Standardization of the dot enzyme-lynked immunosorbent assay (dot-ELISA) for experimental plague

A dot enzyme linked immunosorbent assay (dot-ELISA) was previously developed to detect specific antibodies in rabbits sera immunized against FIA protein obtained from Yersina pestis. This antigen was covalently linked onto the surface of dacron (polyethyleneterephthalate). Here, standard conditions are described for the optimization of this procedure: an amount of 20 ng of FIA protein was fixed onto dacron; anti-rabbit IgG peroxidase conjugate diluted 1:8,000 and 30% non-fat instant milk as blocking substance were used throughout the method. This procedure was compared with that employing nitrocellulose as solid-phase which showed to be more sensitive. However, the method based on dacron did not show false positive reactions against non-immunized rabbits sera at low antigen amount and diluted anti-IgG peroxidase conjugate.

Cloning and Characterization of a Schistosoma mansoni cDNA Clone with a Specific Antigenic Expression during Development in a Vertebrate Host

Approximately 2.0 x 10 cDNA clones of an Schistosoma mansoni lgt11 cDNA library were screened in duplicate with serum from infected mice corresponding to distinct phases of infection. A cDNA clone (7/1) was isolated and recognized only by seven week serum. The clone was subcloned in pGEX-2T and Western-blot studies showed a specific antigenic expression confirming that only serum from the chronic phase is capable of recognizing this antigen. Dot-hybridization with RNA from different developmental phases of the parasite showed that the corresponding 7/1 RNA is expressed in all phases of parasite development in vertebrate hosts

Positive IgG Western Blot for Borrelia burgdorferi in Colombia

In order to evaluate the presence of specific IgG antibodies to Borrelia burgdorferi in patients with clinical manifestations associated with Lyme borreliosis in Cali, Colombia, 20 serum samples from patients with dermatologic signs, one cerebrospinal fluid (CSF) sample from a patient with chronic neurologic and arthritic manifestations, and twelve serum samples from individuals without clinical signs associated with Lyme borreliosis were analyzed by IgG Western blot. The results were interpreted following the recommendations of the Centers for Diseases Control and Prevention (CDC) for IgG Western blots. Four samples fulfilled the CDC criteria: two serum specimens from patients with morphea (localized scleroderma), the CSF from the patient with neurologic and arthritic manifestations, and one of the controls. Interpretation of positive serology for Lyme disease in non-endemic countries must be cautious. However these results suggest that the putative "Lyme-like" disease may correlate with positivity on Western blots, thus raising the possibility that a spirochete genospecies distinct from B. burgdorferi sensu stricto, or a Borrelia species other than B. burgdorferi sensu lato is the causative agent. Future work will focus on a survey of the local tick and rodent population for evidence of spirochete species that could be incriminated as the etiologic agent.

Problemàtiques en l'estudi i investigació del "Dot Painting o Dot Art Movement" Aborigen austràlià

L'objecte d'estudi de la següent memòria són les obres de l'anomenat, en la literatura anglosaxona, "Dot Art Movement o Dot Painting"; un moviment artístic índigena contemporani engendrat entre la dècada dels 70 i els 80 del segle passat a les comunitats de Papunya i Yuendumu en el cor del Desert Australià. La memòria s'articula entorn les principals problemàtiques a les que s'enfronta la definició i descripció de l'anomenat "Dot Art Movement o Dot Painting" dins el context de consum i recepció de les obres a Europa. Aquestes problemàtiques, sobretot conceptuals però també metodològiques, són descrites entorn tres grans apartats: el fet "primitiu", el fet "aborigen" i el fet del "Dot Art Movement o Dot painting". A través d'aquests tres àmbits la memòria presenta diverses herències històriques de caràcter eurocèntric, amb base al període colonial, que dificulten la proliferació d'interpretacions i noves aproximacions vers la recepció europea de les obres d'aquest moviment artístic.

