Over a period of 6 1/2 years between January 1986 and May 1992, 135 unselected primary breast cancers were cultured and of these 10 developed into cell lines. Six of the lines grew in defined serum-free medium, while the other four required supplementation with 0.5% fetal calf serum. Two of the lines are from the same breast, being derived from a local excision specimen and from a mastectomy specimen 12 months later. In addition, 12 lymph nodes containing metastatic breast cancer were cultured; one of these cultures became permanent in a defined serum-free medium. Oestrogen receptor (ER) status was negative in all but one of the tumours which grew successfully, and even in this case the derived cell line is ER negative. The epithelial nature of the lines has been confirmed by immunocytochemistry and by electron microscopy (EM), while their malignant nature is shown by morphology, unattached growth, chromosome analysis, and, in the case of the line from a lymph node metastasis, the absence of any benign source of epithelial cells.