Ab86497

Western blot assay was performed as previously described^{20 (link)}. Tissue or cell extracts were prepared using a dounce homogenizer in cold RIPA buffer supplemented with cocktail. The homogenates were centrifuged at 12,000 rpm for 15 min, and the protein concentrations were determined using a protein assay from Thermo. The protein lysates were separated by SDS-PAGE and electrotransferred onto a nitrocellulose membrane. The membrane was scanned using the Image Lab statistical software (Bio-Rad). Antibodies against PAR (Trevigen, 4335-MC-100), PARP1 (Trevigen, 4338-MC-50), p-γH2AX (Ser139) (Trevigen, 4418-APC-020), Gabarapl1 (Abcam, ab86497), ATG12 (Abcam, ab109491), P62 (Cell signaling Technology, 39749), LC3 (Cell Signaling Technology CST, 3868), p-P53 (Ser15) (Cell signaling technology, 9284), p-ATM (Ser1981) (Thermo Fisher MA1-2020),FoxO3a (Cell signaling technology, 12829), pFoxO3a (Cell signaling technology, 9466), β-actin (Cell signaling technology, 3700), PGC1α (Abcam, ab54481), TFAM (Abcam, ab131607), NRF1 (Abcam, ab175932), Lamin B1 (Cell signaling technology,13435), α-tubulin (Cell signaling technology,3873) were used as primary antibodies.

Tissues were derived from clinical specimens. The tissue sections were deparaffinized, hydrated, repaired and blocked with citric acid antigen. Subsequently, the tissue sections were probed with an ULK2 antibody (1:50, Omnimabs, OM294638) and GABARAPL1 antibody (1:200, Abcam, ab86497) at 4°C overnight. The sections were washed with PBS for 5 times, and then a secondary antibody was added and incubated at room temperature for 10 minutes. DAB and hematoxylin were added for visualization.