Enzyme-linked immunosorbent assay and Western blot antibody determination in sera from patients diagnosed with different helminthic infections with Anisakis simplex antigen purified by affinity chromatography

An evaluation of the sensitivity and the specificity of the Anisakis simplex antigens purified by affinity chromatography was performed using sera from patients diagnosed with Anisakis sensitisation and sera from patients previously diagnosed with different helminthic infections. Only the sera of the patients diagnosed with Schistosoma mansoni or Onchocerca volvulus parasitic infections were negative against the A. simplex antigen and its purified fractions (PAK antigen: A. simplex antigen purified using columns prepared with anti-A. simplex rabbit IgG and PAS antigen: PAK antigen purified using columns prepared with anti-Ascaris suum rabbit IgG). However all the sera were positive against the A. suum antigen. In all the sera from the patients diagnosed with Anisakis sensitisation, the antibody levels detected using the purified antigens (PAK and PAS antigens) were lower than the observed using the A. simplex crude extract with the highest diminution in the case of the IgG. When these same sera were tested against the A. simplex crude extract by Western blot, several bands of high molecular masses were observed as well as, intense bands at 60 and/or 40 kDa. A concentration of these last proteins was observed in the PAK and the PAS antigens. When the sensitivity and the specificity determinations were performed, only seven of the 38 patients diagnosed of Anisakis sensitisation were positive, as well as, the sera from the patients diagnosed with parasitisms by Echinococcus granulosus or Fasciola hepatica.

Evaluation of the dot enzyme-linked immunosorbent assay in comparison with standard ELISA for the immunodiagnosis of human toxocariasis

A dot enzyme-linked immunosorbent assay (dot-ELISA) was standardized using excretory-secretory antigens of Toxocara canis for the rapid immunodiagnosis of human toxocariasis. Thirty patients with clinical signs of toxocariasis, 20 cases with other parasitic diseases, and 40 healthy subjects were tested. A total of 0.2 ng of antigen per dot, serum dilution of 1:160 and dilution conjugate of 1:1000 were found optimal. The sensitivity and specificity of the assay were 100 and 95%, respectively. Comparable sensitivity of dot-ELISA and the standard ELISA was obtained, but only 3 cross-reactions occurred in the dot-ELISA, compared with 6 in the standard ELISA. Dot-ELISA is simple to perform, rapid, and low cost. Large-scale screening studies should be done to evaluate its usefulness under field conditions.

Evaluation of an enzyme-linked immunoelectrotransfer blot test for the confirmatory serodiagnosis of human toxocariasis

To improve the serodiagnosis of human toxocariasis, a sensitive and specific enzyme-linked immunoelectrotransfer blot (EITB-IgG) test was developed and evaluated using Toxocara canislarvae excretory-secretory antigens for detecting anti-Toxocara IgG antibodies. The EITB-IgG profile of toxocariasis was characterized by comparing 27 sera from patients with toxocariasis, 110 sera from healthy subjects and 186 sera from patients with other helminth diseases (ascariasis, ancylostomiasis, trichuriasis, enterobiasis, strongyloidiasis, hymenolepiasis, diphyllobothriasis, taeniasis, cysticercosis, hydatidosis and fascioliasis). Antigenic bands of 24, 28, 30, 35, 56, 117, 136 and 152 kDa were predominantly recognized in sera from all patients with toxocariasis. However, only bands of 24-35 kDa were highly specific for Toxocara infection (98.3%), whereas other antigenic bands observed displayed cross-reactivity. Additionally, when the results of the EITB-IgG test were compared to those of the ELISA-IgG test, a 100% concordance was observed for positive results in human toxocariasis cases. The concordance for negative results between the two tests for healthy subjects and patients with other helminth diseases were 96.3% and 53.7%, respectively, showing that the EITB-IgG test has a higher specificity than ELISA. In conclusion, the EITB-IgG test is a very useful tool to confirm the serological diagnosis of human toxocariasis.

IgG and IgM western blot assay for diagnosis of congenital toxoplasmosis

The aim of this study was to evaluate the utility of western blot (WB) analysis as a diagnostic tool for congenital toxoplasmosis in 215 newborn infants. The children were submitted to clinical examinations to assess macular, neurological and hearing signals. The WB results obtained were compared to the persistence of IgG antibodies at the end of 12 months, which is regarded as the "gold standard" diagnosis of congenital toxoplasmosis. Association between the WB results and the clinical signs presented by the infants was also assessed. Of the 215 children, 177 had a confirmed congenital toxoplasmosis diagnosis and 38 were uninfected. IgG-WB showed a sensitivity of 73.5% and a specificity of 97.4%. IgM-WB showed a sensitivity of 54.8% and a specificity of 94.7%. The IgG-WB and IgM-WB combination increased the sensitivity to 86.5%. The IgM-WB-positive children had a 1.4-fold greater risk of presenting active macular lesions than did those that were IgM-WB-negative. This study showed that the WB assay is a useful tool to confirm a diagnosis of congenital toxoplasmosis and that the IgM-WB-positive results can indicate active macular lesions in newborn infants.

