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1

Using combined recombinant protein in the diagnosis of bovine brucellosis

100%
Bulashev A., Jakubowski T., Mukantayev K., Tursunov K., Kiyan V., Zhumalin A.
Medycyna Weterynaryjna
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2018
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tom 74
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nr 03
The aim of this research was to obtain a combined recombinant protein consisting of immunodominant regions of Brucella outer membrane proteins (Omps) with the molecular weights of 25 kDa (Omp25) and 31 kDa (Omp31). The search for candidate proteins was carried out using NCBI PubMed and NCBI GenBank databases. The two most immunodominant regions of Omps were selected. The first region was in a position from 48 to 83 amino acids of Omp31, while the second one was in the position from 180 to 224 amino acids of Omp25. This combined sequence was designated as OmpBm-Ba. The pET32 vector was used for cloning and the expression of the combined recombinant protein in E. coli BL21(DE3). The antigenicity of recombinant OmpBm-Ba (rOmpBm-Ba) as compared with rOmp25 and rOmp31 has been tested on the sera of 109 cattle with positive results to brucellosis according to classical serological tests. The rOmpBm-Ba had a higher antigenicity than the single ones, since it confirmed the presence of Brucella-antibody in all serum samples with positive results of i-ELISA/rOmp25 and/or i-ELISA/rOmp31, as well as additionally revealing 7.3% and 31.2% seropositive animals, respectively. A comparative study of the diagnostic value of i-ELISA/rOmpBm-Ba and conventional tests on blood sera of 24 cattle subjected to a bacteriological examination at slaughter showed a higher reliability of enzyme immunoassay. Further research is necessary to get a more objective evaluation of the diagnostic accuracy of Brucella rOmpBm-Ba by challenging laboratory animals.
2

Diagnosis of bovine brucellosis using traditional serological techniques and fluorescence polarisation assay

80%
Weiner M., Iwaniak W., Zlotnicka J., Szulowski K.
Bulletin of the Veterinary Institute in Puławy
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2010
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tom 54
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nr 4
The aim of this study was the application of fluorescence polarisation assay (FPA) for the examination of bovine sera and comparison of the results of the assay with the results of Rose Bengal test (RBT), serum agglutination test (SAT), complement fixation test (CFT), and ELISA. Six hundred thirty-five sera from cattle, including 300 sera from healthy animals, 32 sera from animals regarded as serologically positive for brucellosis and culled, and 303 sera originated from confirmatory investigations were used. All sera originating from healthy animals, negative in ELISA, RBT, SAT, and CFT were also negative in FPA. Among 303 sera from confirmatory investigations, 269 were positive in both RBT and SAT, 21 were positive in SAT and remaining 13 were RBT-positive only. Only two sera, one positive in both tests (RBT, SAT) and one SAT-positive, were also positive in FPA. Among 32 sera originated from animals regarded as serologically positive, which reacted in RBT, SAT, and CFT, 14 gave positive results in ELISA, whereas 18 were negative. Among these ELISA-positive sera, 13 were also positive in FPA. All samples positive in SAT, RBT, and CFT, and negative in ELISA, were also negative in FPA.
3

Evaluation of the ELISA in diagnosis of bovine brucellosis. Part I - Examination of sera

80%
Szulowski K.
Polish Journal of Veterinary Sciences
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1998
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tom 01
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nr 2
The investigation aimed at preparing an ELISA kit for examination of bovine sera for brucellosis and evaluating the diagnostic properties of the test. Lipopolysaccharide extract (LPS) was used as the antigen for microplate coating, antibodies against IgG of bovine sera with horseradish peroxidase were used as the conjugate and ABTS with H202 as the substrate. A weak-positive serum prepared from the second International Standard of anti-Brucella abortus Serum (ISaBaS II) and a negative serum obtained by mixing sera from Brucella-free cows were used as controls. Evaluation of the diagnostic usefulness of the ELISA was performed by comparison of the results obtained with those in the rose bengal test (RBT) and complement fixation test (CFT). The examinations involved 212 bovine sera positive in the RBT, 60 sera positive in the CFT, 5 standard sera, and 1005 sera negative in the RBT. A high correlation was found between the ELISA and CFT results (55 sera out 60 sera) and no correlation between the ELISA and RBT results. Only 60 out of 212 sera positive in RBT gave positive results in the ELISA. All standard sera were positive in the ELISA and only 1 serum from those negative in the RBT was doubtful in the ELISA. The ELISA proved to be a suitable method for diagnosis of brucellosis in cattle.
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