Evidence from preclinical research and clinical trials demonstrates the use of the stromal vascular fraction (SVF) as therapy for numerous indications. These results demonstrate that autologous SVF is not only safe and effective but provides robust anti-inflammatory, immunomodulatory, and reparative effects in vivo. The potency of the SVF is attributed to the cellular composition which includes adipose-derived stem cells (ASCs), adipocytes, endothelial cells, and various immune cells. As the name would suggest, these SVF cells are derived from the stromal compartment of adipose, or fat. Once digested, the cells that constitute adipose are released and collected as the SVF. The cellular frequencies within the SVF can then be assessed using a fluorescent antibody-based technique known as flow cytometry. The following chapter provides a standard operating protocol that describes the procedures from harvesting the fat tissue from experimental mice to isolating and characterizing the SVF.