An evaluation of the dot-ELISA procedure as a diagnostic test in an area with a high prevalence of human Toxocara canis infection

The aim of this work was to evaluate a dot-enzyme-linked immunosorbent assay (dot-ELISA) using excretory-secretory antigens from the larval stages of Toxocara canis for the diagnosis of toxocariasis. A secondary aim was to establish the optimal conditions for its use in an area with a high prevalence of human T. canis infection. The dot-ELISA test was standardised using different concentrations of the antigen fixed on nitrocellulose paper strips and increasing dilutions of the serum and conjugate. Both the dot-ELISA and standard ELISA methods were tested in parallel with the same batch of sera from controls and from individuals living in the problem area. The best results were obtained with 1.33 µg/mL of antigen, dilutions of 1/80 for the samples and controls and a dilution of 1/5,000 for the anti-human IgG-peroxidase conjugate. All steps of the procedure were performed at room temperature. The coincidence between ELISA and dot-ELISA was 85% and the kappa index was 0.72. The dot-ELISA test described here is rapid, easy to perform and does not require expensive equipment. Thus, this test is suitable for the serological diagnosis of human T. canis infection in field surveys and in the primary health care centres of endemic regions.

Synthesis of Best Practice for Increasing Protection and Visibility of Highway Maintenance Vehicles, CTRE Final Report Iowa DOT Project TR-475, August 2002

The purpose of this research project is to study current practices in enhancing visibility and protection of highway maintenance vehicles involved in moving operations such as snow removal and shoulder operations, crack sealing, and pothole patching. The results will enable the maintenance staff to adequately assess the applicability and impact of each strategy to their use and budget. The report’s literature review chapter examines the use of maintenance vehicle warning lights, retroreflective tapes, shadow vehicles and truck-mounted attenuators, and advanced vehicle control systems, as well as other practices to improve visibility for both snowplow operators and vehicles. The chapter concludes that the Manual on Uniform Traffic Control Devices does not specify what color or kind of warning lights to use. Thus, a wide variety of lights are being used on maintenance vehicles. The study of the relevant literatures also suggests that there are no clear guidelines for moving work zones at this time. Two types of surveys were conducted to determine current practices to improve visibility and safety in moving work zones across the country and in the state of Iowa. In the first survey of state departments of transportation, most indicated using amber warning lights on their maintenance vehicles. Almost all the responding states indicated using some form of reflective material on their vehicles to make them more visible. Most participating states indicated that the color of their vehicles is orange. Most states indicated using more warning lights on snow removal vehicles than their other maintenance vehicles. All responding state agencies indicated using shadow vehicles and/or truck-mounted attenuators during their moving operations. In the second survey of Iowa counties, most indicated using very similar traffic control and warning devices during their granular road maintenance and snow removal operations. Mounting warning signs and rotating or strobe lights on the rear of maintenance vehicles is common for Iowa counties. The most common warning devices used during the counties’ snow removal operations are reflective tapes, warning flags, strobe lights, and auxiliary headlamps.

Identifying and Implementing Effective Training for Hispanic Craft Workers, American Supervisors, and DOT Inspectors: Phase III of the Hispanic Workforce Research Project, May 2006

The number of Hispanic workers in the U.S. construction industry has been steadily increasing, and language and cultural barriers have sometimes arisen on the jobsite. Due in part to these barriers, the number of fatalities among Hispanics at construction sites in 2001 jumped 24%, while construction fatalities overall dropped 3%. This study, which constitutes Phase III of the Hispanic Workforce Research Project, addresses these language and cultural barriers by investigating the most effective way to deliver training material developed in Phases I and II to Hispanic workers, American supervisors, and department of transportation (DOT) inspectors. The research methodology consisted of assessing the needs and interests of potential and current course participants in terms of exploring innovative ways to deliver the training. The training courses were then adapted and delivered to fit the specific needs of each audience. During Phase III of this project, the research team delivered the courses described in the Phase I and II reports to eight highway construction companies and two DOT groups. The courses developed in Phases I and II consist of four construction-focused language training courses that can be part of an effective training program to facilitate integration among U.S. and Hispanic workers, increase productivity and motivation at the jobsite, and decrease the existing high mortality rate for Hispanic workers. Moreover, the research team developed a course for the construction season called Toolbox Integration Course for Hispanic workers and American supervisors (TICHA), which consists of nine 45-minute modules delivered to one construction company over 11 weeks in the summer of 2005.

