Isolation of regulatory T cells from healthy murine mammary glands

Regulatory T (Treg) cells are a subset of CD4⁺ lymphocytes broadly recognized for their critical role in maintaining peripheral tolerance. More recent findings have highlighted their growing importance in maintaining homeostasis in several non-lymphoid tissues. These functions have been characterized in a number of tissues, but Treg cell physiologic function in mammary glands remains unexplored. Due to their low abundance in this tissue, Treg cells can be challenging to isolate for study; thus, here we describe a protocol for isolating Treg cells from murine mammary glands through fluorescence-activated cell sorting (FACS).