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 gondii to be used as diagnosis reagents in a recombinant form was evaluated. Both proteins were expressed in Escherichia coli as glutathione-S-transferase (GST) fusions. The GST-GRA1 fusion comprises the entire GRA1 sequence devoid 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 hydrophilic part of the protein was recognized by a pool of positive human sera in an immunoblot. One hundred T. gondii-positive and 98 negative human sera were tested in two separate immunoglobulin G (IgG)-direct enzyme-linked immunosorbent assays (ELISAs) using either GST-GRA1 or GST-GRA6-Nt recombinant protein. Whereas the sensitivity of the GST-GRA1 IgG ELISA was low (68%), the GST-GRA6-Nt IgG ELISA reached a sensitivity of 96%. The reactivity to GRA6-Nt was shown to be high even with human sera of low IgG titers. In addition, comparison of the optical density values for each serum revealed that GRA1 may complement GRA6-Nt to reach an overall sensitivity of 98%. Therefore, the GST-GRA6-Nt ELISA could be used together with another antigen like GRA1 for the development of a recombinant antigen-based test for serodiagnosis of toxoplasmosis.
Lecordier
L
L
Mécanismes Moléculaires de la Pathogénèse des Sporozoaires, Institut Pasteur de Lille, IBL, 59019 Lille Cedex, France.
Fourmaux
M P
MP
Mercier
C
C
Dehecq
E
E
Masy
E
E
Cesbron-Delauw
M F
MF
D000818:Animals; D000913:Antibodies, Protozoan / Q000032:analysis* / Q000276:immunology*; D000953:Antigens, Protozoan / Q000235:genetics / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D007074:Immunoglobulin G / Q000032:analysis / Q000276:immunology; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity; D014122:Toxoplasma / Q000235:genetics / Q000276:immunology*; D014123:Toxoplasmosis / Q000175:diagnosis* / Q000276:immunology*
D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; C412913:GRA1 antigen; C095047:GRA6 protein, Toxoplasma gondii; D007074:Immunoglobulin G; D015800:Protozoan Proteins; D011994:Recombinant Proteins