Preparation Of A Monoclonal Antibody Against Human 4IgB7-H3 Molecule And Its Primary Application

B7-H3(CD276),a member of the B7 family,is expressed in several nonlymphoid tissues of humans and mice and in several human tumor cell lines,and can be induced in monocytes and dendritic cells by proinflammatory cytokines.The receptor of B7-H3 has not been identified yet,but is located on activated T cells.The functional role of the B7-H3 system is discussed controversially.B7-H3 still has some dispute on the function in immune system.In addition,different splicing of human B7-H3 leads to a 4Ig domain (VCVC) transcript[also called B7-H3b and 4Ig-B7-H3]and a 2Ig domain(containing V1 and C2) transcript.Both isoforms are not clarified,there has been much discussion on their expression patterns,T-cell responses,etc.So the further study about the biological function of B7-H3 and the existence of the two isoforms is valuable.Objective:To prepare functional monoclonal antibodies against human 4IgB7-H3 molecule and analysis of their primary application.Methods:Female BALB/c mice of 6-8 weeks old were immunized with human 293T cell as immunogen.The spleen B cells of the mice were fused with SP2/0 and hybridoma cells were screened with L929/4IgB7-H3 and L929/2IgB7-H3 transfected cells by FCM. The specificitis of antibody were investigated by competitive inhibition test.Furthermore, to analyze the expression patterns of 4IgB7-H3 on human different cell lines by FCM.We also use the immunohistochemistry to analyze the expression of 4IgB7-H3 on human glioma tissues.Results:One hybridoma cell line named 9C3 was obtained,which had the property of secreting anti-human 4IgB7-H3 molecule continuously and steadily.This mAb specifically recognize human 4IgBT-H3 transfected cells but not 2IgB7-H3 transfected cells.The expression pattern of 4IgB7-H3 on different cell lines indicates that 4IgB7-H3 molecule is not the major pattern and expressed limited although the ealier study think that the expression of 4IgB7-H3 is higher than 2IgB7-H3 on mRNA level.The 4IgB7-H3 molecule expresses on human glioma tissues specifically,and increases as the pathology's grades.Conclusion:One hybridoma cell line which pecifically recognize human 4IgB7-H3 molecule was obtained.Our study gives a clue to recognize the expression patterns and biological function of B7-H3.