Preparation and application of mouse polyclonal antibodies against human Shisa like 1 (SSL1)

: Objective To prepare polyclonal antibodies against Shisa like 1 protein (SSL1) and study the localization of SSL1 in hepatocellular carcinoma SMMC-7721 cells. Methods Human SSL1 gene was cloned from HepG2 cells by reverse transcription PCR, and then inserted into prokaryotic expression vector pET-28a to generate the SSL1 expression vector. The recombinant plasmid pET28a-SSL1 was then transformed into E. coli BL21 (DE3) and induced to express by IPTG. Polyclonal antibody against SSL1 was generated by immunizing Kunming mouse with the purified protein by the routine method. The specificity of polyclonal antibody was verified by Western blot analysis. The expression of SSL1 in SMMC-7721 cells was detected by immunofluorescent cytochemistry. Golgi complexes were signed by Golgi-Tracker Red to analyze the subcellular localization of SSL1 protein in SMMC-7721 cells. Results The SSL1 gene was cloned and the recombinant vector pET28a-SSL1 was successfully constructed. Pure SSL1 protein expression in E. coli BL21 was confirmed and polyclonal antibodies against protein SSL1 was obtained in immunized Kunming mice. Immunofluorescent cytochemistry showed that SSL1 was expressed in the cytoplasm, and was co-localized with Golgi-Tracker Red in SMMC-7721 cells. Conclusion We have obtained SSL1 polyclonal antibodies with high specificity, which was proved situated in Golgi bodies of SMMC-7721 cells.