The expression and application of human Fas ligand in E.coli

AIM: To express recombinant human FasL molecule in E.coli. METHODS; RT-PCR was applied to amplify FasL cDNA from the total RNA extracted from activated human peripheral blood lymphocytes. The DNA fragment was cloned into PCR2. 1 vector. After sequencing, the FasL gene was inserted into pQE-31 vector and expressed in E. coli M15. The FasL protein was purified through Ni-ATA affinity chromatography column and identified by SDS-PAGE and Western blot. The mice were immunized with the FasL protein and the specific anti-serum was harvested 6 weeks after immunization. The serum level of FasL from with different kinds of diseases patients were detected using the anti-FasL antibodies from the immunized mice. RESULTS: The expressed protein could be recognized by anti-human FasL antibody in Western-blot analysis with M, 40 000. This protein could induce Jurket cells apoptosis. anti-FasL serum prepared from mouse could detect the serum FasL as sensitive as commercial ELISA kits. CONCLUSION: The human FasL protein is obtained. It lays the foundation for the further detecting the concentration of FasL and sFasL of patients.