Anti ape1

Liver mitochondria were isolated in a medium containing 220 mM mannitol, 70 mM sucrose, 20 mM Tris–HCl, 1 mM EDTA, and 5 mM EGTA, pH 7.4, at 4 °C according to [16 (link)]. In addition, 10 μg of mitochondrial proteins were used for Western immunoblotting analysis. Anti-TFAM (1:50,000), anti-VDAC (1:50,000, Abcam, Cambridge, UK), anti-OGG1 (1:2500, Abcam, Cambridge, UK), anti-APE1 (1:5000, Abcam, Cambridge, UK), anti-MFN2 (1:5000, Abnova, Taipei, Taiwan), anti-DRP1 (1:2500, Abnova, Taipei, Taiwan), anti-Cyt c (1:500, Pharmingen, San Diego, CA, USA), and anti-Lon Protease (1:10,000) were used as primary antibodies. The antibodies against TFAM and Lon were custom-made and kindly donated, respectively, by Dr. H. Hinagaki (Department of Chemistry, National Industrial Research Institute of Nagoya, Nagoya-shi, Aichi, Japan) and Dr. C. Suzuki (Department of Biochemistry and Molecular Biology, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, NJ, USA). The proteins were detected by chemiluminescence and immunoreactive bands were quantified using the Image Lab Software (BioRad Laboratories Inc., Hercules, CA, USA) and normalized against VDAC-expression.

Anti-hnRNP-U/SAF-A, anti-APE1, anti-Ku 70, anti-DNA-PKcs, anti-beta tubulin, anti-beta actin, and anti-Lig3 Abs were purchased from Abcam (Cambridge, MA). anti-APE1 was obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Anti-PARP1, anti-Ku 80, anti-XRCC1, anti-XRCC4, anti-GST, anti-His, anti-H3, and anti-NBS1 were purchased from Cell Signaling (Beverly, MA). Anti-FLAG Ab was purchased from Sigma (St. Louis, MO). The rabbit anti-NEIL1 and anti-pS59 SAF-A Abs were custom-generated [26 (link), 33 (link), 50 (link)].