Ablation of c-Myc in mammary glands results in impaired lactation due to reduced milk volume. (a) Growth analysis of pups nursed by wild type (WT) or mutant mothers. Data are shown as average body weight plus standard deviation. Left panel: analysis of three littermate mothers nursing seven WT pups each. *, P = 2.2 × 10-5; **, P = 1.1 × 10-9. Right panel: comparison of a WT mother with six pups to a mutant mother nursing two pups (all pups WT littermates). NS = not significant, P = 0.52. (b) Milk protein composition in milk obtained from WT (W) or mutant (M) mice at lactation day 15.5. Freshly collected milk was diluted 1:20 in phosphate-buffered saline and 5 or 10 μl loaded on a 15% SDS-gel for Coomassie staining. (c) Measurement of lactose concentration in milk collected at lactation day 14.5. In a colorimetric assay, lactose concentration is determined as concentration of free galactose in lactase-treated skim milk. Results from five animals with the indicated genotype are shown as average value of duplicate measurements. (d) Analysis of fat content in the same milk samples used in (c), presented as percentage of fat layer length over total length of milk. Results are the mean ± standard deviation of three measurements per animal. (e) Whole mounts of WT and mutant mammary glands collected on lactation day 0.5. Mothers were sacrificed directly after removing them from pups (a, b), or after 2 hours without pups to allow filling of glands with milk (c, d). Arrows point to distended alveoli in WT gland. Scale bar, 500 μm. (f) Western analysis of milk proteins (loading: 1 μg per lane) and α-tubulin (loading: 9 μg per lane of the identical lysates) of WT and mutant mammary gland lysates at lactation day 5.5, 10.5, and 15.5. Blot was probed with anti-milk serum; two different exposure times are shown.