Serodiagnosis of viral hepatitis A: detection of acute-phase immunoglobulin M anti-hepatitis A virus by radioimmunoassay

Abstract

A modified micro solid-phase radioimmunoassay (RIA) for antibody to hepatitis A virus (anti-HAV) was developed. This double antibody procedure was performed by coating the surface of a polyvinyl microtiter plate "well" with 200 microliter of a 1:1,000 dilution of a patient's test serum. Purified HAV and 125I-labeled immunoglobulin G (IgG) anti-HAV were then sequentially added to form an antibody sandwich. The specificity and sensitivity of the RIA procedure for anti-HAV were verified by examination of coded human and chimpanzee serum specimens. Radioimmunoassay of early-acute-phase serum specimens from human cases of hepatitis A revealed the presence of anti-HAV activity. Differential examination by RIA of IgG and IgM fractions of acute-phase sera from experimentally infected chimpanzees demonstrated that IgM contained the bulk of the anti-HAV activity. A modification of the RIA procedure for anti-HAV (RIA-IgM blocking), incorporating an incubation step with anti-IgM (Mu chain specific), was further shown to differentiate acute- from convalescent-phase hepatitis A sera. This adapted RIA-IgM blocking procedure required less than 1 microliter of a single acute-phase serum specimen for the diagnosis of viral hepatitis A.