Heart specimens were homogenized in lysis buffer (100 mmol/L NaCl, 0.1% Triton X-100, 50 mmol/L Tris; pH 8.3) loaded on 8% SDS-polyacrylamide gel, separated by gel electrophoresis, transferred onto a nitrocellulose membrane and probed with a relevant primary antibody.
Types and sources of the antibody used for the immunoblotting procedure are as follows: the antibody against α-actin (sc-32251), LC3-I (sc-134226), and LC3-II (sc-271625) were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Secondary antibodies were conjugated to horseradish peroxidase and the ensuing immunoreactive bands were developed using a Western Lightning Chemiluminescence (WLC) kit (PerkinElmer Life Science, MA, USA). The nitrocellulose membrane used for western blotting was obtained from Bio-Rad Laboratories (Hercules, CA, USA). Immunoblots were digitalized and quantified using NIH ImageJ software (National Institute of Health, Bethesda, MD, USA).
Types and sources of the antibody used for the immunoblotting procedure are as follows: the antibody against α-actin (sc-32251), LC3-I (sc-134226), and LC3-II (sc-271625) were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Secondary antibodies were conjugated to horseradish peroxidase and the ensuing immunoreactive bands were developed using a Western Lightning Chemiluminescence (WLC) kit (PerkinElmer Life Science, MA, USA). The nitrocellulose membrane used for western blotting was obtained from Bio-Rad Laboratories (Hercules, CA, USA). Immunoblots were digitalized and quantified using NIH ImageJ software (National Institute of Health, Bethesda, MD, USA).