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Fluorescence Imaging (fluorescence + imaging)
Kinds of Fluorescence Imaging Selected AbstractsSilica-Coated Manganese Oxide Nanoparticles as a Platform for Targeted Magnetic Resonance and Fluorescence Imaging of Cancer CellsADVANCED FUNCTIONAL MATERIALS, Issue 11 2010Hong Yang Abstract Monodisperse silica-coated manganese oxide nanoparticles (NPs) with a diameter of ,35,nm are synthesized and are aminated through silanization. The amine-functionalized core,shell NPs enable the covalent conjugation of a fluorescent dye, Rhodamine B isothiocyanate (RBITC), and folate (FA) onto their surface. The formed Mn3O4@SiO2(RBITC),FA core,shell nanocomposites are water-dispersible, stable, and biocompatible when the Mn concentration is below 50,µg mL,1 as confirmed by a cytotoxicity assay. Relaxivity measurements show that the core,shell NPs have a T1 relaxivity (r1) of 0.50,mM,1,s,1 on the 0.5 T scanner and 0.47,mM,1,s,1 on the 3.0 T scanner, suggesting the possibility of using the particles as a T1 contrast agent. Combined flow cytometry, confocal microscopy, and magnetic resonance imaging studies show that the Mn3O4@SiO2(RBITC),FA nanocomposites can specifically target cancer cells overexpressing FA receptors (FARs). Findings from this study suggest that the silica-coated Mn3O4 core,shell NPs could be used as a platform for bimodal imaging (both magnetic resonance and fluorescence) in various biological systems. [source] Fluorescent Imaging: Surface Modification of Exfoliated Layered Gadolinium Hydroxide for the Development of Multimodal Contrast Agents for MRI and Fluorescence Imaging (Adv. Funct.ADVANCED FUNCTIONAL MATERIALS, Issue 21 2009Mater. The synthesis of a new stable colloid of fluorescent LGdH layers through a newly developed surface modification method is reported by Y.-S. Yoon et al. on page 3375. This new method involves layer exfoliation, anion exchange, and PEG coating. The efficient MRI contrast-enhancement of these LGdH layers, both in vitro and in vivo, demonstrates their potential utility as a multimodal probe combining optical and MR imaging. [source] Surface Modification of Exfoliated Layered Gadolinium Hydroxide for the Development of Multimodal Contrast Agents for MRI and Fluorescence ImagingADVANCED FUNCTIONAL MATERIALS, Issue 21 2009Young-su Yoon Abstract A novel method for modifying the surface of magnetic-resonance-contrasting layered gadolinium hydroxide (LGdH) is developed providing them with water- and bio-compatibility and acid-resistance, all of which are essential for medical applications. A stable colloid of exfoliated layers is synthesized by exchanging interlayer anions of LGdH with oleate ions. The delaminated layers are successively coated with phospholipids with poly(ethylene glycol) tail groups, and their effectiveness as a contrast agent for magnetic resonance imaging (MRI) is demonstrated. The adaptability of this surface modification approach for incorporating functional molecules and fabricating a fluorescent colloid of LGdH, which has the potential utility as a multimodal probe, is also demonstrated. This result provides a novel approach for expanding the applications of layered inorganic materials and developing a new class of MRI contrast agents. [source] A New Series of Quadrupolar Type Two-Photon Absorption Chromophores Bearing 11, 12-Dibutoxydibenzo[a,c]-phenazine Bridged Amines; Their Applications in Two-Photon Fluorescence Imaging and Two-Photon Photodynamic TherapyADVANCED FUNCTIONAL MATERIALS, Issue 15 2009Marappan Velusamy Abstract A new series of quadrupolar type two-photon absorption (2PA) chromophores 3,9 bearing a core arylamine-[a,c]phenazine-arylamine motif are synthesized in high yields. Palladium-catalyzed Stille coupling and CN coupling reactions are utilized to prepare target chromophores. Detailed characterization and systematic studies of these molecules, including absorption and fluorescence emission, are conducted. These compounds are found to exhibit very large 2PA cross section values, for example, ,7000 GM at 800,nm for 8 in toluene. Two-photon-induced fluorescence imaging is successfully demonstrated in vitro using compound- 8 -encapsulated silica nanoparticles with excellent bio-compatibility. In combination with the capability of both one- and two-photon singlet-oxygen sensitizations, this nanocomposite demonstrates its promising potential in dual functionality toward two-photon fluorescence imaging and two-photon photodynamic therapy. [source] Cellular Imaging: Conjugated Oligoelectrolyte Harnessed Polyhedral Oligomeric Silsesquioxane as Light-Up Hybrid Nanodot for Two-Photon Fluorescence Imaging of Cellular Nucleus (Adv. Mater.ADVANCED MATERIALS, Issue 37 201037/2010) Bin Liu and co-workers report on p. 4186 on a water-soluble organic/inorganic hybrid nanodot based on polyhedral oligomeric silsesquioxane (POSS) and conjugated oligoelectrolytes for twophoton fluorescence imaging of cellular nuclei. The small size of the dots (,3.3 nm) impart nucleus permeability and substantial DNA-enhanced twophoton excited fluorescence that allows highcontrast illumination of the nucleus. [source] Conjugated Oligoelectrolyte Harnessed Polyhedral Oligomeric Silsesquioxane as Light-Up Hybrid Nanodot for Two-Photon Fluorescence Imaging of Cellular NucleusADVANCED MATERIALS, Issue 37 2010Kan-Yi Pu A water-soluble organic/inorganic hybrid nanodot based on polyhedral oligomeric silsesquioxane (POSS) and conjugated oligoelectrolyte