We have examined the growth and differentiation properties of the transplantable mammary tumor line of the A X C rat, MCCLX. The MCCLX tumor grows rapidly and exhibits high activity of the milk protein alpha-lactalbumin (alpha LA) (40% of the level in midlactating mammary gland) during serial passage in estrogen-supplemented (E+) intact males. Hypophysectomy of hosts bearing established tumors results in rapid tumor regression and rapid loss of alpha LA protein activity. Treatment with ovine prolactin three times daily (total of 2 mg/day) completely restores alpha LA activity and results in a partial restoration of tumor growth. In females, MCCLX grows only in E+ virgins or in pregnant hosts. During lactation, tumor growth ceases. However, alpha LA activity is equally high in E+, pregnant, and lactating females. Behavior of MCCLX apparently depends on circulating lactogenic hormones. When hormone levels are very high, as in E+ intact males or pregnant females, both growth and alpha LA activity are stimulated. In lactating hosts, having lower serum lactogen, alpha LA activity could be maintained in the absence of growth. In hypophysectomized hosts, even with continuous estrogen supplementation, alpha LA activity disappears and growth ceases, although ovine prolactin can replace the pituitary in restoring these traits. MCCLX contains prolactin-binding capacity; thus, direct action of the lactogenic hormones on the tumor is possible. Our results indicate that MCCLX is a stably hormone-responsive tumor in terms of growth and differentiation.