Characterization of a reduction-sensitive factor from human plasma responsible for apparent false activity in competitive assays for antibody to hepatitis B core antigen

Abstract

Addition of reducing agents to competitive assays for antibody to hepatitis B core antigen (anti-HBc) eliminates apparent false reactivity of specimens obtained from individuals with no prior history of hepatitis B virus (HBV) infection and without other serological markers of HBV infection. We have purified and characterized a reduction-sensitive factor (RSF) isolated from the plasma of several volunteer blood donors. Column fractions were assayed for anti-HBc by using a highly sensitive chemiluminescence assay with a detection of 0.15 Paul Ehrlich Institute units per ml at 50% inhibition. Gel filtration on Sephacryl S-300 indicated that reductant-sensitive samples possessed anti-HBc activity that was associated with immunoglobulin M (IgM), whereas reductant-stable activity was associated with IgG. Gel filtration followed by metal chelate affinity chromatography resulted in a 55-fold purification and demonstrated that RSF activity copurifies with IgM. RSF was recovered from a recombinant hepatitis B core antigen matrix and shown to be an IgM species by immunoblot. In addition, RSF activity coeluted with IgM protein from anti-μ-chain Sepharose. Discrepancies between enzyme immunoassay and radioimmunoassay procedures for anti-HBc (Corzyme and Corab, respectively: Abbott Laboratories, North Chicago, Ill.) appear to be due to the relative sensitivity of the enzyme immunoassay for IgM anti-HBc (sevenfold greater than the radioimmunoassay using a specific panel). The biological basis for the occurrence of low levels of nonspecific IgM anti-HBc reactivity in individuals not previously exposed to HBV remains to be elucidated.