An enzyme-linked immunosorbent assay (elisa) for the detection of antibodies against babesia boris in cattle

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Padronização e aplicação da técnica de ELISA (Enzyme-linked immunosorbent assay) indireto com anticorpo de captura para a detecção de anticoepos contra o cicovírus suíno tipo 2

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

A liquid-phase-blocking concanavalin A enzyme-linked immunosorbent assay for the detection of antibodies against Newcastle disease virus in serum of free-ranging pigeons

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Production of recombinant EMA-1 protein and its application for the diagnosis of Theileria equi using an enzyme immuno assay in horses from São Paulo State, Brazil

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

Evaluation of the double agar gel immunodiffusion test and of the enzyme-linked immunosorbent assay in the diagnosis and follow-up of patients with chronic pulmonary aspergillosis

Coordenação de Aperfeiçoamento do Pessoal de Nível Superior (CAPES)

Improvement of an enzyme immunosorbent assay for detecting antibodies against Dioctophyma renale

An available enzyme-linked immunosorbent assay (ELISA) was studied for the detection of anti-Dioctophyma renale antibodies in the sera of dogs using, detection of parasite eggs in urine sediment as a reference test. ELISA uses a soluble antigenic preparation of esophagus of D. renale and the optimal dilutions of the antigen, serum and conjugate were determined by means of checker board titration, using positive (n=13) and negative (n=27) reference serum. The specificity and sensitivity of the ELISA were 93.8% and 92.3% respectively and the kappa index was good (0.76). These results suggest that ELISA described may prove to be an effective serological test for detecting dogs infected and exposed to this parasite mainly dogs that are not eliminating parasite eggs through their urine.

Selective activity of butyrylcholinesterase in serum by a chemiluminescent assay

In a previous study, we showed that purified commercial esterase activity can be detected in a chemiluminescent assay based on the hydrolysis of 2-methyl-1-propenylbenzoate (MPB) to 2-methyl-1-propenol, which is subsequently oxidized by the horseradish peroxidase (HRP)-H2O2 system. The purpose of this study was to verify the applicability of this assay to human serum. The existence of an esterase activity capable of hydrolysing MPB is indicated by the fact that the MPB-scruin-HRP-H2O2 System consumes oxygen and emits light. Both signals were abolished by prior serum heat inactivation and were preserved when serum was stored at less than or equal to4 degreesC. Addition of aliesterase inhibitors, such as fluoride ion and trichlorfon or the cholinesterase inhibitor eserine, totally prevents light emission. The butyrylcholinesterase-specific substrate benzoylcholine causes a delay in both O-2 uptake and light emission, while the specific acetylcholinesterase substrate, acetyl-beta -methylcholine, had practically no effect. Purified butyrylcholinesterase, but not acetylcholinesterase, triggered light emission. The finding that butyryleholinesterase is responsible for the hydrolysis of MPB in serum should serve as the basis for the development of a specific chemiluminescent assay for this enzyme. Copyright (C) 2001 John Wiley & Sons, Ltd.

Serological diagnosis of visceral leishmaniasis by an enzyme immunoassay using protein A in naturally infected dogs

Um ensaio de imunoadsorção enzimática para detecção de anticorpos contra Leishmania chagasi, utilizando antígeno total de formas promastigotas lisados foi desenvolvido. Cinqüenta cães com sintomas clínicos de leishmaniose visceral foram examinados. Esta técnica utilizou anti-IgG de cão conjugado a peroxidase ou proteína A conjugado a peroxidase. Foi verificado que nos animais positivos diagnosticados por exame parasitológico direto o ensaio ELISA utilizando proteína A conjugada a peroxidase (média da densidade óptica ± desvio padrão 2,078 ± 0,631) detecta mais anticorpos do que o sistema utilizando anti-IgG de cão conjugado a peroxidase (média da densidade óptica ± desvio padrão 1,008 ± 0,437), enquanto para os animais negativos o resultado obtido nos dois sistemas de detecção são similares. Esse resultado sugere que o sistema de ELISA utilizando proteína A conjugado a peroxidase pode ser útil na detecção de animais na fase aguda da infecção e desta forma auxiliar na identificação dos animais positivos e no controle desta importante zoonose.

