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fluorescence lifetime imaging microscopy

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https://www.readbyqxmd.com/read/28211922/applying-phasor-approach-analysis-of-multiphoton-flim-measurements-to-probe-the-metabolic-activity-of-three-dimensional-in-vitro-cell-culture-models
#1
Pirmin H Lakner, Michael G Monaghan, Yvonne Möller, Monilola A Olayioye, Katja Schenke-Layland
Fluorescence lifetime imaging microscopy (FLIM) can measure and discriminate endogenous fluorophores present in biological samples. This study seeks to identify FLIM as a suitable method to non-invasively detect a shift in cellular metabolic activity towards glycolysis or oxidative phosphorylation in 3D Caco-2 models of colorectal carcinoma. These models were treated with potassium cyanide or hydrogen peroxide as controls, and epidermal growth factor (EGF) as a physiologically-relevant influencer of cell metabolic behaviour...
February 13, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28199849/quantitative-flim-fret-microscopy-to-monitor-nanoscale-chromatin-compaction-in%C3%A2-vivo-reveals-structural-roles-of-condensin-complexes
#2
David Llères, Aymeric P Bailly, Aurélien Perrin, David G Norman, Dimitris P Xirodimas, Robert Feil
How metazoan genomes are structured at the nanoscale in living cells and tissues remains unknown. Here, we adapted a quantitative FRET (Förster resonance energy transfer)-based fluorescence lifetime imaging microscopy (FLIM) approach to assay nanoscale chromatin compaction in living organisms. Caenorhabditis elegans was chosen as a model system. By measuring FRET between histone-tagged fluorescent proteins, we visualized distinct chromosomal regions and quantified the different levels of nanoscale compaction in meiotic cells...
February 14, 2017: Cell Reports
https://www.readbyqxmd.com/read/28198734/new-frontiers-in-intravital-microscopy-of-the-kidney
#3
Andrew M Hall, Claus D Schuh, Dominik Haenni
PURPOSE OF REVIEW: Intravital imaging with multiphoton microscopy enables the detailed study of dynamic cellular processes within functioning organs in living animals, in ways that would not otherwise be possible. It therefore represents a powerful tool in translational kidney research. In this article, we will discuss several new technical developments that have significantly increased the capabilities of kidney imaging. RECENT FINDINGS: Important contemporary advances in biomedical imaging technology include longer wavelength excitation lasers, far-red emitting fluorescent reporters, highly sensitive detectors, fluorescence lifetime measurements, adaptive optics, microendoscopes, high-throughput automated analysis algorithms and tissue clearing techniques...
February 13, 2017: Current Opinion in Nephrology and Hypertension
https://www.readbyqxmd.com/read/28193235/dynamic-presenilin-1-and-synaptotagmin-1-interaction-modulates-exocytosis-and-amyloid-%C3%AE-production
#4
Katarzyna Marta Zoltowska, Masato Maesako, Iryna Lushnikova, Shuko Takeda, Laura J Keller, Galina Skibo, Bradley T Hyman, Oksana Berezovska
BACKGROUND: Alzheimer's disease (AD)-linked protein, presenilin 1 (PS1), is present at the synapse, and the knock-out of presenilin in mice leads to synaptic dysfunction. On the other hand, synaptic activity was shown to influence PS1-dependent generation of distinct amyloid β (Aβ) species. However, the precise nature of these regulations remains unclear. The current study reveals novel role of PS1 at the synapse, and deciphers how PS1 and synaptic vesicle-associated protein, synaptotagmin 1 (Syt1) modulate each other functions in neurons via direct activity-triggered interaction...
February 13, 2017: Molecular Neurodegeneration
https://www.readbyqxmd.com/read/28168255/do-grain-boundaries-dominate-non-radiative-recombination-in-ch3nh3pbi3-perovskite-thin-films
#5
Mengjin Yang, Yining Zeng, Zhen Li, Dong Hoe Kim, Chun-Sheng Jiang, Jao van de Lagemaat, Kai Zhu
Here, we examine grain boundaries (GBs) with respect to non-GB regions (grain surfaces (GSs) and grain interiors (GIs)) in high-quality micrometer-sized perovskite CH3NH3PbI3 (or MAPbI3) thin films using high-resolution confocal fluorescence-lifetime imaging microscopy in conjunction with kinetic modeling of charge-transport and recombination processes. We show that, contrary to previous studies, GBs in our perovskite MAPbI3 thin films do not lead to increased recombination but that recombination in these films happens primarily in the non-GB regions (i...