is designed and synthesized for two-photon fluorescence imaging of cellular nucleus, which takes advantage of its small size (,3.3 nm) that imparts nucleus permeability and substantial DNA-enhanced two-photon excited fluorescence that allows illuminating the nucleus with a high contrast. [source] Ex Vivo Fluorescence Imaging of Normal and Malignant Urothelial Cells to Enhance Early DiagnosisPHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 5 2007Karine Steenkeste ABSTRACT Urinary cytology is a noninvasive and unconstraining technique for urothelial cancer diagnosis but lacks sensitivity for detecting low-grade lesions. In this study, the fluorescence properties of classical Papanicolaou-stained urothelial cytological slides from patients or from cell lines were monitored to investigate metabolic changes in normal and tumoral cells. Time- and spectrally-resolved fluorescence imaging was performed at the single cell level to assess the spectral and temporal properties as well as the spatial distribution of the fluorescence emitted by urothelial cells. The results reveal quite different fluorescence distributions between tumoral urothelial cells, characterized by a perimembrane fluorescence localization, and the normal cells which exhibit an intracellular fluorescence. This is not caused by differences in the fluorescence emission of the endogenous fluorophores NAD(P)H, flavoproteins or porphyrins but by various localization of the EA 50 Papanicolaou stain as revealed by both the spectral and time-resolved parameters. The present results demonstrate that the use of single-cell endofluorescence emission of Papanicolaou-stained urothelial cytological slides can allow an early ex vivo diagnosis of low-grade bladder cancers. [source] ChemInform Abstract: Photobleaching of Asymmetric Cyanines Used for Fluorescence Imaging of Single DNA Molecules.CHEMINFORM, Issue 49 2001Claire Kanony Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source] Electrochemical Modulation of Remote Fluorescence Imaging at an Ordered Opto-electrochemical Nanoaperture ArrayCHEMPHYSCHEM, Issue 8 2004Arnaud Chovin Abstract An array of nanometer-sized apertures capable of electrochemically modulating the fluorescence of a model analyte is presented. The device, which combines near-field optical methods and ultramicroelectrode properties in an array format, is based on an etched coherent optical fiber bundle. Indeed, the fabrication steps produced an ordered array where each optical nanoaperture is surrounded by a ring-shaped gold nanoelectrode. The chronoamperometric behavior of the array shows stable diffusion-limited quasi-steady-state response. The model analyte, tris(2,2,-bipyridine) ruthenium, emits fluorescence in the Ru(II) state, but not in the oxidized Ru(III) state. Fluorescence is excited by visible light exiting from each nanoaperture since light is confined to the tip apex by the gold coating. A fraction of the isotropically emitted luminescence is collected by the same nanoaperture, transmitted by the corresponding fiber core and eventually detected by a charge-coupled device (CCD) camera. The array format provides a fluorescence image resolved at the nanometric scale which covers a large micrometric area. Therefore the high-density array plays a bridging role between these two fundamental scales. We established that the opto-electrochemical nanoapertures are optically independent. Fluorescence of the sample collected by each nanoaperture is modulated by changing the potential of the nanoring electrodes. Reversible electrochemical switching of remote fluorescence imaging is performed through the opto-electrochemical nanoaperture array itself. Eventually this ordered structure of nanometer light sources which are electrochemically manipulated provides promising photonic or electro-optical devices for various future applications. For example, such an array has potential in the development of a combined SNOM-electrochemical nanoprobe array to image a real sample concomitantly at the nanometer and micrometer scale. [source] Alcohol and Mitochondria in Cardiac Apoptosis: Mechanisms and VisualizationALCOHOLISM, Issue 5 2005György Hajnóczky Apoptosis of myocytes is likely to contribute to a variety of heart conditions and could also be important in the development of alcoholic heart disease. A fundamental pathway to apoptosis is through mitochondrial membrane permeabilization and release of proapoptotic factors from the mitochondrial intermembrane space to the cytosol. The authors' results show that prolonged exposure of cultured cardiac cells to ethanol (35 mM for 48 hr) promotes Ca2+ -induced activation of the mitochondrial permeability transition pore (PTP). PTP-dependent mitochondrial membrane permeabilization is followed by release of cytochrome c and execution of apoptosis. The authors propose that chronic ethanol exposure, in combination with other stress signals, may allow for activation of the PTP by physiological calcium oscillations, providing a trigger for cardiac apoptosis during chronic alcohol abuse. Coincidence of apoptosis promoting factors occurs in only a small fraction of myocytes, but because of the absence of regeneration, even a modest increase in the rate of cell death may contribute to a decrease in cardiac contractility. Detection of apoptotic changes that are present in only a few myocytes at a certain time in the heart is not feasible with most of the apoptotic assays. Fluorescence imaging is a powerful