Immunochemical Demonstration of αs1- and β-Casein in Mouse Mammary Glands at Early Stages of Pregnancy

We generated monoclonal antibodies (MAbs) against mouse αs1- and β-casein and used them to survey casein immunochemically in mammary glands of mice at pericoitous and pregnant stages. Two MAb-producing hybridoma cells, designated MCα1 cell and MCβ1 cell, were established. Each antibody, when used in Western blotting, recognized specifically mouse αs1- and β-casein among a wide spectrum of proteins of both a lactating mammary homogenate and mouse skim milk. Immunohistochemistry revealed αs1- and β-casein in sections of lactating mammary glands. Staining was found in substances in the lumen and cytoplasm of duct and alveolar cells, particularly in rough endoplasmic reticulum and the Golgi apparatus. Mammary glands at Days 2, 4, 6, 8, and 14 of pregnancy showed positive staining specific to both αs1- and β-casein in the lumen and cytoplasm of duct cells, whereas the glands at estrus and Day 0 of pregnancy were positive mainly for αs1-casein. Semiquantitative Western blotting analysis of both casein components in epithelial cell fractions from glands during pregnancy confirmed that intra-epithelial αs1- and β-casein changed during three phases, elevated from trace levels to detectable levels during initial stages of pregnancy (Days 0, 2, and 4), declined to lower levels during mid-pregnancy (Days 6 and 8), and then rose to high levels during late pregnancy (Day 14).