microfabrication, as well as other applications, such as two-photon fluorescence imaging and 3D optical data storage. These materials are based on a donor- (pi) -donor, donor-acceptor-donor, or acceptor-donor-acceptor structural motif. The magnitude of the two-photon absorption cross-section, (delta) , and the position of the two-photon absorption maximum, (lambda) (2)max, can be controlled by varying the length of the conjugated bridge and by varying the strength of the donor/acceptor groups. In this way, chromophores have been developed which exhibit strong two- photon absorption in the range of 500 - 975 nm, in some cases as high as 4400 X 10-50 cm4 s/photon-molecule. In the case of donor-(pi) -donor structures, quantum-chemical calculations show that the large absorption cross-sections arise from the symmetric re-distribution of charge from the donor end-groups to the conjugated bridge, resulting in an electronic excited-state which is more delocalized than the ground state. For many of these molecules, two-photon excitation populates a state which is sufficiently reducing that a charge transfer reaction can occur with acrylate monomers. The efficiency of these processes can be described using Marcus theory. Under suitable conditions, such reactions can induce radical polymerization of acrylate resins. Polymerization rates have been measured, and we show that these two-photon

  4. Fano interference in two-photon transport

    Science.gov (United States)

    Xu, Shanshan; Fan, Shanhui

    2016-10-01

    We present a general input-output formalism for the few-photon transport in multiple waveguide channels coupled to a local cavity. Using this formalism, we study the effect of Fano interference in two-photon quantum transport. We show that the physics of Fano interference can manifest as an asymmetric spectral line shape in the frequency dependence of the two-photon correlation function. The two-photon fluorescence spectrum, on the other hand, does not exhibit the physics of Fano interference.

  5. Integrating Terrestrial Time-Lapse Photography with Laser Scanning to Distinguish the Drivers of Movement at Sólheimajökull, Iceland

    Science.gov (United States)

    How, P.; James, M. R.; Wynn, P.

    2014-12-01

    Glacier movement is attributed to a sensitive configuration of driving forces. Here, we present an approach designed to evaluate the drivers of movement at Sólheimajökull, an outlet glacier from the Myrdalsjökull ice cap, Iceland, through combining terrestrial time-lapse photography and laser scanning (TLS). A time-lapse camera (a dSLR with intervalometer and solar-recharged battery power supply) collected hourly data over the summer of 2013. The data are subject to all the difficulties that are usually present in long time-lapse sequences, such as highly variable illumination and visibility conditions, evolving surfaces, and camera instabilities. Feature-tracking software [1] was used to: 1) track regions of static topography (e.g. the skyline) from which camera alignment could be continuously updated throughout the sequence; and 2) track glacial surface features for velocity estimation. Absolute georeferencing of the image sequence was carried out by registering the camera to a TLS survey acquired at the beginning of the monitoring period. A second TLS survey (July 2013) provided an additional 3D surface. By assuming glacial features moved in approximately planimetrically straight lines between the two survey dates, combining the two TLS surfaces with the monoscopic feature tracking allows 3D feature tracks to be derived. Such tracks will enable contributions from different drivers (e.g. surface melting) to be extracted, even in imagery that is acquired not perpendicular to glacier motion. At Sólheimajökull, our aim is to elucidate any volcanic contribution to the observed movement.[1] http://www.lancaster.ac.uk/staff/jamesm/software/pointcatcher.htm

  6. Adiabatic following in two-photon transition

    Energy Technology Data Exchange (ETDEWEB)

    Nayfeh, M.H.; Nayfeh, A.H.

    1977-01-01

    There has been much interest recently in coherent multiphoton transitions in many-level systems. The present work considers the effect of relaxation in the response of a three-level system to a smoothly varying, near-resonant, two-photon field. The relaxation-dependent contributions to the nonlinear refractive index are calculated. It is shown that the coherent interaction of two smoothly varying, near-resonant, two-photon pulses with a three-level system can be described by ''two-photon damped Bloch equations'' which are analogous to those for a one-photon transition in a two-level system except for the presence of a two-photon coupling and a frequency shift. 1 figure. (RWR)

  7. Two-Photon Physics in Hadronic Processes

    Energy Technology Data Exchange (ETDEWEB)

    Carl Carlson; Marc Vanderhaeghen

    2007-11-01

    Two-photon exchange contributions to elastic electron-scattering are reviewed. The apparent discrepancy in the extraction of elastic nucleon form factors between unpolarized Rosenbluth and polarization transfer experiments is discussed, as well as the understanding of this puzzle in terms of two-photon exchange corrections. Calculations of such corrections both within partonic and hadronic frameworks are reviewed. In view of recent spin-dependent electron scattering data, the relation of the two-photon exchange process to the hyperfine splitting in hydrogen is critically examined. The imaginary part of the two-photon exchange amplitude as can be accessed from the beam normal spin asymmetry in elastic electron-nucleon scattering is reviewed. Further extensions and open issues in this field are outlined.

  8. Sideband-Induced Two-Photon Transparency

    Institute of Scientific and Technical Information of China (English)

    CHENG Guang-Ling; HU Xiang-Ming

    2006-01-01

    @@ We show that it is possible to use a single sideband to induce two-photon transparency in a three-level cascade medium. The medium simultaneously absorbs two photons as a one-step process when the middle level is far off one-photon resonance. A resonant sideband coupling on the upper transition and the two-photon one-step process drive the medium into a trapped state, and the dominant component is the ground state. Thus almost all population is trapped in the ground state and the two-photon absorption is dramatically suppressed. We present a numerical calculation for arbitrary values of the atomic and field parameters and also provide an analytic description for the required conditions.

  9. Time-lapse analysis reveals different modes of primordial germ cell migration in the medaka Oryzias latipes.

    Science.gov (United States)

    Kurokawa, Hiromi; Aoki, Yumiko; Nakamura, Shuhei; Ebe, Youko; Kobayashi, Daisuke; Tanaka, Minoru

    2006-04-01

    Previous studies have shown that medaka primordial germ cells (PGC) are first distinguishable by olvas expression during late gastrulation, and that they migrate to the gonadal region through the lateral plate mesoderm. Here, we demonstrate that medaka nanos expression marks the germ line at early gastrulation stage. By marking the germ line with green fluorescent protein (GFP) fused to the nanos 3' untranslated region, we were able to visualize the behavior of PGC using time-lapse imaging. We show that there are three distinct modes of PGC migration that function at different stages of development. At early gastrulation stage, PGC actively migrate towards the marginal zone, a process that requires the function of a chemokine receptor, CXCR4. However, at late gastrulation stage, PGC change the mode and direction of their movement, as they are carried towards the midline along with somatic cells undergoing convergent movements. After aligning bilaterally, PGC actively migrate to the posterior end of the lateral plate mesoderm. This posterior movement depends on the activity of both HMGCoAR and a ligand of CXCR4, SDF-1a. These results demonstrate that PGC undergo different modes of migration to reach the prospective gonadal region of the embryo.

  10. Automated segmentation and tracking of non-rigid objects in time-lapse microscopy videos of polymorphonuclear neutrophils.

    Science.gov (United States)

    Brandes, Susanne; Mokhtari, Zeinab; Essig, Fabian; Hünniger, Kerstin; Kurzai, Oliver; Figge, Marc Thilo

    2015-02-01

    Time-lapse microscopy is an important technique to study the dynamics of various biological processes. The labor-intensive manual analysis of microscopy videos is increasingly replaced by automated segmentation and tracking methods. These methods are often limited to certain cell morphologies and/or cell stainings. In this paper, we present an automated segmentation and tracking framework that does not have these restrictions. In particular, our framework handles highly variable cell shapes and does not rely on any cell stainings. Our segmentation approach is based on a combination of spatial and temporal image variations to detect moving cells in microscopy videos. This method yields a sensitivity of 99% and a precision of 95% in object detection. The tracking of cells consists of different steps, starting from single-cell tracking based on a nearest-neighbor-approach, detection of cell-cell interactions and splitting of cell clusters, and finally combining tracklets using methods from graph theory. The segmentation and tracking framework was applied to synthetic as well as experimental datasets with varying cell densities implying different numbers of cell-cell interactions. We established a validation framework to measure the performance of our tracking technique. The cell tracking accuracy was found to be >99% for all datasets indicating a high accuracy for connecting the detected cells between different time points.

  11. The Relationship between Cell Number, Division Behavior and Developmental Potential of Cleavage Stage Human Embryos: A Time-Lapse Study.

    Science.gov (United States)

    Kong, Xiangyi; Yang, Shuting; Gong, Fei; Lu, Changfu; Zhang, Shuoping; Lu, Guangxiu; Lin, Ge

    2016-01-01

    Day 3 cleavage embryo transfer is routine in many assisted reproductive technology centers today. Embryos are usually selected according to cell number, cell symmetry and fragmentation for transfer. Many studies have showed the relationship between cell number and embryo developmental potential. However, there is limited understanding of embryo division behavior and their association with embryo cell number and developmental potential. A retrospective and observational study was conducted to investigate how different division behaviors affect cell number and developmental potential of day 3 embryos by time-lapse imaging. Based on cell number at day 3, the embryos (from 104 IVF/intracytoplasmic sperm injection (ICSI) treatment cycles, n = 799) were classified as follows: less than 5 cells (10C; n = 42). Division behavior, morphokinetic parameters and blastocyst formation rate were analyzed in 5 groups of day 3 embryos with different cell numbers. In 10C embryos increased compared to 7-8C embryos (45.8%, 33.3% vs. 11.1%, respectively). In ≥5C embryos, FR and DC significantly reduced developmental potential, whereas division behaviors. In NB embryos, the blastocyst formation rate increased with cell number from 7.4% (10C). In NB embryos, the cell cycle elongation or shortening was the main cause for abnormally low or high cell number, respectively. After excluding embryos with abnormal division behaviors, the developmental potential, implantation rate and live birth rate of day 3 embryos increased with cell number.

  12. Correlations of two photons at hadron colliders

    OpenAIRE

    Kozlov, G. A.

    2011-01-01

    We study the Bose-Einstein correlations of two photons and their coherent properties that can provide the information about the space-time structure of the emitting source through the Higgs-boson decays into two photons. We argue that such an investigation could probe the Higgs-boson mass. The model is rather sensitive to the temperature of the environment and to the external distortion effect in medium.

  13. Platinum Acetylide Two-Photon Chromophores (Preprint)

    Science.gov (United States)

    2007-04-01

    the higher energy range that lead to its photodegradation . Secondly, because there is a quadratic dependence of two-photon absorption (2PA) on the...to either an electron donating amino- fluorenyl or electron withdrawing benzothiazolyl-fluorene that are themselves known as two-photon absorbing dyes ...groups in place of phenyl groups have shown a doubling of the intrinsic cr2value at 740 nm.40,41In this paper we describe novel platinum dyes that

  14. Two-photon absorbing porphyrins for oxygen microscopy (Conference Presentation)

    Science.gov (United States)

    Esipova, Tatiana V.; Vinogradov, Sergei A.

    2016-03-01

    The ability to quantify oxygen in vivo in 3D with high spatial and temporal resolution is invaluable for many areas of the biomedical science, including ophthalmology, neuroscience, cancer and stem biology. An optical method based on oxygen-dependent quenching of phosphorescence is being developed, that allows quantitative minimally invasive real-time imaging of partial pressure of oxygen (pO2) in tissue. In the past, dendritically protected phosphorescent oxygen probes with controllable quenching parameters and defined bio-distributions have been developed. More recently our probe strategy has extended to encompass two-photon excitable oxygen probes, which brought about first demonstrations of two-photon phosphorescence lifetime microscopy (2PLM) of oxygen in vivo, providing new valuable information for neuroscience and stem cell biology. However, current two-photon oxygen probes suffer from a number of limitations, such as low brightness and high cost of synthesis, which dramatically reduce imaging performance and limit usability of the method. Here we present an approach to new bright phosphorescent chromophores with internally enhanced two-photon absorption cross-sections, which pave a way to novel proves for 2PLM. In addition to substantial increase in performance, the new probes can be synthesized by much more efficient methods, thereby greatly reducing the cost of the synthesis and making the technique accessible to a broader range of researchers across different fields.

  15. Time-lapse cinematography of the capillary tube cell migration inhibition test.

    Science.gov (United States)

    Bray, M A

    1980-01-01

    The kinetics of human and guinea pig cell migration inhibition have been studied using time-lapse cinematography of cells migrating from capillary tubes. Guinea pig and human cells exhibit markedly different kinetics in the absence of inhibitors. Specific antigen causes a dose-related inhibition of migration for up to 60 h using guinea pig cells and a peak of inhibition after 18 h using the human leucocyte system. The timing of measurement of maximum activity more critical for the latter test. The kinetics of lymphokine generation have been examined and the migration inhibitory activity of the plant mitogen (PHA), a Kurloff cell product and a continuous cell line supernatant have been compared with the inhibitory profiles of lymphokine preparations and specific antigen.

  16. Effect of oxygen concentration on human embryo development evaluated by time-lapse monitoring

    DEFF Research Database (Denmark)

    Ingerslev, Hans Jakob; Hindkjær, Johnny Juhl; Kirkegaard, Kirstine

    2012-01-01

    -points are given as hours after fertilisation Results: The timing of the first two cleavage cycles resulting in a 4-cell embryo was not significantly different between the groups. The timing of the third cleavage cycle, i.e. division to 5, 6, 7 and 8 cells was delayed for embryos cultured in 20% oxygen (P5cell =0......Introduction: Data from a number of studies indicate -but not unequivocally- that culture of embryos in 5% O2 compared to 20% O2 improves blastocyst formation in humans and various animal species and may yield better pregnancy rates in IVF. The detrimental effects of atmospheric oxygen were...... recently demonstrated to occur from first cleavage cycle in mice using time-lapse microscopy, with the largest impact on the pre-compaction stages. However, embryonic development in mice differs in many aspects from human embryonic development. The objective of this retrospective, descriptive study...

  17. Time-lapse cinematographic analysis of beryllium--lung fibroblast interactions.

    Science.gov (United States)

    Absher, M; Sylwester, D; Hart, B A

    1983-02-01

    The proliferative response to beryllium chloride of cells in a population of human lung fibroblasts was quantitatively assessed using time-lapse cinematography. A dose of 0.02 microgram Be/ml, known to decrease the growth rate of fibroblasts, affects an estimated 75% of the cells in the population, increasing their interdivision time (IDT) by approximately 5 hr. The differences in mean 1n(IDT) between treated and control cells were essentially constant for comparable culture sizes ranging from 25 to 250 cells. There was no correlation between mother and daughter cell IDTs in control or treated culture at any culture size. IDTs of sister pairs were highly correlated in control cultures at selected culture sizes while sister pair IDTs of treated cultures were not. The data suggest that while beryllium alters the IDT of fibroblasts, an effect not related to culture size, any given cell affected by beryllium does not impart effects of the mineral to its progeny.

  18. Time-lapse seismic analysis of the North Sea Fulmar Field

    Energy Technology Data Exchange (ETDEWEB)

    Johnston, David H.; McKenny, Robert S.; Burkhart, Tucker D.

    1998-12-31

    Time-lapse seismic analysis has been applied to two 3-D seismic surveys acquired over the central North Sea Fulmar field in a pre-production survey shot in 1977, reprocessed in 1987, and a survey in 1992. The Upper Jurassic reservoirs in the field have been under production since 1982. Differences in averaged impedance between the 1977 and 1992 surveys clearly show the effects of water influx and pressure decline. The changes observed in the seismic data are overall consistent with predictions obtained from a full-field, history-matched simulation. Differences in details may suggest areas of bypassed oil. Dta quality is not sufficient to serve as the sole basis for drilling decisions. 1 ref., 6 figs.

  19. Time-lapse cinematography of dynamic changes occurring during in vitro development of human embryos.

    Science.gov (United States)

    Mio, Yasuyuki; Maeda, Kazuo

    2008-12-01

    The purpose of this study was to clarify developmental changes of early human embryos by using time-lapse cinematography (TLC). For human ova, fertilization and cleavage, development of the blastocyst, and hatching, as well as consequent changes were repeatedly photographed at intervals of 5-6 days by using an inverse microscope under stabilized temperature and pH. Photographs were taken at 30 frames per second and the movies were studied. Cinematography has increased our understanding of the morphologic mechanisms of fertilization, development, and behavior of early human embryos, and has identified the increased risk of monozygotic twin pregnancy based on prolonged incubation in vitro to the blastocyst stage. Using TLC, we observed the fertilization of an ovum by a single spermatozoon, followed by early cleavages, formation of the morula, blastocyst hatching, changes in the embryonic plates, and the development of monozygotic twins from the incubated blastocysts.

  20. In vivo amyloid aggregation kinetics tracked by time-lapse confocal microscopy in real-time.

    Science.gov (United States)

    Villar-Piqué, Anna; Espargaró, Alba; Ventura, Salvador; Sabate, Raimon

    2016-01-01

    Amyloid polymerization underlies an increasing number of human diseases. Despite this process having been studied extensively in vitro, aggregation is a difficult process to track in vivo due to methodological limitations and the slow kinetics of aggregation reactions in cells and tissues. Herein we exploit the amyloid properties of the inclusions bodies (IBs) formed by amyloidogenic proteins in bacteria to address the kinetics of in vivo amyloid aggregation. To this aim we used time-lapse confocal microscopy and a fusion of the amyloid-beta peptide (A β42) with a fluorescent reporter. This strategy allowed us to follow the intracellular kinetics of amyloid-like aggregation in real-time and to discriminate between variants exhibiting different in vivo aggregation propensity. Overall, the approach opens the possibility to assess the impact of point mutations as well as potential anti-aggregation drugs in the process of amyloid formation in living cells.

  1. Applicability of time-lapse refraction seismic tomography for the detection of ground ice degradation

    Directory of Open Access Journals (Sweden)

    C. Hilbich

    2010-01-01

    Full Text Available The ice content of the subsurface is a major factor controlling the natural hazard potential of permafrost degradation in alpine terrain. Monitoring of changes in ground ice content is therefore similarly important as temperature monitoring in mountain permafrost. Although electrical resistivity tomography monitoring (ERTM has proved to be a valuable tool for the observation of ground ice degradation, results are often ambiguous or contaminated by inversion artefacts. In theory, the P-wave velocity of seismic waves is similarly sensitive to phase changes between unfrozen water and ice. Provided that the general conditions (lithology, stratigraphy, state of weathering, pore space remain unchanged over the observation period, temporal changes in the observed travel times of repeated seismic measurements should indicate changes in the ice and water content within the pores and fractures of the subsurface material. In this paper, the applicability of refraction seismic tomography monitoring (RSTM as an independent and complementary method to ERTM is analysed for two test sites in the Swiss Alps. The development and validation of an appropriate RSTM approach involves a the comparison of time-lapse seismograms and analysis of reproducibility of the seismic signal, b the analysis of time-lapse travel time curves with respect to shifts in travel times and changes in P-wave velocities, and c the comparison of inverted tomograms including the quantification of velocity changes. Results show a high potential of the RSTM approach concerning the detection of altered subsurface conditions caused by freezing and thawing processes. For velocity changes on the order of 3000 m/s even an unambiguous identification of significant ground ice loss is possible.