technology to visualize changes that are confined to a minor fraction of cells in a tissue, and the use of multiphoton excitation permits imaging in situ deep in the wall of the intact heart. This article discusses potential mechanisms of the effect of alcohol on mitochondrial membrane permeabilization and visualization of mitochondria-dependent apoptosis in cardiac muscle. [source] Chondroprotective role of the osmotically sensitive ion channel transient receptor potential vanilloid 4: Age- and sex-dependent progression of osteoarthritis in Trpv4 -deficient miceARTHRITIS & RHEUMATISM, Issue 10 2010Andrea L. Clark Objective Mechanical loading significantly influences the physiology and pathology of articular cartilage, although the mechanisms of mechanical signal transduction are not fully understood. Transient receptor potential vanilloid 4 (TRPV4) is a Ca++ -permeable ion channel that is highly expressed by articular chondrocytes and can be gated by osmotic and mechanical stimuli. The goal of this study was to determine the role of Trpv4 in the structure of the mouse knee joint and to determine whether Trpv4,/, mice exhibit altered Ca++ signaling in response to osmotic challenge. Methods Knee joints of Trpv4,/, mice were examined histologically and by microfocal computed tomography for osteoarthritic changes and bone structure at ages 4, 6, 9, and 12 months. Fluorescence imaging was used to quantify chondrocytic Ca++ signaling within intact femoral cartilage in response to osmotic stimuli. Results Deletion of Trpv4 resulted in severe osteoarthritic changes, including cartilage fibrillation, eburnation, and loss of proteoglycans, that were dependent on age and male sex. Subchondral bone volume and calcified meniscal volume were greatly increased, again in male mice. Chondrocytes from Trpv4+/+ mice demonstrated significant Ca++ responses to hypo-osmotic stress but not to hyperosmotic stress. The response to hypo-osmotic stress or to the TRPV4 agonist 4,-phorbol 12,13-didecanoate was eliminated in Trpv4,/, mice. Conclusion Deletion of Trpv4 leads to a lack of osmotically induced Ca++ signaling in articular chondrocytes, accompanied by progressive, sex-dependent increases in bone density and osteoarthritic joint degeneration. These findings suggest a critical role for TRPV4-mediated Ca++ signaling in the maintenance of joint health and normal skeletal structure. [source] Bispectral fluorescence imaging of aggressive basal cell carcinoma combined with histopathological mapping: a preliminary study indicating a possible adjunct to Mohs micrographic surgeryBRITISH JOURNAL OF DERMATOLOGY, Issue 2 2006B. Stenquist Summary Background, Fluorescence imaging is an attractive diagnostic technique for skin tumour demarcation with potential to move to clinical use. Bispectral fluorescence imaging combines skin autofluorescence with ,-aminolaevulinic acid-induced fluorescence. To evaluate the technique, fluorescence data must be compared with the histopathological extent of the tumour, which is the purpose of the current study. Objectives, To investigate the agreement between bispectral fluorescence images and the histopathological tumour boundary of ill-defined basal cell carcinomas (BCCs). After fluorescence imaging the tumours were removed using Mohs micrographic surgery (MMS) to obtain histopathological maps of the tumour boundaries. Methods, Twelve patients with aggressive BCC of mean diameter 16 mm (range 5,32) in the face were included in the study. The patients were subjected to bispectral fluorescence imaging within the 2 months prior to MMS. The fluorescence images and histopathological maps were aligned using image warping. Results, Five patients (42%) showed good agreement with the histopathological mapping and the remaining seven patients (58%) showed partial agreement. Bispectral investigation combining autofluorescence with protoporphyrin IX (PpIX) fluorescence generally yielded better agreement with the histopathological boundaries of the tumours compared with using only the PpIX fluorescence. Conclusions, In this preliminary study the fluorescence has been compared with the histopathological tumour boundaries. The result implies that the technique can be applied as a useful tool for indicating tumour boundary of aggressive BCCs. Further refinement is needed to be able to indicate the exact tumour border. [source] Antitumor effect of photodynamic therapy with chlorin-based photosensitizer DH-II-24 in colorectal carcinomaCANCER SCIENCE, Issue 12 2009Young-Cheol Lim While photodynamic therapy (PDT) has been recognized as a promising therapeutic modality for the treatment of various cancers and diseases, developments of effective photosensitizers are highly desired to improve the prospect for the use of PDT. In this study, we evaluated DH-II-24, a new photosensitizer, for antitumor PDT in vitro and in vivo. Loaded into human colorectal carcinoma cells (HCT116), DH-II-24 was primarily accumulated in mitochondria, lysosomes, and endoplasmic reticula. Administration of DH-II-24 followed by light exposure induced necrotic cell death in a dose-dependent manner, whereas DH-II-24 in the absence of light induced minimal cell death. In order to investigate the distribution and phamacokinetics of the photosensitizer in vivo, DH-II-24 was intravenously injected to female BALB/c nude mice. Fluorescence imaging in vivo showed that DH-II-24 was rapidly distributed across the entire body and then mostly eliminated at 24 h. Next, effectiveness