February 7, 2017: Physical Chemistry Chemical Physics: PCCP
https://www.readbyqxmd.com/read/28134273/imaging-tumor-microscopic-viscosity-in-vivo-using-molecular-rotors
#6
Lyubov' E Shimolina, Maria Angeles Izquierdo, Ismael López-Duarte, James A Bull, Marina V Shirmanova, Larisa G Klapshina, Elena V Zagaynova, Marina K Kuimova
The microscopic viscosity plays an essential role in cellular biophysics by controlling the rates of diffusion and bimolecular reactions within the cell interior. While several approaches have emerged that have allowed the measurement of viscosity and diffusion on a single cell level in vitro, the in vivo viscosity monitoring has not yet been realized. Here we report the use of fluorescent molecular rotors in combination with Fluorescence Lifetime Imaging Microscopy (FLIM) to image microscopic viscosity in vivo, both on a single cell level and in connecting tissues of subcutaneous tumors in mice...
January 30, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28129796/two-photon-flim-of-nad-p-h-and-fad-in-mesenchymal-stem-cells-undergoing-either-osteogenic-or-chondrogenic-differentiation
#7
Aleksandra V Meleshina, Varvara V Dudenkova, Alena S Bystrova, Daria S Kuznetsova, Marina V Shirmanova, Elena V Zagaynova
BACKGROUND: Metabolic plasticity and the versatility of different lineages of stem cells as they satisfy their energy demands are not completely understood. In this study we investigated the metabolic changes in mesenchymal stem cells (MSCs) undergoing differentiation in two directions, osteogenic and chondrogenic, using two-photon fluorescence microscopy combined with FLIM. METHODS: Differentiation was induced by incubating the human bone marrow MSCs in osteogenic or chondrogenic mediums...
January 28, 2017: Stem Cell Research & Therapy
https://www.readbyqxmd.com/read/28125231/improved-fluorescent-protein-contrast-and-discrimination-by-optically-controlling-dark-state-lifetimes
#8
Yen-Cheng Chen, Robert M Dickson
Modulation and optical control of photoswitchable fluorescent protein (PS-FP) dark state lifetimes drastically improves sensitivity and selectivity in fluorescence imaging. The dark state population of PS-FPs generates an out-of-phase fluorescence component relative to the sinusoidally modulated 488 nm laser excitation. Because this apparent phase advanced emission results from slow recovery to the fluorescent manifold, we hasten recovery and, therefore, modulation frequency by varying coillumination intensity at 405 nm...
February 16, 2017: Journal of Physical Chemistry Letters
https://www.readbyqxmd.com/read/28115230/time-resolved-fluorescence-microscopy-flim-as-an-analytical-tool-in-skin-nanomedicine
#9
REVIEW
Ulrike Alexiev, Pierre Volz, Alexander Boreham, Robert Brodwolf
The emerging field of nanomedicine provides new approaches for the diagnosis and treatment of diseases, for symptom relief, and for monitoring of disease progression. Topical application of drug-loaded nanoparticles for the treatment of skin disorders is a promising strategy to overcome the stratum corneum, the upper layer of the skin, which represents an effective physical and biochemical barrier. The understanding of drug penetration into skin and enhanced penetration into skin facilitated by nanocarriers requires analytical tools that ideally allow to visualize the skin, its morphology, the drug carriers, drugs, their transport across the skin and possible interactions, as well as effects of the nanocarriers within the different skin layers...
January 21, 2017: European Journal of Pharmaceutics and Biopharmaceutics
https://www.readbyqxmd.com/read/28114413/visualization-of-bri1-and-serk3-bak1-nanoclusters-in-arabidopsis-roots
#10
Stefan J Hutten, Danny S Hamers, Marije Aan den Toorn, Wilma van Esse, Antsje Nolles, Christoph A Bücherl, Sacco C de Vries, Johannes Hohlbein, Jan Willem Borst
Brassinosteroids (BRs) are plant hormones that are perceived at the plasma membrane (PM) by the ligand binding receptor BRASSINOSTEROID-INSENSITIVE1 (BRI1) and the co-receptor SOMATIC EMBRYOGENESIS RECEPTOR LIKE KINASE 3/BRI1 ASSOCIATED KINASE 1 (SERK3/BAK1). To visualize BRI1-GFP and SERK3/BAK1-mCherry in the plane of the PM, variable-angle epifluorescence microscopy (VAEM) was employed, which allows selective illumination of a thin surface layer. VAEM revealed an inhomogeneous distribution of BRI1-GFP and SERK3/BAK1-mCherry at the PM, which we attribute to the presence of distinct nanoclusters...
