Preparation and characterization of immunogen for monoclonal antibody production against infectious bursal disease virus serotype 1 of poultry

Infectious bursal disease (IBD), defined as a list B disease by the Office International des Epizootics (OIE), caused by IBD virus of birnaviridae, shows antigenic variation and recently very virulent field isolates have been identified in U.S.A. and European countries. Monoclonal antibodies (MAbs) are valuable tools in characterizing virion's antigenic sites and detecting antigenic variation. Preparation of immunogen is one of the initial hurdles of MAb production against a virus. The present study was undertaken with the objective to produce suitable IBD virus immunogen for production of MAbs. For this a field isolate of IBD virus was adapted to chick embryo fibroblast (CEF) culture and the virus produced in bulk was partially purified by polyethylene glycol precipitation and trichloro-trifluoro ethane (Freon) extraction followed by discontinuous sucrose gradient (60 and 30%) ultracentifugation. The partially purified virus was confirmed by AGPT, virus neutralization test and characterized by SDS-PAGE analysis and then used as immunogen for inmmnizing the Balb/c mice. The splenocytes obtained from the immunized mice were used for producing hybridoma and then the hybridoma colonies were screened by indirect ELISA for secretion ofIBDV specific antibodies. A 23.2% (26 out of 112 hybridoma colonies screened) positive hybridoma colonies specific for IBD virus were obtained in one ofthe hybridization experiments. The relatively high percentage of IBD virus specific hybridomas obtained might be due to immunization of the mice with purified IBD virus antigen, which was propagated in CEF culture. The CEF culture system is advantageous as the purification is easier and at the same time it is a homologous system for IBD virus propagation.