TY - JOUR ID - 7180 TI - Production and evaluation of polyclonal rabbit antihuman p53 antibody using bacterially expressed glutathione S-transferase-p53 fusion protein JO - Iranian Journal of Biotechnology JA - IJB LA - en SN - 1728-3043 AU - Rasti, Mozhgan AU - Khatooni, Zahed AU - Mostafavi-Pour, Zoherh AD - Recombinant Protein Laboratory, Department of Biochemistry, Medical School, Shiraz University of Medical Sciences, P.O. Box 7134-845794, Shiraz, I.R. Iran. Y1 - 2012 PY - 2012 VL - 10 IS - 4 SP - 240 EP - 248 KW - GST-p53 KW - Tumor KW - Immunoassays KW - Antip53 antibody DO - N2 - p53 is a key tumor suppressor gene that is targeted for inactivation during human tumorigenesis. In this study, we produced and characterized polyclonal antihuman p53 antibody. The cDNA encoding the completehuman p53 protein was cloned into pGEX-4T-1 and expressed in Escherichia coli as a fusion protein with Schistosoma japonicum glutathione S-transferase (GST). The rabbits were immunized with the purifiedp53 recombinant protein. The obtained antisera were purified to increase the specificity of recognition. Thesensitivity and specificity of the produced antibody was analyzed by enzyme-linked immunosorbent,immunoblot, immunofluorescence and chromatin immunoprecipitation assays. Enzyme-linked immunosorbent assay showed that immunization with purified GST-p53 produced the high titer (1:10000) polyclonal antibodies with high specificity. Anti-p53 antibody allowed the sensitive detection of native p53 protein in immunoblotting, immunofluorescence and chromatin immunoprecipitation assays. Our results showed that anti-GST-p53 antibody provides a good means for studying the p53 expression pattern and its binding ability to other proteins in tumors. UR - https://www.ijbiotech.com/article_7180.html L1 - https://www.ijbiotech.com/article_7180_c389df12c31ca6d18c3d23bb57b226ed.pdf ER -