Enzyme-linked immunosorbent assays using the recombinant dense granule antigens GRA6 and GRA1 of Toxoplasma gondii for detection of immunoglobulin G antibodies

The potential of the dense granule antigens GRA1 and GRA6 of Toxoplasma gon dii to be used as diagnosis reagents in a recombinant form was evaluated. B oth proteins were expressed in Escherichia coli as glutathione-S-transferas e (GST) fusions, The GST-GRA1 fusion comprises the entire GRA1 sequence dev oid of its N-terminal signal peptide. Separate expression of the two N- and C-terminal hydrophilic regions of GRA6 showed that only the N-terminal hyd rophilic part of the protein was recognized by a pool of positive human ser a in an immunoblot. One hundred T. gondii-positive and 98 negative human se ra were tested in two separate immunoglobulin G (IgG)-direct enzyme-linked immunosorbent assays (ELISAs) using either GST-GRA1 or GST-GRA6-Nt recombin ant protein. Whereas the sensitivity of the GST-GEA1 IgG ELISA was low (68% ), the GST-GRA6-Nt IgG ELISA reached a sensitivity of 96%. The reactivity t o GRA6-Nt was shown to be high even with human sera of low IgG titers, In a ddition, comparison of the optical density values for each serum revealed t hat GRA1 may complement GRA6-Nt to reach an overall sensitivity of 98%. The refore, the GST-GRA6-Nt ELISA could be used together with another antigen l ike GRA1 for the development of a recombinant antigen-based test for serodi agnosis of toxoplasmosis.