Le multiple evanescent white dot syndrome: une prédisposition génétique [Multiple evanescent white dot syndrome: a genetic predisposition?]

BACKGROUND: Multiple evanescent white dot syndrome (MEWDS) is a benign acquired isolated chorioretinal disorder. Symptoms include photopsia, visual blur and scotomas. Ocular examination reveals multiple white dots at the level of the deep retina. A parainfectious disorder was suggested but the exact mechanism of MEWDS is still unknown. Postulating that MEWDS might be an antigen driven inflammatory reaction, we analyzed HLA subtypes in patients with MEWDS. PATIENTS AND METHODS: Sixteen patients were diagnosed with MEWDS in Lausanne from 1985 to 1994. Blood was withdrawn in 9/16 patients. HLA-A, -B and -DR were sought. RESULTS: HLA-B51 was detected in 4/9 patients (44.4%). Other HLA subtypes were detected sporadically. CONCLUSIONS: The frequency of HLA-B51 haplotype was found to be 3.7 times more elevated than in a normal control caucasian group. This suggests the possibility that MEWDS might be a genetically determined disorder as it is the case for other ocular diseases like Birdshot chorioretinopathy (HLA-A29), Harada's disease (HLA-DRMT3), acute anterior uveitis (HLA-B27) or Behçet's disease (HLA-B51). We have no explanation for the presence of HLA-B51 in both Behçet's disease and MEWDS. The association of HLA-B51 and MEWDS needs confirmation by further testing.

Review of Inconsistencies Between SUDAS and Iowa DOT Specifications, May 2006

The objectives of this multi-part project were to review the Iowa DOT Specifications and SUDAS Specifications section by section and develop recommendations for possible changes that will allow the two specifications to be used together.

Hepatites Virais Frequência do antígeno de superfície da hepatite viral tipo B nos portadores do HIV e grávidas atendidos no Laboratório Elisa-Blot do Hospital Agostinho Neto Praia em 2006 e 2007

Com a finalidade de fazer uma avaliação da frequência do antígeno de superfície da hepatite viral do tipo B (AgHBs) nos portadores HIV e grávidas atendidos no Laboratório Elisa-Blot do Hospital Agostinho Neto em 2006 e 2007, foi feito um estudo descritivo retrospectivo, com base na recolha de dados entre 1 de Janeiro de 2006 a 31 de Dezembro de 2007. Os dados foram recolhidos com base em fichas e no banco de dados informático, existentes no Laboratório Elisa-Blot do Hospital Agostinho Neto. Os dados foram colectados segundo as variáveis de tempo, pessoa e lugar. Os resultados demonstraram uma frequência do AgHBs durante o período de estudo de 4,96% (25/504). Esse valor corresponde a 11% (15/135) dos portadores do HIV e 3% em grávidas com sorologia positiva para HBsAg. Em relação ao perfil dos portadores do HIV, estes tinham uma idade média de 40,6 anos, a maioria do sexo feminino e residentes no concelho da Praia. As grávidas tinham uma idade média de 28,4 anos, a maioria residente também no concelho da Praia. As análises também demonstraram que o perfil dos casos com sorologia positiva para o AgHBs nos portadores do HIV era na sua maioria do sexo masculino com uma proporção de 60% durante o período de estudo. A maior ocorrência ocorreu na faixa etária 30 a 49 anos com uma proporção de 67% e 50% relativamente ao período de estudo. O concelho da Praia foi o que apresentou maior proporção com 67% e 58% em 2006 e 2007 respectivamente. Em relação as grávidas a maior proporção de positividade do AgHBs ocorreu também na faixa etária 30 a 49 anos com 75% e 67% e no concelho da Praia com uma proporção de 50% e 67% referentes aos anos 2006 e 2007 respectivamente.