Use of a TT virus ORF1 recombinant protein to detect anti-TT virus antibodies in human sera

TT virus (TTV), isolated initially from a Japanese patient with hepatitis o f unknown aetiology, has since been found to infect both healthy and diseas ed individuals and numerous prevalence studies have raised questions about its role in unexplained hepatitis. In order to determine the prevalence of ongoing TTV infection as well as resolved infection, a serological study wa s performed with a recombinant protein generated from the open reading fram e 1 (ORF1) sequence isolated from a French patient infected by TTV. The C-t erminal end of the ORF1 protein, containing a particularly hydrophilic regi on, was retained to be used as antigen to detect the presence of anti-TTV a ntibodies in serum samples by a Western blot analysis. For this purpose, th e C-terminal ORF1 region was expressed in fusion with a hexahistidine tail in E. coli and purified by metal-chelate affinity chromatography. The serol ogical screening of 70 human sera, including 30 patients with hepatitis of unknown aetiology, 30 blood donors and 10 healthy children, allowed the imm une response of infected hosts to be identified by the detection of TTV-spe cific antibodies, with a very high prevalence of 98.6% in the human sera te sted. In contrast, TTV DNA was detected by PCR in only 76.1% of the tested sera. The use of the ORF1 C-terminal recombinant protein thereby provided a diagnostic tool to follow the immune response of the host against TTV.