IL-25–elicited MPP^type2 cells, but not ILC2, possess multipotent potential. BALB/c WT mice (The Jackson Laboratory) were treated with 0.3 µg of recombinant IL-25 daily for 7 d. MLNs and PECs were harvested from IL-25–treated mice, and ILC2 and MPP^type2 cells were sort-purified and cultured in the presence of IL-33 alone or SCF plus IL-3. (a and d) FACS purification gating of ILC2 (Lin^neg T1/ST2^pos IL-7Rα^pos; a) or MPP^type2 cells (Lin^neg T1/ST2^neg IL-7Rα^neg c-kit^pos; d) from IL-25–treated mice. Plots shown are gated on live, Lin^neg cells (CD3ε, CD8α, CD19, CD11b, CD11c, Gr-1). (b and c) Flow cytometric analysis of myeloid, granulocyte, and ILC2 cell differentiation of day 8–12 cultures seeded with FACS-purified ILC2 in the presence of IL-33 (b) or SCF and IL-3 (c). (e and f) Flow cytometric analysis of macrophage, granulocyte, and ILC2 cell differentiation of day 8–12 cultures seeded with FACS-purified MPP^type2 cells in the presence of IL-33 (e) or SCF and IL-3 (f). (g) Culture supernatants from b, c, e, and f were collected, and IL-4, IL-5, and IL-13 protein levels were measured by ELISA. Data in a–g are representative of at least three independent experiments. Error bars indicate SEM.