Enzyme-Linked Immunosorbent Assay for Detection of Hepatitis A Antigen in Stool and Antibody to Hepatitis A Antigen in Sera: Comparison with Solid-Phase Radioimmunoassay, Immune Electron Microscopy, and Immune Adherence Hemagglutination Assay

Abstract

Previously described techniques for detection of hepatitis A antigen (HA Ag) and antibody (anti-HA) have required purified HA Ag and expensive equipment. Herein is described an enzyme-linked immunosorbent assay (ELISA) for specific detection of HA Ag in human stool filtrates and of anti-HA in sera by using selected HA Ag-containing human stool filtrates as the antigen source. Because human stools often react nonspecifically in serological tests for HA Ag, blocking with preexposure and hyperimmune anti-HA sera from a chimpanzee inoculated with hepatitis A virus was used to confirm specific detection of HA Ag. The sensitivity of ELISA was found to be comparable to that of solid-phase radioimmunoassay (SPRIA) and immune electron microscopy (IEM). Of 37 acute-phase stools collected from nine patients, 16 were positive for HA Ag by ELISA. In 13 of these, HA Ag particles were found by IEM, and an additional 3 stools negative by ELISA contained HA Ag particles by IEM. Eight control stools were negative by both ELISA and IEM. Anti-HA was measured in sera by demonstrating its ability to block binding of the enzyme conjugate to HA Ag in a stool without detectable nonspecificity. This test (blocking ELISA) was as sensitive and specific as blocking SPIRA, IEM, and immune adherence hemagglutination and, like SPRIA and IEM, detected early-developing antibody. The ELISA is simple to perform and requires only a minimum of equipment. It is useful for screening stools for HA Ag and for monitoring HA Ag during purification, as well as for detecting early and late anti-HA in sera.