Alterations in alveolar density and secretory activity. (a) Immunohistochemistry against cytokeratin (CK) 18 on sections of wild type (WT) and mutant glands to visualize epithelium (brown). For optimal comparison, a central part of each gland containing the lymph node was taken. Lymph nodes (blue) and stroma display no CK18 staining. (b) Quantification of alveolar area per total organ area (excluding lymph node) from sections processed as shown in (a). Two sections per animal were quantified and the average was used for calculations. Results are the mean ± standard deviation of four and three pairs of animals quantified on lactation day 3.5 and 10.5, respectively. *, P = 0.00057; **, P = 0.0043. (c) Number of alveoli (including mean ± standard deviation) measured in four pairs of animals at lactation day 3.5, which were used for area measurement in (b). Numbers were calculated per analyzed area, and values from WT glands were set to 1. NS = not significant, P = 0.25. (d) Electron microscopy pictures from day 7.5 lactating WT and mutant glands. Arrows point to regions of dilated, vesicle forming endoplasmic reticulum. Scale bar, 500 nm.