Remove mammary glands from mice and place in sterile PBS to rinse.
Chop/mince glands with razor blade and place into 15ml conical with 10mls collagenase solution. (Typically 4-8 virgin glands/tube ~500-1000mg tissue/tube)
Digest tissue in collagenase solution for 2-3 hours at 37oC on a rotator.
Transfer supernatant (this contains single cells/stromal cells) to a new tube and leave the settled material in the tube (epithelial cells). Wash the remaining settled material by adding 10mls of wash solution. Spin cells.
Repeat the wash 3X.
The primary cells are now ready to expand in tissue culture in appropriate media.
Charlotte Kuperwasser, PhD
Associate Professor of Developmental, Molecular & Chemical Biology
Tufts University School of Medicine
Molecular Oncology Research Institute (MORI)
Tufts Medical Center
Boston, MA 02111