Mab1501

The following mouse antibodies were used for immunoblotting (WB) in this study: ATP5A (Abcam, ab14748, 1:10,000), actin (Millipore, MAB1501, 1:5,000), Ubiquitin (clone P4D1, Cell Signalling Technology, 3936, 1:1,000), Ubiquitin (clone FK2, MBL, D058-3, 1:1,000). The following rabbit antibodies were used in this study: pS65-Ub (Cell Signalling Technologies, 62802S, 1:1,000), Marf ([16 (link)], 1:1,000), GABARAP/Atg8a (Abcam ab109364, 1:1,000), ref(2)P/p62 (Abcam ab178440, 1:1,000). The following secondary antibodies were used: sheep anti-mouse (HRP-conjugated, GE Healthcare, NXA931V, 1:10,000), donkey anti-rabbit (HRP-conjugated, GE Healthcare, NA934V, 1:10,000).

Rabbit anti-Drosophila Parkin polyclonal antibody was raised against recombinant MBP-tagged Drosophila Parkin (275–482 aa) produced in the E. coli strain Rosetta 2 (Novagen). The antibodies used in the western blot analysis were as follows: anti-Parkin (1∶5,000 dilution), anti-Mfn (1∶2,000 dilution; a kind gift of Dr A. Whitworth), anti-Miro (1∶2,000 dilution; a kind gift of Dr E. Zinsmaier), anti-Drp1 (1∶2,000 dilution; a kind gift of Dr L. Pallanck), anti-ATP5A (1∶10,000 dilution; Abcam, 15H4C4), anti-NDUFS3 (1∶10,000 dilution; Abcam, 17D95), anti-Actin (1∶10,000 dilution; Millipore, MAb1501) and anti-Hsp60 (1∶1,000 dilution; Cell Signaling, D307). The antibody used in immunocytochemistry was anti-TH (1∶1,000 dilution; ref. [11] (link)). Complementary DNAs for human and Drosophila Parkin and PINK1 were described in previous studies [11] (link), [14] (link), [45] (link). Parkin phospho-mutants were generated by PCR-based mutagenesis followed by sequence confirmation of the entire gene.