High fluorescence quantum efficiency of CdSe/ZnS quantum dots embedded in GPTS/TEOS-derived organic/silica hybrid colloids

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Fluorescence Intensity of Composite Layering Combined with Surface Sealant Submitted to Staining Solutions

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Surface-spreading technique of meiotic cells and immunodetection of synaptonemal complex proteins in teleostean fishes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Toxicogenetic effects of low concentrations of the pesticides imidacloprid and sulfentrazone individually and in combination in in vitro tests with HepG2 cells and Salmonella typhimurium

The insecticide imidacloprid and the herbicide sulfentrazone are two different classes of pesticides that are used for pest control in sugarcane agriculture. To evaluate the genotoxic potential of low concentrations of these two pesticides alone and in mixture, the comet assay and the micronucleus (MN) test employing fluorescence in situ hybridization (FISH) with a centromeric probe were applied in human hepatoma cell lines (HepG2), in a 24-h assay. Mutagenicity was assessed by Salmonella/microsome assay with TA98 and TA100 strains in the absence and presence of an exogenous metabolizing system (S9). The results showed significant inductions of MN in HepG2 cells by both pesticides, for all the tested concentrations. As evidenced in the comet assay, only the imidacloprid presented significant responses. When the two pesticides were associated, a significant induction of damage was observed in the HepG2 cells by the comet assay, but not by the MN test. Moreover, the MN induced by the mixtures of the pesticides appeared at lower levels than those induced by sulfentrazone and imidacloprid when tested alone. According to the FISH results, the damage induced by imidacloprid in the HepG2 cells resulted from a clastogenic action of this insecticide (76.6% of the MN did not present a centromeric signal). For the herbicide sulfentrazone and for the mixture of the pesticides, a similar frequency of MN with and without the presence of the centromeric signal (herbicide: 52.45% of the MN without centromeric signal and 47.54% of the MN with centromeric signal; mixture: 48.71% of the MN without centromeric signal and 51.42% of the MN with centromeric signal) was verified. Based on these results, it was concluded that each one of the pesticides evaluated interacts with the DNA of HepG2 cells and causes irreparable alterations in the cells. However, the combination of the pesticides showed an antagonistic effect on the cells and the damage induced was milder and not persistent in HepG2 cells. The results obtained by the Ames test did not point out significant results.

In situ selective determination of methylmercury in river water by diffusive gradient in thin films technique (DGT) using baker's yeast (Saccharomyces cerevisiae) immobilized in agarose gel as binding phase

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Constraining H-0 in general dark energy models from Sunyaev-Zeldovich/X-ray technique and complementary probes

In accelerating dark energy models, the estimates of the Hubble constant, Ho, from Sunyaev-Zerdovich effect (SZE) and X-ray surface brightness of galaxy clusters may depend on the matter content (Omega(M)), the curvature (Omega(K)) and the equation of state parameter GO. In this article, by using a sample of 25 angular diameter distances of galaxy clusters described by the elliptical beta model obtained through the SZE/X-ray technique, we constrain Ho in the framework of a general ACDM model (arbitrary curvature) and a flat XCDM model with a constant equation of state parameter omega = p(x)/rho(x). In order to avoid the use of priors in the cosmological parameters, we apply a joint analysis involving the baryon acoustic oscillations (BA()) and the (MB Shift Parameter signature. By taking into account the statistical and systematic errors of the SZE/X-ray technique we obtain for nonflat ACDM model H-0 = 74(-4.0)(+5.0) km s(-1) Mpc(-1) (1 sigma) whereas for a fiat universe with constant equation of state parameter we find H-0 = 72(-4.0)(+5.5) km s(-1) Mpc(-1)(1 sigma). By assuming that galaxy clusters are described by a spherical beta model these results change to H-0 = 6(-7.0)(+8.0) and H-0 = 59(-6.0)(+9.0) km s(-1) Mpc(-1)(1 sigma), respectively. The results from elliptical description are in good agreement with independent studies from the Hubble Space Telescope key project and recent estimates based on the Wilkinson Microwave Anisotropy Probe, thereby suggesting that the combination of these three independent phenomena provides an interesting method to constrain the Bubble constant. As an extra bonus, the adoption of the elliptical description is revealed to be a quite realistic assumption. Finally, by comparing these results with a recent determination for a, flat ACDM model using only the SZE/X-ray technique and BAO, we see that the geometry has a very weak influence on H-0 estimates for this combination of data.

