Application of immunoperoxidase-based techniques to detect herpesvirus infection in tortoises

Abstract

Indirect (IIP) and direct (DIP) immunoperoxidase assays were developed for the serological and histological diagnoses of herpesvirus infection in tortoises, respectively. A mouse monoclonal antibody (MAb HL1546), specific for the heavy chain of tortoise IgY, was used as the secondary antibody in the IIP assay. Rabbit polyclonal antisera raised against 2 sucrose gradient-purified tortoise herpesvirus isolates (HV4295/7R/ 95 and HV1976) were used as primary antibodies for the detection of herpesvirus antigen either in infected cell cultures or in formalin-fixed, paraffin-embedded tissues. The IIP and DIP assays could detect either the presence of anti-herpesvirus antibody in the plasma of exposed tortoises or the presence of herpesvirus antigen in infected tissues, respectively. Although the IIP test complements the enzyme-linked immunosorbent assay and the serum neutralization test already available for measuring herpesvirus-specific antibody in tortoises, the DIP test is useful for the histological diagnosis of herpesvirus infection in tortoises.