Induction of CD316/EWI-2 on DCs. (a to d) Three different novel anti-EWI-2 mAbs were characterized by FACS staining of BaF/3 transfectants expressing full-length EWI-2: 5E8 (a), 6H5 (b), and 8D3 (c) (filled profiles; empty profiles are the isotype controls). (d) On Western blots, clone 5E8 specifically recognizes EWI-2 from PEER cells (lane 3) and from transfected SP2/0 cells (lane 2), whereas parental SP2/0 cells are negative (lane 1). The expected size (molecular mass [M]) of EWI-2 is ∼65 kDa, as indicated by the black arrow. (e to n) Expression of EWI-2 was analyzed by FACS on monocytes (e and j), MoDCs (f and k), myeloid DCs (g and l), plasmacytoid DCs (h and m), and in vitro generated Langerhans-like cells (i and n) before (upper row) and after (lower row) stimulation. Filled profiles, mAb 5E8; empty profiles, isotype controls.