Design and synthesis of a new coumarin-based 'turn-on' fluorescent probe selective for Cu+2

The novel coumarin-based 'turn-on' fluorescent probe (E)-3-(2,5-dimethoxybenzylideneamino)-7-hydroxy-2H-chromen-2-one (MGM) was designed, synthesized, and characterized. This compound shows high selectivity for Cu+2, combined with a large fluorescence enhancement upon binding to Cu2+. Benesi-Hildebrand and Job plots demonstrate that the stoichiometry of the Cu+2 complex formed is 2:1. Preliminary studies employing epifluorescence microscopy demonstrated that Cu+2 could be imaged in human neuroblastoma SH-SY5Y cells treated with MGM. (c) 2012 Elsevier Ltd. All rights reserved.

Fluorescence spectroscopy as a tool to detect and evaluate glucocorticoid-induced skin atrophy

Topical glucocorticoid (GC) therapy has been successfully used in the treatment of several common cutaneous diseases in clinical practice for a long time, and skin atrophy is one of the most typical cutaneous side effects of this therapy. The aim of this study was to evaluate the potential of noninvasive fluorescence spectroscopy (FS) technique in the detection and classification of GC-induced skin atrophy. A total of 20 male Wistar rats were used in the experimental protocol under controlled environmental conditions and with free access to food. One group received topical application of clobetasol propionate 0.05% for 14 days to induce cutaneous atrophy (atrophic group) and the other (control) group received only vehicle application following the same protocol and schedule. Histological analyses and FS measurements with laser excitation at both 532 nm and 408 nm were obtained on days 1 and 15. The FS results were classified as "normal" or "atrophic" according by histological analysis. Fluorescence spectra obtained with excitation at 408 nm allowed a clear distinction between the control and atrophic groups, and were more informative than the those obtained at 532 nm. Our results reveal that, if correctly applied, FS allows noninvasive evaluation of corticosteroid-induced skin atrophy, and thus represents an important step towards better monitoring of undesirable side effects of cutaneous therapy.

A light in the shadow: the use of Lucifer Yellow technique to demonstrate nectar reabsorption

Abstract Background Nectar reabsorption is a widely known phenomenon, related to the strategy of resource-recovery and also to maintain the nectar homeostasis at the nectary. The method currently performed to demonstrate nectar being reabsorbed involves the use of radioactive tracers applied to the nectary. Although this method works perfectly, it is complex and requires specific supplies and equipment. Therefore, here we propose an efficient method to obtain a visual demonstration of nectar reabsorption, adapting the use of Lucifer Yellow CH (LYCH), a fluorescent membrane-impermeable dye that can enter the vacuole by endocytosis. Results We applied a LYCH solution to the floral nectary (FN) of Cucurbita pepo L., which is a species known for its ability of nectar reabsorption, and to the extrafloral nectary (EFN) of Passiflora edulis Sims which does not reabsorb the secreted nectar. In all tests performed, we observed that LYCH stained the nectary tissues differentially according to the reabsorption ability of the nectary. The treated FN of C. pepo presented a concentrated fluorescence at the epidermis that decreased at the deeper nectary parenchyma, until reaching the vascular bundles, indicating nectar reabsorption in the flowers of the species. In contrast, treated EFN of P. edulis presented fluorescence only at the cuticle surface, indicating that nectar is not reabsorbed by that particular tissue. Conclusion LYCH is an efficient marker to demonstrate nectar reabsorption.

Copy number variation in Williams-Beuren syndrome: suitable diagnostic strategy for developing countries

Abstract Background Williams-Beuren syndrome (WBS; OMIM 194050) is caused by a hemizygous contiguous gene microdeletion at 7q11.23. Supravalvular aortic stenosis (SVAS), mental retardation, and overfriendliness comprise typical symptoms of WBS. Although fluorescence in situ hybridization (FISH) is considered the gold standard technique, the microsatellite DNA markers and multiplex ligation-dependent probe amplification (MLPA) could be used for to confirm the diagnosis of WBS. Results We have evaluated a total cohort of 88 patients with a suspicion clinical diagnosis of WBS using a collection of five markers (D7S1870, D7S489, D7S613, D7S2476, and D7S489_A) and a commercial MLPA kit (P029). The microdeletion was present in 64 (72.7%) patients and absent in 24 (27.3%) patients. The parental origin of deletion was maternal in 36 of 64 patients (56.3%) paternal in 28 of 64 patients (43.7%). The deletion size was 1.55 Mb in 57 of 64 patients (89.1%) and 1.84 Mb in 7 of 64 patients (10.9%). The results were concordant using both techniques, except for four patients whose microsatellite markers were uninformative. There were no clinical differences in relation to either the size or parental origin of the deletion. Conclusion MLPA was considered a faster and more economical method in a single assay, whereas the microsatellite markers could determine both the size and parental origin of the deletion in WBS. The microsatellite marker and MLPA techniques are effective in deletion detection in WBS, and both methods provide a useful diagnostic strategy mainly for developing countries.

