Development of a sandwich ELISA for the detection of bovine herpesvirus type 1

Objective:To develop a standard enzyme-linked immunosorbent assay(ELJSA) for the detection of bovine herpesvirus type 1(BHV-1).Methods:The assay was based on hyperimmune rabbit and guinea pig antisera raised against purified BHV-1.Polyethylene glycol precipitation and sucrose density gradient methods were adopted for viral concentration and purification.Antisera were raised using Freund’s adjuvant followed by extraction of IgG of high purity.Results: Optimum antisera dilutions as determined by titrations were chosen as 14 000,whereas the conjugate was used at 1:2 000 dilution.Using 95 clinical specimens,the ELISA test showed a sensitivity and specificity of 91.90%and 93.10%,respectively when compared to PCR.The cutoff value was fixed at 0.15) and a P/N ratio of>1.30 indicated a significant positive reaction. Conclusions:The results have demonstrated that this ELISA could efficiently detect BHV-1 and can be used as an important diagnostic tool.