Preparation and identification of the polyclonal antibodies with immunohistochemical analysis from GST-p12~(DOC-1) fusion protein

Objective: We expressed p12DOC-1 in E.coli and prepared antibody against it for further study on function and modulation of doc-1 and its protein p12DOC-1 in squamous cell carcinoma of the human oral cavity. Methods: The constructed vector of pGEX-4T-1-doc-1 was transformed into E.coli BL21(DE3). GST-p12 fusion protein was expressed under IPTG induction and purified by affinity column and SDS-PAGE. The interest protein band was cut out as antigen, immunized the New Zealand rabbit to produce polyclonal antibodies. The specific antibody against p12DOC-1 was identified by enzyme-linked immunosorbent assay(ELISA),Western blot and immunohistochemistry. Results: The high purity GST-p12 fusion protein was obtained. GST-p12 antibody was generated in rabbit. Western blot and immunohistochemistry analyses showed that the antibody had high titer and specifically recognized GST-p12 fusion protein in vitro. Conclusion: The rabbit antiserum against human p12DOC-1 is prepared successfully, which will facilitate the study of the function and modulation of p12DOC-1 protein in oral cancer.