Ab55474

The following antibodies were used: Rabbit anti-Eva1a/Tmem166 (GeneTex, Irvine, CA, USA, GTX32925), Rabbit anti-Lc3b (Sigma Aldrich, St. Louis, MO, USA, L7543), Mouse anti-Gapdh (Sungene, Tianjin, China, KM9002). Antibodies against Atg5 (12994), caspase3 (39665), cleaved caspase3 (39664), Parp (9532), and Ubiquitin (3936) were purchased from Cell Signaling Technology (Danvers, MA, USA). Antibodies against Ulk1 (ab128859), Atg16l1 (ab187671), Nbr1 (ab55474), Caspase 6 (ab185645), Bnip3 (ab109362), Drp1 (ab184247), Mfn2 (ab124773), Parkin (ab179812), and Tomm20 (ab186734) were purchased from Abcam (Cambridge, UK). Antibodies against p62/SQSTM1 (PM045) and Beclin1 (PD017) were purchased from MBL International (Woburn, MA, USA). Secondary antibodies included DyLight 800/DyLight 680-conjugated IgG against mouse (Rockland, Philadelphia, PA, USA, 610-145-002/610-144-002) or rabbit (Rockland, 611-145-002/611-144-002).

Tissue samples were homogenised with the Precellys 24 tissue homogeniser in Laemmli buffer and samples ran on 12.5 or 15% gels. Protein was transferred to PVDF membranes (Immobilon, Millipore), which was subsequently blocked for 1 h at room temperature (5% milk solution in TBS-Tween 0.1%) before incubating with primary antibody at 4 °C overnight. An appropriate HRP-conjugated secondary antibody was incubated at room temperature for 1 h. Western blots were visualised with chemiluminescence reagents (Sigma, RPN2106). Antibodies were used at the following concentrations: Anti-ATG5 (Abcam, ab108327; 1:1000), anti-LC3 (Abcam, ab192890; 1:1000), anti-ACTIN (Santa Cruz Biotechnology, I-19; 1:5000 [no longer commercially available]), anti-P53 (Cell Signalling Technologies, Clone 1C12; 1:1000), anti-P21 (Santa Cruz, SC-6246; 1:1000), anti-Histone H3 (Abcam, ab1791; 1:5000), anti-P16 (Santa Cruz, SC-1207; 1:1000), anti-HMGA1 (Abcam, ab129153; 1:1000), anti-NBR1 (Abcam, ab55474; 1:1000).