Semiconductor quantum dots as fluorescent probes for in vitro and in vivo bio-molecular and cellular imaging

Over the years, biological imaging has seen many advances, allowing scientists to unfold many of the mysteries surrounding biological processes. The ideal imaging resolution would be in nanometres, as most biological processes occur at this scale. Nanotechnology has made this possible with functionalised nanoparticles that can bind to specific targets and trace processes at the cellular and molecular level. Quantum dots (QDs) or semiconductor nanocrystals are luminescent particles that have the potential to be the next generation fluorophores. This paper is an overview of the basics of QDs and their role as fluorescent probes for various biological imaging applications. Their potential clinical applications and the limitations that need to be overcome have also been discussed

Nanotechnology and molecular cytogenetics: the future has not yet arrived

Quantum dots (QDs) are a novel class of inorganic fluorochromes composed of nanometer-scale crystals made of a semiconductor material. They are resistant to photo-bleaching, have narrow excitation and emission wavelengths that can be controlled by particle size and thus have the potential for multiplexing experiments. Given the remarkable optical properties that quantum dots possess, they have been proposed as an ideal material for use in molecular cytogenetics, specifically the technique of fluorescent in situ hybridisation (FISH). In this review, we provide an account of the current QD-FISH literature, and speculate as to why QDs are not yet optimised for FISH in their current form

Neurocranial osteology and neuroanatomy of a late Cretaceous Titanosaurian Sauropod from Spain (Ampelosaurus sp.)

Titanosaurians were a flourishing group of sauropod dinosaurs during Cretaceous times. Fossils of titanosaurians have been found on all continents and their remains are abundant in a number of Late Cretaceous sites. Nonetheless, the cranial anatomy of titanosaurians is still very poorly known. The Spanish latest Cretaceous locality of "Lo Hueco" yielded a relatively well preserved, titanosaurian braincase, which shares a number of phylogenetically restricted characters with Ampelosaurus atacis from France such as a flat occipital region. However, it appears to differ from A. atacis in some traits such as the greater degree of dorsoventral compression and the presence of proatlas facets. The specimen is, therefore, provisionally identified as Ampelosaurus sp. It was CT scanned, and 3D renderings of the cranial endocast and inner-ear system were generated. Our investigation highlights that, although titanosaurs were derived sauropods with a successful evolutionary history, they present a remarkably modest level of paleoneurological organization. Compared with the condition in the basal titanosauriform Giraffatitan brancai, the labyrinth of Ampelosaurus sp. shows a reduced morphology. The latter feature is possibly related to a restricted range of head-turning movementsThis is a contribution to the research project CGL2009-12143 (Ministerio de Economía y Competitividad, Madrid), of which FK, who is currently supported by the Ramón y Cajal Program, is Principal Investigator. LMW and RCR acknowledge funding support from the United States National Science Foundation (IBN-9601174, IBN-0343744, IOB-0517257, IOS-1050154) and the Ohio University Heritage College of Osteopathic Medicine. The Ohio Supercomputing Center also provided suppor

The Braincase of the Basal Sauropod Dinosaur Spinophorosaurus and 3D Reconstructions of the Cranial Endocast and Inner Ear

