Production of monoclonal antibodies to polyhedrin of Monodon Baculovirus (MBV) from shrimp

Abstract

Monodon baculovirus (MBV) is a member of the nucleopolyhedrosis virus (NPV) group that produces polyhedral protein occlusion bodies that contain virions in epithelial cells of the hepatopancreas of the peinaeid shrimp Penaeus monodon. The major constituent protein of occlusion bodies is polyhedrin. Polyhedrin protein of MBV was extracted from the infected hepatopancreas of P. monodon post larvae by hydroforce and partially purified using Urografin gradient ultracentrifugation. A fraction at 50% Urografin was examined by electron microscopy and shown to be dominated by polyhedrin protein particles of 20-23 nm diameter. By SDS-PAGE, this fraction yielded a single protein band at a molecular weight of 58 kDa corresponding to the published size of MBV polyhedrin. This fraction was used to produce 17 monoclonal antibodies (MAb) that were specific to MBV and without cross-reactivity to other common shrimp viruses (i.e, hepatopancreatic parvovirus or HPV, yellow head virus or YHV and white spot syndrome virus or WSSV) by immunohistochemistry. These antibodies could be used to detect purified-polyhedrin with high sensitivity up to 0.2 μg/ml by dot blot immunoassay. These MAb are candidates for sensitive MBV immunodetection methods.