Isolation of gE gene deleted pseudorabies virus by using a gE specific monoclonal antibody

A simple, convenient method employing a gE-specific neutralizing monoclonal antibody (mAb; FTpn3) for isolation of the gE gene-deleted recombinant pseudorabies virus (PRV) is described. FTpn3 secreting hybridoma was obtained by fusing PRV-immunized BALB/c splenocytes with myeloma cells. To construct gE gene deleted PRV, a 5.7 kbp DNA fragment with deletion of the gE gene was engineered and cloned. The plasmid was then used for cotransfecting Vero cells with wild-type PRV genome. The resulting viruses were subjected to FTpn3 neutralization. The FTpn3 resistant virus was isolated and plaque purified further. By DNA fingerprinting and Western blotting analysis, the virus resistant to FTpn3 neutralization was proved to be the gE-deleted recombinant virus.