Fig. 2. Electroporation-based gene transfer in the flat whole-mount culture of the embryonic rat medulla. (A) Schematic diagrams showing the procedure to make the flat whole-mount preparation of the hindbrain into which expression vectors are transferred. The DNA solution containing expression vectors and a dye is injected into the fourth ventricle. The head is cut, immersed in PBS and electroporated using forceps-shaped electrodes. The hindbrain is dissected out, cut at the dorsal midline and opened in the medium. The medulla is excised from it by cutting at the red lines and placed on the membrane in an open book configuration. Fixed preparations are sectioned along the blue line to make cryostat sections for immunostaining. (B-D) Transverse sections of 2 div explants immunostained with an anti-pan-cadherin Ab. An egfp cDNA is introduced into the explants. Lower (B,C) and higher (D) magnification. EGFP-positive migrating neurons express cadherin proteins. VM, ventral midline. (E-G) Transverse sections of 2 div explants expressing EGFP+CAD11-FLAG (E), EGFP+cN390-FLAG (F) and EGFP-NCAD(t)+DsRed (G). FLAG signal is detected with an anti-FLAG antibody. Most of the cells labeled with fluorescent reporter proteins express exogenous cadherin constructs. Scale bars: 200 µm in B and C; 50 µm in D-G.