Lactational function in the mammary epithelial cell is subject to complex regulation, most probably involving multiple extracellular and intracellular proteins that act at any of a number of levels. Although some of these proteins have been identified it is likely that additional controllers of lactation exist, but have yet to be discovered. In an effort to identify such proteins, a search was made for non-milk lactation-associated or prolactin-responsive proteins in primary mouse mammary epithelial cells and the mouse mammary epithelial cell line, COMMA-D using two-dimensional electrophoresis on large-format gels. These analyses revealed 12 proteins whose rate of synthesis was dependent on lactation state or on response to prolactin. Two of these (p77 and p63) were lactation-associated in primary cells and prolactin-responsive in COMMA-D cells. These two proteins were identified by amino acid sequencing as glucose-regulated protein 78 (GRP78) and protein disulphide isomerase (PDI). The localization of these proteins in the endoplasmic reticulum and their presence in other secretory cell types and tissues suggests that they have a function in the processing or secretion of milk proteins.