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Two photon microscopy

Sven Peters, Max Griebsch, Matthias Klemm, Jens Haueisen, Martin Hammer
In this study the influence of hydrogen peroxide (H2 O2 ) on the redox state, NADH protein binding, and mitochondrial membrane potential in Müller cells is investigated. Cultures of permanent human Müller cells MIO-M1 were exposed to H2 O2 in 75 µM and 150 µM concentration for two hours. Fluorescence emission spectra and lifetimes were measured by two-photon microscopy (excitation wavelength: 740 nm) at the mitochondria which were identified in the microscopic images by their fluorescence properties (spectra and intensity)...
November 29, 2016: Journal of Biophotonics
Xiaoqing Xiong, Yanyan Xu, Laijiu Zheng, Jun Yan, Hongjuan Zhao, Juan Zhang, Yanfeng Sun
As one of the most important coumarin-like dyes, disperse fluorescent Yellow 82 exhibits exceptionally large two-photon effects. Here, it was firstly introduced into the supercritical CO2 dyeing polyester fabrics in this work. Results of the present work showed that the dyeing parameters such as the dyeing time, pressure and temperature had remarkable influences on the color strength of fabrics. The optimized dyeing condition in supercritical CO2 dyeing has been proposed that the dyeing time was 60 min; the pressure was 25 MPa and the temperature was 120 °C...
November 28, 2016: Journal of Fluorescence
Kuen-Che Li, Lynn L H Huang, Jhih-Hao Liang, Ming-Che Chan
A simple approach to multi-color two-photon microscopy of the red, green, and blue fluorescent indicators was reported based on an ultra-compact 1.03-μm femtosecond laser and a nonlinear fiber. Inside the nonlinear fiber, the 1.03-μm laser pulses were simultaneously blue-shifted to 0.6~0.8 μm and red-shifted to 1.2~1.4 μm region by the Cherenkov radiation and fiber Raman gain effects. The wavelength-shifted 0.6~0.8 μm and 1.2~1.4 μm radiations were co-propagated with the residual non-converted 1.03-μm pulses inside the same nonlinear fiber to form a fiber-output three-color femtosecond source...
November 1, 2016: Biomedical Optics Express
Mihaela Balu, Hideharu Mikami, Jue Hou, Eric O Potma, Bruce J Tromberg
We present a multiphoton microscope designed for mesoscale imaging of human skin. The system is based on two-photon excited fluorescence and second-harmonic generation, and images areas of ~0.8x0.8 mm(2) at speeds of 0.8 fps (800x800 pixels; 12 frame averages) for high signal-to-noise ratio, with lateral and axial resolutions of 0.5µm and 3.3µm, respectively. The main novelty of this instrument is the design of the scan head, which includes a fast galvanometric scanner, optimized relay optics, a beam expander and high NA objective lens...
November 1, 2016: Biomedical Optics Express
C Fink, H A Haenssle
BACKGROUND: While the excisional biopsy and histological examination of suspicious lesions remains the current gold standard for diagnosing cutaneous melanoma (CM), there is a demand for more objective and non-invasive examination methods that may support clinicians in their decision when to biopsy or not. METHODS: This review is based on publications and guidelines retrieved by a selective search in PubMed and MEDLINE and focused on non-invasive diagnostic strategies for detecting melanoma...
November 22, 2016: Skin Research and Technology
Anupam Srivastav, Anuradha Verma, Anamika Banerjee, Saif A Khan, Mukul Gupta, Vibha Rani Satsangi, Rohit Shrivastav, Sahab Dass
The present study investigates the effect of gradient doping on modifying the photoelectrochemical response of Ti-doped Fe2O3 photoanodes for their use in sunlight based water splitting for hydrogen evolution. The deposition of a thin film over the ITO (tin doped indium oxide) substrate was carried out using a spray pyrolysis method. The concentration of dopant was varied from 0.5-8.0 at% and two sets of samples were also prepared with low to high (0.5-8%) and high to low (8-0.5%) dopant concentrations in the direction towards the substrate...
November 23, 2016: Physical Chemistry Chemical Physics: PCCP
Skylar M Spangler, Michael R Bruchas
Optogenetics has revolutionized neuroscience by providing means to control cell signaling with spatiotemporal control in discrete cell types. In this review, we summarize four major classes of optical tools to manipulate neuromodulatory GPCR signaling: opsins (including engineered chimeric receptors); photoactivatable proteins; photopharmacology through caging-photoswitchable molecules; fluorescent protein based reporters and biosensors. Additionally, we highlight technologies to utilize these tools in vitro and in vivo, including Cre dependent viral vector expression and two-photon microscopy...