Clinicopathological and molecular characterization of gastroesophageal junction (GEJ) adenocarcinoma before age of 40 years

Gastroesophageal junction (GEJ) adenocarcinoma are uncommon before age of 40 years. While certain clinical, pathological and molecular features of GEJ adenocarcinoma in older patients have been extensively studied, these characteristics in the younger population remain to be determined. In the recent literature, a high sensitivity and specificity for the detection of dysplasia and esophageal adenocarcinoma was demonstrated by using multicolor fluorescence in situ hybridization (FISH) DNA probe set specific for the locus specific regions 9p21 (p16), 20q13.2 and Y chromosome. We evaluated 663 patients with GEJ adenocarcinoma and further divided them into 2 age-groups of or= 50 years, rispectively. FISH with selected DNA probe for Y chromosome, locus 9p21 (p16), and locus 20q13.2 was investigated with formalin fixed and parassin embedded tissue from surgical resections of 17 younger and 11 older patients. Signals were counted in > 100 cells with each given histopathological category. The chromosomal aberrations were then compared in the 2 age-groups with the focus on uninvolved squamous and columnar epithelium, intestinal metaplasia (Barrett's mucosa), glandular dysplasia, and adenocarcinoma. Comparisons were performed by the X2 test, Fisher's exact test, Student's t-test and Mann-Whitney U-test as appropriate. Survival was estimated by the Kaplan-Meier method with univariate analysis by the log-rank. Significance was taken at the 5% level. There was no difference in the surgical technique applied in both age groups and most patients underwent Ivor Lewis esophagectomy. Among clinical variables there was a higher incidence of smocking history in older patient group. We identified a progressive loss of Y chromosome from benign squamos epithelium to Barrett's mucosa and glandular dysplasia, and, ultimately, to a near complete loss in adenocarcinoma in both age groups. The young group revealed significantly more losses of 9p21 in both benign and neoplastic cells when compared to the older patients group. In addition, we demonstrated an increase in the percentage of cells showing gain of locus 20q13.2 with progression from benign epithelium through dysplasia to adenocarcinoma with almost the same trend in both the young and the older patients. When compared with the older age-group, younger patients with GEJ adenocarcinoma possess similar known demographics, environmental factors, clinical, and pathologic characteristics. The most commonly detected genetic aberrations of progressive Y chromosomal loss, 9p21 locus loss, and 20q13 gains were similar in the younger and older patients. However the rate of loss of 9p21 is significantly higher in young patients, in both the benign and the neoplastic cells. The loss of 9p21, and possibly, the subsequent inactivation of p16 gene may be one of the molecular mechanisms responsible for the accelerated neoplastic process in young patients.

Strategies for detecting DNA hybridisation using surface plasmon fluorescence spectroscopy

Die Anregung und Emission von Fluorophoren nahe planaren Metalloberflächen und schiefen Gittern wurde mittels Oberflächenplasmonen Fluoreszenz Spektroskopie (SPFS) untersucht. Die Fluorophore konnten durch das evaneszente Plasmonenfeld angeregt und die einzelnen Abregungskanäle identifiziert werden.Die Sensorarchitektur für den Nachweis der Hybridisierung bestand aus auf einer Streptavidin-Matrix immobilisierten unmarkierten Sondensträngen. Cy5 markierte Zielsequenzen wurden aus der Lösung hybridisiert und die Adsorptionskinetiken konnten oberflächensensitiv detektiert werden.Ein neues Detektionsschema für unmarkierte Zielstränge wurde mittels fluoreszenzmarkirten Sondensträngen realisiert. Die Hybridisierung führte zu der Bildung von steifen helikalen Bereichen in der Probe und separierte den Farbstoff von der Metalloberfläche. Reduzierte Fluorezenzlöschung zeigte daher das Hybridisierungsereignis an.Die Verwendung eines potentiellen Förster-Paares zur Detektion von DNA Hybridisierung wurde untersucht. Donor und Akzeptor wurden an Ziel- und Sondenstrang immobilisiert und das Hybridisierungsereignis konnte anhand der Auslöschung der Donor-Fluorezenz nachgewiesen werden.Schließlich wurde der Einsatz von einzelstrangbindenden Proteinen (SSB) zur Steigerung der Sensitivität bezüglich Basenfehlpaarungen betrachtet. Verdrängungsreaktionen zwischen Proteinen und markierten Zielsträngen wurden anhand von SPS und Fluorezenzkinetiken studiert.

