DIAGNOSTIC EVALUATION OF CHROMATOGRAPHIC FRACTIONS OF FASCIOLA-HEPATICA BY WESTERN-BLOT AND ELISA IN INFECTED ANIMALS

The antigenic components of excretory-secretory products of adult F. h epatica, were separated by gel filtration chromatography (Sephacryl S- 300) and then analized by polyacrylamide gel electrophoresis (SDS-PAGE ), followed by Western Blot (WE). In order to evaluate the sensitivity , specificity and predictive value of the selected fractions an enzyme -linked immunosorbent assay (ELISA) was used with sera from sheep, swi ne and horses infected with F. hepatica, as well as with control sera (uninfected animals). The chromatographic curve presented up to 5 peak s, representing polypeptides with a molecular weight of 2000, 400, 150 , 29 and less than 29 kDa, according to the interpolation with a stand ard curve of commercial polypeptide molecular weights. The results obt ained with SDS-PAGE and WE using sera from the three species, indicate d that those fractions of 400, 150, 29 and <29 kDa contained polypepti des, specifically recognized only by infected animals. When these frac tions were evaluated with sheep, horse and swine sera by means of ELIS A, the most efficient fraction was the 29 kDa, exhibiting an average s ensitivity and specificity of 94.5% and 93.5%, respectively in the thr ee species. The SDS-PAGE and WE assay of this fraction showed the pres ence of two immunoreactive bands, a 29 kDa and another of 14 kDa. Acco rding to these results, it is concluded that the polypeptides containe d in the 29 kDa chromatographic fraction would be suitable specific an tigens, since they discriminate between infected and non infected F. h epatica animals. This fraction was also efficient in diagnosing fascio losis in the prepatent stage of the infection in sheep. Therefore this fraction should be further studied and purified as it constitutes a p rominent immunodiagnostic fraction which may be used for mass screenin g studies of fasciolosis using an ELISA test.