Detection of circulating polysaccharide antigen of Schistosoma mansoni in hamster sera by crossed immunoelectrophoresis

Crossed immunoelectrophoresis (IEC) was used for detection of free and complexed circulating polisaccharide anodic antigen (AgCA) of Schistosoma mansoni in sera of infected hamsters. An attempt was also done to detect AgCA in human sera from patients infected with S. mansoni. The conditions for isolation and detection of complexed AgCA were established. The sensitivity of IEC was increased by incorporation of 2% polyethylene glicol (PEG) to the agarose and by maintaining the system at 4°C during the electrophoretic run. Free AgCA was detected in 12 and the complexed in 30 of the 37 hamsters sera analysed. Correlation between AgCA (free and complexed) and the parasite load was observed. AgCA was not detected, under the experimental conditions used, in human sera from 7 patients in the acute and 23 in the chronic phase of infection.

Using device detection techniques in M-learning scenarios

Recent studies of mobile Web trends show the continued explosion of mobile-friend content. However, the wide number and heterogeneity of mobile devices poses several challenges for Web programmers, who want automatic delivery of context and adaptation of the content to mobile devices. Hence, the device detection phase assumes an important role in this process. In this chapter, the authors compare the most used approaches for mobile device detection. Based on this study, they present an architecture for detecting and delivering uniform m-Learning content to students in a Higher School. The authors focus mainly on the XML device capabilities repository and on the REST API Web Service for dealing with device data. In the former, the authors detail the respective capabilities schema and present a new caching approach. In the latter, they present an extension of the current API for dealing with it. Finally, the authors validate their approach by presenting the overall data and statistics collected through the Google Analytics service, in order to better understand the adherence to the mobile Web interface, its evolution over time, and the main weaknesses.

Simplification of Immune Adherence Hemagglutination test for detection of rabies antibodies in human serum

In the present work the immune adherence hemagglutination test (IAHA) was standardized in a simplified procedure. This test showed good reproducibility, better than the classical mice serum neutralization test (SN). The tests showed high correlation degree: high titers in one test corresponded to high titers in the other one, and the same occured with low titers. The IAHA test is extremely simple, fast to perform, and of low cost when compared to tests such as SN or indirect immunofluorescense (IIF). It also proved to be useful in less sophisticated laboratories or even as a screening test for the titration of rabies antibodies.

Evaluation of enzyme-linked immunosorbent and alternative assays for detection of HIV antibodies using panels of Brasilian sera

Sera from 472 Brazilian subjects, confirmed to be either positive or negative for HIV antibodies and comprising the total clinical spectrum of HIV infection, were utilized in the evaluation of six commercially available enzyme-linked immunosorbent assays (ELISA), as well as of four alternative assays, namely indirect immunofluorescence (IIF), passive hemagglutination (PHA), dot blot and Karpas AIDS cell test. The sensitivities ranged from 100% (Abbott and Roche ELISA) to 84.2% (PHA) and the specificities ranged from 99.3% (IIF) to 80.2% (PHA). The sensitivity and specificity of the PHA and the sensitivity of the Karpas cell test were significantly lower than those of the other tests. Although the IFF and dot blot had good sensitivities and specificities, the six ELISA were more attractive than those tests when other parameters such as ease of reading and duration of assay were considered.

DOT-ELISA for detection of anti-Cysticercus cellulosae antibodies in human cerebrospinal fluid using a new solid-phase (resin-treated polyester fabrics): preliminary report

A dot enzyme-linked immunosorbent assay (DOT-ELISA) was developed to detect specific antibodies in cerebrospinal fluid (CSF) for human neurocysticercosis immunodiagnosis, with Cysticercus cellulosae antigen dotted on a new solid-phase. This was represented by sheets of a synthetic polyester fabric impregnated with a polymerized resin (N-methylol-acrylamide). A very stable preparation was thus obtained, the antigen being covalently bound by cross-linking with free N-methylol groups on the resin. Since robust, no special care was necessary for handling the solid-phase. The test could be performed at room-temperature. From 30 CSF samples assayed, 14 were positive, from a group of 15 cases of neurocysticercosis, with titers from 1 to 128; 15 other samples, from normals or other neurological diseases, were all negative. Test characteristics seem to indicate it as adequate for epidemiological surveys. A more detailed study on sensitivity, specificity, reproducibility and the use in serum samples is being conducted.

