Mesenchymal stromal/stem cells (MSCs) are currently being used in clinical trials as proposed treatments for a large range of genetic, immunological, orthopaedic, cardiovascular, endocrine and neurological disorders. MSCs are potent anti-inflammatory mediators which are considered immune evasive and employ a large range of secreted vesicles to communicate and repair damaged tissue. Despite their prolific use in therapy, sex specific mechanism of action is rarely considered as a potential confounding factor for use.The purpose of this study was to examine the potency and functionality of both female and male adipose derived MSCs in order to gain further insights into donor selection. Using functional immune assays, ELISA, multiplex and immunophenotyping we showed female MSCs (fMSC), consistently suppressed Peripheral blood mononuclear cell (PBMC) proliferation significantly (p<0.0001) more than male MSC (mMSC). In co-culture mPBMCs, showed 60.7+/-15.6% suppression with fMSCs compared with 22.5+/-13.6% suppression with mMSCs. Similarly, fPBMCs were suppressed by 67.9+/-10.4% with fMSCs compared to 29.4+/-9.3% with mMSCs. The enhanced immunosuppression of fMSCs was attributed to the production of higher concentrations of the anti-inflammatory IL-1RA (1025 pg/ml vs 701 pg/ml), PGE-2 (6142pg/ml vs 2448 pg/ml), IDO (3301pg/ml vs 1699pg/ml) and prolonged expression of VCAM-1 post activation relative to mMSCs. In contrast, mMSCs produces more inflammatory G-CSF than fMSCs (806pg/mL vs 503 pg/mL). Moreover, fMSCs, but not mMSCs induced downregulation of the IL-2 receptor and sustained expression of the early T cell activation marker, CD69 in PBMCs further highlighting the differences in immunomodulation potentials between the sexes. In conclusion, our data shows that female MSC are more potent in vitro than their male counterparts. The inability of male MSC to match female MSC driven immunomodulation and to use the inflammatory microenvironment to their advantage is evident and is likely a red flag when using allogeneic male MSC as a therapeutic for disease states.