Figure 2

Rapid onset of gene expression using scAAV in the murine heart. CD-1 mice (Charles Rivers) were injected 4 days after birth with 1.85 × 10¹¹ vector genomes (vg) of scAAV using previously established methods [14]. Animals were returned to their dams for 4 days (A) or 6 days (B) at which time their hearts were harvested, cut into thirds representing the apical region (apex), the mid-region (mid), or base of the heart (base). The tissues were fixed in 4% paraformaldehyde for 12 hours, rinsed in PBS and allowed to equilibrate in 20% sucrose for 12–24 hours. The next day, the hearts were frozen in cryomolds containing OCT compound (Tissue-Tek) to prepare for cryostat sectioning into 5 μm sections. Slides were mounted using Vectashield mounting media containing DAPI (Vector Labs) to counter stain the nuclei. A Leica TCS SP2 AOBS Spectral Confocal Microscope with a 10× objective was used to obtain fluorescent images. Staining was documented using the Leica Confocal Software (LCS) Version 2.61. Sections with GFP expression can be seen in the top panels while merged images containing GFP and DAPI signals can be seen in the lower panels. The bar represents 150 μm in the 6 day apex and 300 μm in all other panels.