Evaluation of Kinetics and Single-Read Enzyme-Linked Immunoassays for Detection of Toxoplasma Gondii Antibodies in Sheep

Abstract

A kinetics enzyme-linked immunosorbent assay (ELISA) and a single-read ELISA for the detection of ovine anti- Toxoplasma gondii IgG were developed and optimized. During the kinetics assay, 3 optical densities were obtained for each serum sample at intervals of 45 seconds, and the results were presented as average slopes (replicates of 2) of the reaction rate between bound enzyme and substrate solution. The kinetics ELISA was stopped 5 minutes after dispensing the substrate to constitute the single-read ELISA, and the results were presented as average optical densities for duplicates of each sample. Performance of the assays was evaluated using the modified agglutination test (MAT) as the “gold standard.” There was a high level of agreement between both ELISAs and the MAT, as measured by Pearson correlation coefficients, kappa statistics, and the area under the receiver operating characteristics curves. The single-read ELISA was as accurate as the kinetics ELISA, with a sensitivity of 89% and a specificity of 96%.