2017: PloS One
https://www.readbyqxmd.com/read/28096507/de-novo-phosphorylation-and-conformational-opening-of-the-tyrosine-kinase-lck-act-in-concert-to-initiate-t-cell-receptor-signaling
#11
Lars Philipsen, Amarendra V Reddycherla, Roland Hartig, Janine Gumz, Matthias Kästle, Andreas Kritikos, Mateusz P Poltorak, Yury Prokazov, Evgeny Turbin, André Weber, Werner Zuschratter, Burkhart Schraven, Luca Simeoni, Andreas J Müller
The enzymatic activity of the Src family tyrosine kinase p56(Lck) (Lck) is tightly controlled by differential phosphorylation of two tyrosine residues, Tyr(394) and Tyr(505) Phosphorylation of Tyr(394) and the conformational opening of Lck are believed to activate the kinase, whereas Tyr(505) phosphorylation is thought to generate a closed, inactive conformation of Lck. We investigated whether the conformation of Lck and its phosphorylation state act in concert to regulate the initiation of T cell receptor (TCR) signaling...
January 17, 2017: Science Signaling
https://www.readbyqxmd.com/read/28073262/fluorescence-self-quenching-from-reporter-dyes-informs-on-the-structural-properties-of-amyloid-clusters-formed-in-vitro-and-in-cells
#12
WeiYue Chen, Laurence J Young, Meng Lu, Alessio Zaccone, Florian Ströhl, Na Yu, Gabriele S Kaminski Schierle, Clemens F Kaminski
The characterization of the aggregation kinetics of protein amyloids and the structural properties of the ensuing aggregates are vital in the study of the pathogenesis of many neurodegenerative diseases and the discovery of therapeutic targets. In this article, we show that the fluorescence lifetime of synthetic dyes covalently attached to amyloid proteins informs on the structural properties of amyloid clusters formed both in vitro and in cells. We demonstrate that the mechanism behind such a "lifetime sensor" of protein aggregation is based on fluorescence self-quenching and that it offers a good dynamic range to report on various stages of aggregation without significantly perturbing the process under investigation...
January 11, 2017: Nano Letters
https://www.readbyqxmd.com/read/28067047/solvatochromic-and-fluorogenic-dyes-as-environment-sensitive-probes-design-and-biological-applications
#13
Andrey S Klymchenko
Fluorescent environment-sensitive probes are specially designed dyes that change their fluorescence intensity (fluorogenic dyes) or color (e.g., solvatochromic dyes) in response to change in their microenvironment polarity, viscosity, and molecular order. The studies of the past decade, including those of our group, have shown that these molecules become universal tools in fluorescence sensing and imaging. In fact, any biomolecular interaction or change in biomolecular organization results in modification of the local microenvironment, which can be directly monitored by these types of probes...
January 9, 2017: Accounts of Chemical Research
https://www.readbyqxmd.com/read/28063999/relationship-between-intracellular-ph-metabolic-co-factors-and-caspase-3-activation-in-cancer-cells-during-apoptosis
#14
Tatiana F Sergeeva, Marina V Shirmanova, Olga A Zlobovskaya, Alena I Gavrina, Varvara V Dudenkova, Maria M Lukina, Konstantin A Lukyanov, Elena V Zagaynova
A complex cascade of molecular events occurs in apoptotic cells but cell-to-cell variability significantly complicates determination of the order and interconnections between different processes. For better understanding of the mechanisms of programmed cell death, dynamic simultaneous registration of several parameters is required. In this paper we used multiparameter fluorescence microscopy to analyze energy metabolism, intracellular pH and caspase-3 activation in living cancer cells in vitro during staurosporine-induced apoptosis...
January 4, 2017: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/28060890/developing-and-testing-a-bayesian-analysis-of-fluorescence-lifetime-measurements
#15
Bryan Kaye, Peter J Foster, Tae Yeon Yoo, Daniel J Needleman
FRET measurements can provide dynamic spatial information on length scales smaller than the diffraction limit of light. Several methods exist to measure FRET between fluorophores, including Fluorescence Lifetime Imaging Microscopy (FLIM), which relies on the reduction of fluorescence lifetime when a fluorophore is undergoing FRET. FLIM measurements take the form of histograms of photon arrival times, containing contributions from a mixed population of fluorophores both undergoing and not undergoing FRET, with the measured distribution being a mixture of exponentials of different lifetimes...