November 19, 2016: Current Opinion in Pharmacology
Grazyna Palczewska, Akiko Maeda, Marcin Golczak, Eisuke Arai, Zhiqian Dong, Lindsay Perusek, Brian Kevany, Krzysztof Palczewski
Accumulation of bis-retinoids in the retinal pigmented epithelium (RPE) is a hallmark of aging and retinal disorders such as Stargardt disease and age-related macular degeneration. These aberrant fluorescent condensation products including A2E are thought to be transferred to RPE cells primarily through phagocytosis of the photoreceptor outer segments. However, we observed by two-photon microscopy that mouse retinas incapable of phagocytosis due to a deficiency of the c-mer proto-oncogene tyrosine kinase (Mertk) nonetheless contained fluorescent retinoid condensation material in their RPE...
November 14, 2016: Journal of Biological Chemistry
Sarah Baldivia, Alexander Levy, Shylaja Hegde, Stijn J A Aper, Maarten Merkx, Rafael Grytz
Increasing evidence suggests that unknown collagen remodeling mechanisms in the sclera underlie myopia development. We are proposing a novel organ culture system in combination with two-photon fluorescence imaging to quantify collagen remodeling at the tissue- and lamella-level. Tree shrew scleral shells were cultured up to 7 days in serum-free media and cellular viability was investigated under: (i) minimal tissue manipulations; (ii) removal of intraocular tissues; gluing the eye to a washer using (iii) 50 μL and (iv) 200 μL of cyanoacrylate adhesive; (v) supplementing media with Ham's F-12 Nutrient Mixture; and (vi) culturing eyes subjected to 15 mmHg intraocular pressure in our new bioreactor...
2016: PloS One
Yu Ding, Chunqiang Li
Nanoscale particle tracking in three dimensions is crucial to directly observe dynamics of molecules and nanoparticles in living cells. Here we present a three-dimensional particle tracking method based on temporally focused two-photon excitation. Multiple particles are imaged at 30 frames/s in volume up to 180 × 180 × 100 µm(3). The spatial localization precision can reach 50 nm. We demonstrate its capability of tracking fast swimming microbes at speed of ~200 µm/s. Two-photon dual-color tracking is achieved by simultaneously exciting two kinds of fluorescent beads at 800 nm to demonstrate its potential in molecular interaction studies...
October 1, 2016: Biomedical Optics Express
Nathan A Jones, Sywe-Ren Chang, William J Troske, Brian H Clarkson, Joerg Lahann
Although dental caries is the most prevalent oral disease worldwide, currently, many dentists continue to use the traditional mirror and probe (dental explorer) method of caries diagnosis. This method of caries detection has the drawback that it is often difficult to distinguish between active and inactive carious lesions. In this work, novel bio-based nanoparticles are developed to specifically detect active caries in vitro. The nanoparticles are made from a cationic fluorescein-labeled food-grade starch in order to fluoresce when illuminated by a standard dental curing light, and to degrade in the oral cavity into nontoxic compounds after detecting the active carious lesion...
November 15, 2016: Advanced Healthcare Materials
Chuanming Luo, Fengyin Liang, Huixia Ren, Xiaoli Yao, Qiang Liu, Mingyue Li, Dajiang Qin, Ti-Fei Yuan, Zhong Pei, Huanxing Su
Collateral blood flow as vascular adaptions to focal cerebral ischemia is well recognized. However, few studies directly investigate the dynamics of collateral vessel recruitment in vivo and little is known about the effect of collateral blood flow in different cerebrovascular hierarchy on the neuropathology after focal ischemic stroke. Here, we report that collateral blood flow is critically involved in blood vessel compensations following regional ischemia. We occluded a pial arteriole using femtosecond laser ablating under the intact thinned skull and documented the changes of collateral flow around the surface communication network and between the surface communication network and subsurface microcirculation network using in vivo two photon microscopy imaging...
November 15, 2016: Brain Pathology
Gioia Della Giustina, Stefano Giulitti, Laura Brigo, Michele Zanatta, Maximilian Tromayer, Robert Liska, Nicola Elvassore, Giovanna Brusatin
In this work, an engineered hydrogel system with a 2D and 3D tunable cross-linking degree is presented. A precise chemical design by the introduction of cross-linkable units, having reaction orthogonality, allows to control the network formation both in time and space and to selectively alter the hydrogel physical properties. Hydrogel chemistry has been tailored in order to produce spatially controlled stiffness changes and drive cell morphology through mechanical cues. Elastic modulus rises by more than double after photocross-linking, as shown by atomic force microscopy measurements...
November 17, 2016: Macromolecular Rapid Communications
Cosmo Lupo, Stefano Pirandola
We determine the ultimate potential of quantum imaging for boosting the resolution of a far-field, diffraction-limited, linear imaging device within the paraxial approximation. First, we show that the problem of estimating the separation between two pointlike sources is equivalent to the estimation of the loss parameters of two lossy bosonic channels, i.e., the transmissivities of two beam splitters. Using this representation, we establish the ultimate precision bound for resolving two pointlike sources in an arbitrary quantum state, with a simple formula for the specific case of two thermal sources...
