Prostate stem cell antigen (PSCA) is a 1998 Reiter et al study gene expression in prostate cancer discovered during a tumor-associated antigen, belonging Ly-6/Thy-1 family, is a 123 amino acid glycoprotein. Because PSCA with stem cell antigen -2 (SCA-2) having a homology of 30%, which is called the prostate "stem cell antigen", but the current study, PSCA expressed mainly in cell differentiation and maturation of stem cells than the surface of the surface, their specific function is unclear. PSCA in prostate cancer, bladder cancer, and pancreatic cancer, increased expression and found its expression levels and prostate cancer clinical stage and metastasis, so PSCA has become of these tumor diagnosis and prognosis of biological markers. In addition to PSCA also become the target of these tumors immunotherapy Points, particularly in the treatment of prostate cancer, immune show great potential for clinical application. PSCA as a target in prostate cancer immunotherapy preclinical studies, the use of transgenic mice or mouse prostate cancer model for preclinical evaluation. To further explore the purpose of PSCA in prostate cancer immune RT-PCR amplification of mouse prostate stem cell antigen (mPSCA) gene prokaryotic expression vector pET-42a-mPSCA, induced mPSCA protein expression and purification, and to detect mPSCA antigenic activity. Methods RT-PCR method from the mouse prostate cancer cell line RM-1 cells was amplified mPSCA gene sequencing PCR method to remove mPSCA after the signal peptide sequence was cloned into the prokaryotic expression vector pET-42a to construct recombinant plasmid pET -42a-mPSCA. To convert To E. coli BL21 (DE3) induced expression, the fusion protein is then purified. Analyzed by SDS-PAGE, Western blot detection of purified protein, the purified protein further package board after its antigenic activity by ELISA were evaluated. Results sequencing results confirmed length mPSCA successfully amplified gene; digestion and sequencing Results confirmed pET-42a-mPSCA prokaryotic expression vector was successfully constructed; transformation can be successfully induced and purified proteins consistent with the expected size; Western blot test confirmed the purified protein could react with the specific antibody; ELISA assay showed purified mPSCA immunogenic antigens. Conclusion successfully amplified full-length mouse PSCA gene was successfully constructed mPSCA prokaryotic expression vector obtained purified mPSCA protein, which has a good antigenicity for further research to PSCA as a target for immunotherapy of prostate cancer foundation.
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From:Cytokines products
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