  2. Assessment of Saturation Patterns on Agricultural Land Using Time-lapse Photography

    Science.gov (United States)

    Silasari, R.; Bloeschl, G.

    2015-12-01

    Agricultural land generates overland flow differently from natural environment due to features from anthropogenic activities such as cultivated soil layer and tile drain pipe. During rainfall events, the formation of overland flow may happen from infiltration excess and/or saturation excess according to the threshold processes which are influenced by rainfall characteristics and soil hydraulic parameters. The dynamics of threshold processes in varying rainfall and soil hydraulic conditions will affect the surface runoff response which can be inversely analyzed by visually observing the generated saturation patterns. This study aims to explore the use of time-lapse photographs of saturated plot during rainfall events to observe and understand the threshold processes of overland flow generation. The observation was conducted at Hydrological Open Air Laboratory (HOAL) in Lower Austria with a 2 megapixels surveillance camera overlooking a 1.8 ha tile-drained agricultural field situated on a hillslope. The main tile drain pipe extends from the higher ground into the riparian area - creating a depression line which generates the main saturation track. The time-lapse photographs are able to capture the spatial and temporal dynamics of 0.1 ha saturated plot (117 m long and 10 m wide in average) during three big rainfall events in 2014 which produced measurable overland flow. The photographs also manage to capture the behavior of overland flow on tractor tracks which were generated faster than on the main saturated plot - due to the more compacted soil - and contribute significantly to the overall overland flow discharge and movements. Comparison of the photographs with on-field manual plotting shows good accuracy of the captured saturation plot and the possibility of calculating the plot area digitally. This method gives opportunity to observe overland flow generation on visual basis as a complement of the customary discharge measurements.

  3. On the Resolvability of Steam Assisted Gravity Drainage Reservoirs Using Time-Lapse Gravity Gradiometry

    Science.gov (United States)

    Elliott, E. Judith; Braun, Alexander

    2017-08-01

    Unconventional heavy oil resource plays are important contributors to oil and gas production, as well as controversial for posing environmental hazards. Monitoring those reservoirs before, during, and after operations would assist both the optimization of economic benefits and the mitigation of potential environmental hazards. This study investigates how gravity gradiometry using superconducting gravimeters could resolve depletion areas in steam assisted gravity drainage (SAGD) reservoirs. This is achieved through modelling of a SAGD reservoir at 1.25 and 5 years of operation. Specifically, the density change structure identified from geological, petrological, and seismic observations is forward modelled for gravity and gradients. Three main parameters have an impact on the resolvability of bitumen depletion volumes and are varied through a suitable parameter space: well pair separation, depth to the well pairs, and survey grid sampling. The results include a resolvability matrix, which identifies reservoirs that could benefit from time-lapse gravity gradiometry monitoring. After 1.25 years of operation, during the rising phase, the resolvable maximum reservoir depth ranges between the surface and 230 m, considering a well pair separation between 80 and 200 m. After 5 years of production, during the spreading phase, the resolvability of depletion volumes around single well pairs is greatly compromised as the depletion volume is closer to the surface, which translates to a larger portion of the gravity signal. The modelled resolvability matrices were derived from visual inspection and spectral analysis of the gravity gradient signatures and can be used to assess the applicability of time-lapse gradiometry to monitor reservoir density changes.

  4. Two-photon microscopy using fiber-based nanosecond excitation.

    Science.gov (United States)

    Karpf, Sebastian; Eibl, Matthias; Sauer, Benjamin; Reinholz, Fred; Hüttmann, Gereon; Huber, Robert

    2016-07-01

    Two-photon excitation fluorescence (TPEF) microscopy is a powerful technique for sensitive tissue imaging at depths of up to 1000 micrometers. However, due to the shallow penetration, for in vivo imaging of internal organs in patients beam delivery by an endoscope is crucial. Until today, this is hindered by linear and non-linear pulse broadening of the femtosecond pulses in the optical fibers of the endoscopes. Here we present an endoscope-ready, fiber-based TPEF microscope, using nanosecond pulses at low repetition rates instead of femtosecond pulses. These nanosecond pulses lack most of the problems connected with femtosecond pulses but are equally suited for TPEF imaging. We derive and demonstrate that at given cw-power the TPEF signal only depends on the duty cycle of the laser source. Due to the higher pulse energy at the same peak power we can also demonstrate single shot two-photon fluorescence lifetime measurements.

  5. Two-photon STED spectral determination for a new V-shaped organic fluorescent probe with efficient two-photon absorption.

    Science.gov (United States)

    Belfield, Kevin D; Bondar, Mykhailo V; Morales, Alma R; Padilha, Lazaro A; Przhonska, Olga V; Wang, Xuhua

    2011-10-24

    Two-photon stimulated emission depletion (STED) cross sections were determined over a broad spectral range for a novel two-photon absorbing organic molecule, representing the first such report. The synthesis, comprehensive linear photophysical, two-photon absorption (2PA), and stimulated emission properties of a new fluorene-based compound, (E)-2-{3-[2-(7-(diphenylamino)-9,9-diethyl-9H-fluoren-2-yl)vinyl]-5-methyl-4-oxocyclohexa-2,5-dienylidene} malononitrile (1), are presented. Linear spectral parameters, including excitation anisotropy and fluorescence lifetimes, were obtained over a broad range of organic solvents at room temperature. The degenerate two-photon absorption (2PA) spectrum of 1 was determined with a combination of the direct open-aperture Z-scan and relative two-photon-induced fluorescence methods using 1 kHz femtosecond excitation. The maximum value of the 2PA cross section ~1700 GM was observed in the main, long wavelength, one-photon absorption band. One- and two-photon stimulated emission spectra of 1 were obtained over a broad spectral range using a femtosecond pump-probe technique, resulting in relatively high two-photon stimulated emission depletion cross sections (~1200 GM). A potential application of 1 in bioimaging was demonstrated through one- and two-photon fluorescence microscopy images of HCT 116 cells incubated with micelle-encapsulated dye.

  6. MONITORING EROSION OF STONE SURFACES USING TIME-LAPSE AND PTM PHOTOGRAPHY: FIELD STUDY OF A 14TH CENTURY MONASTERY IN YORKSHIRE

    Science.gov (United States)

    Doehne, E.; Pinchin, S.

    2009-12-01

    Evaluating stone weathering rates and their relationship to environmental fluctuations is an important challenge in understanding the critical zone and also in efforts to prevent the loss of important cultural heritage in stone, such as monuments, sculpture and archaeological sites. Repeat photography has been widely used to evaluate geological processes such as the retreat of glaciers and the weathering of stone surfaces. However, a fundamental difficulty is that the images are often shot under differing lighting conditions, making the interpretation of stone surface loss particularly challenging. Two developments in photographic documentation show promise for improving the situation. One is the use of digital time-lapse methods to provide more frequent images to correlate stone surface loss with ongoing environmental changes. The other is a relatively new method known as polynomial transform mapping (PTM), which integrates multiple photographs taken at different angles to document more comprehensively the texture of stone surfaces. Using Java-based software, the viewer can control the precise angle of the light source in an interpolated, high-quality image. PTM can produce raking light images from any angle, as well as images with ‘normal’ illumination. We present here results based on several years of macro-photography, time-lapse imaging, and PTM imaging of rapidly eroding stone surfaces at the site of Howden Minster in Yorkshire, UK, which suffers from salt weathering. The images show that surface loss is episodic rather than continuous and in some cases is related to unusual environmental conditions, such as high winds and condensation events. Damage was also found to be synchronous, with surface change (flaking, granular disintegration, and loss of flakes) occurring at the same time in different stone blocks. Crystallization pressure from phase transitions in magnesium sulfate salts appears to be the main cause of the loss of stone surfaces.

  7. Robust joint full-waveform inversion of time-lapse seismic data sets with total-variation regularization

    CERN Document Server

    Maharramov, Musa

    2014-01-01

    We present a technique for reconstructing subsurface velocity model changes from time-lapse seismic survey data using full-waveform inversion (FWI). The technique is based on simultaneously inverting multiple survey vintages, with model difference regularization using the total variation (TV) seminorm. We compare the new TV-regularized time-lapse FWI with the $L_2$-regularized joint inversion proposed in our earlier work, using synthetic data sets that exhibit survey repeatability issues. The results demonstrate clear advantages of the proposed TV-regularized joint inversion over alternatives methods for recovering production-induced model changes that are due to both fluid substitution and geomechanical effects.

  8. A new approach to dual-color two-photon microscopy with fluorescent proteins

    Directory of Open Access Journals (Sweden)

    Rebane Aleks

    2010-02-01

    Full Text Available Abstract Background Two-photon dual-color imaging of tissues and cells labeled with fluorescent proteins (FPs is challenging because most two-photon microscopes only provide one laser excitation wavelength at a time. At present, methods for two-photon dual-color imaging are limited due to the requirement of large differences in Stokes shifts between the FPs used and their low two-photon absorption (2PA efficiency. Results Here we present a new method of dual-color two-photon microscopy that uses the simultaneous excitation of the lowest-energy electronic transition of a blue fluorescent protein and a higher-energy electronic transition of a red fluorescent protein. Conclusion Our method does not require large differences in Stokes shifts and can be extended to a variety of FP pairs with larger 2PA efficiency and more optimal imaging properties.

  9. Medical prototyping using two photon polymerization

    Directory of Open Access Journals (Sweden)

    Roger J Narayan

    2010-12-01

    Full Text Available Two photon polymerization involves nearly simultaneous absorption of ultrashort laser pulses for selective curing of photosensitive material. This process has recently been used to create small-scale medical devices out of several classes of photosensitive materials, such as acrylate-based polymers, organically-modified ceramic materials, zirconium sol-gels, and titanium-containing hybrid materials. In this review, the use of two photon polymerization for fabrication of several types of small-scale medical devices, including microneedles, artificial tissues, microfluidic devices, pumps, sensors, and valves, from computer models is described. Necessary steps in the development of two photon polymerization as a commercially viable medical device manufacturing method are also considered.

  10. Two Photon Couplings of Hybrid Mesons

    CERN Document Server

    Page, P R

    1996-01-01

    A new formalism is developed for the two photon production of hybrid mesons via intermediate hadronic decays. In an adiabatic and non--relativistic context with spin 1 pair creation we obtain the first absolute estimates of unmixed hybrid production strengths to be small (0.03 - 3 eV) in relation to experimental meson widths (0.1 - 5 keV). Within this context, two photon collisions therefore strongly discriminate between hybrid and conventional meson wave function components at BaBar, Cleo II, LEP2 and LHC, filtering out non--gluonic components. Decay widths of unmixed hybrids are tiny. The formalism also induces conventional meson two photon widths roughly in agreement with experiment.

  11. Two-photon induced photoluminescence and singlet oxygen generation from aggregated gold nanoparticles.

    Science.gov (United States)

    Jiang, Cuifeng; Zhao, Tingting; Yuan, Peiyan; Gao, Nengyue; Pan, Yanlin; Guan, Zhenping; Zhou, Na; Xu, Qing-Hua

    2013-06-12

    Metal nanoparticles have potential applications as bioimaging and photosensitizing agents. Aggregation effects are generally believed to be adverse to their biomedical applications. Here we have studied the aggregation effects on two-photon induced photoluminescence and singlet oxygen generation of Au nanospheres and Au nanorods of two different aspect ratios. Aggregated Au nanospheres and short Au nanorods were found to display enhanced two-photon induced photoluminescence and singlet oxygen generation capabilities compared to the unaggregated ones. The two-photon photoluminescence of Au nanospheres and short Au nanorods were enhanced by up to 15.0- and 2.0-fold upon aggregation, and the corresponding two-photon induced singlet oxygen generation capabilities were enhanced by 8.3 and 1.8-fold, respectively. The two-photon induced photoluminescence and singlet oxygen generation of the aggregated long Au nanorods were found to be lower than the unaggregated ones. These results support that the change in their two-photon induced photoluminescence and singlet oxygen generation originate from aggregation modulated two-photon excitation efficiency. This finding is expected to foster more biomedical applications of metal nanoparticles as Au nanoparticles normally exist in an aggregated form in the biological environments. Considering their excellent biocompatibility, high inertness, ready conjugation, and easy preparation, Au nanoparticles are expected to find more applications in two-photon imaging and two-photon photodynamic therapy.

  12. Time-lapse electrical resistivity tomography: a powerful tool for landslide monitoring?

    Science.gov (United States)

    Perrone, A.

    2011-12-01

    The extreme rainfall events and the quick snowmelt occurrences play an important role in the triggering of the landslides. The occurrence of one of these factors can determine the variation of water content in the first layers of the subsoil and as a consequence a quick soil saturation inducing both an increase in pore-water pressures and the overloaded of the slopes progressively collapsing. The electrical resistivity, self-potential, electromagnetic induction and GPR methods can be considered as the most appropriate for assessing the presence of water in the underground. Such methods allow us to study the behavior of water content over much wider and deeper areas than those offered by traditional methods (thermo-gravimetric, tensiometric, TDR, etc) based on spot measures and concerning small volumes. In particular, the Electrical Resistivity Tomography (ERT), which has already proved to be a powerful tool both for the geometrical reconstruction of a landslide body (location of sliding surface, estimation of the thickness of the slide material) and the individuation of high water content areas, can be considered as an alternative tool to be employed for a qualitative and quantitative water content monitoring in the first layers of the subsoil. Indeed, time-lapse 2D ERT can be tested in order to gather information on the temporal and spatial patterns of water infiltration processes and water content variation. This work reports the preliminary results from a new prototype system planned to obtain time-lapse 2D ERTs, TDR and precipitation measurements in two landslide areas located in the Southern Apennine chain (Italy). The system was planned with the aim to estimate the variation of the resistivity parameter on a long period considering the water content variation, the rain water infiltration and the seasonal changes. The prototype system, linked to a pc used for storing data and managing the time interval acquisition, consists of: a resistivimeter connected to a

  13. Geoelectrical time-lapse analysis for improved interpretation of data in a contaminated area

    Science.gov (United States)

    Chitea, Florina; Serban, Adrian; Ioane, Dumitru; Georgescu, Paul

    2014-05-01

    Non invasive geoelectrical studies are useful in the preliminary assessment of areas suspected to be contaminated but also in the investigation stage. Correctly adapted to the site specific situation, they are used to detect and investigate buried sources of pollution, to characterize the geology of the area, to detect the contaminated plume or to study the attenuation of pollution in case the appliance of an site-specific remediation techniques. Despite the improved technological acquisition techniques and the optimized inversion data algorithms, interpretation of geoelectrical data in still a challenging task, especially in a contaminated hydrogeological context. Beside the soil physical properties (composition, porosity, texture, etc.), moisture content and chemical composition of the pollutant are also influencing the measured parameter. Apparent electrical resistivity method was use in an area located near an Oil Refinery. Electrical measurements performed on profiles (transverse and along the direction of water flow -according to hydrological data) revealed the presence of contaminants by means of high resistivity anomalies. Using the same acquisition technique (Schlumberger array, same VES points, injection - AB - and voltage - MN - lines extension), measurements were repeated during time, along the same profiles. On the resulted electrical sections from 2006 to 2013, a dynamic situation regarding the pollution plume was observed. Time - lapse analysis, based on the calculation of resistivity differences between sets of data acquired along the same profile was applied, and data interpretation was made using the resulted sections. Significant variation between data sets (> 17% of apparent resistivity normalized differences) observed along the main profile were mainly ranging from the near surface (1.5 m) to an approximated depth (AB/2) of 10m. Using the time-lapse method, changes in the lateral and in depth extension of polluted areas could be observed and

  14. Fast history matching of time-lapse seismic and production data for high resolution models

    Science.gov (United States)

    Jimenez Arismendi, Eduardo Antonio

    Integrated reservoir modeling has become an important part of day-to-day decision analysis in oil and gas management practices. A very attractive and promising technology is the use of time-lapse or 4D seismic as an essential component in subsurface modeling. Today, 4D seismic is enabling oil companies to optimize production and increase recovery through monitoring fluid movements throughout the reservoir. 4D seismic advances are also being driven by an increased need by the petroleum engineering community to become more quantitative and accurate in our ability to monitor reservoir processes. Qualitative interpretations of time-lapse anomalies are being replaced by quantitative inversions of 4D seismic data to produce accurate maps of fluid saturations, pore pressure, temperature, among others. Within all steps involved in this subsurface modeling process, the most demanding one is integrating the geologic model with dynamic field data, including 4Dseismic when available. The validation of the geologic model with observed dynamic data is accomplished through a "history matching" (HM) process typically carried out with well-based measurements. Due to low resolution of production data, the validation process is severely limited in its reservoir areal coverage, compromising the quality of the model and any subsequent predictive exercise. This research will aim to provide a novel history matching approach that can use information from high-resolution seismic data to supplement the areally sparse production data. The proposed approach will utilize streamline-derived sensitivities as means of relating the forward model performance with the prior geologic model. The essential ideas underlying this approach are similar to those used for high-frequency approximations in seismic wave propagation. In both cases, this leads to solutions that are defined along "streamlines" (fluid flow), or "rays" (seismic wave propagation). Synthetic and field data examples will be used

  15. Generalized focusing of time-lapse changes with applications to direct current and time-domain induced polarization inversions

    Science.gov (United States)

    Fiandaca, Gianluca; Doetsch, Joseph; Vignoli, Giulio; Auken, Esben

    2015-11-01

    Often in geophysical monitoring experiments time-lapse inversion models vary too smoothly with time, owing to the strong imprint of regularization. Several methods have been proposed for focusing the spatiotemporal changes of the model parameters. In this study, we present two generalizations of the minimum support norm, which favour compact time-lapse changes and can be adapted to the specific problem requirements. Inversion results from synthetic direct current resistivity models that mimic developing plumes show that the focusing scheme significantly improves size, shape and magnitude estimates of the time-lapse changes. Inversions of the synthetic data also illustrate that the focused inversion gives robust results and that the focusing settings are easily chosen. Inversions of full-decay time-domain induced polarization (IP) field data from a CO2 monitoring injection experiment show that the focusing scheme performs well for field data and inversions for all four Cole-Cole polarization parameters. Our tests show that the generalized minimum support norms react in an intuitive and predictable way to the norm settings, implying that they can be used in time-lapse experiments for obtaining reliable and robust results.

  16. Bayesian Markov chain Monte Carlo Inversion of Time-Lapse Geophysical Data To Characterize the Vadose Zone

    DEFF Research Database (Denmark)

    Scholer, Marie; Irving, James; Zibar, Majken Caroline Looms;

    Geophysical methods have the potential to provide valuable information on hydrological properties in the unsaturated zone. In particular, time-lapse geophysical data, when coupled with a hydrological model and inverted stochastically, may allow for the effective estimation of subsurface hydraulic...

