Test for bleed-through between channels using bench-made tyramides. (A-I) Single-color FISH experiments were performed with digoxigenin-labeled dlx2a (A-C), dinitrophenyl-labeled shha (D-F) and fluorescein-labeled dbx1a (G-I) RNA probes and visualized by FAM-, TAMRA- and DyLight633- tyramide, respectively. Lateral views of 1-dpf forebrains are shown with anterior to the left. Only the appropriate detection channel showed a bright signal and no significant bleed-through between channels was observed. Images were recorded with identical settings to those of Figure 7 on a LSM510 microscope (Carl Zeiss) and false colored in ImageJ. Scale bar = 50 μm.