Human mast cells differentiated from genetically modified hES cells. (A) A reporter gene expressing GFP under EF-1 α promoter was expressed in hES cell clone H1 by lentiviral transfection. hES cells double-positive for GFP and pluripotent marker TRA-1-81 (region R5) were sorted by MoFlow sorter. Left panel: Nontransfected cells, isotype control. Right panel: Transfected cells stained with TRA-1-81. (B) hES cell lines expressing GFP were established. (C) Differentiating EBs formed from GFP-hES cells. (D) GFP-expressing mast cell progenitors releasing from differentiated EBs to medium. Images were captured using an Olympus IX81 inverted fluorescence microscope with a 4× objective. (E) Mast cell progenitors express GFP and CD43. Left panel: Isotype control. Right panel: CD43 antibody. (F) Mast cell progenitors differentiated to mast cells expressing c-Kit, anti–c-kit-APC (gray), and isotype control (empty). (G) Mast cells derived from GFP-hES express tryptase. Top panel: GFP-hES cells. Bottom panels: Mast cells derived from nontransfected H1 clone. Figures were captured by an Olympus BX61 upright fluorescence microscope with a 40× objective with a Hamamatsu ORCA RC camera, operated by Velocity software (PerkinElmer Life and Analytical Sciences).