Development of a new galaxy classification technique and its application to the zCOSMOS survey

The purpose of this Thesis is to develop a robust and powerful method to classify galaxies from large surveys, in order to establish and confirm the connections between the principal observational parameters of the galaxies (spectral features, colours, morphological indices), and help unveil the evolution of these parameters from $z \sim 1$ to the local Universe. Within the framework of zCOSMOS-bright survey, and making use of its large database of objects ($\sim 10\,000$ galaxies in the redshift range $0 < z \lesssim 1.2$) and its great reliability in redshift and spectral properties determinations, first we adopt and extend the \emph{classification cube method}, as developed by Mignoli et al. (2009), to exploit the bimodal properties of galaxies (spectral, photometric and morphologic) separately, and then combining together these three subclassifications. We use this classification method as a test for a newly devised statistical classification, based on Principal Component Analysis and Unsupervised Fuzzy Partition clustering method (PCA+UFP), which is able to define the galaxy population exploiting their natural global bimodality, considering simultaneously up to 8 different properties. The PCA+UFP analysis is a very powerful and robust tool to probe the nature and the evolution of galaxies in a survey. It allows to define with less uncertainties the classification of galaxies, adding the flexibility to be adapted to different parameters: being a fuzzy classification it avoids the problems due to a hard classification, such as the classification cube presented in the first part of the article. The PCA+UFP method can be easily applied to different datasets: it does not rely on the nature of the data and for this reason it can be successfully employed with others observables (magnitudes, colours) or derived properties (masses, luminosities, SFRs, etc.). The agreement between the two classification cluster definitions is very high. ``Early'' and ``late'' type galaxies are well defined by the spectral, photometric and morphological properties, both considering them in a separate way and then combining the classifications (classification cube) and treating them as a whole (PCA+UFP cluster analysis). Differences arise in the definition of outliers: the classification cube is much more sensitive to single measurement errors or misclassifications in one property than the PCA+UFP cluster analysis, in which errors are ``averaged out'' during the process. This method allowed us to behold the \emph{downsizing} effect taking place in the PC spaces: the migration between the blue cloud towards the red clump happens at higher redshifts for galaxies of larger mass. The determination of $M_{\mathrm{cross}}$ the transition mass is in significant agreement with others values in literature.

