Enzyme-linked immunosorbent assay (ELISA) for imunodiagnostic of bovine cysticercosis and kinetics of antibodies production against-Cysticercus bovis

摘要

> size="2" face="Verdana, Arial, Helvetica, sans-serif">An indirect ELISA (ENZYME-LINKED IMMUNOSORBENT ASSAY) was developed for searching of antibodies against-Cysticercus bovis in bovine. Three antigens were studied: partial antigen of C. cellulosae, total antigen of C. bovis, and total antigen of C. longicollis. In the standardization of the ELISA the following combinations were analyzed: antigen 250 and 500 ng of protein/well, dilution of the sera 50, 100, 200, and 400 times, dilution of the conjugated (anti bovine -IgG conjugated IgG of goat with peroxidase) 400 and 800 times. The crossing of the conditions above resulted in the following standardization: antigen 250 ng/well, sera and conjugated diluted 100 and 400 times respectively. The reaction cut-off between reagents and non-reagents animals was determined by the average of the optic densities of 54 negative sera plus three standard deviation resulting in the value of 0,303. The reactivity of the three antigens used in the ELISA test was compared using sera from experimentally infected calves, using sera dilutions and conjugated standardized previously. Using the antigen of C. bovis was verified high correlation with the test standardized with C. cellulosae. However, the absorbance values were significantly smaller. With C. longicollis was observed low reactivity, but increasing the amount of antigen, up to 3000ng/well, there was a proportional increase of the response. The kinetics of antibodies anti-C. bovis production was studied in ten calves experimentally infected with 2 x 104 T. saginata eggs. Six non-infected calves were used as control. After 90 days from the infection date, the animals were killed. Thirteen samples of sera of each animal were analyzed. The first was picked in the day of the infection and the remaining at each 15 days. The maximum production of antibody was observed between 30 and 60 days post infection. With the standardized test it was detected antibodies against-C. bovis, in 2 from 20 cattle considerated as non-holder of cyst by the inspection service. These animals could be considered possible cyst holders.