  17. Joint inversion of time-lapse VSP data for monitoring CO2 injection at the Farnsworth EOR field in Texas

    Science.gov (United States)

    Zhang, M.; Gao, K.; Balch, R. S.; Huang, L.

    2016-12-01

    During the Development Phase (Phase III) of the U.S. Southwest Regional Partnership on Carbon Sequestration (SWP), time-lapse 3D vertical seismic profiling (VSP) data were acquired to monitor CO2 injection/migration at the Farnsworth Enhanced Oil Recovery (EOR) field, in partnership with the industrial partner Chaparral Energy. The project is to inject a million tons of carbon dioxide into the target formation, the deep oil-bearing Morrow Formation in the Farnsworth Unit EOR field. Quantitative time-lapse seismic monitoring has the potential to track CO2 movement in geologic carbon storage sites. Los Alamos National Laboratory (LANL) has recently developed new full-waveform inversion methods to jointly invert time-lapse seismic data for changes in elastic and anisotropic parameters in target monitoring regions such as a CO2 reservoir. We apply our new joint inversion methods to time-lapse VSP data acquired at the Farnsworth EOR filed, and present some preliminary results showing geophysical properties changes in the reservoir.

  18. Mass conservative three-dimensional water tracer distribution from MCMC inversion of time-lapse GPR data

    NARCIS (Netherlands)

    Lalov, E.; Linde, N.; Vrugt, J.A.

    2012-01-01

    Time-lapse geophysical measurements are widely used to monitor the movement of water and solutes through the subsurface. Yet commonly used deterministic least squares inversions typically suffer from relatively poor mass recovery, spread overestimation, and limited ability to appropriately estimate

  19. PFLOTRAN-E4D: A parallel open source PFLOTRAN module for simulating time-lapse electrical resistivity data

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, Timothy C.; Hammond, Glenn E.; Chen, Xingyuan

    2017-02-01

    Time-lapse electrical resistivity tomography (ERT) is finding increased application for remotely monitoring processes occurring in the near subsurface in three-dimensions (i.e. 4D monitoring). However, there are few codes capable of simulating the evolution of subsurface resistivity and corresponding tomographic measurements arising from a particular process, particularly in parallel and with an open source license. Herein we describe and demonstrate an electrical resistivity tomography module for the PFLOTRAN subsurface simulation code, named PFLOTRAN-E4D. The PFLOTRAN-E4D module operates in parallel using a dedicated set of compute cores in a master-slave configuration. At each time step, the master processes receives subsurface states from PFLOTRAN, converts those states to bulk electrical conductivity, and instructs the slave processes to simulate a tomographic data set. The resulting multi-physics simulation capability enables accurate feasibility studies for ERT imaging, the identification of the ERT signatures that are unique to a given process, and facilitates the joint inversion of ERT data with hydrogeological data for subsurface characterization. PFLOTRAN-E4D is demonstrated herein using a field study of stage-driven groundwater/river water interaction ERT monitoring along the Columbia River, Washington, USA. Results demonstrate the complex nature of changes subsurface electrical conductivity, in both the saturated and unsaturated zones, arising from water table changes and from river water intrusion into the aquifer. The results also demonstrate the sensitivity of surface based ERT measurements to those changes over time. PFLOTRAN-E4D is available with the PFLOTRAN development version with an open-source license at https://bitbucket.org/pflotran/pflotran-dev .

  20. PFLOTRAN-E4D: A parallel open source PFLOTRAN module for simulating time-lapse electrical resistivity data

    Science.gov (United States)

    Johnson, Timothy C.; Hammond, Glenn E.; Chen, Xingyuan

    2017-02-01

    Time-lapse electrical resistivity tomography (ERT) is finding increased application for remotely monitoring processes occurring in the near subsurface in three-dimensions (i.e. 4D monitoring). However, there are few codes capable of simulating the evolution of subsurface resistivity and corresponding tomographic measurements arising from a particular process, particularly in parallel and with an open source license. Herein we describe and demonstrate an electrical resistivity tomography module for the PFLOTRAN subsurface flow and reactive transport simulation code, named PFLOTRAN-E4D. The PFLOTRAN-E4D module operates in parallel using a dedicated set of compute cores in a master-slave configuration. At each time step, the master processes receives subsurface states from PFLOTRAN, converts those states to bulk electrical conductivity, and instructs the slave processes to simulate a tomographic data set. The resulting multi-physics simulation capability enables accurate feasibility studies for ERT imaging, the identification of the ERT signatures that are unique to a given process, and facilitates the joint inversion of ERT data with hydrogeological data for subsurface characterization. PFLOTRAN-E4D is demonstrated herein using a field study of stage-driven groundwater/river water interaction ERT monitoring along the Columbia River, Washington, USA. Results demonstrate the complex nature of subsurface electrical conductivity changes, in both the saturated and unsaturated zones, arising from river stage fluctuations and associated river water intrusion into the aquifer. The results also demonstrate the sensitivity of surface based ERT measurements to those changes over time. PFLOTRAN-E4D is available with the PFLOTRAN development version with an open-source license at https://bitbucket.org/pflotran/pflotran-dev.

  1. Two-photon physics at LEP2

    Energy Technology Data Exchange (ETDEWEB)

    Cartwright, Susan; Lehto, Mark [University of Sheffield Department of Physics, Sheffield S3 7RH (United Kingdom); Seymour, Michael H.; Close, Frank; Wright, Alison [Rutherford Appleton Laboratory, Chilton, Didcot, Oxfordshire OX11 0QX (United Kingdom); Affholderbach, Klaus; Cowan, Glen [Universitaet Siegen, Fachbereich Physik, D-57068 Siegen (Germany); Finch, Alex [University of Lancaster, Lancaster LA1 4YB (United Kingdom); Lauber, Jan [University College London, Gower Street, London WC1E 6BT (United Kingdom)

    1998-02-01

    The working group on two-photon physics concentrated on three main subtopics: modelling the hadronic final state of deep inelastic scattering on a photon; unfolding the deep inelastic scattering data to obtain the photon structure function; and resonant production of exclusive final states, particularly of glueball candidates. In all three areas, new results were presented. (author)

  2. Mathematical analysis of endothelial sibling pair cell-cell interactions using time-lapse cinematography data.

    Science.gov (United States)

    Brown, L M; Ryan, U S; Absher, M; Olazabal, B M

    1982-01-01

    The sibling pairs from two different endothelial cell cultures were analysed by time-lapse cinematography. It was shown that wounded and regular (low density seeded) cultures differed in the behaviour patterns of their siblings. The cultures differed most significantly in the minimum interdivision time (IDT) which was 27% lower for the wounded culture. In the wounded culture there was a greater correlation of IDT values between sibling pairs. IDT values recorded both for paired and for unpaired cells were shorter for the wounded than for the regular culture. The mean IDT for unpaired cells was longer than the mean IDT for paired cells in the regular culture. Thus paired cells in the regular culture, had shorter IDTs, but not as short as in the wounded culture. It was significant that in the wounded culture the first generation of siblings were very close (less than 150 microns apart) at division. Overall the behaviour differences between the two cultures resulted in a higher rate of increase in cell numbers, and thus faster repair, of the wounded monolayer.

  3. Time-Lapse Dynamics of the Mouse Oocyte Chromatin Organisation during Meiotic Resumption

    Directory of Open Access Journals (Sweden)

    Martina Belli

    2014-01-01

    Full Text Available In the mammalian oocyte, distinct patterns of centromeres and pericentromeric heterochromatin localisation correlate with the gamete’s developmental competence. Mouse antral oocytes display two main types of chromatin organisation: SN oocytes, with a ring of Hoechst-positive chromatin surrounding the nucleolus, and NSN oocytes lacking this ring. When matured to MII and fertilised, only SN oocytes develop beyond the 2-cell, and reach full term. To give detailed information on the dynamics of the SN or NSN chromatin during meiosis resumption, we performed a 9 hr time-lapse observation. The main significant differences recorded are: (1 reduction of the nuclear area only in SN oocytes; (2 ~17 min delay of GVBD in NSN oocytes; (3 chromatin condensation, after GVBD, in SN oocytes; (4 formation of 4-5 CHCs in SN oocytes; (5 increase of the perivitelline space, ~57 min later in NSN oocytes; (6 formation of a rosette-like disposition of CHCs, ~84 min later in SN oocytes; (7 appearance of the MI plate ~40 min later in NSN oocytes. Overall, we described a pathway of transition from the GV to the MII stage that is punctuated of discrete recordable events showing their specificity and occurring with different time kinetics in the two types of oocytes.

  4. Landslide Monitoring in Southwestern China via Time-lapse Electrical Resistivity Tomography

    Science.gov (United States)

    Xu, D.; Hu, X.; Shan, C.

    2016-12-01

    The dynamic monitoring of landslides in engineering geology has focused on the correlation among landslide stability, rainwater infiltration, and subsurface hydrogeology. However, the understanding of this complicated correlation is still poor and inadequate. Thus, in this study, we investigated a typical landslide in southwestern China via time-lapse electrical resistivity tomography (TLERT) in November 2013, August 2014 and May 2016. From the data, shallow sediments showed short-term resistivity variability due to evaporation and rainfall, whereas deep zone exhibited seasonal fluctuations related to dry season, rainy season and snow melting during springtime. We also studied landslide mechanisms based on the spatiotemporal characteristics of surface water infiltration and flow within the landslide body. Combined with borehole data, inverted resistivity models accurately defined the interface between Quaternary sediments and bedrock. Preferential flow pathways attributed to fracture zones and fissures were also delineated. In addition, we found that surface water permeates through these pathways into the slipping mass and drains away as fissure water in the fractured bedrock, probably causing the weakly weathered layer to gradually soften and erode, eventually leading to a landslide. Clearly, TLERT dynamic monitoring can provide precursory information of critical sliding and can be used in landslide stability analysis and prediction.

  5. Time lapse investigation of antibiotic susceptibility using a microfluidic linear gradient 3D culture device.

    Science.gov (United States)

    Hou, Zining; An, Yu; Hjort, Karin; Hjort, Klas; Sandegren, Linus; Wu, Zhigang

    2014-09-01

    This study reports a novel approach to quantitatively investigate the antibacterial effect of antibiotics on bacteria using a three-dimensional microfluidic culture device. In particular, our approach is suitable for studying the pharmacodynamics effects of antibiotics on bacterial cells temporally and with a continuous range of concentrations in a single experiment. The responses of bacterial cells to a linear concentration gradient of antibiotics were observed using time-lapse photography, by encapsulating bacterial cells in an agarose-based gel located in a commercially available microfluidics chamber. This approach generates dynamic information with high resolution, in a single operation, e.g., growth curves and antibiotic pharmacodynamics, in a well-controlled environment. No pre-labelling of the cells is needed and therefore any bacterial sample can be tested in this setup. It also provides static information comparable to that of standard techniques for measuring minimum inhibitory concentration (MIC). Five antibiotics with different mechanisms were analysed against wild-type Escherichia coli, Staphylococcus aureus and Salmonella Typhimurium. The entire process, including data analysis, took 2.5-4 h and from the same analysis, high-resolution growth curves were obtained. As a proof of principle, a pharmacodynamic model of streptomycin against Salmonella Typhimurium was built based on the maximal effect model, which agreed well with the experimental results. Our approach has the potential to be a simple and flexible solution to study responding behaviours of microbial cells under different selection pressures both temporally and in a range of concentrations.

  6. Thermal erosion of a permafrost coastline: Improving process-based models using time-lapse photography

    Science.gov (United States)

    Wobus, C.; Anderson, R.; Overeem, I.; Matell, N.; Clow, G.; Urban, F.

    2011-01-01

    Coastal erosion rates locally exceeding 30 m y-1 have been documented along Alaska's Beaufort Sea coastline, and a number of studies suggest that these erosion rates have accelerated as a result of climate change. However, a lack of direct observational evidence has limited our progress in quantifying the specific processes that connect climate change to coastal erosion rates in the Arctic. In particular, while longer ice-free periods are likely to lead to both warmer surface waters and longer fetch, the relative roles of thermal and mechanical (wave) erosion in driving coastal retreat have not been comprehensively quantified. We focus on a permafrost coastline in the northern National Petroleum Reserve-Alaska (NPR-A), where coastal erosion rates have averaged 10-15 m y-1 over two years of direct monitoring. We take advantage of these extraordinary rates of coastal erosion to observe and quantify coastal erosion directly via time-lapse photography in combination with meteorological observations. Our observations indicate that the erosion of these bluffs is largely thermally driven, but that surface winds play a crucial role in exposing the frozen bluffs to the radiatively warmed seawater that drives melting of interstitial ice. To first order, erosion in this setting can be modeled using formulations developed to describe iceberg deterioration in the open ocean. These simple models provide a conceptual framework for evaluating how climate-induced changes in thermal and wave energy might influence future erosion rates in this setting.

  7. Time-Lapse Dynamics of the Mouse Oocyte Chromatin Organisation during Meiotic Resumption

    Science.gov (United States)

    Redi, Carlo Alberto; Zuccotti, Maurizio

    2014-01-01

    In the mammalian oocyte, distinct patterns of centromeres and pericentromeric heterochromatin localisation correlate with the gamete's developmental competence. Mouse antral oocytes display two main types of chromatin organisation: SN oocytes, with a ring of Hoechst-positive chromatin surrounding the nucleolus, and NSN oocytes lacking this ring. When matured to MII and fertilised, only SN oocytes develop beyond the 2-cell, and reach full term. To give detailed information on the dynamics of the SN or NSN chromatin during meiosis resumption, we performed a 9 hr time-lapse observation. The main significant differences recorded are: (1) reduction of the nuclear area only in SN oocytes; (2) ~17 min delay of GVBD in NSN oocytes; (3) chromatin condensation, after GVBD, in SN oocytes; (4) formation of 4-5 CHCs in SN oocytes; (5) increase of the perivitelline space, ~57 min later in NSN oocytes; (6) formation of a rosette-like disposition of CHCs, ~84 min later in SN oocytes; (7) appearance of the MI plate ~40 min later in NSN oocytes. Overall, we described a pathway of transition from the GV to the MII stage that is punctuated of discrete recordable events showing their specificity and occurring with different time kinetics in the two types of oocytes. PMID:24864231

  8. Time-lapse analysis of potential cellular responsiveness to Johrei, a Japanese healing technique

    Directory of Open Access Journals (Sweden)

    Moore Dan

    2005-01-01

    Full Text Available Abstract Background Johrei is an alternative healing practice which involves the channeling of a purported universal healing energy to influence the health of another person. Despite little evidence to support the efficacy of such practices the use of such treatments is on the rise. Methods We assessed cultured human cancer cells for potential responsiveness to Johrei treatment from a short distance. Johrei treatment was delivered by practitioners who participated in teams of two, alternating every half hour for a total of four hours of treatment. The practitioners followed a defined set of mental procedures to minimize variability in mental states between experiments. An environmental chamber maintained optimal growth conditions for cells throughout the experiments. Computerized time-lapse microscopy allowed documentation of cancer cell proliferation and cell death before, during and after Johrei treatments. Results Comparing eight control experiments with eight Johrei intervention experiments, we found no evidence of a reproducible cellular response to Johrei treatment. Conclusion Cell death and proliferation rates of cultured human cancer cells do not appear responsive to Johrei treatment from a short distance.

  9. ENVIRONMENTAL MONITORING OF LEAKS USING TIME LAPSED LONG ELECTRODE ELECTRICAL RESISTIVITY

    Energy Technology Data Exchange (ETDEWEB)

    RUCKER DF; FINK JB; LOKE MH; MYERS DA

    2009-11-05

    Highly industrialized areas pose significant challenges for surface based electrical resistivity characterization and monitoring due to the high degree of metallic infrastructure. The infrastructure is typically several orders of magnitude more conductive than the desired targets, preventing the geophysicist from obtaining a clear picture of the subsurface. These challenges may be minimized if steel-cased wells are used as long electrodes. We demonstrate a method of using long electrodes in a complex nuclear waste facility to monitor a simulated leak from an underground storage tank. The leak was simulated by injecting high conductivity fluid in a perforated well and the resistivity measurements were made before and after the leak test. The data were processed in four dimensions, where a regularization procedure was applied in both the time and space domains. The results showed a lowered resistivity feature develop south of the injection site. The time lapsed regularization parameter had a strong influence on the differences in inverted resistivity between the pre and post datasets, potentially making calibration of the results to specific hydrogeologic parameters difficult.

  10. Landslide monitoring in southwestern China via time-lapse electrical resistivity tomography

    Institute of Scientific and Technical Information of China (English)

    Xu Dong; Hu Xiang-Yun; Shan Chun-Ling; Li Rui-Heng

    2016-01-01

    The dynamic monitoring of landslides in engineering geology has focused on the correlation among landslide stability, rainwater infiltration, and subsurface hydrogeology. However, the understanding of this complicated correlation is still poor and inadequate. Thus, in this study, we investigated a typical landslide in southwestern China via time-lapse electrical resistivity tomography (TLERT) in November 2013 and August 2014. We studied landslide mechanisms based on the spatiotemporal characteristics of surface water infi ltration andfl ow within the landslide body. Combined with borehole data, inverted resistivity models accurately defined the interface between Quaternary sediments and bedrock. Preferential fl ow pathways attributed to fracture zones andfi ssures were also delineated. In addition, we found that surface water permeates through these pathways into the slipping mass and drains away asfi ssure water in the fractured bedrock, probably causing the weakly weathered layer to gradually soften and erode, eventually leading to a landslide. Clearly, TLERT dynamic monitoring can provide precursory information of critical sliding and can be used in landslide stability analysis and prediction.

  11. Time lapse 3D geoelectric measurements for monitoring of in-situ remediation

    Science.gov (United States)

    Tildy, Péter; Neducza, Boriszláv; Nagy, Péter; Kanli, Ali Ismet; Hegymegi, Csaba

    2017-01-01

    In the last decade, different kinds of in-situ methods have been increasingly used for hydrocarbon contamination remediation due to their effectiveness. One of these techniques operates by injection of chemical oxidant solution to remove (degrade) the subsurface contaminants. Our aim was to develop a surface (non-destructive) measurement strategy to monitor oxidative in-situ remediation processes. The difficulties of the presented study originate from the small volume of conductive solution that can be used due to environmental considerations, the effect of conductive groundwater and the high clay content of the targeted layer. Therefore a site specific synthetic modelling was necessary for measurement design involving the results of preliminary 2D ERT measurements, electrical conductivity measurements of different active agents and expected resistivity changes calculated by soil resistivity modelling. The results of soil resistivity modelling have suggested that the reagent have complex effects on contaminated soils because of chemical biodegradation. As a result the plume of resistivity changes caused by the injected agent was determined showing strong fracturing effect because of the high pressure of injection. Based on the sophisticated tests and synthetic modelling 3D time-lapse geo-electric measurements were proven to provide a usable monitoring tool for in-situ remediation to help in-field design of such techniques.

