Two spontaneous mouse leukemias were adapted to culture. In agreement with most reported observations on spontaneous tumors, injection of irradiated cells of the malignant culture cell lines failed to protect mice against these leukemias. These cell lines were treated in vitro with the mutagen N-methyl-N'-nitro-N-nitrosoguanidine and stable immunogenic variants (tum-) were obtained, that failed to form progressive tumors in syngeneic CBA/Ht mice. Mice that had rejected tum- variants showed a significant degree of resistance to challenge not only with the original malignant cell line but also with the original transplantable tumor. No protection was observed against syngeneic tumor cells other than those of the parental tumor. These results indicate that these two spontaneous leukemias carry a specific transplantation antigen that can be the target of a rejection response by syngeneic mice. In confirmation of this, we found that lymphocytes of CBA/Ht mice that had rejected tum- variants could be restimulated in vitro so as to develop a cytolytic activity directed against an antigen that was specific for the original tumor cell line.

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