Cancer Immunity 6:7 (2006)

Generation of monoclonal antibodies to cancer/testis (CT) antigen CT10/MAGE-C2

Ran Zhuang^1*, Yong Zhu^1*, Liang Fang¹, Xue-Song Liu¹, Ying Tian¹, Li-Hua Chen¹, Wei-Ming Ouyang¹, Xiao-Guang Xu¹, Jin-Long Jian¹, Ali O. Güre², Sheila Fortunato², Gerd Ritter², Lloyd J. Old², Andrew J. G. Simpson², Yao T. Chen³, Boquan Jin¹, and Achim A. Jungbluth²

¹Department of Immunology, Fourth Military Medical University, Xi’an 710032, P. R. China
²Ludwig Institute for Cancer Research, New York Branch at Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, Box 32, New York, NY 10021, USA
³Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10158, USA
^*These authors contributed equally to this work

Keywords: cancer/testis, tumor antigens, CT10/MAGE-C2, monoclonal antibodies, ELISA, immunohistochemistry

Abstract

CT10/MAGE-C2 is a recently identified antigen that, typically of cancer/testis (CT) antigens, can be found in various malignant tumors and in normal adult testis. As with many other CT antigens, our knowledge is based mainly on mRNA expression data. In the present study, we describe the generation of mAbs to CT10/MAGE-C2 for the analysis of its protein expression. Newly generated clones were chosen based on their reactivity in ELISA, immunoblotting, and immunohistochemistry (IHC). Emphasis was put on the reactivity of newly generated reagents on formalin-fixed, paraffin-embedded tissue to ensure their applicability to archival material. Eventually we selected two clones, LX-CT10.5 and LX-CT10.9, that showed intense reactivity to CT10/MAGE-C2 protein and CT10/MAGE-C2 mRNA-positive cell lines, but no cross-reactivity with other CT antigens. Both mAbs show superior staining characteristics in IHC and are applicable to frozen and paraffin sections. In testis, CT10/MAGE-C2 displays the typical CT pattern with regard to staining of germ cells, which is intense during the early maturation stages. In tumors, we analyzed a limited number of cases displaying the typical heterogeneous CT expression pattern. Interestingly, immunoreactivity was seen solely in the nucleus: No staining was seen in the cytoplasm of tumor cells.