of DH-II-24-mediated PDT was examined on colorectal carcinoma xenografts established subcutaneously in BALB/c nude mice. DH-II-24 (1 mg/kg, i.v. administration) followed by light exposure significantly suppressed growth of xenograft tumors, compared to light exposure or DH-II-24 alone. Histological examination revealed necrotic damage in PDT-treated tumors, concomitantly with severe damage of tumor vasculature. These results suggest that DH-II-24 is a potential photosensitizer of photodynamic therapy for cancer. (Cancer Sci 2009; 100: 2431,2436) [source] Effects of short-term food deprivation on orexin-A-induced intestinal bicarbonate secretion in comparison with related secretagoguesACTA PHYSIOLOGICA, Issue 3 2010G. Flemström Abstract Studies of gastrointestinal physiology in humans and intact animals are usually conducted after overnight fast. We compared the effects of orexin-A, vasoactive intestinal polypeptide (VIP), melatonin, serotonin, uroguanylin, ghrelin and prostaglandin E2 (PGE2) on duodenal bicarbonate secretion in fed and overnight fasted animals. This review is a summary of our findings. Secretagogues were administered by intra-arterial infusion or luminally (PGE2). Enterocyte intracellular calcium ([Ca2+]i) signalling was studied by fluorescence imaging. Total RNA was extracted, reverse transcripted to cDNA and expression of orexin receptors measured by quantitative real-time PCR. Orexin-A stimulates the duodenal secretion in continuously fed animals but not in food-deprived animals. Similarly, short-term fasting causes a 100-fold decrease in the amount of the muscarinic agonist bethanechol required for stimulation of secretion. In contrast, fasting does not affect secretory responses to intra-arterial VIP, melatonin, serotonin, uroguanylin and ghrelin, or that to luminal PGE2. Orexin-A induces [Ca2+]i signalling in enterocytes from fed rats but no significant [Ca2+]i responses occur in enterocytes from fasted animals. In addition, overnight fasting decreases the expression of mucosal orexin receptors. Short-term food deprivation thus decreases duodenal expression of orexin receptors and abolishes the secretory response to orexin-A as well as orexin-A-induced [Ca2+]i signalling. Fasting, furthermore, decreases mucosal sensitivity to bethanechol. The absence of declines in secretory responses to other secretagogues tested strongly suggests that short-term fasting does not affect the secretory capacity of the duodenal mucosa in general. Studies of intestinal secretion require particular evaluation with respect to feeding status. [source] In vivo analysis of MT-based vesicle transport by confocal reflection microscopyCYTOSKELETON, Issue 2 2009Imre Gáspár Abstract The use of confocal reflection microscopy (CRM) for the in vivo analysis of microtubule (MT) mediated transport of lipid droplets in the developing Drosophila egg primordia is described here. The developing Drosophila oocytes are ideal objects to study MT-mediated transport in vivo: transport of e.g. the lipid droplets can be conveniently, selectively and sensitively monitored through CRM and the egg primordia are readily available for physical, chemical and/or genetic manipulations. CRM is a non-destructive way to follow vesicle movement and allows high frame rate image recording. When combined with fluorescence imaging, CRM offers simultaneous visualization of the cargo and the protein(s) of interest, i.e. a motor or a cargo adapter, thus allowing a better understanding of MT-mediated transport and spatiotemporal coordination of the transport machinery. Cell Motil. Cytoskeleton 2009. © 2009 Wiley-Liss, Inc. [source] Visualized Sclerotherapy of Varicose VeinsDERMATOLOGIC SURGERY, Issue 2010MAMORU KIKUCHI MD BACKGROUND The spread and movement of sclerosant after injection during sclerotherapy is difficult to monitor. OBJECTIVE To develop a new visualization method that allows monitoring of sclerosant dosage and flow during sclerotherapy. METHODS We used a photodynamic eye (PDE) to perform indocyanine green (ICG) imaging. ICG produces strong fluorescence detectable using PDE and allows monitoring of sclerosant spread through blood vessels in real time. We performed visualized sclerotherapy on 50 limbs, comprising high ligation and sclerotherapy (35 limbs), stripping and sclerotherapy (10 limbs), and sclerotherapy alone (5 limbs). RESULTS In all cases, fluorescence imaging of the injected sclerosant was possible. No complications resulted from combining ICG and polidocanol in any of the patients, all of whom received follow-up evaluations at 1 week, 1 month, and 3 months after treatment. CONCLUSIONS Our new method not only avoids the risk of radiation exposure, but also allows for simple observation of sclerosant range of access, determination of the dosage for each lesion, and accurate administration of therapy to target lesions. This method will contribute to further advances in sclerotherapy, given that it allows administration of sclerosant and visual confirmation of optimal injection dosage, speed, and movement of sclerosant after injection. The authors have indicated no significant interest with commercial supporters. [source] Diagnosis of Nonmelanoma Skin Cancer/Keratinocyte Carcinoma: A Review of Diagnostic Accuracy of Nonmelanoma Skin Cancer Diagnostic Tests and TechnologiesDERMATOLOGIC SURGERY, Issue 10 2007METTE MOGENSEN MD BACKGROUND Nonmelanoma skin cancer (NMSC) is the most prevalent cancer in the light-skinned population. Noninvasive