  12. Time-lapse-Embryomonitoring und Oozytenvitrifikation: Neue Verfahren in der Reproduktionsmedizin aus ethischer Sicht

    Directory of Open Access Journals (Sweden)

    Kreß H

    2013-01-01

    Full Text Available Zurzeit zeigt sich der stetige technische Fortschritt der Reproduktionsmedizin an zwei neuen Verfahren. Es geht um die Möglichkeit, außerkörperlich erzeugte Embryonen in neuartigen Inkubatorsystemen zu beobachten (Time-lapse-Monitoring, und um die Kryokonservierung von Eizellen (Oozytenvitrifikation. Das Einfrieren von Eizellen kann in Frage kommen, wenn eine Frau einen Kinderwunsch aus persönlichen oder sozialen Gründen auf einen späteren Zeitpunkt verschieben möchte („social egg freezing“ oder wenn sie sich aus Krankheitsgründen einer keimzellschädigenden Behandlung aussetzen lassen muss (medizinisch indizierte Vitrifikation. Der Artikel beleuchtet diese unterschiedlichen Handlungsoptionen unter ethischen Aspekten. Er wägt jeweils ihre normative Verträglichkeit, ihre Humanverträglichkeit und ihre Sozialverträglichkeit ab und hält sie – bei manchen Differenzen, die im Einzelnen bestehen – insgesamt für ethisch vertretbar. Für konkrete Entscheidungen kommt es auf den Einzelfall und auf die Einzelumstände an.

  13. A clustering approach applied to time-lapse ERT interpretation - Case study of Lascaux cave

    Science.gov (United States)

    Xu, Shan; Sirieix, Colette; Riss, Joëlle; Malaurent, Philippe

    2017-09-01

    The Lascaux cave, located in southwest France, is one of the most important prehistoric cave in the world that shows Paleolithic paintings. This study aims to characterize the structure of the weathered epikarst setting located above the cave using Time-Lapse Electrical Resistivity Tomography (ERT) combined with local hydrogeological and climatic environmental data. Twenty ERT profiles were carried out for two years and helped us to record the seasonal and spatial variations of the electrical resistivity of the hydraulic upstream area of the Lascaux cave. The 20 interpreted resistivity models were merged into a single synthetic model using a multidimensional statistical method (Hierarchical Agglomerative Clustering). The individual blocks from the synthetic model associated with a similar resistivity variability were gathered into 7 clusters. We combined the resistivity temporal variations with climatic and hydrogeological data to propose a geo-electrical model that relates to a conceptual geological model. We provide a geological interpretation for each cluster regarding epikarst features. The superficial clusters (no 1 & 2) are linked to effective rainfall and trees, probably a fractured limestone. Another two clusters (no 6 & 7) are linked to detrital formations (sand and clay respectively). The cluster 3 may correspond to a marly limestone that forms a non-permeable horizon. Finally, the electrical behavior of the last two clusters (no 4 & 5) is correlated with the variation of flow rate; they may be a privileged feed zone of the flow in the cave.

  14. Time-lapse characterization of hydrothermal seawater and microbial interactions with basaltic tephra at Surtsey Volcano

    Science.gov (United States)

    Jackson, M. D.; Gudmundsson, M. T.; Bach, W.; Cappelletti, P.; Coleman, N. J.; Ivarsson, M.; Jónasson, K.; Jørgensen, S. L.; Marteinsson, V.; McPhie, J.; Moore, J. G.; Nielson, D.; Rhodes, J. M.; Rispoli, C.; Schiffman, P.; Stefánsson, A.; Türke, A.; Vanorio, T.; Weisenberger, T. B.; White, J. D. L.; Zierenberg, R.; Zimanowski, B.

    2015-12-01

    A new International Continental Drilling Program (ICDP) project will drill through the 50-year-old edifice of Surtsey Volcano, the youngest of the Vestmannaeyjar Islands along the south coast of Iceland, to perform interdisciplinary time-lapse investigations of hydrothermal and microbial interactions with basaltic tephra. The volcano, created in 1963-1967 by submarine and subaerial basaltic eruptions, was first drilled in 1979. In October 2014, a workshop funded by the ICDP convened 24 scientists from 10 countries for 3 and a half days on Heimaey Island to develop scientific objectives, site the drill holes, and organize logistical support. Representatives of the Surtsey Research Society and Environment Agency of Iceland also participated. Scientific themes focus on further determinations of the structure and eruptive processes of the type locality of Surtseyan volcanism, descriptions of changes in fluid geochemistry and microbial colonization of the subterrestrial deposits since drilling 35 years ago, and monitoring the evolution of hydrothermal and biological processes within the tephra deposits far into the future through the installation of a Surtsey subsurface observatory. The tephra deposits provide a geologic analog for developing specialty concretes with pyroclastic rock and evaluating their long-term performance under diverse hydrothermal conditions. Abstracts of research projects are posted at http://surtsey.icdp-online.org.

  15. Time lapse seismic signal analysis for Cranfield, MS, EOR and CCS site

    Science.gov (United States)

    Ditkof, J.; Caspari, E.; Pevzner, R.; Urosevic, M.; Meckel, T. A.; Hovorka, S. D.

    2012-12-01

    The Cranfield field located in Southwest Mississippi is an EOR and CCS project which has been under continuous CO2 injection by Denbury Onshore LLC since 2008. To date, more than 3 million tons of CO2 remain in the subsurface. In 2007 and 2010, 3D seismic surveys were shot and an initial 4D seismic response was characterized showing coherent amplitude anomalies in some areas which received large amounts of CO2, but not in others. Previous work used Gassmann fluid substitution at two different wells, 31F-2 observation well and the 28-1 injection well to predict a post-injection saturation curves and acoustic impedance change through the reservoir. Since this writing, a second injection well, the 44-2 well, was added to the analysis to improve the practically unconstrained inversion. The two seismic volumes were cross-equalized with an appropriate correlation coefficient through well ties. Acoustic impedance inversions were carried out on each survey resulting with higher acoustic impedance changes than predicted by Gassmann for the 28-1 and 44-2 injection wells. The time-lapse acoustic impedance however is similar to the difference calculated from a time-delay along a horizon below the reservoir.

  16. Aberrant behavior of mouse embryo development after blastomere biopsy as observed through time-lapse cinematography.

    Science.gov (United States)

    Ugajin, Tomohisa; Terada, Yukihiro; Hasegawa, Hisataka; Velayo, Clarissa L; Nabeshima, Hiroshi; Yaegashi, Nobuo

    2010-05-15

    To analyze whether blastomere biopsy affects early embryonal growth as observed through time-lapse cinematography. Comparative prospective study between embryos in which a blastomere was removed and embryos in which a blastomere was not removed. An experimental laboratory of the university. We calculated the time between blastocele formation and the end of hatching, the time between the start and end of hatching, the number of contractions and expansions between blastocyst formation and the end of hatching, and the maximum diameter of the expanded blastocyst. In blastomere removal embryos, compaction began at the six-cell stage instead of at the eight-cell stage. We also found that hatching was delayed in these embryos as compared with matched controls. Moreover, the frequency of contraction and expansion movements after blastocyst formation was significantly higher in the blastomere removal group as compared with the control group. Finally, the maximum diameter of the expanded blastocyst just before hatching was not significantly different between both groups. These findings suggested that blastomere removal has an adverse effect on embryonic development around the time of hatching. Thus, future developments in preimplantation genetic diagnosis and screening should involve further consideration and caution in light of the influence of blastomere biopsy on embryonal growth. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  17. 4D inversion of time-lapse magnetotelluric data sets for monitoring geothermal reservoir

    Science.gov (United States)

    Nam, Myung Jin; Song, Yoonho; Jang, Hannuree; Kim, Bitnarae

    2017-06-01

    The productivity of a geothermal reservoir, which is a function of the pore-space and fluid-flow path of the reservoir, varies since the properties of the reservoir changes with geothermal reservoir production. Because the variation in the reservoir properties causes changes in electrical resistivity, time-lapse (TL) three-dimensional (3D) magnetotelluric (MT) methods can be applied to monitor the productivity variation of a geothermal reservoir thanks to not only its sensitivity to the electrical resistivity but also its deep depth of survey penetration. For an accurate interpretation of TL MT-data sets, a four-dimensional (4D) MT inversion algorithm has been developed to simultaneously invert all vintage data considering time-coupling between vintages. However, the changes in electrical resistivity of deep geothermal reservoirs are usually small generating minimum variation in TL MT responses. Maximizing the sensitivity of inversion to the changes in resistivity is critical in the success of 4D MT inversion. Thus, we further developed a focused 4D MT inversion method by considering not only the location of a reservoir but also the distribution of newly-generated fractures during the production. For the evaluation of the 4D MT algorithm, we tested our 4D inversion algorithms using synthetic TL MT-data sets.

  18. Interaction of poxvirus intracellular mature virion proteins with the TPR domain of kinesin light chain in live infected cells revealed by two-photon-induced fluorescence resonance energy transfer fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Jeshtadi, Ananya; Burgos, Pierre; Stubbs, Christopher D; Parker, Anthony W; King, Linda A; Skinner, Michael A; Botchway, Stanley W

    2010-12-01

    Using two-photon-induced fluorescence lifetime imaging microscopy, we corroborate an interaction (previously demonstrated by yeast two-hybrid domain analysis) of full-length vaccinia virus (VACV; an orthopoxvirus) A36 protein with the cellular microtubule motor protein kinesin. Quenching of enhanced green fluorescent protein (EGFP), fused to the C terminus of VACV A36, by monomeric red fluorescent protein (mDsRed), fused to the tetratricopeptide repeat (TPR) domain of kinesin, was observed in live chicken embryo fibroblasts infected with either modified vaccinia virus Ankara (MVA) or wild-type fowlpox virus (FWPV; an avipoxvirus), and the excited-state fluorescence lifetime of EGFP was reduced from 2.5 ± 0.1 ns to 2.1 ± 0.1 ns due to resonance energy transfer to mDsRed. FWPV does not encode an equivalent of intracellular enveloped virion surface protein A36, yet it is likely that this virus too must interact with kinesin to facilitate intracellular virion transport. To investigate possible interactions between innate FWPV proteins and kinesin, recombinant FWPVs expressing EGFP fused to the N termini of FWPV structural proteins Fpv140, Fpv168, Fpv191, and Fpv198 (equivalent to VACV H3, A4, p4c, and A34, respectively) were generated. EGFP fusions of intracellular mature virion (IMV) surface protein Fpv140 and type II membrane protein Fpv198 were quenched by mDsRed-TPR in recombinant FWPV-infected cells, indicating that these virion proteins are found within 10 nm of mDsRed-TPR. In contrast, and as expected, EGFP fusions of the IMV core protein Fpv168 did not show any quenching. Interestingly, the p4c-like protein Fpv191, which demonstrates late association with preassembled IMV, also did not show any quenching.

  19. Two-photon excited ultraviolet photoluminescence of zinc oxide nanorods.

    Science.gov (United States)

    Zhu, Guangping; Xu, Chunxiang; Zhu, Jing; Lu, Changgui; Cui, Yiping; Sun, Xiaowei

    2008-11-01

    High density zinc oxide nanorods with uniform size were synthesized on (100) silicon substrate by vapor-phase transport method. The scanning electron microscopy images reveal that the nanorods have an average diameter of about 400 nm. The X-ray diffraction pattern demonstrates the wurtzite crystalline structure of the ZnO nanorods growing along [0001] direction. The single-photon excited photoluminescence presents a strong ultraviolet emission band at 394 nm and a weak visible emission band at 600 nm. When the ZnO nanorods were respectively pumped by various wavelength lasers from 520 nm to 700 nm, two-photon excited ultraviolet photoluminescence was observed. The dependence of the two-photon excited photoluminescence intensity on the excitation wavelength and power was investigated in detail.

  20. High-resolution three-dimensional imaging of a depleted CMOS sensor using an edge Transient Current Technique based on the Two Photon Absorption process (TPA-eTCT)

    CERN Document Server

    García, Marcos Fernández; Echeverría, Richard Jaramillo; Moll, Michael; Santos, Raúl Montero; Moya, David; Pinto, Rogelio Palomo; Vila, Iván

    2016-01-01

    For the first time, the deep n-well (DNW) depletion space of a High Voltage CMOS sensor has been characterized using a Transient Current Technique based on the simultaneous absorption of two photons. This novel approach has allowed to resolve the DNW implant boundaries and therefore to accurately determine the real depleted volume and the effective doping concentration of the substrate. The unprecedented spatial resolution of this new method comes from the fact that measurable free carrier generation in two photon mode only occurs in a micrometric scale voxel around the focus of the beam. Real three-dimensional spatial resolution is achieved by scanning the beam focus within the sample.

  1. Time-lapse monitoring of soil water content using electromagnetic conductivity imaging

    Science.gov (United States)

    The volumetric soil water content (VWC) is fundamental to agriculture. Unfortunately, the universally accepted thermogravimetric method is labour intensive and time-consuming to use for field-scale monitoring. Electromagnetic (EM) induction instruments have proven to be useful in mapping the spatio-...

  2. Live Cell Imaging of Bacillus subtilis and Streptococcus pneumoniae using Automated Time-lapse Microscopy

    NARCIS (Netherlands)

    Jong, Imke G. de; Beilharz, Katrin; Kuipers, Oscar P.; Veening, Jan-Willem

    2011-01-01

    During the last few years scientists became increasingly aware that average data obtained from microbial population based experiments are not representative of the behavior, status or phenotype of single cells. Due to this new insight the number of single cell studies rises continuously. However, ma

  3. Transparency induced by two photon interference in a beam splitter

    Institute of Scientific and Technical Information of China (English)

    Wang Kai-Ge; Yang Guo-Jian

    2004-01-01

    We propose a special two-photon state which is completely transparent in a 50/50 beam splitter. This effect is caused by the destructive two-photon interference and shows the signature of photon entanglement. We find that the symmetry of the two-photon spectrum plays the key role for the properties of two-photon interference.

  4. ENVIRONMENTAL MONITORING OF LEAKS USING TIME LAPSED LONG ELECTRODE ELECTRICAL RESISTIVITY

    Energy Technology Data Exchange (ETDEWEB)

    MYERS DA; RUCKER DF; FINK JB; LOKE MH

    2009-12-16

    Highly industrialized areas pose challenges for surface electrical resistivity characterization due to metallic infrastructure. The infrastructure is typically more conductive than the desired targets and will mask the deeper subsurface information. These challenges may be minimized if steel-cased wells are used as long electrodes in the area near the target. We demonstrate a method of using long electrodes to electrically monitor a simulated leak from an underground storage tank with both synthetic examples and a field demonstration. The synthetic examples place a simple target of varying electrical properties beneath a very low resistivity layer. The layer is meant to replicate the effects of infrastructure. Both surface and long electrodes are tested on the synthetic domain. The leak demonstration for the field experiment is simulated by injecting a high conductivity fluid in a perforated well within the S tank farm at Hanford, and the resistivity measurements are made before and after the leak test. All data are processed in four dimensions, where a regularization procedure is applied in both the time and space domains. The synthetic test case shows that the long electrode ERM could detect relative changes in resistivity that are commensurate with the differing target properties. The surface electrodes, on the other hand, had a more difficult time matching the original target's footprint. The field results shows a lowered resistivity feature develop south of the injection site after cessation of the injections. The time lapsed regularization parameter has a strong influence on the differences in inverted resistivity between the pre and post injection datasets, but the interpretation of the target is consistent across all values of the parameter. The long electrode ERM method may provide a tool for near real-time monitoring of leaking underground storage tanks.

  5. Time Lapse Gravity and Seismic Monitoring of CO2 Injection at the West Hastings Field, Texas

    Science.gov (United States)

    Ferguson, J. F.; Richards, T.; Klopping, F.; MacQueen, J.; Hosseini, S. A.

    2015-12-01

    Time lapse or 4D gravity and seismic reflection surveys are being conducted at the West Hastings Field near Houston, Texas to monitor the progress of CO2 injection. This Department of Energy supported CO2 sequestration experiment is conducted in conjunction with a Denbury Onshore, LLC tertiary recovery project. The reservoir is at a depth of 1.8 km in the Oligocene Frio sands and has been produced since the 1930s. Goals are an accounting and mapping of the injected CO2 and to determine if migration occurs along intra-reservoir faults. An integrated interpretation of the geophysical surveys will be made together with well logs and engineering data. Gravity monitoring of water versus gas replacement has been very successful, but liquid phase CO2 monitoring is problematic due to the smaller density contrast with respect to oil and water. This reservoir has a small volume to depth ratio and hence only a small gravity difference signal is expected on the surface. New borehole gravity technology introduced by Micro-g-Lacoste can make gravity measurements at near reservoir depths with a much higher signal to noise ratio. This method has been successfully evaluated on a simulation of the Hastings project. Field operations have been conducted for repeated surface and borehole gravity surveys beginning in 2013. The surface survey of 95 stations covers an area of 3 by 5 km and 22 borehole gravity logs are run in the interval above the Frio formation. 4D seismic reflection surveys are being made at 6 month intervals on the surface and in 3 VSP wells. CO2 injection into the targeted portion of the reservoir only began in early 2015 and monitoring will continue into 2017. To date only the baseline reservoir conditions have been assessed. The overall success of the gravity monitoring will not be determined until 2017.

  6. Two-photon cooling of magnesium atoms

    DEFF Research Database (Denmark)

    Malossi, N.; Damkjær, S.; Hansen, P. L.

    2005-01-01

    A two-photon mechanism for cooling atoms below the Doppler temperature is analyzed. We consider the magnesium ladder system (3s2)S01¿(3s3p)P11 at 285.2nm followed by the (3s3p)P11¿(3s3d)D21 transition at 880.7nm . For the ladder system quantum coherence effects may become important. Combined...... with the basic two-level Doppler cooling process this allows for reduction of the atomic sample temperature by more than a factor of 10 over a broad frequency range. First experimental evidence for the two-photon cooling process is presented and compared to model calculations. Agreement between theory...... and experiment is excellent. In addition, by properly choosing the Rabi frequencies of the two optical transitions a velocity independent atomic dark state is observed....

  7. Magnetic two-photon scattering and two-photon emission - Cross sections and redistribution functions

    Science.gov (United States)

    Alexander, S. G.; Meszaros, P.

    1991-01-01

    The magnetic two-photon scattering cross section is discussed within the framework of QED, and the corresponding scattering redistribution function for this process and its inverse, as well as the scattering source function are calculated explicitly. In a similar way, the magnetic two-photon emission process which follows the radiative excitation of Landau levels above ground is calculated. The two-photon scattering and two-photon emission are of the same order as the single-photon magnetic scattering. All three of these processes, and in optically thick cases also their inverses, are included in radiative transport calculations modeling accreting pulsars and gamma-ray bursters. These processes play a prominent role in determining the relative strength of the first two cyclotron harmonics, and their effects extend also to the higher harmonics.

