We recently established transfectants of MCF-7 human breast cancer cells with fibroblast growth factor 4 (fgf-4) that showed rapid growth and spontaneous metastasis in ovariectomized and tamoxifen-treated nude mice. To establish a spontaneous metastatic model of human breast cancer cells in nude mice with a sensitive marker for detection of micrometastasis, the transfection of fgf-4 was combined with transfection of the bacterial lacZ gene encoding beta-galactosidase. MKL-4 cells, a lacZ transfectant of an fgf-4-transfected cell line, showed the same level of fgf-4 expression as parental cells and expressed a high level of beta-galactosidase activity. When MKL-4 cells were injected s.c. into female nude mice, rapidly growing tumors developed. Whole organ staining for beta-galactosidase activity was able to detect even small numbers of metastatic tumor cells. Micrometastases in lymph nodes, lung, and brain were detected 3 weeks after the tumor cell injections, the first time point tested. Within 12 weeks, metastases were observed in lymph nodes, lung, brain, kidney, perirenal fatty tissues, liver, spleen, retroperitoneum, heart, and gallbladder. The frequency of metastasis and number of foci were correlated with the volume of the primary tumors. The distribution of metastatic sites was similar to that in breast cancer patients. MKL-4 cells may be a useful model for studying the malignant progression of hormone-dependent breast cancer, antimetastatic drugs, or early events in metastasis.