treatment is increasingly used for NMSC patients with superficial lesions, making the development of noninvasive diagnostic technologies highly relevant. OBJECTIVE The scope of this review is to present data on the current state-of-the-art diagnostic methods for keratinocyte carcinoma: basal cell carcinoma, squamous cell carcinoma, and actinic keratosis. METHODS AND MATERIALS MEDLINE, BIOSIS, and EMBASE searches on NMSC and physical and clinical examination, biopsy, molecular marker, ultrasonography, Doppler, optical coherence tomography, dermoscopy, spectroscopy, fluorescence imaging, confocal microscopy, positron emission tomography, computed tomography, magnetic resonance imaging, terahertz imaging, electrical impedance and sensitivity, specificity, and diagnostic accuracy. RESULTS State-of-the-art diagnostic research has been limited in this field, but encouraging results from the reviewed diagnostic trials have suggested a high diagnostic accuracy for many of the technologies. Most of the studies, however, were pilot or small studies and the results would need to be validated in larger trials. CONCLUSIONS Some of these new imaging technologies have the capability of providing new, three-dimensional in vivo, in situ understanding of NMSC development over time. Some of the new technologies described here have the potential to make it from the bench to the clinic. [source] Comparison of Different Strategies on DNA Chip Fabrication and DNA-Sensing: Optical and Electrochemical ApproachesELECTROANALYSIS, Issue 22 2005Sabine Szunerits Abstract New strategies for the construction of DNA chips and the detection of DNA hybridization will be discussed in this review. The focus will be on the use of polypyrrole as a linker between a substrate and oligonucleotide probes. The modification step is based on the electrochemical copolymerization of pyrrole and oligonucleotides bearing a pyrrole group on its 5, end. This strategy was employed for the immobilization of oligonucleotides on millimeter-sized electrodes, microelectrode arrays, as well as for the local structuring of homogeneous gold surfaces. Our approaches for the localized patterning of gold surfaces will be also discussed. Localized immobilization was achieved by using an electrospotting technique, where a micropipette served as an electrochemical cell where spot sizes with 800,,m diameters were fabricated. The use of a microcell using a Teflon covered metal needle with a cavity of 100,,m resulted in immobilized probe spots of 300,,m. Scanning electrochemical microscopy (SECM) was also used, and surface modifications of 100,,m were obtained depending on the experimental conditions. Different detection methods were employed for the reading of the hybridization event: fluorescence imaging, surface plasmon resonance imaging (SPRI), photocurrent measurements, and voltamperometric measurements using intercalators. Their advantages concerning the various immobilization strategies will also be discussed. [source] A novel approach to tag and identify geranylgeranylated proteinsELECTROPHORESIS, Issue 20 2009Lai N. Chan Abstract A recently developed proteomic strategy, the "GG-azide"-labeling approach, is described for the detection and proteomic analysis of geranylgeranylated proteins. This approach involves metabolic incorporation of a synthetic azido-geranylgeranyl analog and chemoselective derivatization of azido-geranylgeranyl-modified proteins by the "click" chemistry, using a tetramethylrhodamine-alkyne. The resulting conjugated proteins can be separated by 1-D or 2-D and pH fractionation, and detected by fluorescence imaging. This method is compatible with downstream LC-MS/MS analysis. Proteomic analysis of conjugated proteins by this approach identified several known geranylgeranylated proteins as well as Rap2c, a novel member of the Ras family. Furthermore, prenylation of progerin in mouse embryonic fibroblast cells was examined using this approach, demonstrating that this strategy can be used to study prenylation of specific proteins. The "GG-azide"-labeling approach provides a new tool for the detection and proteomic analysis of geranylgeranylated proteins, and it can readily be extended to other post-translational modifications. [source] Dissolved copper triggers cell death in the peripheral mechanosensory system of larval fishENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 2 2006Tiffany L. Linbo Abstract Dissolved copper is an increasingly common non,point source contaminant in urban and urbanizing watersheds. In the present study, we investigated the sublethal effects of dissolved copper on the peripheral mechanosensory system, or lateral line, of larval zebrafish (Danio rerio). Zebrafish larvae were exposed to copper (0,65 ,g/L), and the cytotoxic responses of individual lateral line receptor neurons were examined using a combination of in vivo fluorescence imaging, confocal microscopy, scanning electron microscopy, and conventional histology. Dissolved copper triggered a dose-dependent loss of neurons in identified lateral line neuromasts at concentrations ,20 ,g/L. The onset of cell death in the larval mechanosensory system was rapid (<1 h). When copper-exposed zebrafish were transferred to clean water, the lateral line regenerated over the course of 2 d. In contrast, the lateral line of larvae exposed continuously to dissolved copper (50 ,g/L) for 3 d did not recover. Collectively, these results show that peripheral mechanosensory neurons are vulnerable