  8. Two-photon ionization of colliding atoms

    Energy Technology Data Exchange (ETDEWEB)

    Nayfeh, M.H.

    1977-09-01

    Semiclassical expressions of two-photon ionization of two colliding atoms are derived for a wide range of electromagnetic field intensity and detunings from the isolated atom line. The dependence of the ionization yield on the details of the interaction potential of the system is derived. This process promises an extremely sensitive method for studying line broadening on the far wing, especially when absorption or fluorescence becomes very weak.

  9. Two-photon cooling of magnesium atoms

    DEFF Research Database (Denmark)

    Malossi, N.; Damkjær, S.; Hansen, P. L.;

    2005-01-01

    A two-photon mechanism for cooling atoms below the Doppler temperature is analyzed. We consider the magnesium ladder system (3s2)S01¿(3s3p)P11 at 285.2nm followed by the (3s3p)P11¿(3s3d)D21 transition at 880.7nm . For the ladder system quantum coherence effects may become important. Combined...

  10. Two-Photon Collective Atomic Recoil Lasing

    Directory of Open Access Journals (Sweden)

    James A. McKelvie

    2015-11-01

    Full Text Available We present a theoretical study of the interaction between light and a cold gasof three-level, ladder configuration atoms close to two-photon resonance. In particular, weinvestigate the existence of collective atomic recoil lasing (CARL instabilities in differentregimes of internal atomic excitation and compare to previous studies of the CARL instabilityinvolving two-level atoms. In the case of two-level atoms, the CARL instability is quenchedat high pump rates with significant atomic excitation by saturation of the (one-photoncoherence, which produces the optical forces responsible for the instability and rapid heatingdue to high spontaneous emission rates. We show that in the two-photon CARL schemestudied here involving three-level atoms, CARL instabilities can survive at high pump rateswhen the atoms have significant excitation, due to the contributions to the optical forces frommultiple coherences and the reduction of spontaneous emission due to transitions betweenthe populated states being dipole forbidden. This two-photon CARL scheme may form thebasis of methods to increase the effective nonlinear optical response of cold atomic gases.

  11. How to connect time-lapse recorded trajectories of motile microorganisms with dynamical models in continuous time

    Science.gov (United States)

    Pedersen, Jonas N.; Li, Liang; Grǎdinaru, Cristian; Austin, Robert H.; Cox, Edward C.; Flyvbjerg, Henrik

    2016-12-01

    We provide a tool for data-driven modeling of motility, data being time-lapse recorded trajectories. Several mathematical properties of a model to be found can be gleaned from appropriate model-independent experimental statistics, if one understands how such statistics are distorted by the finite sampling frequency of time-lapse recording, by experimental errors on recorded positions, and by conditional averaging. We give exact analytical expressions for these effects in the simplest possible model for persistent random motion, the Ornstein-Uhlenbeck process. Then we describe those aspects of these effects that are valid for any reasonable model for persistent random motion. Our findings are illustrated with experimental data and Monte Carlo simulations.

  12. Harmonic Oscillator in Heat Bath: Exact simulation of time-lapse-recorded data, exact analytical benchmark statistics

    CERN Document Server

    Norrelykke, Simon F

    2011-01-01

    The stochastic dynamics of the damped harmonic oscillator in a heat bath is simulated with an algorithm that is exact for time steps of arbitrary size. Exact analytical results are given for correlation functions and power spectra in the form they acquire when computed from experimental time-lapse recordings. Three applications are discussed: (i) Effects of finite sampling-rate and -time, described exactly here, are similar for other stochastic dynamical systems-e.g. motile micro-organisms and their time-lapse recorded trajectories. (ii) The same statistics is satisfied by any experimental system to the extent it is interpreted as a damped harmonic oscillator at finite temperature-such as an AFM cantilever. (iii) Three other models of fundamental interest are limiting cases of the damped harmonic oscillator at finite temperature; it consequently bridges their differences and describes effects of finite sampling rate and sampling time for these models as well. Finally, we give a brief discussion of nondimensio...

  13. A time-series method for automated measurement of changes in mitotic and interphase duration from time-lapse movies.

    Directory of Open Access Journals (Sweden)

    Frederic D Sigoillot

    Full Text Available BACKGROUND: Automated time-lapse microscopy can visualize proliferation of large numbers of individual cells, enabling accurate measurement of the frequency of cell division and the duration of interphase and mitosis. However, extraction of quantitative information by manual inspection of time-lapse movies is too time-consuming to be useful for analysis of large experiments. METHODOLOGY/PRINCIPAL FINDINGS: Here we present an automated time-series approach that can measure changes in the duration of mitosis and interphase in individual cells expressing fluorescent histone 2B. The approach requires analysis of only 2 features, nuclear area and average intensity. Compared to supervised learning approaches, this method reduces processing time and does not require generation of training data sets. We demonstrate that this method is as sensitive as manual analysis in identifying small changes in interphase or mitotic duration induced by drug or siRNA treatment. CONCLUSIONS/SIGNIFICANCE: This approach should facilitate automated analysis of high-throughput time-lapse data sets to identify small molecules or gene products that influence timing of cell division.

  14. Realtime infiltration process monitoring in macroporous soil - a plot-scale experiment accompanied by high-resolution time-lapse 3D GPR

    Science.gov (United States)

    Jackisch, Conrad; Allroggen, Niklas

    2016-04-01

    Infiltration and quick vertical redistribution of event water through rapid subsurface flow in soil structures is one of the key issues in hydrology. Although the importance of preferential flow is broadly recognised, our theories, observation techniques and modelling approaches lose grounds when the assumption of well-mixed states in REVs collapses. To characterise the combination of advective and diffusive flow is especially challenging. We have shown in earlier studies that a combination of TDR monitoring, dye- and salt-tracer recovery and time-lapse 3D GPR in irrigation experiments provides means to characterise infiltration dynamics at the plot- and hillslope-scale also in highly structured soils. We pinpointed that the spatial and temporal resolution requires special attention and improvement - particularly owing to the facts of high velocity (10-3 ms-1) of advective flow and small scale (10-2 m) of the respective flow structures. We present insights from a novel technique of continuous high-resolution time-lapse 3D GPR measurements during and after a plot-scale (1 m x 1 m) irrigation experiment. Continuous TDR soil moisture measurements, dye tracer excavation and salt-tracer samples are used as qualitative and quantitative references. While classical infiltration experiments either look at spatial patterns or temporal dynamics at singular gauges, we highlight the advantage of combining both to achieve a more complete image of the infiltration process. Although operating at the limits of the techniques this setup enables non-invasive observation of preferential flow processes in the field and allows to explore and characterise macropore matrix exchange.

  15. Preliminary Seismic Time-lapse Results of the First Post-injection Monitoring at the Ketzin Pilot Site for CO2 Storage

    Science.gov (United States)

    Huang, Fei; Monika, Ivandic; Julin, Christopher; Lüth, Stefan; Bergmann, Peter

    2016-04-01

    Time-lapse surface seismic monitoring methods have proven to be notably successful in imaging the development of the CO2 plume injected into the 630-650 meters deep saline aquifer at the Ketzin pilot site. A 3D baseline survey was acquired in the autumn of 2005 prior to CO2 injection, followed by two 3D repeat surveys conducted in the same season of 2009 and 2012 after injection of about 22 and 61 kt of CO2, respectively. These repeat surveys showed that the CO2 plume was concentrated around the injection well with a preferred WNW propagating trend due to reservoir heterogeneity. The CO2 induced amplitude anomaly continued to grow with more CO2 injected into the reservoir. The CO2 injection ended on August 29, 2013 after a total of about 67 kt injected CO2. In the autumn of 2015, a third 3D repeat survey with a nominal fold of 25 was acquired during the post-injection phase. The acquisition was implemented with an identical template scheme and acquisition parameters as in the previous surveys. About 5500 source points were acquired during the 57 days of active acquisition. The same processing steps as used before were carried out from pre-stack to post-stack. In order to minimize changes unrelated to the site operations, cross-calibration was applied to the data. Preliminary results of the time-lapse analysis show that the obtained amplitude anomaly at the reservoir level is smaller in size than the one observed at the time of the second repeat survey. This may indicate that CO2 dissolution is active, especially east of the injection site, where permeability and flow rate appear to be lower. In comparison to the previous surveys, the maximum amplitude anomaly has moved towards the west, which is consistent with the previously observed propagating tendency of the CO2 plume within the reservoir. No CO2 leakage is observed within the overburden.

  16. Time-lapse electromagnetic induction surveys under olive tree canopies reveal soil moisture dynamics and controls

    Science.gov (United States)

    Martínez, Gonzalo; Giraldez Cervera, Juan Vicente; Vanderlinden, Karl

    2015-04-01

    Soil moisture (θ) is a critical variable that exerts an important control on plant status and development. Soil sampling, neutron attenuation and electromagnetic methods such as TDR or FDR have been used widely to measure θ and provide point data at a possible range of temporal resolutions. However, these methods require either destructive sampling or permanently installed devices with often limiting measurement depths, or are extremely time-consuming. Moreover, the small support of such measurements compromises its value in heterogeneous soils. To overcome such limitations electromagnetic induction (EMI) can be tested to monitor θ at different spatial and temporal scales. This work investigates the potential of EMI to characterize the spatio-temporal variability of soil moisture from apparent electrical conductivity (ECa) under the canopy of individual olive trees. During one year we measured θ with a frequency of 5 min and ECa on an approximately weekly basis along transects from the tree trunk towards the inter-row area. CS-616 soil moisture sensors where horizontally installed in the walls of a trench at depths of 0.1, 0.2, 0.4, 0.6 and 0.8 m at five locations along the transect, with a separation of 0.8 m. The Dualem-21S sensor was used to measure weekly the ECa at 0.2 m increments, from the tree trunk to a distance of 4.4 m. The results showed similar drying and wetting patterns for θ and ECa. Both variables showed a decreasing pattern from the tree trunk towards the drip line, followed by a sharp increment and constant values towards the center of the inter-row space. This pattern reflects clearly the influence of root-zone water uptake under the tree canopy and higher θ values in the inter-row area where root-water uptake is smaller. Time-lapse ECa data responded to evaporation and infiltration fluxes with the highest sensitivity for the 1 and 1.5 m ECa signals, as compared to the 0.5 and 3.0 m signals. Overall these preliminary results revealed the

  17. Penicillin induced persistence in Chlamydia trachomatis: high quality time lapse video analysis of the developmental cycle.

    Directory of Open Access Journals (Sweden)

    Rachel J Skilton

    Full Text Available BACKGROUND: Chlamydia trachomatis is a major human pathogen with a unique obligate intracellular developmental cycle that takes place inside a modified cytoplasmic structure known as an inclusion. Following entry into a cell, the infectious elementary body (EB differentiates into a non-infectious replicative form known as a reticulate body (RB. RBs divide by binary fission and at the end of the cycle they redifferentiate into EBs. Treatment of C.trachomatis with penicillin prevents maturation of RBs which survive and enlarge to become aberrant RBs within the inclusion in a non-infective persistent state. Persistently infected individuals may be a reservoir for chlamydial infection. The C.trachomatis genome encodes the enzymes for peptidoglycan (PG biosynthesis but a PG sacculus has never been detected. This coupled to the action of penicillin is known as the chlamydial anomaly. We have applied video microscopy and quantitative DNA assays to the chlamydial developmental cycle to assess the effects of penicillin treatment and establish a framework for investigating penicillin induced chlamydial persistence. PRINCIPAL FINDINGS: Addition of penicillin at the time of cell infection does not prevent uptake and the establishment of an inclusion. EB to RB transition occurs but bacterial cytokinesis is arrested by the second binary fission. RBs continue to enlarge but not divide in the presence of penicillin. The normal developmental cycle can be recovered by the removal of penicillin although the large, aberrant RBs do not revert to the normal smaller size but remain present to the completion of the developmental cycle. Chromosomal and plasmid DNA replication is unaffected by the addition of penicillin but the arrest of bacterial cytokinesis under these conditions results in RBs accumulating multiple copies of the genome. CONCLUSIONS: We have applied video time lapse microscopy to the study of the chlamydial developmental cycle. Linked with accurate

  18. Developmental pathway of somatic embryogenesis in Picea abies as revealed by time-lapse tracking.

    Science.gov (United States)

    Filonova, L H; Bozhkov, P V; von Arnold, S

    2000-02-01

    Several coniferous species can be propagated via somatic embryogenesis. This is a useful method for clonal propagation, but it can also be used for studying how embryo development is regulated in conifers. However, in conifers it is not known to what extent somatic and zygotic embryos develop similarly, because there has been little research on the origin and development of somatic embryos. A time-lapse tracking technique has been set up, and the development of more than 2000 single cells and few-celled aggregates isolated from embryogenic suspension cultures of Norway spruce (Picea abies L. Karst.) and embedded in thin layers of agarose has been traced. Experiments have shown that somatic embryos develop from proembryogenic masses which pass through a series of three characteristic stages distinguished by cellular organization and cell number (stages I, II and III) to transdifferentiate to somatic embryos. Microscopic inspection of different types of structures has revealed that proembryogenic masses are characterized by high interclonal variation of shape and cellular constitution. In contrast, somatic embryos are morphologically conservative structures, possessing a distinct protoderm-like cell layer as well as embryonal tube cells and suspensor. The lack of staining of the arabinogalactan protein epitope recognized by the monoclonal antibody JIM13 was shown to be an efficient marker for distinguishing proembryogenic masses from somatic embryos. The vast majority of cells in proembryogenic masses expressed this epitope and none of cells in the early somatic embryos. The conditions that promote cell proliferation (i.e. the presence of exogenous auxin and cytokinin), inhibit somatic embryo formation; instead, continuous multiplication of stage I proembryogenic masses by unequal division of embryogenic cells with dense cytoplasm is the prevailing process. Once somatic embryos have formed, their further development to mature forms requires abscisic acid and shares a

  19. Oriented cell divisions and cellular morphogenesis in the zebrafish gastrula and neurula: a time-lapse analysis.

    Science.gov (United States)

    Concha, M L; Adams, R J

    1998-03-01

    We have taken advantage of the optical transparency of zebrafish embryos to investigate the patterns of cell division, movement and shape during early stages of development of the central nervous system. The surface-most epiblast cells of gastrula and neurula stage embryos were imaged and analysed using a computer-based, time-lapse acquisition system attached to a differential interference contrast (DIC) microscope. We find that the onset of gastrulation is accompanied by major changes in cell behaviour. Cells collect into a cohesive sheet, apparently losing independent motility and integrating their behaviour to move coherently over the yolk in a direction that is the result of two influences: towards the vegetal pole in the movements of epiboly and towards the dorsal midline in convergent movements that strengthen throughout gastrulation. Coincidentally, the plane of cell division becomes aligned to the surface plane of the embryo and oriented in the anterior-posterior (AP) direction. These behaviours begin at the blastoderm margin and propagate in a gradient towards the animal pole. Later in gastrulation, cells undergo increasingly mediolateral-directed elongation and autonomous convergence movements towards the dorsal midline leading to an enormous extension of the neural axis. Around the equator and along the dorsal midline of the gastrula, persistent AP orientation of divisions suggests that a common mechanism may be involved but that neither oriented cell movements nor shape can account for this alignment. When the neural plate begins to differentiate, there is a gradual transition in the direction of cell division from AP to the mediolateral circumference (ML). ML divisions occur in both the ventral epidermis and dorsal neural plate. In the neural plate, ML becomes the predominant orientation of division during neural keel and nerve rod stages and, from late neural keel stage, divisions are concentrated at the dorsal midline and generate bilateral progeny

  20. Adiabatic following in two-photon transition

    Energy Technology Data Exchange (ETDEWEB)

    Nayfeh, M.H.; Nayfeh, A.H.

    1977-03-01

    The coherent interaction of two smoothly varying, near-resonant, two-photon pulses with a three-level system can be described by ''two-photon damped Bloch equations'' which are analogous to those for a one-photon transition in a two-level system except for the presence of a two-photon coupling and a frequency shift. These equations are solved for the cases ..gamma../sub 1/, ..gamma../sub 2/ very-much-less-than ..cap omega.., ..gamma../sub 1/ = ..gamma../sub 2/, and ..gamma../sub 2/k/sup 2/epsilon/sup 4//..cap omega../sup 2/, ..gamma../sub 1/ very-much-less-than ..cap omega.., where ..gamma../sub 1/ and ..gamma../sub 2/ are the atomic energy and phase relaxation widths, respectively, and ..cap omega.. is the Rabi frequency. The leading contribution to the refractive index is intensity dependent, caused by the level shifts inherent in multiphoton processes; it includes a relaxation dependent part which is important at times shorter than ..gamma../sup -1//sub 1/. The second-order contributions depend on the square of the intensity and the time-integrated square of the intensity. The latter contribution, which is relaxation dependent, causes line asymmetry at the long-wavelength wing; it consists of a term proportional to ..gamma../sub 2/-..gamma../sub 1/ and only important at early times and a term proportional to 2..gamma../sub 2/-..gamma../sub 1/.

