Anti par 10h

Hippocampi were dissected on ice and homogenized in lysis buffer (NaCl 150 mM, Tris 10 mM, 1% Triton X-100, 0.5% nonidet p-40) with protease inhibitor cocktail (EMD Milipore). Fifty μg of proteins were gel separated, transferred to immobilon PVDF-FL membrane. Membranes were incubated in Odyssey blocking buffer (Licor Biosciences) for an hour at room temperature. This was followed by incubation of membranes with primary antibody overnight at 4 °C (anti-PAR (10H), 1:1000 Enzo; anti-Nampt (PBEF), 1:1000 abcam; anti-Nmnat2, 1:1000 abcam, anti-β-actin at 1:10,000; Cell Signaling). After washing the membranes in PBS with 0.1% tween-20, they were incubated with the appropriate infrared fluorophore conjugated secondary antibody (Licor) for 30 min at room temperature. The membranes were scanned and the bands were quantified with Odyssey infrared image system (Licor). The β-actin signal was used for normalization of the data detected from bands of the proteins of interest.