Automated illustration of multimedia stories

Submitted in part fulfillment of the requirements for the degree of Master in Computer Science

PINPIN a-Si: H based structures for X-ray image detection using the laser scanning technique

Conventional film based X-ray imaging systems are being replaced by their digital equivalents. Different approaches are being followed by considering direct or indirect conversion, with the later technique dominating. The typical, indirect conversion, X-ray panel detector uses a phosphor for X-ray conversion coupled to a large area array of amorphous silicon based optical sensors and a couple of switching thin film transistors (TFT). The pixel information can then be readout by switching the correspondent line and column transistors, routing the signal to an external amplifier. In this work we follow an alternative approach, where the electrical switching performed by the TFT is replaced by optical scanning using a low power laser beam and a sensing/switching PINPIN structure, thus resulting in a simpler device. The optically active device is a PINPIN array, sharing both front and back electrical contacts, deposited over a glass substrate. During X-ray exposure, each sensing side photodiode collects photons generated by the scintillator screen (560 nm), charging its internal capacitance. Subsequently a laser beam (445 nm) scans the switching diodes (back side) retrieving the stored charge in a sequential way, reconstructing the image. In this paper we present recent work on the optoelectronic characterization of the PINPIN structure to be incorporated in the X-ray image sensor. The results from the optoelectronic characterization of the device and the dependence on scanning beam parameters are presented and discussed. Preliminary results of line scans are also presented. (C) 2014 Elsevier B.V. All rights reserved.

LADAR based mapping and obstacle detection system for service robots

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para a obtenção do grau de Mestre em Engenharia Electrotécnica e de Computadores

Damage detection in structures using a transmissibility-based Mahalanobis distance

In this paper, a damage-detection approach using the Mahalanobis distance with structural forced dynamic response data, in the form of transmissibility, is proposed. Transmissibility, as a damage-sensitive feature, varies in accordance with the damage level. Besides, Mahalanobis distance can distinguish the damaged structural state condition from the undamaged one by condensing the baseline data. For comparison reasons, the Mahalanobis distance results using transmissibility are compared with those using frequency response functions. The experiment results reveal quite a significant capacity for damage detection, and the comparison between the use of transmissibility and frequency response functions shows that, in both cases, the different damage scenarios could be well detected. Copyright (c) 2015 John Wiley & Sons, Ltd.

Enzyme-linked immunosorbent assay ELISA for the detection of antibodies in the human leptospirosis

An Enzyme-linked immunosorbent assay ELISA was evaluated for the detection of IgA antibodies in the human leptospirosis. The assay proved to be sensitive and specific when compared with the ELISA-IgM, in the examinated serum samples. The results found suggest that IgA antibodies became positive later in leptospirosis, and will can be an evolutive indicator in the development of the disease

Human papillomavirus detection in cervical dysplasias or neoplasias and in condylomata acuminaata by in situ hybridization with biotinylated DNA probes

Specimens from cervical dysplasias or carcinomas and genital condylomata acuminata were retrospectively analysed by in situ hybridization (ISH) with bioti-nylated DNA probes for human papillomavirus (HPV) types 6, 11, 16 and 18. In the control group no case was positive for HPV DNA. In mild/moderate dysplasias, 4 cases (14%) were positive for HPV 6 or 11 and 2 cases (7%), for HPV 16. In the severe dysplasia/in situ carcinoma group, 9 cases (31%) showed presence of DNA of HPV types 16 or 18. Six invasive carcinomas (20%) were positive for HPV type 16 or 18. Among condylomata acuminata, 22 cases (73%) were positive for HPV types 6 or 11. In all ISH-positive cases only one viral type was detected. No correlation between HPV DNA positivity and histological findings of HPV infection was observed. Although less sensitive than some other molecular biology techniques, in situ hybridization with biotinylated DNA probes proved to be simple and useful for detecting and typing HPV in samples routinely received for histopathological analysis.

