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Preparation and identification of monoclonal antibody against human GCRG213

WU Yin-qiao,WU Ben-yan,WANG Gang-shi,YOU Wei-di,WANG Wei-hua,WANG Meng-wei Department of Gastroenterology,South Building,General Hospital of PLA,Beijing 100853,China  
AIM:To prepare and characterize the monoclonal antibody against human GCRG213.METHODS:The HIS-GCRG213 fusion protein was expressed in E.coli.Mice were immunized with the purified HIS-GCRG213 protein.Hybridoma cell lines secreting monoclonal antibodies against GCRG213 were screened by regular cell fusion and subcloning approach.The titer and specificity of the antibody was characterized by ELISA and Western blot,respectively.The expression of GCRG213 was determined using immunohistochemistry technique on paraffin-embedded tissue sections from normal gastric mucosal tissues and advanced gastric cancer.RESULTS:The HIS-GCRG213 fusion protein with relative molecular mass of 20 800 was over expressed in E.coli.Two hybridoma cell lines which secreted monoclonal antibody specifically against human GCRG213 fusion protein were successfully obtained.The ascite titers of this monoclonal antibody reached 1∶ 106.Western blot analysis showed that the monoclonal antibody could bind to the recombinant HIS-GCRG213 protein specifically.The immunohistochemistry showed that GCRG213 were expressed higher in gastric cancer tissues than in normal ones.CONCLUSION:The monoclonal antibody against human GCRG213 with high titer and specificity has been successfully prepared,which could be utilized as a useful reagent for further studying the biological function of the GCRG213.
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