Generation and testing of a highly specific anti-serum directed against porcine endogenous retrovirus nucleocapsid

Advances in xenotransplantation offer chances to alleviate the shortage of human donor organs. The discovery that pig endogenous retroviruses (PERV) c an infect human cells in vitro has stimulated the discussion on infectious risk in xenotransplantation. A molecular and immunologic monitoring of xeno graft recipients and of donor animals for putative infection with PERV and other microorganisms is inevitable. In this report, we describe the generat ion and testing of a highly specific anti-serum directed against the PERV n ucleocapsid protein. The Gag amino acid (aa) sequence of PERV class B was u sed to define immunogenic domains by computer analysis. A peptide correspon ding to the C-terminal 19 aa of the 10 kDa (p10) nucleocapsid (NC) portion of the Gag polyprotein was used to immunize rabbits. The generated serum wa s tested using recombinant PERV Gag protein expressed in insect cells, puri fied PERV virus particles and human 293 cells transfected or infected with PERV, respectively. Test methods included Western blotting, indirect immuno fluorescence, immunoperoxidase assay and ELISA. The PERV anti-serum provide s a tool that is instrumental for detection of a potential agent of zoonosi s. It can be used for screening of donor animals and xenograft recipients i n the course of xenotransplantation procedures.