Troposphere Calibration Techniques for Deep Space Probe Tracking

Ground-based Earth troposphere calibration systems play an important role in planetary exploration, especially to carry out radio science experiments aimed at the estimation of planetary gravity fields. In these experiments, the main observable is the spacecraft (S/C) range rate, measured from the Doppler shift of an electromagnetic wave transmitted from ground, received by the spacecraft and coherently retransmitted back to ground. If the solar corona and interplanetary plasma noise is already removed from Doppler data, the Earth troposphere remains one of the main error sources in tracking observables. Current Earth media calibration systems at NASA’s Deep Space Network (DSN) stations are based upon a combination of weather data and multidirectional, dual frequency GPS measurements acquired at each station complex. In order to support Cassini’s cruise radio science experiments, a new generation of media calibration systems were developed, driven by the need to achieve the goal of an end-to-end Allan deviation of the radio link in the order of 3×〖10〗^(-15) at 1000 s integration time. The future ESA’s Bepi Colombo mission to Mercury carries scientific instrumentation for radio science experiments (a Ka-band transponder and a three-axis accelerometer) which, in combination with the S/C telecommunication system (a X/X/Ka transponder) will provide the most advanced tracking system ever flown on an interplanetary probe. Current error budget for MORE (Mercury Orbiter Radioscience Experiment) allows the residual uncalibrated troposphere to contribute with a value of 8×〖10〗^(-15) to the two-way Allan deviation at 1000 s integration time. The current standard ESA/ESTRACK calibration system is based on a combination of surface meteorological measurements and mathematical algorithms, capable to reconstruct the Earth troposphere path delay, leaving an uncalibrated component of about 1-2% of the total delay. In order to satisfy the stringent MORE requirements, the short time-scale variations of the Earth troposphere water vapor content must be calibrated at ESA deep space antennas (DSA) with more precise and stable instruments (microwave radiometers). In parallel to this high performance instruments, ESA ground stations should be upgraded to media calibration systems at least capable to calibrate both troposphere path delay components (dry and wet) at sub-centimetre level, in order to reduce S/C navigation uncertainties. The natural choice is to provide a continuous troposphere calibration by processing GNSS data acquired at each complex by dual frequency receivers already installed for station location purposes. The work presented here outlines the troposphere calibration technique to support both Deep Space probe navigation and radio science experiments. After an introduction to deep space tracking techniques, observables and error sources, in Chapter 2 the troposphere path delay is widely investigated, reporting the estimation techniques and the state of the art of the ESA and NASA troposphere calibrations. Chapter 3 deals with an analysis of the status and the performances of the NASA Advanced Media Calibration (AMC) system referred to the Cassini data analysis. Chapter 4 describes the current release of a developed GNSS software (S/W) to estimate the troposphere calibration to be used for ESA S/C navigation purposes. During the development phase of the S/W a test campaign has been undertaken in order to evaluate the S/W performances. A description of the campaign and the main results are reported in Chapter 5. Chapter 6 presents a preliminary analysis of microwave radiometers to be used to support radio science experiments. The analysis has been carried out considering radiometric measurements of the ESA/ESTEC instruments installed in Cabauw (NL) and compared with the requirements of MORE. Finally, Chapter 7 summarizes the results obtained and defines some key technical aspects to be evaluated and taken into account for the development phase of future instrumentation.

New discotic liquid crystals based on large polycyclic aromatic hydrocarbons as materials for molecular electronics

A series of new columnar discotic liquid crystalline materials based on the superphenalene (C96) core has been synthesized by oxidative cyclodehydrogenation with iron(III) chloride of suitable three-dimensional oligophenylene precursors. These compounds were investigated by means of differential scanning calorimetry (DSC), polarized optical microscopy (POM) and wide angle X-ray scattering (WAXS), and showed highly ordered supramolecular arrays and mesophase behavior over a broad temperature range. Good solubility, through the introduction of long alkyl chains, and the fact that these new superphenalene derivatives were found to be liquid crystalline at room temperature enabled the formation of highly ordered films (using the zone-casting technique), a requirement for application in organic electronic devices. The one-dimensional, intracolumnar charge carrier mobilities of superphenalene derivatives were determined using the pulse-radiolysis time-resolved microwave conductivity technique (PR-TRMC). Electrical properties of different C96-C12 architectures on mica surfaces were examined by using Electrostatic Force Microscopy (EFM) and Kelvin Probe Force Microscopy (KPFM). Hexa-peri-hexabenzocoronene (C42) derivatives substituted at the periphery with six branched alkyl ether chains were also synthesized. It was found that the introduction of ether groups within the side chains enhances the affinity of the discotic molecules towards polar surfaces, resulting in homeotropic self-assembly (as shown by POM and 2D-WAXS) when the compounds are processed from the isotropic state between two surfaces. A new, insoluble, superphenalene building block bearing six reactive sites was prepared, and was further used for the preparation of dendronized superphenalenes with bulky dendritic substituents around the core. UV/Vis and fluorescence experiments suggest reduced π-π stacking of the superphenalene cores as a result of steric hindrance between the peripheral dendritic units. A new family of graphitic molecules with partial ”zig-zag” periphery has been established. The incorporation of ”zig-zag” edges was shown to have a strong influence on the electronic properties of the new molecules (as studied by solution and solid-state UV/Vis, and fluorescence spectroscopy), leading to a significant bathochromic shift with respect to the parent PAHs (C42 and C96). The reactivity of the additional double bonds was examined. The attachment of long alkyl chains to a ”zig-zag” superphenalene core afforded a new, processable, liquid crystalline material.