*2.14. Western Blot Analysis*

Western blot analysis was executed on brain and sciatic nerve harvested 21 h after reserpine injection. Cytosolic proteins were extracts as described previously [50]. Membranes were probed with specific Abs: with anti-VEGF (1:500; Santa Cruz Biotechnology, Heidelberg, Germany), or with anti-NGF (1:500; Santa Cruz Biotechnology) in 1× PBS (Phosphate buffered saline), 5% *w*/*v* nonfat dried milk, 0.1% Tween-20 at 4 ◦C, overnight. To control the equal amounts of proteins, blots also were probed with antibody against b-actin protein (cytosolic fraction 1:500; Santa Cruz Biotechnology). Signals were examined with enhanced chemiluminescence (ECL) detection system reagen<sup>t</sup> according to the manufacturer's instructions (Thermo Fisher, Waltham, MA, USA). The relative expression of the protein bands was quantified by densitometry with BIORAD ChemiDocTM XRS + software and standardized to b-actin and lamin A/C levels. The blot was stripped with glycine 2% and re-incubated several times to optimize detection of proteins and to visualize other proteins minimizing the number of gels and transfers.