Sensitive bi-enzymatic biosensor based on polyphenoloxidases–gold nanoparticles–chitosan hybrid film–graphene doped carbon paste electrode for carbamates detection

A bi-enzymatic biosensor (LACC–TYR–AuNPs–CS/GPE) for carbamates was prepared in a single step by electrodeposition of a hybrid film onto a graphene doped carbon paste electrode (GPE). Graphene and the gold nanoparticles (AuNPs) were morphologically characterized by transmission electron microscopy, X-ray photoelectron spectroscopy, dynamic light scattering and laser Doppler velocimetry. The electrodeposited hybrid film was composed of laccase (LACC), tyrosinase (TYR) and AuNPs entrapped in a chitosan (CS) polymeric matrix. Experimental parameters, namely graphene redox state, AuNPs:CS ratio, enzymes concentration, pH and inhibition time were evaluated. LACC–TYR–AuNPs–CS/GPE exhibited an improved Michaelis–Menten kinetic constant (26.9 ± 0.5 M) when compared with LACC–AuNPs–CS/GPE (37.8 ± 0.2 M) and TYR–AuNPs–CS/GPE (52.3 ± 0.4 M). Using 4-aminophenol as substrate at pH 5.5, the device presented wide linear ranges, low detection limits (1.68×10− 9 ± 1.18×10− 10 – 2.15×10− 7 ± 3.41×10− 9 M), high accuracy, sensitivity (1.13×106 ± 8.11×104 – 2.19×108 ± 2.51×107 %inhibition M− 1), repeatability (1.2–5.8% RSD), reproducibility (3.2–6.5% RSD) and stability (ca. twenty days) to determine carbaryl, formetanate hydrochloride, propoxur and ziram in citrus fruits based on their inhibitory capacity on the polyphenoloxidases activity. Recoveries at two fortified levels ranged from 93.8 ± 0.3% (lemon) to 97.8 ± 0.3% (orange). Glucose, citric acid and ascorbic acid do not interfere significantly in the electroanalysis. The proposed electroanalytical procedure can be a promising tool for food safety control.

Fast-to-fed shift in glucose homeostasis: clues to an earlier detection of human prediabetic states

FCM: UC Bioquímica I - PhD Thesis

The role of the efflux mechanisms in multidrug resistance in Mycobacterium tuberculosis

