An ELISA was developed for detection of antibodies to GB virus C (GBV-C) using a recombinant E2 protein expressed in CHO cells. Seroconversion to anti-E2 positivity was noted among several persons infected with GBV-C RNA-positive blood through transfusion. Of 6 blood recipients infected by GBV-C RNA-positive donors, 4 (67%) became anti-E2 positive and cleared their viremia. Thus, anti-E2 seroconversion is associated with viral clearance. The prevalence of antibodies to E2 was relatively low (3.0%-8.1%) in volunteer blood donors but was higher in several other groups, including plasmapheresis donors (34.0%), intravenous drug users (85.2%), and West African subjects (13.3%), all of whom tested negative by GBV-C reverse-transcription polymerase chain reaction (RT-PCR). These data demonstrate that testing for anti-E2 should greatly extend the ability of RT-PCR to define the epidemiology and clinical significance of GBV-C.

The increasing use of hepatitis B vaccination has had an overwhelming positive impact on the prevention of hepatitis B viral infection. Mutations in the hepatitis B surface antigen (HBsAg) gene occur as a result of vaccine escape mutants, anti-hepatitis B surface antigen immunotherapy, or in chronic hepatitis B viral infection. These mutants may present a challenge to immunoassay detection. Evaluation of the immunodetection of various HBsAg mutants has been sporadic, as the occurrence of these mutants is not common, and sufficient volume of serum samples is difficult to obtain. To investigate mutant detection, recombinant antigens were constructed to reflect mutations described in the literature occurring throughout the S gene. A limited number of serum samples exhibiting discordant immunoassay reactivity were also used to construct recombinant antigens. The evaluation of 25 HBsAg mutants across nine commercial assays of differing formats is described. Mutations affecting immunoassay performance were characterized as occurring mainly in loop 2 of the "a" determinant of HBsAg. It was determined that reagent epitope recognition was more significant for mutant detection than assay format.

Hepatitis B viral mutants can emerge in patients as a result of selection pressure from either immune response or treatment options. Mutations that occur within the immunodominant epitopes of hepatitis B surface antigen (HBsAg) allow mutant virus to propagate in the presence of a neutralizing immune response, while wild-type virus is reduced to undetectable levels. HBsAg mutants present as false-negative results in some immunoassays. An understanding of immunoassay reactivity with HBsAg mutants is key to establishing an appropriate testing algorithm for hepatitis B virus detection programs.

A 336-amino-acid segment of the GB virus C second envelope protein (E2) has been produced in BHK-21 cells using the Semliki Forest virus vector system. Secretion of this protein was facilitated by deletion of a hydrophobic region at the C-terminus that may represent the membrane anchoring domain. The E2 protein recovered from the culture supernatant exhibited a molecular mass of approximately 52 kDa, with the increase in size relative to the polyprotein backbone being contributed by N-linked glycosylation. A radioimmunoprecipitation assay using GBV-C E2 was developed to test for the presence of antibodies against this protein in human sera. The prevalence of antibodies to E2 was high among injection drug users and other individuals at risk for acquiring parenterally transmitted agents. There was a much higher percentage of anti-E2 seropositivity in GBV-C RT-PCR negative compared to GBV-C RT-PCR positive samples from these populations. In addition, serial samples from patients transfused with blood containing GBV-C showed seroconversion to anti-E2 positivity and loss of GBV-C viremia as measured by RT-PCR within 11 months of transfusion in five of seven individuals. Thus, this system provided a rapid means to identify GBV-C E2 as a useful antigen for the study of GBV-C exposure.