Immunotherapy of breast cancer at an early stage of the disease increases the likelihood of success. Here, in a mouse model, we report a new strategy that enables vaccines to be prepared from microgram amounts of tumor tissue. The vaccine is prepared by transfer of a cDNA expression library from relatively small numbers of breast cancer cells into a highly immunogenic cell line, where genes specifying TAA are expressed. As the transferred DNA is integrated and replicated as the recipient cells divide, the number of vaccine cells can be conveniently expanded for repeated immunizations. A cDNA expression library prepared from a breast cancer that arose spontaneously in a C3H/He mouse (H-2(k)) was transferred into a mouse fibroblast cell line derived from C3H/He mice. To augment their nonspecific immunogenic properties, the fibroblasts were genetically modified before DNA transfer to secrete IL-2 and to express allogeneic MHC class I H-2K(b)-determinants. C3H/He mice, highly susceptible to growth of the breast cancer cells, were immunized with the cDNA-transfected cells. Robust breast cancer-specific CD8(+) T-cell-mediated immunity was generated in the mice, raising the possibility that an analogous treatment strategy could be used to treat breast cancer patients at an early stage of the disease.