to the neurotoxic effects of copper. Consequently, dissolved copper in non-point source storm-water runoff has the potential to interfere with rheotaxis, schooling, predator avoidance, and other mechanosensory-mediated behaviors that are important for the migration and survival of fish. [source] Visualization of local Ca2+ dynamics with genetically encoded bioluminescent reportersEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 3 2005Kelly L. Rogers Abstract Measurements of local Ca2+ signalling at different developmental stages and/or in specific cell types is important for understanding aspects of brain functioning. The use of light excitation in fluorescence imaging can cause phototoxicity, photobleaching and auto-fluorescence. In contrast, bioluminescence does not require the input of radiative energy and can therefore be measured over long periods, with very high temporal resolution. Aequorin is a genetically encoded Ca2+ -sensitive bioluminescent protein, however, its low quantum yield prevents dynamic measurements of Ca2+ responses in single cells. To overcome this limitation, we recently reported the bi-functional Ca2+ reporter gene, GFP-aequorin (GA), which was developed specifically to improve the light output and stability of aequorin chimeras [V. Baubet, et al., (2000) PNAS, 97, 7260,7265]. In the current study, we have genetically targeted GA to different microdomains important in synaptic transmission, including to the mitochondrial matrix, endoplasmic reticulum, synaptic vesicles and to the postsynaptic density. We demonstrate that these reporters enable ,real-time' measurements of subcellular Ca2+ changes in single mammalian neurons using bioluminescence. The high signal-to-noise ratio of these reporters is also important in that it affords the visualization of Ca2+ dynamics in cell,cell communication in neuronal cultures and tissue slices. Further, we demonstrate the utility of this approach in ex-vivo preparations of mammalian retina, a paradigm in which external light input should be controlled. This represents a novel molecular imaging approach for non-invasive monitoring of local Ca2+ dynamics and cellular communication in tissue or whole animal studies. [source] Maturation of postsynaptic nicotinic structures on autonomic neurons requires innervation but not cholinergic transmissionEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2002Sergio Kaiser Abstract Postsynaptic development at the neuromuscular junction depends on nicotinic transmission and secreted components from the presynaptic motor nerve terminal. Similarly, secreted components and synaptic activity are both thought to guide development of glutamatergic synapses in the CNS. Nicotinic synapses on chick ciliary neurons are structurally complex: a large presynaptic calyx engulfs the postsynaptic neuron and overlays a series of discrete mats of receptor-rich somatic spines tightly interwoven and folded against the soma. We used fluorescence imaging of ,7-containing nicotinic receptors and the spine constituent drebrin to monitor postsynaptic development. The results show that surgical disruption of the preganglionic input or removal of the ganglionic synaptic target tissue after synapses form in the ganglion does not disrupt the receptor-rich spine mats. Similarly, removal of the target tissue even prior to synapse formation in the ganglion does not prevent subsequent formation of the receptor clusters and associated spine constituents. Postsynaptic development is arrested, however, if normal innervation is prevented by ablating the preganglionic neurons prior to synapse formation. In this case the neurons express reduced levels of nicotinic receptors and cytoskeletal components and organize them only into early-stage clusters. Even low levels of residual innervation, however, can restore much of the normal postsynaptic receptor patterns. Chronic pharmacological blockade of cholinergic synaptic activity fails to replicate the effects of ablating the preganglionic nucleus. The results indicate that ciliary neurons are programmed to express postsynaptic components and can initiate clustering of ,7-containing receptors but need presynaptic guidance for maturation of the postsynaptic structure. [source] Expression of the recombinant bacterial outer surface protein A in tobacco chloroplasts leads to thylakoid localization and loss of photosynthesisFEBS JOURNAL, Issue 21 2007Anna Hennig Bacterial lipoproteins play crucial roles in host,pathogen interactions and pathogenesis and are important targets for the immune system. A prominent example is the outer surface protein A (OspA) of Borrelia burgdorferi, which has been efficiently used as a vaccine for the prevention of Lyme disease. In a previous study, OspA could be produced in tobacco chloroplasts in a lipidated and immunogenic form. To further explore the potential of chloroplasts for the production of bacterial lipoproteins, the role of the N-terminal leader sequence was investigated. The amount of recombinant OspA could be increased up to ten-fold by the variation of the insertion site in the chloroplast genome. Analysis of OspA mutants revealed that replacement of the invariant cysteine residue as well as deletion of the leader sequence abolishes palmitolyation of OspA. Also, decoration of OspA with an N-terminal eukaryotic lipidation motif does not lead to palmitoylation in chloroplasts. Strikingly, the bacterial signal peptide of OspA efficiently targets the protein to thylakoids, and