ELISA for Antibody to Hepatitis E Virus (HEV) Based on Complete Open-Reading Frame-2 Protein Expressed in Insect Cells: Identification of HEV Infection in Primates

Abstract

A recombinant baculovirus containing the complete open-reading frame (ORF)-2 region of the hepatitis E virus (HEV) genome was constructed. The major protein synthesized in insect cells infected with recombinant virus was about the size expected for the complete ORF-2 product. This protein reacted in a Western blot assay with plasma from an HEV-infected chimpanzee. Lysates of the recombinant virus-infected insect cells were used in ELISA to monitor seroconversion of eight primate species (chimpanzees, four species of Old World monkeys, and three species of New World monkeys) inoculated with HEV. Homologous detector anti-immunoglobulin was more sensitive than heterologous anti-immunoglobulin for detecting anti-HEV by ELISA. All primate species except tamarins seroconverted after inoculation with HEV, although anti-HEV titers of Old World monkey species were generally higher than those of New World monkey species. The ELISA with complete ORF-2 antigen appeared to be a sensitive and practical method for detecting anti-HEV.

Recombinant hepatitis E virus genomes infectious for primates: Importance of capping and discovery of a cis-reactive element

“…Baseline levels were calculated as the geometric mean of three weekly samples collected at week zero and before inoculation: serum enzyme levels Ն2 times background were considered elevated and indicative of hepatitis. Antibody to HEV was detected by an in-house ELISA based on recombinant Sar-55 ORF2 protein as previously described, except that the ORF2 protein was more highly purified (22,23). Percutaneous needle biopsies of the liver were obtained weekly and read under code by a pathologist (S.G.).…”

“…Capped transcripts synthesized in vitro from this cDNA were tested for infectivity by intrahepatic inoculation into the liver of rhesus monkeys 471 and 407. Rhesus monkeys were chosen because they are very susceptible to infection by the Sar-55 strain, and much is known about the natural history of the virus in these animals (15,18,22). Although rhesus 407 had elevated serum liver enzymes during 1 week, neither monkey developed antibody to HEV, and the viral genome was not detected in sera by a sensitive manual RT-PCR assay ( Table 1), indicating that neither rhesus monkey was demonstrably infected.…”

Transfusion-associated TT virus infection and its relationship to liver disease

“…Antibodies to HCV were measured by a second-generation enzyme immunoassay according to the manufacturer' s instructions (Abbott HCV EIA 2.0, Abbott Laboratories, North Chicago, IL.). Antibody to hepatitis E virus was measured by the method of Tsarev et al 10 ALT was measured by a 3-point kinetic assay with an automated photometric analyzer (model 917, Hitachi-Boehringer Mannheim, Indianapolis) and values expressed in international units (normal range: 5-41 U/L).…”

Acute Hepatitis E with Elevated Creatine Phosphokinase

“…This was confirmed by conducting an ELISA, which showed a high-titer of anti-HEV antibodies, diagnostic of acute hepatitis E. IgM anti-HEV-Ab usually appears in the serum of infected patients with the onset of symptoms and remains for about 3-4 months. IgG anti-HEV-Ab can be detected promptly after the IgM response in the acute phase and is boosted in the early-convalescent phase and remains positive for several years (1,10). In a study of a nonendemic area, the specificities of IgG and IgM anti-HEV-Ab for diagnosing acute hepatitis E were 92.1% and 98.6%, respectively (ll).…”

“…He had no history of liver dysfunction and liver function tests at a routine checkup 10 Laboratory tests on admission are shown in Table 1. Leukocyte, erythrocyte and thrombocyte counts were within normal ranges.…”