Identification of feline MAGE-1 gene product by monoclonal antibodies

Melanoma antigens (MAGE) are thought to induce a tumor-specific immune response and to be potential therapeutical targets for cancer immunotherapy. We have earlier identified the cDNA of feline melanoma antigen 1 (fMAGE-1), but its product was not characterized in detail. We have expressed the recombinant fMAGE-1 protein and have generated monoclonal antibodies (mAbs) against it, to identify the native fMAGE-1 protein in feline lymphoma cell lines and tumor tissues. The fMAGE-1 protein was found to be approximately 39 kDa in molecular mass on sodium dodecyl-sulphate-polycrylamide gel electrophoresis (SDS-PAGE), and it was found to be located in the cytoplasm of the cells by immunofluorescence. Immunoblotting analysis detected the fMAGE-1 gene product in the fMAGE-1-mRNA-positive cells, but not in the fMAGE-1-mRNA-negative cells. An interesting finding of the present study was the distribution of the fMAGE-1 protein, which was found to have a spindle-like distribution, with filaments twining around the nucleus, suggesting that the fMAGE-1 protein may be associated with or form some cytoplasmic filaments. This type of finding is so far the first report of its kind, and to the best of our knowledge it has not been reported in either human or mouse MAGE proteins until now. It most probably implies the major diversity of the MAGE family genes.