شناسایی پروتئینهای لارو فیلاریفرم (L3) استرونژیلوئیدس استرکورالیس به روش وسترن بلات

Background: Strongyloides stercoralis is prevalent in tropical and subtropical regions of the world. S. stercoralis is the only nematodes with the ability to multiply in its host's body via autoinfection transmission. Larvae detection in faeces is difficult partly because of low egg production and irregular larvae excretion in faeces. Serologic tests (ELISA, IFA) are also diagnostic, however, S. stercoralis antigens are not available as a diagnostic tool. In the present study, we analyzed filariform larva (L3) proteins of S. stercoralis by the immuno blot technique. Materials and methods: Stool samples were examined by direct smear, formalin-ether and agar plate method to identify the patients. Sera were stored at 20°С. Filariform larvae were obtained by agar plate culture, which was incubated for 6-7days at 25ºC, then frozen at – 70°С. Finally, larvae were suspended at a concentration level of 12000 in 250l PBS, containing protease inhibitors and then sonicated. Protein level was measured by Bradford method. Proteins of S. stercoralis filariform larvae were separated by SDS-PAGE, blotted onto nitrocellulose paper. Western blot analysis of these antigens was achieved using infected human sera (0.1, 0.01, 0.001 dilution) with strongyloidiasis, toxocariasis, hydatidosis, amoebiasis and normal human serum as control. Results: Four immunodominant proteins (23, 28, 30, 41 kDa) were recognized with strongyloidiasis sera in 0.1diluted serum. None of the proteins reacted to normal human and amebiasis serum, but some showed reaction with serum of hydatidosis and toxocariosis. Having increased the level of serum dilution, only 41 kDa protein was recognized by strongyloidiasis sera. Other sera did not show any reaction to the parasite’s proteins. Therefore, the 41 kDa protein presents as most important immunodaminant protein in this study.Conclusion: The identification of immunodominant proteins, which have adapted themselves to the physiological and genetically conditions of the host is an appropriate diagnostic approach. Indeed, recombinant protein, such as 41 kDa, could enhance the sensitivity and specificity of serologic tests.