causes a mutant phenotype. Plants accumulating OspA at 10% total soluble protein could not grow without exogenously supplied sugars and rapidly died after transfer to soil under greenhouse conditions. The plants were found to be strongly affected in photosystem II, as revealed by the analyses of temporal and spatial dynamics of photosynthetic activity by chlorophyll fluorescence imaging. Thus, overexpression of OspA in chloroplasts is limited by its concentration-dependent interference with essential functions of chloroplastic membranes required for primary metabolism. [source] A New Series of Quadrupolar Type Two-Photon Absorption Chromophores Bearing 11, 12-Dibutoxydibenzo[a,c]-phenazine Bridged Amines; Their Applications in Two-Photon Fluorescence Imaging and Two-Photon Photodynamic TherapyADVANCED FUNCTIONAL MATERIALS, Issue 15 2009Marappan Velusamy Abstract A new series of quadrupolar type two-photon absorption (2PA) chromophores 3,9 bearing a core arylamine-[a,c]phenazine-arylamine motif are synthesized in high yields. Palladium-catalyzed Stille coupling and CN coupling reactions are utilized to prepare target chromophores. Detailed characterization and systematic studies of these molecules, including absorption and fluorescence emission, are conducted. These compounds are found to exhibit very large 2PA cross section values, for example, ,7000 GM at 800,nm for 8 in toluene. Two-photon-induced fluorescence imaging is successfully demonstrated in vitro using compound- 8 -encapsulated silica nanoparticles with excellent bio-compatibility. In combination with the capability of both one- and two-photon singlet-oxygen sensitizations, this nanocomposite demonstrates its promising potential in dual functionality toward two-photon fluorescence imaging and two-photon photodynamic therapy. [source] Cellular Imaging: Conjugated Oligoelectrolyte Harnessed Polyhedral Oligomeric Silsesquioxane as Light-Up Hybrid Nanodot for Two-Photon Fluorescence Imaging of Cellular Nucleus (Adv. Mater.ADVANCED MATERIALS, Issue 37 201037/2010) Bin Liu and co-workers report on p. 4186 on a water-soluble organic/inorganic hybrid nanodot based on polyhedral oligomeric silsesquioxane (POSS) and conjugated oligoelectrolytes for twophoton fluorescence imaging of cellular nuclei. The small size of the dots (,3.3 nm) impart nucleus permeability and substantial DNA-enhanced twophoton excited fluorescence that allows highcontrast illumination of the nucleus. [source] Conjugated Oligoelectrolyte Harnessed Polyhedral Oligomeric Silsesquioxane as Light-Up Hybrid Nanodot for Two-Photon Fluorescence Imaging of Cellular NucleusADVANCED MATERIALS, Issue 37 2010Kan-Yi Pu A water-soluble organic/inorganic hybrid nanodot based on polyhedral oligomeric silsesquioxane (POSS) and conjugated oligoelectrolyte is designed and synthesized for two-photon fluorescence imaging of cellular nucleus, which takes advantage of its small size (,3.3 nm) that imparts nucleus permeability and substantial DNA-enhanced two-photon excited fluorescence that allows illuminating the nucleus with a high contrast. [source] Disparity between prostate tumor interior versus peripheral vasculature in response to verteporfin-mediated vascular-targeting therapyINTERNATIONAL JOURNAL OF CANCER, Issue 3 2008Bin Chen Abstract Photodynamic therapy (PDT) is a light-based cancer treatment modality. Here we employed both in vivo and ex vivo fluorescence imaging to visualize vascular response and tumor cell survival after verteporfin-mediated PDT designed to target tumor vasculature. EGFP-MatLyLu prostate tumor cells, transduced with EGFP using lentivirus vectors, were implanted in athymic nude mice. Immediately after PDT with different doses of verteporfin, tumor-bearing animals were injected with a fluorochrome-labeled albumin. The extravasation of fluorescent albumin along with tumor EGFP fluorescence was monitored noninvasively with a whole-body fluorescence imaging system. Ex vivo fluorescence microscopy was performed on frozen sections of tumor tissues taken at different times after treatment. Both in vivo and ex vivo imaging demonstrated that vascular-targeting PDT with verteporfin significantly increased the extravasation of fluorochrome-labeled albumin in the tumor tissue, especially in the tumor periphery. Although PDT induced substantial vascular shutdown in interior blood vessels, some peripheral tumor vessels were able to maintain perfusion function up to 24 hr after treatment. As a result, viable tumor cells were typically detected in the tumor periphery in spite of extensive tumor cell death. Our results demonstrate that vascular-targeting PDT with verteporfin causes a dose- and time-dependent increase in vascular permeability and decrease in blood perfusion. However, compared to the interior blood vessels, peripheral tumor blood vessels were found less sensitive to PDT-induced vascular shutdown, which was associated with subsequent tumor recurrence in the tumor periphery. © 2008 Wiley-Liss, Inc. [source] Early Detection of Bone Metastases in a Murine Model Using Fluorescent Human Breast Cancer Cells: Application to the Use of the Bisphosphonate Zoledronic Acid in the Treatment of Osteolytic LesionsJOURNAL OF BONE AND MINERAL RESEARCH, Issue 11 2001Olivier Peyruchaud Abstract A very common metastatic site for human breast cancer is bone. The traditional bone metastasis model requires human MDA-MB-231 breast carcinoma cell inoculation into the left heart ventricle of nude mice. MDA-MB-231 