EVALUATION BYAN ENZIMATIC IMMUNOASSAY (ELISA) OFToxocara canis SEUDOCOELOMATIC FLUID ANTIGENS FOR DETECT SPECIFIC ANTIBODIES

Abstract

Toxocara canis is a causative agent of human toxocariasis, a zoonosis with a worldwide distribution with high prevalences in Neotropical areas. Toxocariasis is diagnosed by indirect enzyme immunoassay (ELISA) with excretion-secretion antigens (TES), considered the gold standard test; however, there are other types of antigens not yet investigated. In the present investigation, the performance (capacity) of an indirect ELISA test was evaluated using the supernatant of the pseudocoelomic fluid of the adult female forms of Toxocara canis (TFS) as antigen. The parasites were collected from naturally infected dog puppies (Canis lupus familiaris) and the TFS was obtained under sterile conditions by cutting the end of the parasite and then centrifuging. Protein concentration was measured with a Bradford kit (KB) and antigenicity by Western blot technique (Wb) with experimentally immunized rabbit hyperimmune serum. The ELISA tests were made with the TFS and 20 sera positive for Toxocara, 10 sera positive for other parasites and negative for Toxocara, and 52 sera from children from a rural area of the Pataz province (La Libertad, Peru). The cut-off point was 3.0 at optical densities of 450 nm in an ELISA reader (Perkin- Elmer). A protein concentration of 3500 ug/uL was found by KB and 18 reactive bands (between 9.1 and 87.2 KDa) were found by Wb. All the positive sera from the seroteca were also positive and those positive for other parasites were negative. A percentage of 7.8% (4/52) was found in the lot of sera from children in rural areas. In conclusion, the indirect ELISA test using as antigen the supernatant of the pseudocoelomic fluid of the adult female forms of Toxocara canis obtained from the naturally infected dog, Canis lupus familiaris, performs adequately (is effective) in the detection of specific antibodies.