EXPRESSION AND IMMUNOLOGICAL CHARACTERIZATION OF ECHINOCOCCUS-GRANULOSUS RECOMBINANT ANTIGEN-B FOR IGG4 SUBCLASS DETECTION IN HUMAN CYSTIC ECHINOCOCCOSIS

A 165bp DNA fragment derived from the 12 kDa subunit of Echinococcus g ranulosus antigen B (AgB), a major hydatid cyst fluid antigen was clon ed in the pMal-c2 expression vector. A 52 kDa maltose binding-AgB fusi on protein (rAgB.MBP) was produced and inclusion bodies containing the fusion protein were solubilised in urea and affinity purified on an a mylose-Sepharose 6B column. The immunogenicity of the purified recombi nant antigen for IgG4 antibody detection was tested with human serum u sing immunoblotting, ELISA and dot-ELISA assays and compared to native AgB. Both recombinant and native AgB preparations were highly reactiv e for human IgG4 antibodies in serum of cystic echinococcus (CE) patie nts. Recombinant AgB.MBP (rAgB.MBP) showed approximate to 65% sensitiv ity in detection of IgG4 serum antibodies by ELISA from confirmed CE p atients. Cross-reactivity (33%) occurred with alveolar echinococcosis (E. multilocularis) sera but recombinant AgB showed no seroreactivity with sera from other helminth infections tested (schistosomsis, onchoc ercsis, cysticercosis) or from uninfected individuals residing in CE e ndemic or non-endemic regions. The serologic sensitivity (63%) for IgG 4 antibodies of a native AgB fraction enriched from human hydatid cyst fluid was similar to that for recombinant AgB (65%) though specificit y was slightly lower (81%). A dot-ELISA for detection of total IgG, in corporating the rAgB.MBP resulted in 74% sensitivity and 88% specifici ty for human CE and 93% sensitivity and 65% specificity for native AgB . Recombinant AgB is a potential replacement for native antigens curre ntly being used and could provide a better standardised E. granulosus specific test for clinical confirmation for CE especially for IgG4 ant ibody detection which appears to be predominantly associated with adva nced disease. (C) 1997 Elsevier Science B.V.