Isolation and Characterization of Human Breast Epithelial Cells

Abstract

Breast cancer is one of the most common cancers among women in the United States and it is the second leading cause of cancer deaths in this population. Roughly 232,000 women in the U.S. were diagnosed with malignant breast cancer in 2013 and approximately 40,000 of these patients will die. The majority of breast cancers originate in the lobular or ductal cells of the milk‐producing glands. In these structures are two main cell types: luminal cells, which are surrounded by basal myoepithelium. These cells are structurally distinct and are the precursors to variable forms of breast cancer. As such, it is important to study them not only separately but also together. Current methods for culturing human mammary epithelial cells select for those of a basal phenotype. Isolation of pure basal and luminal cell populations was accomplished by using antibody‐linked magnetic beads to isolate essentially pure populations of both basal (CALLA/CD10) and luminal mammary (MUC‐1/CD227) epithelial cells from organoid explant cultures. Organoids were isolated and plated on collagen I coated flasks in either DFCI (basal) or MBCM (luminal) media and cultured at 37°C in 5% CO2 to induce cell migration. The media was replenished every 2 – 3 days until the cells reached confluence. Cultured cells were harvested and immunopurified using a CALLA/CD10 or MUC‐1/CD227 antibody and magnetic beads. Positively selected cells were seeded in tissue culture treated flasks and propagated with medium exchange every 2 – 3 days. A second round of immunopurification resulted in an isolated cell population that was greater than 95% MUC1 positive, suggesting a pure luminal epithelial cell population. These isolated cell types can now be tested individually for drug sensitivity.