Expression and analysis of the diagnostic value of an Echinococcus granulosus antigen gene clone.
A pool of 9 sera from Echinococcus granulosus infected patients (PSP) was used to screen an E. granulosus cDNA library constructed in the expression vector lambda gt11. Ten reactive phage clones were isolated and 8 were confirmed in spot-lysis arrays probed with PSP. The insert of 1 of these clones (lambda AgEg4) previously characterized as an E. granulosus cytosolic malate dehydrogenase encoding gene was subcloned into the plasmid vector pGEX-1 and expressed as a fusion with glutathione S-transferase. The fusion peptide (Ag4-GST) was produced in Escherichia coli and its antigenicity was confirmed in colony immunoassay and in immunoblot using nondenaturing conditions. The lack of antigenicity of Ag4-GST in immunoblot using denaturing conditions suggests that the recognized epitopes are conformational. Ag4-GST was purified by affinity chromatography and tested in ELISA and immunodots to access its sensitivity and specificity in the diagnosis of human cystic hydatid disease. An overall sensitivity of 53.6% was obtained. Cross-reactions were observed with some sera from patients infected with Schistosoma mansoni and Wuchereria bancrofti. Ag4-GST was not recognized by any of the sera from Taenia solium infected patients tested. These preliminary results suggest that Ag4-GST could be useful as an accessory antigen to discriminate some cross-reactions with sera from cysticercosis patients, especially in regions like southern Brazil, where schistosomiasis and filariasis are not prevalent.
Ferreira
H B
HB
Departamento de Biotecnologia, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil.
Zaha
A
A
D000818:Animals; D000947:Antigens, Helminth / Q000032:analysis* / Q000096:biosynthesis / Q000097:blood; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary; D004443:Echinococcosis / Q000175:diagnosis* / Q000469:parasitology; D004446:Echinococcus / Q000235:genetics / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015723:Gene Library; D006801:Humans; D015151:Immunoblotting; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis; D012680:Sensitivity and Specificity
D000947:Antigens, Helminth; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins