Starting in the 1940s numerous attempts were made to isolate the causative agent of hepatitis A in cell culture. Immunofluorescence (IF) assay was originally developed to assist studies of the pathogenesis of HAV infection by localizing HAAg in specimens collected from experimentally infected marmosets and chimpanzees. Its potential for detecting HAAg in cell culture was recognized and the technique was rapidly introduced as a means of screening cultures for evidence of HAV replication. These studies led rapidly to isolation of HAV in liver explant tissue and in a cloned line of fetal rhesus monkey kidney cells. Numerous cell lines were studied and the virus eventually propagated in fetal rhesus monkey kidney cells, a cloned line of epithelial-like cells derived from fetal rhesus monkey kidney. Initially extracts of liver tissue obtained from animals infected with each of the three strains were inoculated into African green monkey kidney cells and evidence of virus replication sought by IF.