Fig. 4

Brain parenchyma expression and diffusibility. ScFv-MC1 is expressed and released in the extracellular milieu: anti Myc-tag staining performed 4 months after the one time intracranial injection with AAV5-eGFP (a-g) (n = 15), AAV5-CAG-scFvMC1 (h-n) (n = 15) and AAV5-GFAP-scFvMC1 (o-u) (n = 15). Representative images from the whole hippocampus (Olympus BH-2 bright field microscope, bar: 500 μm), CA1, subiculum, dentate gyrus (DG), entorhinal cortex (EC), frontal cortex and HB (bar: 100 μm). The scFv is visualized as a brown signal and shows extensive expression in hippocampi from treated mice, with spreading in areas distant from the site of injection. (v) Immunoblot analysis of lysates from the hippocampus, cortex and hindbrain: AAV5-eGFP injected (1), AAV5-CAG-scFMC1 injected (2), AAV5-GFAP-scFvMC1 injected (3). Three representative lysates from each group of treatment were pulled together and loaded on SDS-PAGE (40μg proteins of hippocampus; 300μg proteins of cortex; 150μg proteins of HB plus HRP substrate enhancer when developing). Anti Myc-tag was used to track the scFv in the brain parenchyma; actin was used as a housekeeper and N-cadherin as a marker of neuronal integrity. Of note, immunoblots images shown represent optimal exposure samples to visualize differences within the brain areas