Background: Sauropod dinosaurs were the largest animals ever to walk on land, and, as a result, the evolution of their remarkable adaptations has been of great interest. The braincase is of particular interest because it houses the brain and inner ear. However, only a few studies of these structures in sauropods are available to date. Because of the phylogenetic position of Spinophorosaurus nigerensis as a basal eusauropod, the braincase has the potential to provide key evidence on the evolutionary transition relative to other dinosaurs. Methodology/Principal Findings: The only known braincase of Spinophorosaurus (‘Argiles de l'Irhazer’, Irhazer Group; Agadez region, Niger) differs significantly from those of the Jurassic sauropods examined, except potentially for Atlasaurus imelakei (Tilougguit Formation, Morocco). The basisphenoids of Spinophorosaurus and Atlasaurus bear basipterygoid processes that are comparable in being directed strongly caudally. The Spinophorosaurus specimen was CT scanned, and 3D renderings of the cranial endocast and inner-ear system were generated. The endocast resembles that of most other sauropods in having well-marked pontine and cerebral flexures, a large and oblong pituitary fossa, and in having the brain structure obscured by the former existence of relatively thick meninges and dural venous sinuses. The labyrinth is characterized by long and proportionally slender semicircular canals. This condition recalls, in particular, that of the basal non-sauropod sauropodomorph Massospondylus and the basal titanosauriform Giraffatitan. Conclusions/Significance: Spinophorosaurus has a moderately derived paleoneuroanatomical pattern. In contrast to what might be expected early within a lineage leading to plant-eating graviportal quadrupeds, Spinophorosaurus and other (but not all) sauropodomorphs show no reduction of the vestibular apparatus of the inner ear. This character-state is possibly a primitive retention in Spinophorosaurus, but due the scarcity of data it remains unclear whether it is also the case in the various later sauropods in which it is present or whether it has developed homoplastically in these taxa. Any interpretations remain tentative pending the more comprehensive quantitative analysis underway, but the size and morphology of the labyrinth of sauropodomorphs may be related to neck length and mobility, among other factors.The sojourns of Dr. Knoll in the Museum für Naturkunde (Berlin) were partly funded by the Alexander von Humboldt Foundation through a sponsorship of renewed research stay in Germany and by the European Community Research Infrastructure Action under the FP7 “Capacities” Program through a Synthesys grant (http://www.synthesys.info/). Dr. Knoll is currently supported by the Ramón y Cajal Program. This is a contribution to the research project CGL2009-12143, from the Ministerio de Ciencia e Innovación (Madrid), conducted by Dr. Knoll (PI), Dr. Witmer, and Dr. Schwarz-Wings. Dr. Witmer and Dr. Ridgely acknowledge funding support from the United States National Science Foundation (IBN-9601174, IBN-0343744, IOB-0517257) and the Ohio University Heritage College of Osteopathic Medicine. The Ohio Supercomputing Center also provided support.Peer reviewe

Human Mas-related G protein-coupled receptors-X1 induce chemokine receptor 2 expression in rat dorsal root ganglia neurons and release of chemokine ligand 2 from the human LAD-2 mast cell line

Primate-specific Mas-related G protein-coupled receptors-X1 (MRGPR-X1) are highly enriched in dorsal root ganglia (DRG) neurons and induce acute pain. Herein, we analyzed effects of MRGPR-X1 on serum response factors (SRF) or nuclear factors of activated T cells (NFAT), which control expression of various markers of chronic pain. Using HEK293, DRG neuron-derived F11 cells and cultured rat DRG neurons recombinantly expressing human MRGPR-X1, we found activation of a SRF reporter gene construct and induction of the early growth response protein-1 via extracellular signal-regulated kinases-1/2 known to play a significant role in the development of inflammatory pain. Furthermore, we observed MRGPR-X1-induced up-regulation of the chemokine receptor 2 (CCR2) via NFAT, which is considered as a key event in the onset of neuropathic pain and, so far, has not yet been described for any endogenous neuropeptide. Up-regulation of CCR2 is often associated with increased release of its endogenous agonist chemokine ligand 2 (CCL2). We also found MRGPR-X1-promoted release of CCL2 in a human connective tissue mast cell line endogenously expressing MRGPR-X1. Thus, we provide first evidence to suggest that MRGPR-X1 induce expression of chronic pain markers in DRG neurons and propose a so far unidentified signaling circuit that enhances chemokine signaling by acting on two distinct yet functionally co-operating cell types. Given the important role of chemokine signaling in pain chronification, we propose that interruption of this signaling circuit might be a promising new strategy to alleviate chemokine-promoted pain

