Serological evaluation of EgAgB16 kDa, a recombinant antigen from echinococcus granulosus for diagnosis of human hydatidosis

Regarding that accurate diagnosis of human hydatidosis still needs more investigations, the present study was conducted to clone, express, and evaluate the gene encoding AgB subunits [EgAgB16 kDa] from Echinococcus granulosus [Iranian G1 strain] and its evaluation by ELISA test. DNA was extracted from protoscoleces and was utilized by PCR for strain identification. Total RNA was prepared with RNeasy protect mini kit from E. granulosus [Iranian G1 strain] protoscoleces collected from naturally infected sheep with hydatid cyst. Recombinant AgB16 kDa was produced using pETDuet as vector and evaluated by ELISA method. A panel of sera including hydatid cyst-infected individuals [n=72], healthy individual [n=48], toxoplasmosis [n=4], strongyloidosis [n=4], kala-azar [n=5] and tuberculosis [n=5] were examined using this recombinant antigen. Recombinant protein was purified by affinity chromatography using His-Tag column. After purification, recombinant protein was confirmed by western blot analysis using His Tag monoclonal anti body or hydatid positive human serum. The sensitivity, specificity; positive and negative predictive values were calculated as 93.5%, 95.6%, 96% and 92.9%, in that order. The cut-off point was detected 0.3 for rAgB16. While the produced recombinant AgB16 kDa showed promising results in diagnosing human hydatidosis, but more investigations should be implemented to reach an accurate gold standard

Gene cloning of 30 kDa toxoplasma gondii tachyzoites surface antigen [SAG1]

Toxoplasma gondii is an obligate intracellular parasite and its sexual and asexual cycles, respectively take place in the intestinal epithelial cell of definitive host and tissue of intermediate hosts. Congenital toxoplasmosis is more important when the mother acquired the infection during pregnancy period for the first time. Serological tests are the only methods for diagnosis of toxoplasmosis. Among serological tests, ELISA has specific value and availability of parasite specific antigen increases the specificity of test. This study has designed and performed in the aim of availability to specific antigen of Toxoplasma. A pair of forward and reverse primers was designed based on published sequence of T. gondii SAG1 gene. PCR reaction was performed and PCR product was cloned in the pQE-30 expression vector. The gene of 30 kDa protein of Toxoplasma tachyzoites was cloned in expression vector successfully. Recombinant plasmid was confirmed and is ready to express recombinant protein for further studies. In this research we cloned P30 gene of T. gondii tachyzoites surface protein successfully and is ready to express the recombinant protein