cells usually develop osteolytic lesions 3,4 weeks after intracardiac inoculation in these animals. Here, we report a new approach to study the formation of bone metastasis in animals using breast carcinoma cells expressing the bioluminescent jellyfish protein (green fluorescent protein [GFP]). We first established a subclone of MDA-MB-231 cells by repeated in vivo passages in bone using the heart injection model. On stable transfection of this subclone with an expression vector for GFP and subsequent inoculation of GFP-expressing tumor cells (B02/GFP.2) in the mouse tail vein, B02/GFP.2 cells displayed a unique predilection for dissemination to bone. Externally fluorescence imaging of live animals allowed the detection of fluorescent bone metastases approximately 1 week before the occurrence of radiologically distinctive osteolytic lesions. The number, size, and intensity of fluorescent bone metastases increased progressively with time and was indicative of breast cancer cell progression within bone. Histological examination of fluorescent long bones from B02/GFP.2-bearing mice revealed the occurrence of profound bone destruction. Treatment of B02/GFP.2-bearing mice with the bisphosphonate zoledronic acid markedly inhibited the progression of established osteolytic lesions and the expansion of breast cancer cells within bone. Overall, this new bone metastasis model of breast cancer combining both fluorescence imaging and radiography should provide an invaluable tool to study the effectiveness of pharmaceutical agents that could suppress cancer colonization in bone. [source] Molecular imaging in small animals,roles for micro-CTJOURNAL OF CELLULAR BIOCHEMISTRY, Issue S39 2002Erik L. Ritman Abstract X-ray micro-CT is currently used primarily to generate 3D images of micro-architecture (and the function that can be deduced from it) and the regional distribution of administered radiopaque indicators, within intact rodent organs or biopsies from large animals and humans. Current use of X-ray micro-CT can be extended in three ways to increase the quantitative imaging of molecular transport and accumulation within such specimens. (1) By use of heavy elements, other than the usual iodine, attached to molecules of interest or to surrogates for those molecules. The accumulation of the indicator in the physiological compartments, and the transport to and from such compartments, can be quantitated from the imaged spatial distribution of these contrast agents. (2) The high spatial resolution of conventional X-ray attenuation-based CT images can be used to improve the quantitative nature of radionuclide-based tomographic images (SPECT & PET) by providing correction for attenuation of the emitted gamma rays and the accurate delineation of physiological spaces known to selectively accumulate those indicators. Similarly, other imaging modalities which also localize functions in 2D images (such as histological sections subsequently obtained from the same specimen), can provide a synergistic combination with CT-based 3D microstructure. (3) By increasing the sensitivity and specificity of X-ray CT image contrast by use of methods such as: K-edge subtraction imaging, X-ray fluorescence imaging, imaging of the various types of scattered X-ray and the consequences of the change in the speed of X-rays through different tissues, such as refraction and phase shift. These other methods of X-ray imaging can increase contrast by more than an order of magnitude over that due to conventionally-used attenuation of X-ray. To fully exploit their potentials, much development of radiopaque indicators, scanner hardware and image reconstruction and analysis software will be needed. J. Cell. Biochem. Suppl. 39: 116,124, 2002. © 2002 Wiley-Liss, Inc. [source] Detection of Fecal Contamination on Cantaloupes Using Hyperspectral Fluorescence ImageryJOURNAL OF FOOD SCIENCE, Issue 8 2005Angela M. Vargas ABSTRACT To determine whether detection of fecal contamination on cantaloupes is possible using fluorescence imaging, hyperspectral images of cantaloupes artificially contaminated with a range of diluted bovine feces were acquired from 425 to 774 nm in responses to ultraviolet-A (320 to 400 nm) excitation. Evaluation of images at emission peak wavelengths indicated that 675 nm exhibited the greatest contrast between feces contaminated and untreated surface areas. Two-band ratios compared with the single-band images enhanced the contrast between the feces contaminated spots and untreated cantaloupe surfaces. The 595/655-nm, 655/520-nm, and 555/655-nm ratio images provided relatively high detection rates ranging from 79% to 96% across all feces dilutions. However, both single band and ratio methods showed a number of false positives caused by such features as scarred tissues on cantaloupes. Principal component analysis (PCA) was performed using the entire hyperspectral images data; 2nd and 5th principal component (PC) image exhibited differential responses between feces spots and false positives. The combined use of the 2 PC images demonstrated the detection of feces spots (for example, minimum level of 16-,g/mL dry fecal matter) with minimal false positives. Based on the PC weighing coefficients, the dominant wavelengths were 465, 487, 531, 607, 643, and 688 nm. This research demonstrated the potential of multispectral-based fluorescence imaging for online applications for detection of fecal contamination on cantaloupes. [source] | ||||||||||||||||||||||||||||||||||