Fig. 3. Identification of cells expressing XOtx2 and XOtx5b. XOtx2 is expressed in bipolar cells. (A) Sagittal section of a stage 41 GFP-lipofected embryo hybridised with a DIG-labelled XOtx2 anti-sense RNA probe. The arrow points to a cell in the INL stained with XOtx2. (C) Under fluorescence, the same section as in A, stained with a GFP antibody, reveals that the XOtx2-positive cell is a bipolar cell. (B) Superimposition of the image in A onto that in C. The arrow points to the same cell in all three panels. XOtx5b is also expressed in bipolar cells. (D-F) The same result is found in a retina stained for XOtx5b. (D) Sagittal section of a stage 41 GFP-lipofected embryo hybridised with a DIG-labelled XOtx5b anti-sense RNA probe. The arrow points to the same cell in F, which is a bipolar cell. (E) The overlay shows that the same cell is labelled for both XOtx5b and GFP. Cells in the INL that express XOtx5b also express XOtx2. (G-I) Stage 41 GFP-lipofected embryo section labelled with DIG-labelled XOtx5b (G: blue) and fluorescein-labelled XOtx2 (I: red) anti-sense RNA probes. The arrows point to the same cells in all three panels, which are positive for both the XOtx5b and XOtx2 probes, as shown in the overlay (H). (J-O) XOtx5b is also expressed by both rod and cone photoreceptors. In situ hybridisation of XOtx5b (blue) onto stage 41 sagittal sections co-stained for calbindin antibody (J-L) or rod opsin antibody (M-O). (J) Cells expressing XOtx5b also produce calbindin protein (L). The overlay (K) shows that these are the same cells, demonstrating that they are cone photoreceptor cells. (M) Similarly, XOtx5b is found in cells that generate rhodopsin (O). Thus, rod photoreceptors also express XOtx5b. (K,N) Merging panels J with L (K) and M with O (N) confirms this. Cryostat sections cut to 10 µm.