Characterization of a monoclonal antibody specific to the Gag protein of porcine endogenous retrovirus and its application in detecting the virus infection.
The porcine endogenous retrovirus (PERV) has drawn extensive attention recently, due to the widespread use of biomaterials of porcine origin in organ transplantation. This virus is present in all pig strains and has been demonstrated to be capable of infecting human cells in vitro. Therefore, it is imperative to develop a highly sensitive and specific immunoassay for clinical surveillance in patients receiving xenotransplantation. We describe here the generation of a monoclonal antibody (mAb) named A-11 specifically against the Gag protein of PERV. The mAb was found to be able to detect PERV produced from cultured cells. No cross-reaction with Gag proteins of murine leukemia virus (MuLV) and human immunodeficiency virus-1/2 was observed indicating that it is highly specific to PERV. The mAb was characterized as IgG2b subtype and kappa light chain. The region recognized by the mAb A-11 was localized to amino acid 293-336 on the Gag protein, and a synthetic peptide corresponding to amino acid 313-322 effectively competed the binding of the mAb with recombinant Gag proteins. Both immunocytochemistry and flow cytometry showed that the antibody is suitable for detection of PERV infection. By using the assays, we found that PERV-infected cells primarily of epithelial origin, with the highest infection rate in 293 followed by HEp-2 cells. In summary, the A-11 mAb will be useful for the development of quantitative and qualitative immunoassays for monitoring PERV infection in xenotransplantation patients and individuals who have close contact with pigs.
Chiang
Chen-Yi
CY
Insitute of Molecular Medicine, National Tsing-Hua University, 101 Section 2 Kuang Fu Road, Hsin-Chu, Taiwan 300, ROC.
Chang
Jen-Ting
JT
Lin
Meng-Shiue
MS
Wang
Shih-Rong
SR
Chang
Hwan-You
HY
D000818:Animals; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D002460:Cell Line; D020077:Endogenous Retroviruses / Q000276:immunology* / Q000302:isolation & purification*; D018604:Epitope Mapping; D005434:Flow Cytometry; D015683:Gene Products, gag / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology; D015498:HIV-2 / Q000276:immunology; D006801:Humans; D007120:Immunochemistry; D007074:Immunoglobulin G; D009052:Leukemia Virus, Murine / Q000276:immunology; D012192:Retroviridae Infections / Q000175:diagnosis* / Q000821:virology; D012680:Sensitivity and Specificity; D013552:Swine / Q000821:virology
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D015683:Gene Products, gag; D007074:Immunoglobulin G