Colorimetric enzymatic assay of L-malic acid using dehydrogenase from baker's yeast

A colorimetric method has been developed and optimized to measure L-malic acid in samples of fruit juices and wine. This method is based on oxidation of the analyte, catalyzed by malate dehydrogenase (MDH) from dry baker's yeast, and in combination with the reduction of a tetrazolium salt (MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide). In the present study, the method exhibited sensitivity in the range of 500-4000 mu M of L-malic acid in the reaction cuvette, with the lower detection limit of 6.7-10(-2) g/L, the upper limit of 53.6.10(-2) g/L and a maximum standard deviation of only 2.5 % for the analyzed samples. The MDH activity from baker's yeast was also optimized, the enzyme showed a high stability at pH=8.0-9.0 and the activity was maintained completely at temperatures up to 40 degrees C for 1 hour. The results show that the colorimetric method using enzymatic preparations from dry baker's yeast is a simple and low-cost method with possibility of wide application.

Detection of Cryptosporidium parvum oocysts in calf fecal samples by direct immunofluorescence assay

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

Analysis of pesticides in food and environmental samples by enzyme-linked immunosorbent assays

Enzyme-linked immunosorbent assays (ELISAs) are the most extensively studied types of immunoassay and their application in pesticide residue monitoring is an area with enormous potential for growth. In comparison with classical analytical methods, ELISA methods offer the possibility of highly sensitive, relatively rapid, and cost-effective measurements. This review introduces the general ELISA formats used, focusing on their use in pesticide analysis. Identifying and studying the effects of interferences in immunoassays is an active area of research and we discuss the matrix effects observed in several studies involving e.g. food, crop and environmental samples. The procedures to eliminate the matrix interferences are briefly discussed. (C) 1998 Elsevier B.V. B.V.

Colorimetric assay of ethanol using alcohol dehydrogenase from dry baker's yeast

Alcohol dehydrogenases (ADHs) are oxidoreductases present in animal tissues, plants, and microorganisms. These enzymes attract major scientific interest for the evolutionary perspectives, afforded by their wide occurrence in nature, and for their use in synthesis, thanks to their broad substrate specificity and stereoselectivity. In the present study, the standardization of the activity of the alcohol dehydrogenase from baker's yeast was accomplished, and the pH and temperature stability showed, that the enzyme presented a high stability to pH 6.0-7.0 and the thermal stability were completely maintained up to 50 degrees C during 1 h. The assays of ethanol (detection range 1-5 mM or 4.6 x 10(-2) to 23.0 x 10(-2) g/L) in different samples in alcoholic beverages, presented a maximum deviation of only 7.2%. The standard curve and the analytic curve of this method meet the conditions of precision, sensitivity, simplicity, and low cost, required for a useable analytical method. (c) 2006 Elsevier B.V. All rights reserved.

Detection of cross infections by Leishmania spp. and Trypanosoma spp. in dogs using indirect immunoenzyme assay, indirect fluorescent antibody test and polymerase chain reaction

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

Tillage and phosphorus management effects on enzyme-labile bioactive phosphorus availability in Cerrado Oxisols

Phosphorus (P) is an essential element in crop nutrition, which can be growth limiting or an environmental contaminant, if present in excess. Tillage practices have a direct effect on the behavior and availability of soil P. Sorption and availability of various P forms were evaluated in an incubation-fractionation study of three soils, a Typic Paleudults (CR soil) and two Cerrado Oxisols (Latossolo Vermelho-Amarelo [LVA] and Latossolo Vermelho [LV]) with distinct biogeochemical characteristics and tillage management history. Phosphate and myo-inositol hexakisphosphate (mIPH) were strongly sorbed by the soils. Maximum adsorption capacities (S(max)) were 2.2-6.9, 3.3-7.8, and 1.6-19.8 mmol kg(-1) for phosphate in the 0-40 cm depths of the CR, LV, and LVA soils, respectively. For mIPH, S. were 1.2-3.7, 3.7-5.5, and 4.6-5.2 mmol kg(-1). Saturation indices reflected the long-term effect of repeated manure applications on the Paleudults and the near saturation of its P holding capacity, in contrast to the recently cultivated Cerrado soils. Tillage method appeared to have altered P retention characteristics of the near-surface zone very slightly, while increases in ligand-exchangeable (EEP;) and enzyme-labile organic P (EDTA-PHP) forms were observed in no-till Oxisols. In the Paleudults, added manure P increased bioactive P fractions and P saturation of no-till near-surface soil zone. Estimates of all bioactive P fractions using the ligand-based enzymatic assay showed it to be an effective method for assessing P availability in soil and developing sustainable P management strategies, particularly in Cerrado Oxisols that were low in organic matter while having an extensive P-fixing capacity. Published by Elsevier B.V.