  1. STUDY OF TIME LAPSE IN FOREIGN BODY ASPIRATION IN RELATION TO CHEST X - RAY AND TYPE OF FOREIGN BODY

    Directory of Open Access Journals (Sweden)

    Salma

    2015-10-01

    Full Text Available INTRODUCTI ON: Foreign body aspiration in pediatrics is a potentially fatal accident which will continue until children explore their surroundings with their hand and mouth. Pediatric aspirations will persist until mankind exists. Not all foreign body aspirations are witnessed hence chances of delay in diagnosing an aspiration are high. Delay in diagnosis depends on site and character of foreign body aspirated. The chest x - ray findings and type of foreign body extracted vary depending on the duration the foreign body remains in airway . OBJECTIVE: To study the X - ray finding in pediatric airway aspiration and its relation to time lapse, the type and site of lodgment of foreign body extracted via bronchoscopy. The type of foreign body in relation to time lapse in aspiration. MATERIAL AND METHODS: This was a prospective study done in Bapuji child health institute and research center, JJM Medical College, Davangere . History and pre bronchoscopy x - Ray finding were noted for 65 children who were posted for suspicious bronchoscopy from August 2011 to September 2013. 11 children were excluded from study as they showed no foreign body on bronchoscopy. Time lapse in aspir ation and seeking medical care was noted. The bronchoscopic findings regarding site of foreign body lodgment and type of foreign body were recorded. The type of foreign body and variation of x - ray picture in relation to time lapse in aspiration were noted. Data collected was analyzed using descriptive statistics. RESULT: It was observed that mean age was 28 months. About 80% of the cases were between 1 to 3 years age. 82% (n=53/54 were radio lucent foreign body, only 1.5% (n=1/54 were radio o paque. Site of lodgment of foreign body was right main bronchus in 48% (n=26/54, left main bronchus 46% (n=25/54 , tracheal 1.85% (n=1/54, subglottic 1.85% (n=1/54, carinal 1.85% (n=1/54, multiple site i.e. left bronchus +right bronchus+ carinal 1.85% (n=1/54. Groundnut was most common

  2. Understanding infiltration and groundwater flow at an artificial recharge facility using time-lapse gravity data

    Science.gov (United States)

    Kennedy, Jeffrey

    Groundwater provides a fundamental resource for modern life. Throughout the world, groundwater is managed by storing (recharging) it underground in natural aquifers for future withdrawal and consumptive use. In Arizona, over 4 million people benefit from managed aquifer storage, but little effort is made to track the movement of recharged water through the subsurface. Motivated by current limitations in our ability to monitor percolation and groundwater movement at the scale of a recharge facility, an effort to collect time-lapse gravity data was carried out at the Southern Avra Valley Storage and Recovery Project (SAVSARP) operated by the City of Tucson, Arizona. In addition to collecting water-level data 12 wells, there were three primary gravity experiments: (1) five continuously-recording gravity meters (2 iGrav superconducting gravity meters and 3 gPhone gravity meters) were installed semi-permanently in control buildings adjacent to the recharge basins, (2) absolute gravity measurements were made at nine locations over a 17 month period, and (3) three relative-gravity campaigns were carried out on a network of 70 stations. This large-scale controlled experiment, with known infiltration and pumping rates, resulted in one of the most comprehensive datasets of its kind. Gravity data led to several hydrologic insights, both through direct measurement and modeling. First, the infiltration rate could be estimated accurately based on the initial rate of gravity change during infiltration, regardless of the specific yield. Using two gravity meters, the depth, and therefore speed, of the wetting front beneath a recharge basin was observed, including the time at which the water table was reached. Spatial maps of gravity change from relative gravity surveys show areas where infiltration efficiency is highest, and where groundwater accumulates; storage accumulated preferentially to the west of the recharge basins, away from pumping wells. Such information would be

  3. Two-photon interference : spatial aspects of two-photon entanglement, diffraction, and scattering

    NARCIS (Netherlands)

    Peeters, Wouter Herman

    2010-01-01

    This dissertation contains scientific research within the realm of quantum optics, which is a branch of physics. An experimental and theoretical study is made of two-photon interference phenomena in various optical systems. Spatially entangled photon pairs are produced via the nonlinear optical proc

  4. Plasmonic-enhanced two-photon fluorescence with single gold nanoshell

    Science.gov (United States)

    Zhang, TianYue; Lu, GuoWei; Shen, HongMing; Perriat, P.; Martini, M.; Tillement, O.; Gong, QiHuang

    2014-06-01

    Single gold nanoshell with mutilpolar plasmon resonances is proposed to enhance two-photon fluorescence efficiently. The single emitter single nanoshell configuration is studied systematically by employing the finite-difference time-domain method. The emitter located inside or outside the nanoshell at various positions leads to a significantly different enhancement effect. The fluorescent emitter placed outside the nanoshell can achieve large fluorescence intensity given that both the position and orientation of the emission dipole are optimally controlled. In contrast, for the case of the emitter placed inside the nanoshell, it can experience substantial two-photon fluorescence enhancement without strict requirements upon the position and dipole orientations. Metallic nanoshell encapsulating many fluorescent emitters should be a promising nanocomposite configuration for bright two-photon fluorescence label. The results provide a comprehensive understanding about the plasmonic-enhanced two-photon fluorescence behaviors, and the nanocomposite configuration has great potential for optical detecting, imaging and sensing in biological applications.

  5. A spirobifluorene-based two-photon fluorescence probe for mercury ions and its applications in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Xiao, Haibo, E-mail: xiaohb@shnu.edu.cn; Zhang, Yanzhen; Zhang, Wu; Li, Shaozhi; Tan, Jingjing; Han, Zhongying

    2017-05-01

    A novel spirobifluorene derivative SPF-TMS, which containing dithioacetal groups and triphenylamine units, was synthesized. The probing behaviors toward various metal ions were investigated via UV/Vis absorption spectra as well as one-photon fluorescence changes. The results indicated that SPF-TMS exhibits high sensitivity and selectivity for mercury ions. The detection limit was at least 8.6 × 10{sup −8}M, which is excellent comparing with other optical sensors for Hg{sup 2+}. When measured by two-photon excited fluorescence technique in THF at 800 nm, the two-photon cross-section of SPF-TMS is 272 GM. Especially, upon reaction with mercury species, SPF-TMS yielded another two-photon dye SPF-DA. Both SPF-TMS and SPF-DA emit strong two-photon induced fluorescence and can be applied in cell imaging by two-photon microscopy. - Highlights: • We report a spirobifluorene-based molecule as two-photon fluorescent probe with large two-photon cross-section. • The molecule has exclusive selectivity and sensitivity for mercury species. • The molecule has large two-photon emission changes before and after addition of Hg{sup 2+}. • Both the probe and the mercury ion-promoted reaction product can be applied in cell imaging by two-photon microscopy.

  6. Summer-time Mass Balance of Wolverine Glacier, Alaska, Derived from Ground-based Time-lapse Microgravity Measurements

    Science.gov (United States)

    Young, E. V.; Muto, A.; Babcock, E.

    2016-12-01

    Monitoring the mass balance of alpine glaciers is important because alpine glaciers presently account for about half of the cryospheric contribution to the global sea-level rise. Mass balance measurements of alpine glaciers have predominantly relied upon glaciological and hydrological methods. However, these methods can be logistically costly and have potential extrapolation errors. Remote sensing approaches, such as gravimetric methods using data from GRACE satellite missions, have provided monthly mass-balance estimates of aggregates of alpine glaciers but their spatial resolution is far too large for studying a single glacier. On the other hand, ground-based time-lapse microgravity geophysical measurements can potentially circumvent some of the disadvantages of the glaciological and hydrological methods. It may detect the change in a single glacier's mass and its spatial distribution. We conducted ground-based time-lapse microgravity surveys on Wolverine Glacier, Alaska, in May and August of 2016, using a Scintrex CG-5 Autograv gravimeter. We collected data at seventy-nine individual stations on the glacier, roughly five stations per square kilometer. We included repeat-station and base-station measurements made at least twice a day for instrumental drift control. The uncertainty of our gravity measurements is better than 0.03 mGal, which is about 0.7 meters water equivalent of surface mass balance. Our summer-time mass balance of Wolverine Glacier determined from the time-lapse gravity measurements is independent of that derived from the stake-network or stream-gauge measurements, and could provide spatial insight into the mass balance process on Wolverine Glacier and similar glaciers.

  7. Effect of Time Lapse on the Diagnostic Accuracy of Cone Beam Computed Tomography for Detection of Vertical Root Fractures.

    Science.gov (United States)

    Eskandarloo, Amir; Asl, Amin Mahdavi; Jalalzadeh, Mohsen; Tayari, Maryam; Hosseinipanah, Mohammad; Fardmal, Javad; Shokri, Abbas

    2016-01-01

    Accurate and early diagnosis of vertical root fractures (VRFs) is imperative to prevent extensive bone loss and unnecessary endodontic and prosthodontic treatments. The aim of this study was to assess the effect of time lapse on the diagnostic accuracy of cone beam computed tomography (CBCT) for VRFs in endodontically treated dog's teeth. Forty-eight incisors and premolars of three adult male dogs underwent root canal therapy. The teeth were assigned to two groups: VRFs were artificially induced in the first group (n=24) while the teeth in the second group remained intact (n=24). The CBCT scans were obtained by NewTom 3G unit immediately after inducing VRFs and after one, two, three, four, eight, 12 and 16 weeks. Three oral and maxillofacial radiologists blinded to the date of radiographs assessed the presence/absence of VRFs on CBCT scans. The sensitivity, specificity and accuracy values were calculated and data were analyzed using SPSS v.16 software and ANOVA. The total accuracy of detection of VRFs immediately after surgery, one, two, three, four, eight, 12 and 16 weeks was 67.3%, 68.7%, 66.6%, 64.6%, 64.5%, 69.4%, 68.7%, 68% respectively. The effect of time lapse on detection of VRFs was not significant (p>0.05). Overall sensitivity, specificity and accuracy of CBCT for detection of VRFs were 74.3%, 62.2%, 67.2% respectively. Cone beam computed tomography is a valuable tool for detection of VRFs. Time lapse (four months) had no effect on detection of VRFs on CBCT scans.

  8. Effect of time lapse on the diagnostic accuracy of cone beam computed tomography for detection of vertical root fractures

    Energy Technology Data Exchange (ETDEWEB)

    Eskandarloo, Amir; Shokri, Abbas, E-mail: Dr.a.shokri@gmail.com [Dental Research Center, Department of Oral and Maxillofacial Radiology, Hamadan University of Medical Sciences, Hamadan (Iran, Islamic Republic of); Asl, Amin Mahdavi [Department of Oral and Maxillofacial Radiology, Golestan University of Medical Sciences, Gorgan (Iran, Islamic Republic of); Jalalzadeh, Mohsen [Department of Endodontics, Hamadan University of Medical Sciences, Hamadan (Iran, Islamic Republic of); Tayari, Maryam [Department of Pedodontics, Golestan University of Medical Sciences, Gorgan (Iran, Islamic Republic of); Hosseinipanah, Mohammad [Department of Anatomy, School of Medicine, Hamadan University of Medical Sciences, Hamadan (Iran, Islamic Republic of); Fardmal, Javad [Research Center for Health Sciences and Department of Epidemiology and Biostatistics, School of Public Health, Hamadan University of Medical Sciences, Hamadan (Iran, Islamic Republic of)

    2016-01-15

    Accurate and early diagnosis of vertical root fractures (VRFs) is imperative to prevent extensive bone loss and unnecessary endodontic and prosthodontic treatments. The aim of this study was to assess the effect of time lapse on the diagnostic accuracy of cone beam computed tomography (CBCT) for VRFs in endodontically treated dog’s teeth. Forty eight incisors and premolars of three adult male dogs underwent root canal therapy. The teeth were assigned to two groups: VRFs were artificially induced in the first group (n=24) while the teeth in the second group remained intact (n=24). The CBCT scans were obtained by NewTom 3G unit immediately after inducing VRFs and after one, two, three, four, eight, 12 and 16 weeks. Three oral and maxillofacial radiologists blinded to the date of radiographs assessed the presence/absence of VRFs on CBCT scans. The sensitivity, specificity and accuracy values were calculated and data were analyzed using SPSS v.16 software and ANOVA. The total accuracy of detection of VRFs immediately after surgery, one, two, three, four, eight, 12 and 16 weeks was 67.3%, 68.7%, 66.6%, 64.6%, 64.5%, 69.4%, 68.7%, 68% respectively. The effect of time lapse on detection of VRFs was not significant (p>0.05). Overall sensitivity, specificity and accuracy of CBCT for detection of VRFs were 74.3%, 62.2%, 67.2% respectively. Cone beam computed tomography is a valuable tool for detection of VRFs. Time lapse (four months) had no effect on detection of VRFs on CBCT scans. (author)

  9. Two-photon Interference with Non-identical Photons

    CERN Document Server

    Liu, Jianbin; Zheng, Huaibin; Chen, Hui; Li, Fu-Li; Xu, Zhuo

    2014-01-01

    The indistinguishability of non-identical photons is dependent on detection system in quantum physics. If two photons with different wavelengths are indistinguishable for a detection system, there can be two-photon interference when these two photons are incident to two input ports of a Hong-Ou-Mandel interferometer, respectively. The reason why two-photon interference phenomena are different for classical and nonclassical light is not due to interference, but due to the properties of light and detection system. These conclusions are helpful to understand the physics and applications of two-photon interference.

  10. Soundscape and Noise Exposure Monitoring in a Marine Protected Area Using Shipping Data and Time-Lapse Footage.

    Science.gov (United States)

    Merchant, Nathan D; Pirotta, Enrico; Barton, Tim R; Thompson, Paul M

    2016-01-01

    We review recent work that developed new techniques for underwater noise assessment that integrate acoustic monitoring with automatic identification system (AIS) shipping data and time-lapse video, meteorological, and tidal data. Two sites were studied within the Moray Firth Special Area of Conservation (SAC) for bottlenose dolphins, where increased shipping traffic is expected from construction of offshore wind farms outside the SAC. Noise exposure varied markedly between the sites, and natural and anthropogenic contributions were characterized using multiple data sources. At one site, AIS-operating vessels accounted for total cumulative sound exposure (0.1-10 kHz), suggesting that noise modeling using the AIS would be feasible.

  11. IP4DI: A software for time-lapse 2D/3D DC-resistivity and induced polarization tomography

    Science.gov (United States)

    Karaoulis, M.; Revil, A.; Tsourlos, P.; Werkema, D. D.; Minsley, B. J.

    2013-04-01

    We propose a 2D/3D forward modelling and inversion package to invert direct current (DC)-resistivity, time-domain induced polarization (TDIP), and frequency-domain induced polarization (FDIP) data. Each cell used for the discretization of the 2D/3D problems is characterized by a DC-resistivity value and a chargeability or complex conductivity for TDIP/FDIP problems, respectively. The governing elliptic partial differential equations are solved with the finite element method, which can be applied for both real and complex numbers. The inversion can be performed either for a single snapshot of data or for a sequence of snapshots in order to monitor a dynamic process such as a salt tracer test. For the time-lapse inversion, we have developed an active time constrained (ATC) approach that is very efficient in filtering out noise in the data that is not correlated over time. The forward algorithm is benchmarked with simple analytical solutions. The inversion package IP4DI is benchmarked with three tests, two including simple geometries. The last one corresponds to a time-lapse resistivity problem for cross-well tomography during enhanced oil recovery. The algorithms are based on MATLAB® code package and a graphical user interface (GUI).

  12. A laboratory validation study of the time-lapse oscillatory pumping test for leakage detection in geological repositories

    Science.gov (United States)

    Sun, Alexander Y.; Lu, Jiemin; Islam, Akand

    2017-05-01

    Geologic repositories are extensively used for disposing byproducts in mineral and energy industries. The safety and reliability of these repositories are a primary concern to environmental regulators and the public. Time-lapse oscillatory pumping test (OPT) has been introduced recently as a pressure-based technique for detecting potential leakage in geologic repositories. By routinely conducting OPT at a number of pulsing frequencies, an operator may identify the potential repository anomalies in the frequency domain, alleviating the ambiguity caused by reservoir noise and improving the signal-to-noise ratio. Building on previous theoretical and field studies, this work performed a series of laboratory experiments to validate the concept of time-lapse OPT using a custom made, stainless steel tank under relatively high pressures. The experimental configuration simulates a miniature geologic storage repository consisting of three layers (i.e., injection zone, caprock, and above-zone aquifer). Results show that leakage in the injection zone led to deviations in the power spectrum of observed pressure data, and the amplitude of which also increases with decreasing pulsing frequencies. The experimental results are further analyzed by developing a 3D flow model, using which the model parameters are estimated through frequency domain inversion.

  13. Estimation of soil hydraulic parameters by integrated hydrogeophysical inversion of time-lapse GPR data measured at Selhausen, Germany

    KAUST Repository

    Jadoon, Khan

    2012-06-01

    We present an integrated hydrogeophysical inversion approach that uses time-lapse off-ground ground-penetrating radar (GPR) data to estimate soil hydraulic parameters, and apply it to a dataset collected in the field. Off-ground GPR data are mainly sensitive to the near-surface water content profile and dynamics, and are thus related to soil hydraulic parameters, such as the parameters of the hydraulic conductivity and water retention functions. The hydrological simulator HYDRUS 1-D was used with a two-layer single- and dual-porosity model. To monitor the soil water content dynamics, time-lapse GPR and time domain reflectometry (TDR) measurements were performed, whereby only GPR data was used in the inversion. The dual porosity model provided better results compared to the single porosity model for describing the soil water dynamics, which is supported by field observations of macropores. Furthermore, the GPR-derived water content profiles reconstructed from the integrated hydrogeophysical inversion were in good agreement with TDR observations. These results suggest that the proposed method is promising for non-invasive characterization of the shallow subsurface hydraulic properties and monitoring water dynamics at the field scale.

  14. Time-lapse geophysical technology-based study on overburden strata changes induced by modern coal mining

    Institute of Scientific and Technical Information of China (English)

    Wenfeng Du; Suping Peng; Guowei Zhu; Feng Yang

    2014-01-01

    To study the impact of modern coal mining on overlying strata and its water bearing conditions, integrated time-lapse geophysical prospecting integrating 3D seismic, electrical and ground penetrating radar method were used. Through observing and analyzing the geophysical data variations of all stages of pre-mining, mining and post-mining as well as post-mining deposition stable period, impacts of coal mining on stratigraphic structure and its water bearing were studied and modern coal mining induced stratigraphic change pattern was summarized. The research result shows that the stratigraphic structure and the water bearing of surface layer during modern coal mining have self-healing pattern with mining time;the self-healing capability of near-surface strata is relatively strong while the roof weak;water bearing self-healing of near-surface strata is relatively high while the roof strata adjacent to mined coal beds low. Due to integrated time-lapse geophysical prospecting technology has extra time dimension which makes up the deficiency of static analysis of conventional geophysical methods, it can better highlight the dynamic changes of modern coal mining induced over-burden strata and its water bearing conditions.

  15. Two-Photon and Second Harmonic Microscopy in Clinical and Translational Cancer Research

    Science.gov (United States)

    PERRY, SETH W.; BURKE, RYAN M.; BROWN, EDWARD B.

    2012-01-01

    Application of two-photon microscopy (TPM) to translational and clinical cancer research has burgeoned over the last several years, as several avenues of pre-clinical research have come to fruition. In this review, we focus on two forms of TPM—two-photon excitation fluorescence microscopy, and second harmonic generation microscopy—as they have been used for investigating cancer pathology in ex vivo and in vivo human tissue. We begin with discussion of two-photon theory and instrumentation particularly as applicable to cancer research, followed by an overview of some of the relevant cancer research literature in areas that include two-photon imaging of human tissue biopsies, human skin in vivo, and the rapidly developing technology of two-photon microendoscopy. We believe these and other evolving two-photon methodologies will continue to help translate cancer research from the bench to the bedside, and ultimately bring minimally invasive methods for cancer diagnosis and treatment to therapeutic reality. PMID:22258888

  16. Gigavision - A weatherproof, multibillion pixel resolution time-lapse camera system for recording and tracking phenology in every plant in a landscape

    Science.gov (United States)

    Brown, T.; Borevitz, J. O.; Zimmermann, C.

