Immunochemical demonstration of alpha(s1)- and beta-casein in mouse mammary glands at early stages of pregnancy.

We generated monoclonal antibodies (MAbs) against mouse alpha(s1)- and beta-casein and used them to survey casein immunochemically in mammary glands of mice at peri-coitous and pregnant stages. Two MAb-producing hybridoma cells, designated MCalpha1 cell and MCbeta1 cell, were established. Each antibody, when used in Western blotting, recognized specifically mouse alpha(s1)- and beta-casein among a wide spectrum of proteins of both a lactating mammary homogenate and mouse skim milk. Immunohistochemistry revealed alpha(s1)- and beta-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 alpha(s1)- and beta-casein in the lumen and cytoplasm of duct cells, whereas the glands at estrus and Day 0 of pregnancy were positive mainly for alpha(s1)-casein. Semiquantitative Western blotting analysis of both casein components in epithelial cell fractions from glands during pregnancy confirmed that intra-epithelial alpha(s1)- and beta-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).