2017: PloS One
https://www.readbyqxmd.com/read/28059202/saturation-compensated-measurements-for-fluorescence-lifetime-imaging-microscopy
#16
Yide Zhang, Genevieve D Vigil, Lina Cao, Aamir A Khan, David Benirschke, Tahsin Ahmed, Patrick Fay, Scott S Howard
Fluorophore saturation is the key factor limiting the speed and excitation range of fluorescence lifetime imaging microscopy (FLIM). For example, fluorophore saturation causes incorrect lifetime measurements when using conventional frequency-domain FLIM at high excitation powers. In this Letter, we present an analytical theoretical description of this error and present a method for compensating for this error in order to extract correct lifetime measurements in the limit of fluorophore saturation. We perform a series of simulations and experiments to validate our methods...
January 1, 2017: Optics Letters
https://www.readbyqxmd.com/read/28042459/precision-targeted-ruthenium-ii-luminophores-highly-effective-probes-for-cell-imaging-by-stimulated-emission-depletion-sted-microscopy
#17
Aisling Byrne, Christopher S Burke, Tia E Keyes
Fluorescence microscopy has undergone a dramatic evolution over the past two decades with development of super-resolution far-field microscopy methods that break the light diffraction limited resolution of conventional microscopy, offering unprecedented opportunity to interrogate cellular processes at the nanoscale. However, these methods make special demands of the luminescent agents used for contrast and development of probes suited to super-resolution fluorescent methods is still relatively in its infancy...
October 19, 2016: Chemical Science
https://www.readbyqxmd.com/read/28029135/time-resolved-fluorescence-spectroscopy-and-fluorescence-lifetime-imaging-microscopy-for-characterization-of-dendritic-polymer-nanoparticles-and-applications-in-nanomedicine
#18
REVIEW
Alexander Boreham, Robert Brodwolf, Karolina Walker, Rainer Haag, Ulrike Alexiev
The emerging field of nanomedicine provides new approaches for the diagnosis and treatment of diseases, for symptom relief and for monitoring of disease progression. One route of realizing this approach is through carefully constructed nanoparticles. Due to the small size inherent to the nanoparticles a proper characterization is not trivial. This review highlights the application of time-resolved fluorescence spectroscopy and fluorescence lifetime imaging microscopy (FLIM) for the analysis of nanoparticles, covering aspects ranging from molecular properties to particle detection in tissue samples...
December 24, 2016: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28025889/detailed-study-of-bsa-adsorption-on-micro-and-nanocrystalline-diamond-%C3%AE-sic-composite-gradient-films-by-time-resolved-fluorescence-microscopy
#19
Stephan Handschuh-Wang, Tao Wang, Sergey I Druzhinin, Daniel Wesner, Xin Jiang, Holger Schönherr
The adsorption of bovine serum albumin (BSA) on micro- and nanocrystalline diamond/β-SiC composite films synthesized using the hot filament chemical vapor deposition (HFCVD) technique has been investigated by confocal fluorescence lifetime imaging microscopy. BSA labeled with fluorescein isothiocyanate (FITC) was employed as a probe. The BSA(FITC) conjugate was found to preferentially adsorb on both O-/OH-terminated microcrystalline and nanocrystalline diamond compared to the OH-terminated β-SiC, resulting in an increasing amount of BSA adsorbed to the gradient surfaces with an increasing diamond/β-SiC ratio...
January 9, 2017: Langmuir: the ACS Journal of Surfaces and Colloids
https://www.readbyqxmd.com/read/28018724/gpu-accelerated-real-time-confocal-fluorescence-lifetime-imaging-microscopy-flim-based-on-the-analog-mean-delay-amd-method
#20
Byungyeon Kim, Byungjun Park, Seungrag Lee, Youngjae Won
We demonstrated GPU accelerated real-time confocal fluorescence lifetime imaging microscopy (FLIM) based on the analog mean-delay (AMD) method. Our algorithm was verified for various fluorescence lifetimes and photon numbers. The GPU processing time was faster than the physical scanning time for images up to 800 × 800, and more than 149 times faster than a single core CPU. The frame rate of our system was demonstrated to be 13 fps for a 200 × 200 pixel image when observing maize vascular tissue. This system can be utilized for observing dynamic biological reactions, medical diagnosis, and real-time industrial inspection...
December 1, 2016: Biomedical Optics Express
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