    2010-12-01

    retrieved at bi-weekly intervals. Our longer-term goal is to make gigapixel time-lapse datasets available online in an interactive interface that layers plant-level phenology data with gigapixel resolution images, genomic sequence data from individual plants with weather and other abitotic sensor data. Co-visualization of all of these data types provides researchers with a powerful new tool for examining complex ecological interactions across scales from the individual to the ecosystem. We will present detailed phenostage data from more than 100 plants of multiple species from our Gigavision timelapse camera at our “Big Blowout East” field site in the Indiana Dunes State Park, IN. This camera has been recording three to four 700 million pixel images a day since February 28, 2010. The camera field of view covers an area of about 7 hectares resulting in an average image resolution of about 1 pixel per centimeter over the entire site. We will also discuss some of the many technological challenges with developing and maintaining these types of hardware systems, collecting quantitative data from gigapixel resolution time-lapse data and effectively managing terabyte-sized datasets of millions of images.

  17. Enhancement of two-photon photoluminescence and SERS for low-coverage gold films

    DEFF Research Database (Denmark)

    Novikov, Sergey M.; Beermann, Jonas; Frydendahl, Christian

    2016-01-01

    Electromagnetic field enhancement (FE) effects occurring in thin gold films 3-12-nm are investigated with two-photon photoluminescence (TPL) and Raman scanning optical microscopies. The samples are characterized using scanning electron microscopy images and linear optical spectroscopy. TPL images...

  18. Clinical multiphoton tomography and clinical two-photon microendoscopy

    Science.gov (United States)

    König, Karsten; Bückle, Rainer; Weinigel, Martin; Elsner, Peter; Kaatz, Martin

    2009-02-01

    We report on applications of high-resolution clinical multiphoton tomography based on the femtosecond laser system DermaInspectTM with its flexible mirror arm in Australia, Asia, and Europe. Applications include early detection of melanoma, in situ tracing of pharmacological and cosmetical compounds including ZnO nanoparticles in the epidermis and upper dermis, the determination of the skin aging index SAAID as well as the study of the effects of anti-aging products. In addition, first clinical studies with novel rigid high-NA two-photon 1.6 mm GRIN microendoscopes have been conducted to study the effect of wound healing in chronic wounds (ulcus ulcera) as well as to perform intrabody imaging with subcellular resolution in small animals.

  19. Two-photon luminescence microscopy of field enhancement at gold nanoparticles

    DEFF Research Database (Denmark)

    Beermann, Jonas; Bozhevolnyi, Sergey I.

    2005-01-01

    Using a reflection scanning optical microscope detecting two-photon luminescence (TPL) we have imaged square gold bumps positioned in a periodic array either on a smooth gold film or directly on a glass substrate. The second-harmonic (SH) and TPL response from these structures show both polarizat......Using a reflection scanning optical microscope detecting two-photon luminescence (TPL) we have imaged square gold bumps positioned in a periodic array either on a smooth gold film or directly on a glass substrate. The second-harmonic (SH) and TPL response from these structures show both...

  20. Entropy-Bayesian Inversion of Time-Lapse Tomographic GPR data for Monitoring Dielectric Permittivity and Soil Moisture Variations

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Z; Terry, N; Hubbard, S S; Csatho, B

    2013-02-12

    In this study, we evaluate the possibility of monitoring soil moisture variation using tomographic ground penetrating radar travel time data through Bayesian inversion, which is integrated with entropy memory function and pilot point concepts, as well as efficient sampling approaches. It is critical to accurately estimate soil moisture content and variations in vadose zone studies. Many studies have illustrated the promise and value of GPR tomographic data for estimating soil moisture and associated changes, however, challenges still exist in the inversion of GPR tomographic data in a manner that quantifies input and predictive uncertainty, incorporates multiple data types, handles non-uniqueness and nonlinearity, and honors time-lapse tomograms collected in a series. To address these challenges, we develop a minimum relative entropy (MRE)-Bayesian based inverse modeling framework that non-subjectively defines prior probabilities, incorporates information from multiple sources, and quantifies uncertainty. The framework enables us to estimate dielectric permittivity at pilot point locations distributed within the tomogram, as well as the spatial correlation range. In the inversion framework, MRE is first used to derive prior probability distribution functions (pdfs) of dielectric permittivity based on prior information obtained from a straight-ray GPR inversion. The probability distributions are then sampled using a Quasi-Monte Carlo (QMC) approach, and the sample sets provide inputs to a sequential Gaussian simulation (SGSim) algorithm that constructs a highly resolved permittivity/velocity field for evaluation with a curved-ray GPR forward model. The likelihood functions are computed as a function of misfits, and posterior pdfs are constructed using a Gaussian kernel. Inversion of subsequent time-lapse datasets combines the Bayesian estimates from the previous inversion (as a memory function) with new data. The memory function and pilot point design takes

  1. Entropy-Bayesian Inversion of Time-Lapse Tomographic GPR data for Monitoring Dielectric Permittivity and Soil Moisture Variations

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Zhangshuan; Terry, Neil C.; Hubbard, Susan S.

    2013-02-22

    In this study, we evaluate the possibility of monitoring soil moisture variation using tomographic ground penetrating radar travel time data through Bayesian inversion, which is integrated with entropy memory function and pilot point concepts, as well as efficient sampling approaches. It is critical to accurately estimate soil moisture content and variations in vadose zone studies. Many studies have illustrated the promise and value of GPR tomographic data for estimating soil moisture and associated changes, however, challenges still exist in the inversion of GPR tomographic data in a manner that quantifies input and predictive uncertainty, incorporates multiple data types, handles non-uniqueness and nonlinearity, and honors time-lapse tomograms collected in a series. To address these challenges, we develop a minimum relative entropy (MRE)-Bayesian based inverse modeling framework that non-subjectively defines prior probabilities, incorporates information from multiple sources, and quantifies uncertainty. The framework enables us to estimate dielectric permittivity at pilot point locations distributed within the tomogram, as well as the spatial correlation range. In the inversion framework, MRE is first used to derive prior probability density functions (pdfs) of dielectric permittivity based on prior information obtained from a straight-ray GPR inversion. The probability distributions are then sampled using a Quasi-Monte Carlo (QMC) approach, and the sample sets provide inputs to a sequential Gaussian simulation (SGSIM) algorithm that constructs a highly resolved permittivity/velocity field for evaluation with a curved-ray GPR forward model. The likelihood functions are computed as a function of misfits, and posterior pdfs are constructed using a Gaussian kernel. Inversion of subsequent time-lapse datasets combines the Bayesian estimates from the previous inversion (as a memory function) with new data. The memory function and pilot point design takes advantage of

  2. Highlights from the SoilCAM project: Soil Contamination, Advanced integrated characterisation and time-lapse Monitoring

    Science.gov (United States)

    French, H. K.; van der Zee, S. E. A. T. M.; Wehrer, M.; Godio, A.; Pedersen, L. B.; Toscano, G.

    2012-04-01

    The SoilCAM project (Soil Contamination, Advanced integrated characterisation and time-lapse Monitoring 2008-2012, EU-FP7-212663) is aimed at improving current methods for monitoring contaminant distribution and biodegradation in the subsurface. At two test sites, Oslo airport Gardermoen in Norway and the Trecate site in Italy, a number of geophysical techniques, lysimeter and other soil and water sampling techniques as well as numerical flow and transport modelling have been combined at different scales in order to characterise flow transport processes in the unsaturated and saturated zones. Laboratory experiments have provided data on physical and bio-geo-chemical parameters for use in models and to select remediation methods. The geophysical techniques were used to map geological heterogeneities and also conduct time-lapse measurements of processes in the unsaturated zone. Both cross borehole and surface electrodes were used for electrical resistivity and induced polarisation surveys. The geophysical surveys showed clear indications of areas highly affected by de-icing chemicals along the runway at Oslo airport. The time lapse measurements along the runway at the airport show infiltration patterns during snowmelt and are used to validate 2D unsaturated flow and transport simulations using SUTRA. The Orchestra model is used to describe the complex interaction between bio-geo-chemical processes in a 1D profile along the runway. The presence of installations such as a membrane along the runway highly affects the flow pattern and challenges the capacity of the numerical code. Smaller scale field site measurements have revealed the increase of iron and manganese during degradation of de-icing chemicals. The use of Nitrate to increase red-ox potential was tested, but results have not been analysed yet. So far it cannot be concluded that degradation process can be quantified indirectly by geophysical monitoring. At the Trecate site a combination of georadar, electrical

  3. Live Cells as Dynamic Laboratories: Time Lapse Raman Spectral Microscopy of Nanoparticles with Both IgE Targeting and pH-Sensing Functions

    Science.gov (United States)

    Nowak-Lovato, Kristy L.; Rector, Kirk D.

    2012-01-01

    This review captures the use of live cells as dynamic microlaboratories through implementation of labeled nanoparticles (nanosensors) that have both sensing and targeting functions. The addition of 2,4-ε-dinitrophenol-L-lysine (DNP) as a FcεRI targeting ligand and 4-mercaptopyridine (4-MPy) as a pH-sensing ligand enables spatial and temporal monitoring of FcεRI receptors and their pH environment within the endocytic pathway. To ensure reliability, the sensor is calibrated in vivo using the ionophore nigericin and standard buffer solutions to equilibrate the external [H+] concentration with that of the cell compartments. This review highlights the nanosensors, ability to traffic and respond to pH of receptor-bound nanosensors (1) at physiological temperature (37°C) versus room temperature (25°C), (2) after pharmacological treatment with bafilomycin, an H+ ATPase pump inhibitor, or amiloride, an inhibitor of Na+/H+ exchange, and (3) in response to both temperature and pharmacological treatment. Whole-cell, time lapse images are demonstrated to show the ability to transform live cells into dynamic laboratories to monitor temporal and spatial endosomal pH. The versatility of these probes shows promise for future applications relevant to intracellular trafficking and intelligent drug design. PMID:22778738

  4. Time-lapse camera observations of gas piston activity at Pu‘u ‘Ō‘ō, Kīlauea volcano, Hawai‘i

    Science.gov (United States)

    Orr, Tim R.; Rea, James

    2012-01-01

    Gas pistoning is a type of eruptive behavior described first at Kīlauea volcano and characterized by the (commonly) cyclic rise and fall of the lava surface within a volcanic vent or lava lake. Though recognized for decades, its cause continues to be debated, and determining why and when it occurs has important implications for understanding vesiculation and outgassing processes at basaltic volcanoes. Here, we describe gas piston activity that occurred at the Pu‘u ‘Ō‘ō cone, in Kīlauea’s east rift zone, during June 2006. Direct, detailed measurements of lava level, made from time-lapse camera images captured at close range, show that the gas pistons during the study period lasted from 2 to 60 min, had volumes ranging from 14 to 104 m3, displayed a slowing rise rate of the lava surface, and had an average gas release duration of 49 s. Our data are inconsistent with gas pistoning models that invoke gas slug rise or a dynamic pressure balance but are compatible with models which appeal to gas accumulation and loss near the top of the lava column, possibly through the generation and collapse of a foam layer.

  5. Micromechanical Time-Lapse X-ray CT Study of Fatigue Damage in Uni-Directional Fibre Composites

    DEFF Research Database (Denmark)

    Jespersen, Kristine Munk; Lowe, Tristan; Withers, Philip J.

    2015-01-01

    This study considers fatigue damage evolution in a uni-directional (UD) glass fibre composite used for wind turbine blades which is manufactured from a non-crimp fabric. It is the initial part of a time-lapse study where the damage progression is followed in a sample during a fatigue test....... In the current study 3D X-ray Computed Tomography (XCT) is used to characterise the fatigue damage in the material at three different stages of the fatigue life of a tension-tension fatigue test. 3D XCT is performed on rectangular samples (4x4x110mm) cut out from pre-fatigued full-size fatigue test specimens...

  6. Preliminary observations on polar body extrusion and pronuclear formation in human oocytes using time-lapse video cinematography.

    Science.gov (United States)

    Payne, D; Flaherty, S P; Barry, M F; Matthews, C D

    1997-03-01

    In this study, we have used time-lapse video cinematography to study fertilization in 50 human oocytes that had undergone intracytoplasmic sperm injection (ICSI). Time-lapse recording commenced shortly after ICSI and proceeded for 17-20 h. Oocytes were cultured in an environmental chamber which was maintained under standard culture conditions. Overall, 38 oocytes (76%) were fertilized normally, and the fertilization rate and embryo quality were not significantly different from 487 sibling oocytes cultured in a conventional incubator. Normal fertilization followed a defined course of events, although the timing of these events varied markedly between oocytes. In 35 of the 38 fertilized oocytes (92%), there were circular waves of granulation within the ooplasm which had a periodicity of 20-53 min. The sperm head decondensed during this granulation phase. The second polar body was then extruded, and this was followed by the central formation of the male pronucleus. The female pronucleus formed in the cytoplasm adjacent to the second polar body at the same time as, or slightly after, the male pronucleus, and was subsequently drawn towards the male pronucleus until the two abutted. Both pronuclei then increased in size, the nucleoli moved around within the pronuclei and some nucleoli coalesced. During pronuclear growth, the organelles contracted from the cortex towards the centre of the oocyte, leaving a clear cortical zone. The oocyte decreased in diameter from 112 to 106 microm (P cinematography is an excellent tool for studying fertilization and early embryo development, and have demonstrated that human fertilization comprises numerous complex dynamic events.

  7. Time-lapse ERT and DTS for seasonal and short-term monitoring of an alpine river hyporheic zone

    Science.gov (United States)

    Boaga, Jacopo; Laura, Busato; Mariateresa, Perri; Giorgio, Cassiani

    2016-04-01

    The hyporheic zone (HZ) is the area located beneath and adjacent to rivers and streams, where the interactions between surface water and groundwater take place. This complex physical domain allows the transport of several substances from a stream to the unconfined aquifer below, and vice versa, thus playing a fundamental role in the river ecosystem. The importance of the hyporheic zone makes its characterization a goal shared by several disciplines, which range from applied geophysics to biogeochemistry, from hydraulics to ecology. The frontier field of HZ characterization stays in applied non-invasive methodologies as Electrical Resistivity Tomography - ERT - and Distributed Temperature Sensing - DTS. ERT is commonly applied in cross-well configuration or with a superficial electrodes deployment while DTS is used in hydro-geophysics in the last decade, revealing a wide applicability to the typical issues of this field of study. DTS for hydro-geophysics studies is based on Raman scattering and employs heat as tracer and uses a fiber-optic cable to acquire temperature values. We applied both techniques for an alpine river case studies located in Val di Sole, TN, Italy. The collected measurements allow high-resolution characterization of the hyporheic zone, overcoming the critical problem of invasive measurements under riverbeds. In this work, we present the preliminary results regarding the characterization of the hyporheic zone of the alpine river obtained combining ERT and DTS time-lapse measurements. The data collection benefits from an innovative instrumentation deployment, which consists of both an ERT multicore cable and a DTS fiber-optic located in two separated boreholes drilled 5m under the watercourse and perpendicular to it. In particular we present the first year monitoring results and a short time-lapse monitoring experiment conducted during summer 2015. The site and the results here described are part of the EU FP7 CLIMB (Climate Induced Changes on the

  8. Pembina Cardium CO2-EOR monitoring project: Integrated surface seismic and VSP time-lapse seismic analysis

    Science.gov (United States)

    Alshuhail, A. A.

    2009-12-01

    In the Pembina field in west-central Alberta, Canada, approximately 40,000 tons of supercritical CO2 was injected into the 1650 m deep, 20 m thick upper-Cretaceous Cardium Fm. between March 2005 and 2007. A time-lapse seismic program was designed and incorporated into the overall measurement, monitoring and verification program. The objectives were to track the CO2 plume within the reservoir, and to evaluate the integrity of storage. Fluid replacement modeling predicts a decrease in the P-wave velocity and bulk density in the reservoir by about 4% and 1%, respectively. Synthetic seismograms show subtle reflectivity changes at the Cardium Fm. and a traveltime delay at the later high-amplitude Viking event of less than 1 ms. The time-lapse datasets, however, show no significant anomalies in the P-wave seismic data that can be attributed to supercritical CO2 injected into the Cardium Fm. (Figure 1). The converted-wave (P-S) data, on the other hand, showed small traveltime anomalies. The most coherent results were those obtained by the fixed-array VSP dataset (Figure 2) due to higher frequency bandwidth and high signal to noise ratio. The amplitude and traveltime changes observed in the VSP dataset are small but are consistent in magnitude with those predicted from rock physics modeling. The analysis suggests that the inability to clearly detect the CO2 plume in surface seismic data is likely due to the CO2 being contained in thin permeable sandstone members of the Cardium Formation. The seismic signature of the Cardium Fm. in this area may also be degraded by multiples and strong attenuation involving the shallow Ardley coals. However, the lack of a 4D seismic changes above the reservoir indicates that the injected CO2 is not migrating through the caprock into shallower formations.

  9. Analysis of Snow Line and Albedo Conditions By Means of Time-Lapse Photography on Tapado Glacier, Chile

    Science.gov (United States)

    Vivero, S.; MacDonell, S.; McPhee, J. P.

    2014-12-01

    In the semiarid Coquimbo Region of Chile, high-altitude glaciers and seasonal snow are important sources of freshwater for irrigated agriculture and urban consumption. Due to the aridity of the environment, losses due to sublimation are large which means that accurate melt modelling is essential in order to reliably estimate streamflow. Since 2008, the CEAZA glaciology group has been studying the energy and mass balance of the largest glacier in the catchment, the Tapado Glacier using field and remote sensing measurements, and numerical modelling. The Tapado glacier system (30°08' S, 69°55' W) is a complex assemblage of uncovered and debris-covered ice located at the head of the Elqui basin between 4500 and 5536 m a.s.l. Energy balance modelling studies at the site have been limited in scope due to the development of ice pinnacles or penitentes on snow and ice surfaces. These features complicate energy distribution across the surface, due to modifications of parameters such as albedo. In this paper, we use time-lapse photography and automatic weather station (AWS) measurements to investigate how the development of penitentes impacts the spatial and temporal variability of albedo across the glacier surface and whether terrestrial photography is appropriate for use at such locations. Oblique photographs obtained from a high vantage point were georeferenced using a high resolution digital elevation model available for the entire glacier and its environs. By comparing the photographic data with point albedo measurements made at an AWS, distributed albedo maps were produced. Preliminary results suggest that distributed albedo values may be underestimated by the formation and development of penitentes during the ablation season. Moreover, it was observed that the evolution of the snow line during summer was not only topographically controlled but also modified by occasional convective snowfalls. Time-lapse photography provided to be a cost-effective tool for monitoring

  10. Several Organic Salts with High Two-Photon Active

    Institute of Scientific and Technical Information of China (English)

    TIAN, Yu-Peng; JIANG, Min-Hua; WANG, He-Zhou; FANG, Qi

    2001-01-01

    Several organic salts with D-A molecular structure and different counterion have been prepared and experimentally investigated. The two-photon induced frequency-upconverted spectra and two-photon pumped lasing are measured for the organic salt solutions in various solvents. The results indicate that counterions have influence on their stability and lasing property.

