Preparation Of The Monoclonal Antibody Against The GE Protein And Identification Of The B-Cell Epitope In GE Of Pseudorabies Virus

Pseudorabies(PR)or Aujeszky's disease(AD),caused by pseudorabies virus(PRV),is a serious disease of pigs and other animals.PRV or suid herpesvirus ? belongs to the family of the Herpesviridae,subfamily Alphaherpesvirinae.PRV gE gene is one of PRV virulence determinants and a non-essential gene for in vitro replication as well.When the gE gene is deleted,the propagation and antigenicity of PRV are not affected,but its virulence is reduced greatly.These characteristics make glycoprotein E(gE)can be used as a marker to differentiate vaccine against swine and wild virus infected pigs.Since 2011,PR was epidemic outbreaks in pig farms,which has caused significant economic losses to the pig industry in our countries.Therefore,it is imperative to establish a rapid and effective method for diagnosis and detectionTo prepare the monoclonal antibody(MAb)against PRV gE and identify the epitope of the protein,a hybridoma cell line stably secreting the MAb against PRV gE was prepared by fusion of the SP2/0 cells with the spleen cells from the mouse which was immunized with the prokaryotically expressed truncated protein of gE within aa127 to aa232.The MAb was IgG1 subtybe with ? chain.The titers of the MAb in cell culture medium and ascites were 1:80 and 1:2560,respectively.Western blot assay showed that the MAb was able to recognize gE protein specifically,but had no virus neutralizing activity.The targeted epitope of the MAb was mapped by pepscan with a serial overlapping short peptides,and the sequence of the epitope recognized by the MAb was identified as 151IGDYL155,which was the conservative epitope sequences among the gE of PRV strains.This sequence is shorter and more accurate than the main epitope region aa52 to aa238 identified by Jacobs.These results provide a basis for further study of the antigen structure of PRV gE,and provide a theoretical and practical basis for the diagnosis,treatment and vaccine development of PRV.