A Comparative Plasmonic Study of Nanoporous and Evaporated Gold Films

Previously, we have reported that nanoporous gold (NPG) films prepared by a chemical dealloying method have distinctive plasmonic properties, i.e., they can simultaneously support localized and propagating surface plasmon resonance modes (l-SPR and p-SPR, respectively). In this study, the plasmonic properties of NPG are quantified through direct comparison with thermally evaporated gold (EG) films. Cyclic voltammetry and electrochemical impedance spectroscopy experiments reveal that the NPG films have 4–8.5 times more accessible surface area than EG films. Assemblies of streptavidin–latex beads generate p-SPR responses on both NPG and EG films that correlate well with the bead density obtained from scanning electron microscopy (SEM) images. A layer-by-layer assembly experiment on NPG involving biotinylated anti-avidin IgG and avidin, studied by l-SPR and SEM, shows that the l-SPR signal is directly linked to the accessibility of the interior of the NPG porosity, an adjustable experimental parameter that can be set by the dealloying condition and time

ZyFISH: A Simple, Rapid and Reliable Zygosity Assay for Transgenic Mice

Microinjection of DNA constructs into fertilized mouse oocytes typically results in random transgene integration at a single genomic locus. The resulting transgenic founders can be used to establish hemizygous transgenic mouse lines. However, practical and experimental reasons often require that such lines be bred to homozygosity. Transgene zygosity can be determined by progeny testing assays which are expensive and time-consuming, by quantitative Southern blotting which is labor-intensive, or by quantitative PCR (qPCR) which requires transgene-specific design. Here, we describe a zygosity assessment procedure based on fluorescent in situ hybridization (zyFISH). The zyFISH protocol entails the detection of transgenic loci by FISH and the concomitant assignment of homozygosity using a concise and unbiased scoring system. The method requires small volumes of blood, is scalable to at least 40 determinations per assay, and produces results entirely consistent with the progeny testing assay. This combination of reliability, simplicity and cost-effectiveness makes zyFISH a method of choice for transgenic mouse zygosity determinations

The overmethylated genes in Helicobacter pylori-infected gastric mucosa are demethylated in gastric cancers

<p>Abstract</p> <p>Background</p> <p>The transitional-CpG sites between weakly methylated genes and densely methylated retroelements are overmethylated in the gastric mucosa infected with <it>Helicobacter pylori </it>(<it>H. pylori</it>) and they are undermethylated in the gastric cancers depending on the level of loss of heterozygosity (LOH) events. This study delineated the transitional-CpG methylation patterns of CpG-island-containing and -lacking genes in view of the retroelements.</p> <p>Methods</p> <p>The transitional-CpG sites of eight CpG-island-containing genes and six CpG-island-lacking genes were semi-quantitatively examined by performing radioisotope-labelling methylation-specific PCR under stringent conditions. The level of LOH in the gastric cancers was estimated using the 40 microsatellite markers on eight cancer-associated chromosomes. Each gene was scored as overmethylated or undermethylated based on an intermediate level of transitional-CpG methylation common in the <it>H. pylori</it>-negative gastric mucosa.</p> <p>Results</p> <p>The eight CpG-island genes examined were overmethylated depending on the proximity to the nearest retroelement in the <it>H. pylori</it>-positive gastric mucosa. The six CpG-island-lacking genes were similarly methylated in the <it>H. pylori</it>-positive and -negative gastric mucosa. In the gastric cancers, long transitional-CpG segments of the CpG-island genes distant from the retroelements remained overmethylated, whereas the overmethylation of short transitional-CpG segments close to the retroelements was not significant. Both the CpG-island-containing and -lacking genes tended to be decreasingly methylated in a LOH-level-dependent manner.</p> <p>Conclusions</p> <p>The overmethylated genes under the influence of retroelement methylation in the <it>H. pylori</it>-infected stomach are demethylated in the gastric cancers influenced by LOH.</p