November 4, 2016: Physical Review Letters
Marco Lombardo, Mohit Parekh, Sebastiano Serrao, Alessandro Ruzza, Stefano Ferrari, Giuseppe Lombardo
PURPOSE: To investigate the microstructure of endothelial keratoplasty grafts using two-photon optical microscopy. METHODS: Six endothelial keratoplasty grafts obtained from human donor corneoscleral tissues and prepared by submerged hydrodissection technique were imaged by two-photon optical microscopy. In each graft, two liquid bubbles were created in order to investigate the presence of a conserved cleavage plane regardless of the volume of posterior stroma that remained attached to Descemet's membrane (DM); the first bubble (bubble A) was generated under DM and the second bubble (bubble B) injection was done in order to obtain a layer of deep stroma that kept the two bubbles separated...
November 17, 2016: Graefe's Archive for Clinical and Experimental Ophthalmology
Roberta Galli, Ortrud Uckermann, Achim Temme, Elke Leipnitz, Matthias Meinhardt, Edmund Koch, Gabriele Schackert, Gerald Steiner, Matthias Kirsch
Coherent anti-Stokes Raman scattering (CARS) microscopy is an emerging technique for identification of brain tumors. However, tumor identification by CARS microscopy on bulk samples and in vivo has been so far verified retrospectively on histological sections, which only provide a gross reference for the interpretation of CARS images without matching at cellular level. Therefore, fluorescent labels were exploited for direct assessment of the interpretation of CARS images of solid and infiltrative tumors. Glioblastoma cells expressing green fluorescent protein (GFP) were used for induction of tumors in mice (n = 7)...
March 11, 2016: Journal of Biophotonics
Cheol Ho Heo, Myoung Ki Cho, Seunggun Shin, Tae Hyeon Yoo, Hwan Myung Kim
Herein, we developed a ratiometric two-photon probe (BHS3-EGF), derived from a pH sensitive dye and epidermal growth factor (EGF), for real-time monitoring and quantitative analysis of acidic luminal pH values during endocytic pathway activity. Two-photon microscopy imaging with BHS3-EGF allows the quantitative analysis of pH distributions of single vesicles and their dynamics in receptor-mediated endocytosis in real-time.
November 17, 2016: Chemical Communications: Chem Comm
Sophia Franck, Magdalena Paterka, Jerome Birkenstock, Frauke Zipp, Volker Siffrin, Esther Witsch
Multiple sclerosis is a chronic, disseminated inflammation of the central nervous system which is thought to be driven by autoimmune T cells. Genetic association studies in multiple sclerosis and a large number of studies in the animal model of the disease support a role for effector/memory T helper cells. However, the mechanisms underlying relapses, remission and chronic progression in multiple sclerosis or the animal model experimental autoimmune encephalomyelitis, are not clear. In particular, there is only scarce information on the role of central nervous system-invading naive T helper cells in these processes...
November 10, 2016: Journal of Neuroimmune Pharmacology: the Official Journal of the Society on NeuroImmune Pharmacology
Navina L Chrobok, Alexandre Jaouen, Keith K Fenrich, John G J M Bol, Micha M M Wilhelmus, Benjamin Drukarch, Franck Debarbieux, Anne-Marie van Dam
Leukocyte infiltration into the central nervous system (CNS) is a key pathological feature in multiple sclerosis (MS) and the MS animal model experimental autoimmune encephalomyelitis (EAE). Recently, preventing leukocyte influx into the CNS of MS patients is the main target of MS therapies and insight into cell behaviour in the circulation is needed for further elucidation of such therapies. In this study, we aimed at in vivo visualization of monocytes in a time-dependent manner during EAE. Using intravital two-photon microscopy (IVM), we imaged CX3CR1(gfp/gfp) mice during EAE, visualizing CX3CR1-GFP(+) monocytes and their dynamics in the spinal cord vasculature...
November 9, 2016: Amino Acids
Kyle P Quinn, Kelly E Sullivan, Zhiyi Liu, Zachary Ballard, Christos Siokatas, Irene Georgakoudi, Lauren D Black
Understanding the organization and mechanical function of the extracellular matrix (ECM) is critical for the development of therapeutic strategies that regulate wound healing following disease or injury. However, these relationships are challenging to elucidate during remodeling following myocardial infarction (MI) due to rapid changes in cellularity and an inability to characterize both ECM microstructure and function non-destructively. In this study, we overcome those challenges through whole organ decellularization and non-linear optical microscopy to directly relate the microstructure and mechanical properties of myocardial ECM...
November 7, 2016: Scientific Reports
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