  11. Two-photon absorption in arsenic sulfide glasses

    Science.gov (United States)

    Chunaev, D. S.; Snopatin, G. E.; Plotnichenko, V. G.; Karasik, A. Ya.

    2016-10-01

    The two-photon absorption coefficient of 1047-{\\text{nm}} light in {\\text{As}}35{\\text{S}}65 chalcogenide glass has been measured. CW probe radiation has been used to observe the linear absorption in glass induced by two-photon excitation. The induced absorption lifetime was found to be ∼ 2 {\\text{ms}}.

  12. Examining the information content of time-lapse crosshole GPR data collected under different infiltration conditions to estimate unsaturated soil hydraulic properties

    DEFF Research Database (Denmark)

    Scholer, M.; Irving, J.; Zibar, Majken Caroline Looms

    2013-01-01

    by natural or artificial means, have been demonstrated in a number of studies to contain valuable information concerning the hydraulic properties of the unsaturated zone. Previous work in this domain has considered a variety of infiltration conditions and different amounts of time-lapse GPR data......Time-lapse geophysical data acquired during transient hydrological experiments are being increasingly employed to estimate subsurface hydraulic properties at the field scale. In particular, crosshole ground-penetrating radar (GPR) data, collected while water infiltrates into the subsurface either...... subsurface hydraulic properties as a function of depth, with forced infiltration offering the greatest potential for VGM parameter refinement because of the higher stressing of the hydrological system. Considering greater amounts of time-lapse data in the inversion procedure is also found to help refine VGM...

  13. The development of efficient two-photon singlet oxygen sensitizers

    DEFF Research Database (Denmark)

    Nielsen, Christian Benedikt

    The development of efficient two-photon singlet oxygen sensitizers is addressed focusing on organic synthesis. Photophysical measurements were carried out on new lipophilic molecules, where two-photon absorption cross sections and singlet oxygen quantumyields were measured. Design principles...... for making efficient two-photon singlet oxygen sensitizers were then constructed from these results. Charge-transfer in the excited state of the prepared molecules was shown to play a pivotal role in the generationof singlet oxygen. This was established through studies of substituent effects on both...... the singlet oxygen yield and the two-photon absorption cross section, where it was revealed that a careful balancing of the amount of charge transfer present in theexcited state of the sensitizer is necessary to obtain both a high singlet oxygen quantum yield and a high two-photon cross section. An increasing...

  14. Individual cell motility studied by time-lapse video recording: influence of experimental conditions

    DEFF Research Database (Denmark)

    Hartmann-Petersen, R; Walmod, P S; Berezin, A

    2000-01-01

    BACKGROUND: Eukaryotic cell motility plays a key role during development, wound healing, and tumour invasion. Computer-assisted image analysis now makes it a realistic task to quantify individual cell motility of a large number of cells. However, the influence of culture conditions before...... line. Cellular morphology and organization of filamentous actin were assessed by means of phase-contrast and confocal laser scanning microscopy and compared to the corresponding motility data. RESULTS: Cell dissociation procedure, seeding density, time of cultivation, and substrate concentration were...

  15. Time-resolved two-photon photoemission from metal surfaces

    CERN Document Server

    Weinelt, M

    2002-01-01

    The Rydberg-like series of image-potential states is a prototype system for loosely bound electrons at a metal surface. The electronic structure and the femtosecond dynamics of these states is studied by high-resolution energy-and time-resolved two-photon photoemission spectroscopy. The electron trapped in the image potential moves virtually freely laterally to the surface where it is subject to inelastic and quasielastic scattering processes which cause decay of population and phase relaxation. The influence of surface corrugation on these processes has been investigated for adsorbates on Cu(001) and stepped Cu(117) and Cu(119) surfaces which are vicinal to Cu(001). The dynamics depend on both the distance of the electron in front of the surface and the parallel momentum. For CO molecules on Cu(001) inelastic scattering into bulk states and adsorbate-induced resonances determine the decay rate. For small numbers of Cu adatoms on Cu(001) and the vicinal surfaces the decay rate of image-potential states is sig...

  16. Two-photon holographic optogenetics of neural circuits (Conference Presentation)

    Science.gov (United States)

    Yang, Weijian; Carrillo-Reid, Luis; Peterka, Darcy S.; Yuste, Rafael

    2016-03-01

    Optical manipulation of in vivo neural circuits with cellular resolution could be important for understanding cortical function. Despite recent progress, simultaneous optogenetic activation with cellular precision has either been limited to 2D planes, or a very small numbers of neurons over a limited volume. Here we demonstrate a novel paradigm for simultaneous 3D activation using a low repetition rate pulse-amplified fiber laser system and a spatial light modulator (SLM) to project 3D holographic excitation patterns on the cortex of mice in vivo for targeted volumetric 3D photoactivation. This method is compatible with two-photon imaging, and enables the simultaneous activation of multiple cells in 3D, using red-shifted opsins, such as C1V1 or ReaChR, while simultaneously imaging GFP-based sensors such as GCaMP6. This all-optical imaging and 3D manipulation approach achieves simultaneous reading and writing of cortical activity, and should be a powerful tool for the study of neuronal circuits.

  17. Time-lapse electrical resistivity anomalies due to contaminant transport around landfills

    Directory of Open Access Journals (Sweden)

    J. Yang

    2007-06-01

    Full Text Available The extent of landfill leachate can be delineated by geo-electrical imaging as a response to the varying electrical resistivity in the contaminated area. This research was based on a combination of hydrogeological numerical simulation followed by geophysical forward and inversion modeling performed to evaluate the migration of a contaminant plume from a landfill. As a first step, groundwater flow and contaminant transport was simulated using the finite elements numerical modeling software FEFLOW. The extent of the contaminant plume was acquired through a hydrogeological model depicting the distributions of leachate concentration in the system. Next, based on the empirical relationship between the concentration and electrical conductivity of the leachate in the porous media, the corresponding geo-electrical structure was derived from the hydrogeological model. Finally, forward and inversion computations of geo-electrical anomalies were performed using the finite difference numerical modeling software DCIP2D/DCIP3D. The image obtained by geophysical inversion of the electric data was expected to be consistent with the initial hydrogeological model, as described by the distribution of leachate concentration. Numerical case studies were conducted for various geological conditions, hydraulic parameters and electrode arrays, from which conclusions were drawn regarding the suitability of the methodology to assess simple to more complex geo-electrical models. Thus, optimal mapping and monitoring configurations were determined.

  18. Numerical modeling of time-lapse monitoring of CO2 sequestration in a layered basalt reservoir

    Science.gov (United States)

    Khatiwada, M.; Van Wijk, K.; Clement, W.P.; Haney, M.

    2008-01-01

    As part of preparations in plans by The Big Sky Carbon Sequestration Partnership (BSCSP) to inject CO2 in layered basalt, we numerically investigate seismic methods as a noninvasive monitoring technique. Basalt seems to have geochemical advantages as a reservoir for CO2 storage (CO2 mineralizes quite rapidly while exposed to basalt), but poses a considerable challenge in term of seismic monitoring: strong scattering from the layering of the basalt complicates surface seismic imaging. We perform numerical tests using the Spectral Element Method (SEM) to identify possibilities and limitations of seismic monitoring of CO2 sequestration in a basalt reservoir. While surface seismic is unlikely to detect small physical changes in the reservoir due to the injection of CO2, the results from Vertical Seismic Profiling (VSP) simulations are encouraging. As a perturbation, we make a 5%; change in wave velocity, which produces significant changes in VSP images of pre-injection and post-injection conditions. Finally, we perform an analysis using Coda Wave Interferometry (CWI), to quantify these changes in the reservoir properties due to CO2 injection.

  19. Near infrared two-photon excitation cross-sections of voltage-sensitive dyes.

    Science.gov (United States)

    Fisher, Jonathan A N; Salzberg, Brian M; Yodh, Arjun G

    2005-10-15

    Microscopy based on voltage-sensitive dyes has proven effective for revealing spatio-temporal patterns of neuronal activity in vivo and in vitro. Two-photon microscopy using voltage-sensitive dyes offers the possibility of wide-field visualization of membrane potential on sub-cellular length scales, hundreds of microns below the tissue surface. Very little information is available, however, about the utility of voltage-sensitive dyes for two-photon imaging purposes. Here we report on measurements of two-photon fluorescence excitation cross-sections for nine voltage-sensitive dyes in a solvent, octanol, intended to simulate the membrane environment. Ultrashort light pulses from a Ti:sapphire laser were used for excitation from 790 to 960 nm, and fluorescein dye was used as a calibration standard. Overall, dyes RH795, RH421, RH414, di-8-ANEPPS, and di-8-ANEPPDHQ had the largest two-photon excitation cross-sections ( approximately 15 x 10(-50)cm4 s photon(-1)) in this wavelength region and are therefore potentially useful for two-photon microscopy. Interestingly, di-8-ANEPPDHQ, a chimera constructed from the potentiometric dyes RH795 and di-8-ANEPPS, exhibited larger cross-sections than either of its constituents.

  20. Insights into esophagus tissue architecture using two-photon confocal microscopy

    Science.gov (United States)

    Liu, Nenrong; Wang, Yue; Feng, Shangyuan; Chen, Rong

    2013-08-01

    In this paper, microstructures of human esophageal mucosa were evaluated using the two-photon laser scanning confocal microscopy (TPLSCM), based on two-photon excited fluorescence (TPEF) and second harmonic generation (SHG). The distribution of epithelial cells, muscle fibers of muscularis mucosae has been distinctly obtained. Furthermore, esophageal submucosa characteristics with cancer cells invading into were detected. The variation of collagen, elastin and cancer cells is very relevant to the pathology in esophagus, especially early esophageal cancer. Our experimental results indicate that the MPM technique has the much more advantages for label-free imaging, and has the potential application in vivo in the clinical diagnosis and monitoring of early esophageal cancer.

  1. Near IR two photon absorption of cyanines dyes: application to optical power limiting at telecommunication wavelengths

    Science.gov (United States)

    Bouit, Pierre-Antoine; Wetzel, Guillaume; Feneyrou, Patrick; Bretonnière, Yann; Kamada, Kenji; Maury, Olivier; Andraud, Chantal

    2008-02-01

    The design and synthesis of symmetrical and unsymmetrical heptamethine cyanines is reported. These chromophores present significant two-photon cross section in the 1400-1600 nm spectral range. In addition, they display optical power limiting (OPL) properties. OPL curves were interpreted on the basis of two-photon absorption (2PA) followed by excited state absorption (ESA). Finally, these molecules present several relevant properties (nonlinear absorption properties, two-step gram scale synthesis, high solubility, good thermal stability), which could lead to numerous practical applications in material science (solid state optical limiting, signal processing) or in biology (imaging).

  2. New insight in boron chemistry: Application in two-photon absorption

    Science.gov (United States)

    Bolze, F.; Hayek, A.; Sun, X. H.; Baldeck, P. L.; Bourgogne, C.; Nicoud, J.-F.

    2011-07-01

    Two groups of one-dimensional (1D) boron containing two-photon absorbing fluorophores have been prepared and characterized. One group includes boron atoms incorporated in the conjugated or pseudo conjugated central core and the other contain a boron cluster as an acceptor group at one end of the fluorophores. Two boron containing central cores (with two boron atoms) have been explored: the cyclodiborazane and the pyrazabole moieties. The chosen boron cluster, p-carborane, contains 10 boron atoms. All the prepared fluorophores present high two-photon absorption cross-sections. Some water-soluble as well as lipophylic dyes have been prepared and used in bio-imaging.

  3. Fluorenyl porphyrins for combined two-photon excited fluorescence and photosensitization

    Science.gov (United States)

    Mongin, Olivier; Hugues, Vincent; Blanchard-Desce, Mireille; Merhi, Areej; Drouet, Samuel; Yao, Dandan; Paul-Roth, Christine

    2015-04-01

    The two-photon absorption (2PA), the luminescence and the photosensitization properties of porphyrin-cored fluorenyl dendrimers and meso-substituted fluorenylporphyrin monomer, dimer and trimer are described. In comparison with model tetraphenylporphyrin, these compounds combine enhanced (non-resonant) 2PA cross-sections in the near infrared and enhanced fluorescence quantum yields, together with maintained singlet oxygen generation quantum yields. 'Semi-disconnection' between fluorenyl groups and porphyrins (i.e. direct meso substitution) proved to be more efficient than non-conjugated systems (based on efficient FRET between fluorenyl antennae and porphyrins). These results are of interest for combined two-photon imaging and photodynamic therapy.

  4. Time-lapsed assessment of microcrack initiation and propagation in murine cortical bone at submicrometer resolution.

    Science.gov (United States)

    Voide, R; Schneider, P; Stauber, M; Wyss, P; Stampanoni, M; Sennhauser, U; van Lenthe, G H; Müller, R

    2009-08-01

    The strength of bone tissue is not only determined by its mass, but also by other properties usually referred to as bone quality, such as microarchitecture, distribution of bone cells, or microcracks and damage. It has been hypothesized that the bone ultrastructure affects microcrack initiation and propagation. Due to its high resolution, bone assessment by means of synchrotron radiation (SR)-based computed tomography (CT) allows unprecedented three-dimensional (3D) and non-invasive insights into ultrastructural bone phenotypes, such as the canal network and the osteocyte lacunar system. The aims of this study were to describe the initiation and propagation of microcracks and their relation with these ultrastructural phenotypes. To this end, femora from the two genetically distinct inbred mouse strains C3H/He (C3H) and C57BL/6 (B6) were loaded axially under compression, from 0% strain to failure, with 1% strain steps. Between each step, a high-resolution 3D image (700 nm nominal resolution) was acquired at the mid-diaphysis using SR CT for characterization and quantitative analysis of the intracortical porosity, namely the bone canal network, the osteocyte lacunar system and the emerging microcracks. For C3H mice, the canal, lacunar, and microcrack volume densities accounted typically for 1.91%, 2.11%, and 0.27% of the cortical total volume at 2% apparent strain, respectively. Due to its 3D nature, SR CT allowed to visualize and quantify also the volumetric extent of microcracks. At 2% apparent strain, the average microcrack thickness for both mouse strains was 2.0 microm for example. Microcracks initiated at canal and at bone surfaces, whereas osteocyte lacunae provided guidance to the microcracks. Moreover, we observed that microcracks could appear as linear cracks in one plane, but as diffuse cracks in a perpendicular plane. Finally, SR CT images permitted visualization of uncracked ligament bridging, which is thought to be of importance in bone toughening

  5. Dependence of the two-photon photoluminescence yield of gold nanostructures on the laser pulse duration

    DEFF Research Database (Denmark)

    Biagioni, P.; Celebrano, M.; Savoini, M.

    2009-01-01

    Two-photon photoluminescence (TPPL) from gold nanostructures is becoming one of the most relevant tools for plasmon-assisted biological imaging and photothermal therapy as well as for the investigation of plasmonic devices. Here we study the yield of TPPL as a function of the temporal width δ of ...

  6. Two-Photon Absorption Properties of Mn-Doped ZnS Quantum Dots

    Institute of Scientific and Technical Information of China (English)

    ZHENG Jia-Jin; ZHANG Gui-Lan; GUO Yang-Xue; WANG Xiao-Yan; CHEN Wen-Ju; ZHANG Xiao-Song; HUA Yu-Lin

    2006-01-01

    @@ We investigate the two-photon absorption and nonlinear refractive index properties of a quantum dot material based on ZnS nanocrystals doped with Mn isoelectronic impurities, using the Z-scan technique with 532nm picosecond laser pulses. The Mn-doped ZnS quantum dots have an average two-photon absorption cross section as high as 13600 Goeppert-Mayer units, which turn it into a very promising material for fluorescent label and imaging in biological samples. In addition, we also found that the two-photon absorption coeflicient initially increases and then decreases with increasing pulse irradiance, which demonstrates the presence of the higherorder nonlinearity under the strong excitation.

  7. Synthesis of Two-Photon Materials and Two-Photon Liquid Crystals

    Science.gov (United States)

    Subramaniam, Girija

    2001-01-01

    The duration of the grant was interrupted by two major accidents that the PI met with-- an auto accident in Pasadena, CA during her second summer at JPL which took almost eight months for recovery and a second accident during Fall 2000 that left her in crutches for the entire semester. Further, the time released agreed by the University was not given in a timely fashion. The candidate has been given post-grant expire time off. In spite of all these problems, the PI synthesized a number of new two-photon materials and studied the structure-activity correlation to arrive at the best-optimized structure. The PI's design proved to be one of the best in the sense that these materials has a hitherto unreported two-photon absorption cross section. Many materials based on PI's design was later made by the NASA colleague. This is Phase 1. Phase II of this grant is to orate liquid crystalline nature into this potentially useful materials and is currently in progress. Recent observations of nano- and pico-second response time of homeotropically aligned liquid crystals suggest their inherent potentials to act as laser hardening materials, i.e., as protective devices against short laser pulses. The objective of the current project is to exploit this potential by the synthesis of liquid crystals with high optical nonlinearity and optimizing their performance. The PI is trying structural variations to bring in liquid crystalline nature without losing the high two-photon cross section. Both Phase I and Phase II led to many invited presentations and publications in reputed journals like 'Science' and 'Molecular Crystals'. The list of presentations and reprints are enclosed. Another important and satisfying outcome of this grant is the opportunity that this grant offered to the budding undergraduate scientists to get involved in a visible research of international importance. All the students had a chance to learn a lot during research, had the opportunity to present their work at

  8. Two-photon processes in highly charged ions

    Energy Technology Data Exchange (ETDEWEB)

    Jahrsetz, Thorsten

    2015-03-05

    Two-photon processes are atomic processes in which an atom interacts simultaneously with two photons. Such processes describe a wide range of phenomena, such as two-photon decay and elastic or inelastic scattering of photons. In recent years two-photon processes involving highly charged heavy ions have become an active area of research. Such studies do not only consider the total transition or scattering rates but also their angular and polarization dependence. To support such examinations in this thesis I present a theoretical framework to describe these properties in all two-photon processes with bound initial and final states and involving heavy H-like or He-like ions. I demonstrate how this framework can be used in some detailed studies of different two-photon processes. Specifically a detailed analysis of two-photon decay of H-like and He-like ions in strong external electromagnetic fields shows the importance of considering the effect of such fields for the physics of such systems. Furthermore I studied the elastic Rayleigh as well as inelastic Raman scattering by heavy H-like ions. I found a number of previously unobserved phenomena in the angular and polarization dependence of the scattering cross-sections that do not only allow to study interesting details of the electronic structure of the ion but might also be useful for the measurement of weak physical effects in such systems.

  9. Two-photon interference of temporally separated photons

    Science.gov (United States)

    Kim, Heonoh; Lee, Sang Min; Moon, Han Seb

    2016-10-01

    We present expe