DA-3/sec MDSCs are not suppressive, and MDSCs suppress via the enzyme arginase 1. (A) BALB/c^nu/nu mice were implanted with 10⁶ DA-3/TM or DA-3/sec tumor cells. When tumors reached similar volumes, spleens of normal mice and tumor mice were harvested, and MDSCs were purified using a GR-1 antibody, then cocultured with CFSE-labeled DO11.10 splenocytes, and stimulated with OVA protein. Proliferation was measured by analyzing CFSE dilution by flow cytometry. (B) Splenic MDSCs were isolated from DA-3/TM tumor bearers and cultured in a suppression assay as in panel A, with the addition of the arginase inhibitor (NOHA) and/or the NO synthase inhibitor (NMMA). Proliferation (CPM) was measured after providing cultures [³H]thymidine. Error bars representing SEM are provided, and data are representative of at least 3 experiments.