Fig. 3. Stage-specific fate mapping of the Ascl1-lineage. (A-D',H) X-gal staining of Ascl1-CreER^TM;R26R-lacZ embryos harvested at the stages indicated and treated with tamoxifen 24 hours before harvest. Whole-mount stained and cleared embryos (A-D) and vibratome sections through the neural tube are shown (A'-D',H). Arrowheads in A,A',B indicate X-gal staining in the sympathetic ganglia. (E) Cross section of an E10.5 Ascl1-CreER^TM;R26R-YFP embryo treated with tamoxifen at E9.0. Triple-label immunofluorescence with antibodies to YFP (green), Lmx1b (red) and Lhx1/5 (blue) identify the YFP+ cells as interneurons dI3, dI5 and V2. (F,I,J) Immunofluorescence showing Ascl1 in the dorsal neural tube at E11.5 (F) and broadly in the ventricular zone as well as gray and white matter in E16.5 spinal cord (I). The specificity of the Ascl1 antibody is illustrated by the absence of signal in E16.5 spinal cords of an Ascl1 null embryo (J). (G) In situ hybridization showing Cre mRNA in the neural tube at E11.5. cc, central canal; di, diencephalon; dnt, dorsal neural tube; gm, gray matter; hb, hindbrain; mes, mesencephalon; vt, ventral telencephalon; wm, white matter.