PREPARATION OF HYBRIDOMA CELLS FOR PORCINE EPIDEMIC DIARRHEA VIRUS MONOCLONAL ANTIBODY PRODUCTION

Abstract

Porcine epidemic diarrhea (PED), caused by Porcine epidemic diarrhea virus (PEDV) is an enteric disease characterized by watery diarrhea, vomiting, weight loss, dehydration, resulting in 100% mortality in suckling pigs . PED cannot be clinically distinguished from Transmissible gastroenteritis (TGE), therefore the laboratory diagnosis is necessary to identify PED. However, laboratory techniques are expensive, time-consuming and cannot be carried out in the farm. The objective of this study is to prepare hybridoma cells for monoclonal antibody production against Porcine epidemic diarrhea virus. Two BALB/c mice were immunized subcutaneously 3 times at 3 week-intervals with 50 µg purified PEDV strain K-9 per mouse. Finally, 3 days before fusion, the mice were intraveneously injected with purified PEDV. Hybridoma cells were obtained by fusion between X63Ag 8.653 myeloma cells and B- cells from mouse using 50% polyethylene glycol. Hybridoma cells are cultured in HAT selective medium and supernatant are then tested for antibodies against PEDV using ELISA. The hybridoma cells showing PED specific antibody production were subsequently subjected to limiting dilution in order to obtain monoclone. In this study,Four monoclonal antibodies against PEDV strain K-9 were produced with subclass of IgG1, IgG2a and IgG2b.