Development of mammary tumors and detection of disseminated and/or metastatic cells in MMT mice. (a) Mammary glands were harvested from mice double transgenic for PyMT and MUC1 antigen (MMT) or MUC1 transgenic (MUC1.Tg) mice at indicated ages, processed for whole mount or sections and stained with H&E. The solid masses are indicated by circles. (b) Comparison of mammary masses in different age groups (n = three to seven per group) of MMT, MUC1.Tg and MT (mice transgenic for PyMT oncogene) mice. The whole mount was digitized and the solid masses were traced and measured using SPOT advanced software. (c) Detection of disseminated cells or metastasis in lungs. Lungs were harvested from MMT mice at the indicated ages and perfused to remove circulating cells. The lung tissue was minced and digested in a collagenase enzyme cocktail solution. The cells were then cultured on tissue culture plates for two weeks and stained with 0.5% crystal violet (upper panels). Lungs from mice at the indicated ages were processed for histological examination with H&E staining. The green square indicates the enlarged area. (d) Colonies of more than 50 cells from lung cell cultures of each mouse (n = three to seven per group) were counted. Statistical analysis was performed using one-way analysis of variance.