Preparation Of Monoclonal Antibodies Against Newly Defined Cancer/Testis Antigens BORIS And CT45 And Their Distribution At Protein Level

With the development of tumor immunology, more and more tumor associated antigens were used for the research of tumor immunotherapy. As the characteristic of restrict expression in normal testis and tumour tissues,cancer testis antigens(CTAs) have become the focus of the recent research.However, the research mostly remained at DNA and mRNA levels. The main reason for the obstruction is lacking specific monoclonal antibodies (mAbs) against CT antigens.BORIS and CT45 are both newly defined CT antigens. In order to further investigate its expression pattern at protein level, we prepared monoclonal antibodies against BORIS and CT45 for the first time ,and carryied out the primary investigation of the both CT antigen's expression and distribution at protein level. The monoclonal antibodies against BORIS and CT45 were prepared with the lymphocyte hybridoma technology, and the corresponding mAbs with high specificity and sensitivity which recognize native BORIS or CT45 molecules were selected .The optimal working concentrations of the mAbs BORIS and CT45 in immunohistochemistry staining were 4.5μg/ml and 2.6μg/ml respectively. Using the obtained mAbs and the methodology of immunohistochemistry ,the expression of BORIS and CT45 at protein levels in the normal testis and tumor tissues was investigated. We found that both CT antigens were strongly expressed in some of spermatogonia and primary spermatocytes and distributed in cell nucleus . In addition we detected the expression of BORIS in breast cancer, benign breast diseases, normal breast tissue and CT45 in lung cancer tissue.The expression of BORIS antigen is much stronger in breast cancer than that in benign breast disease and normal breast tissues and the positive percentage of BORIS was 85.5% (94/110).while positive percentage of CT45 was only 4.8% (3/62) in lung cancer cases. The expression of both CT antigens was neither related to the grade nor the stage of cancer.The expression pattern of BORIS and CT45 in the normal testis and tumor tissues was consistent with the typical expression pattern of CT antigen . The preparation of monoclonal antibodies against two newly defined CT antigens BORIS and CT45 provides a powerful tool to further investigate them at protein level.