Abstract The emergence of multi and extensively drug resistant tuberculosis (MDRTB and XDRTB) has increased the concern of public health authorities around the world. The World Health Organization has defined MDRTB as tuberculosis (TB) caused by organisms resistant to at least isoniazid and rifampicin, the main first-line drugs used in TB therapy, whereas XDRTB refers to TB resistant not only to isoniazid and rifampicin, but also to a fluoroquinolone and to at least one of the three injectable second-line drugs, kanamycin, amikacin and capreomycin. Resistance in Mycobacterium tuberculosis is mainly due to the occurrence of spontaneous mutations and followed by selection of mutants by subsequent treatment. However, some resistant clinical isolates do not present mutations in any genes associated with resistance to a given antibiotic, which suggests that other mechanism(s) are involved in the development of drug resistance, namely the presence of efflux pump systems that extrude the drug to the exterior of the cell, preventing access to its target. Increased efflux activity can occur in response to prolonged exposure to subinhibitory concentrations of anti-TB drugs, a situation that may result from inadequate TB therapy. The inhibition of efflux activity with a non-antibiotic inhibitor may restore activity of an antibiotic subject to efflux and thus provide a way to enhance the activity of current anti-TB drugs. The work described in this thesis foccus on the study of efflux mechanisms in the development of multidrug resistance in M. tuberculosis and how phenotypic resistance, mediated by efflux pumps, correlates with genetic resistance. In order to accomplish this goal, several experimental protocols were developed using biological models such as Escherichia coli, the fast growing mycobacteria Mycobacterium smegmatis, and Mycobacterium avium, before their application to M. tuberculosis. This approach allowed the study of the mechanisms that result in the physiological adaptation of E. coli to subinhibitory concentrations of tetracycline (Chapter II), the development of a fluorometric method that allows the detection and quantification of efflux of ethidium bromide (Chapter III), the characterization of the ethidium bromide transport in M. smegmatis (Chapter IV) and the contribution of efflux activity to macrolide resistance in Mycobacterium avium complex (Chapter V). Finally, the methods developed allowed the study of the role of efflux pumps in M. tuberculosis strains induced to isoniazid resistance (Chapter VI). By this manner, in Chapter II it was possible to observe that the physiological adaptation of E. coli to tetracycline results from an interplay between events at the genetic level and protein folding that decrease permeability of the cell envelope and increase efflux pump activity. Furthermore, Chapter III describes the development of a semi-automated fluorometric method that allowed the correlation of this efflux activity with the transport kinetics of ethidium bromide (a known efflux pump substrate) in E. coli and the identification of efflux inhibitors. Concerning M. smegmatis, we have compared the wild-type M. smegmatis mc2155 with knockout mutants for LfrA and MspA for their ability to transport ethidium bromide. The results presented in Chapter IV showed that MspA, the major porin in M. smegmatis, plays an important role in the entrance of ethidium bromide and antibiotics into the cell and that efflux via the LfrA pump is involved in low-level resistance to these compounds in M. smegmatis. Chapter V describes the study of the contribution of efflux pumps to macrolide resistance in clinical M. avium complex isolates. It was demonstrated that resistance to clarithromycin was significantly reduced in the presence of efflux inhibitors such as thioridazine, chlorpromazine and verapamil. These same inhibitors decreased efflux of ethidium bromide and increased the retention of [14C]-erythromycin in these isolates. Finaly, the methods developed with the experimental models mentioned above allowed the study of the role of efflux pumps on M. tuberculosis strains induced to isoniazid resistance. This is described in Chapter VI of this Thesis, where it is demonstrated that induced resistance to isoniazid does not involve mutations in any of the genes known to be associated with isoniazid resistance, but an efflux system that is sensitive to efflux inhibitors. These inhibitors decreased the efflux of ethidium bromide and also reduced the minimum inhibitory concentration of isoniazid in these strains. Moreover, expression analysis showed overexpression of genes that code for efflux pumps in the induced strains relatively to the non-induced parental strains. In conclusion, the work described in this thesis demonstrates that efflux pumps play an important role in the development of drug resistance, namely in mycobacteria. A strategy to overcome efflux-mediated resistance may consist on the use of compounds that inhibit efflux activity, restoring the activity of antimicrobials that are efflux pump substrates, a useful approach particularly in TB where the most effective treatment regimens are becoming uneffective due to the increase of MDRTB/XDRTB.

Antimicrobial resistance and plasmid detection in strains of the Bacteroides fragilis group

Resistant populations of the Bacteroides fragilis group bacteria (two reference ones and two isolated from human and Callithrix penicillata marmoset) were obtained by the gradient plate technique, to clindamycin, penicillin G, metronidazole and mercuric chloride. All the four tested strains were originaly susceptible to the four antimicrobial drugs at the breakpoint used in this study. MICs determination for the four cultures gave constant values for each antimicrobial, on the several steps by the gradient plate technique. The intestinal human B. fragilis strains showed three DNA bands, that could be representative of only two plasmids in the closed covalently circular (CCC) form with molecular weights of approximately 25 and 2.5 Md. The results do not permit an association between the presence of plasmid in the human strain with the susceptibility to the studied drugs. The four strains were ß-lactamase negative in the two methods used, and no particular chromosomal genetic resistance marker was demonstred. The resistance (MIC) observed, after contact with penicillin G and mercuric chloride, were two-fold in the four tested strains