Anti hne antibody

All reagents were obtained from Sigma unless otherwise specified. LysoSensor and SYBR Green PCR Master Mix were purchased from Life Technologies Inc. (Grand Island, NY). HNE and bafilomycin A1 (Baf) was from Cayman Chemical Company (Ann Arbor, MI). Rapamycin and wortmannin were purchased from Sigma Chemical Company (St. Louis, MO). DQ-BSA was from Thermo Fisher Scientific. Anti-HNE antibody was from Alpha Diagnostic (San Antonio, TX; catalog #HNE13-M). Cathepsin B and cathepsin D activity assay kits, anti-SQSTM1/p62, anti-LAMP2a (catalog #ab18528) and anti- cathepsin B (catalog #ab58802) antibodies were from Abcam (Cambridge, MA). Anti-cleaved caspase-3 (catalog #9661)) and anti-cathepsin D (catalog # 2284) antibodies were from Cell Signaling Technology (Danvers, MA). Anti-LC3 (catalog #NB100-2220), and Anti-Hsc70 (catalog #NB120-2788) antibodies were from Novus Biologicals (Littleton, CO). Anti mono- and polyubiquitinylated conjugated antibody was from Enzo Life Sciences (Farmingdale, NY; catalog #PW8810). Anti K48-linked and K63-linked polyubiquitin antibodies (catalog number 05-1307 and 05-1308, respectively) were from EMD Millipore (Billerica, MA). MTS assay kit was from Promega Life Sciences (Madison, WI). LDH assay kit was from Roche Diagnostics (Indianapolis, IN).

Paraffin-embedded LV sections (5 μm) were prepared for assessment of oxidative modification of cell injury. The presence of 4-hydroxynonenal (4-HNE), representing lipid peroxide, was assessed by diaminobenzidine (DAB) staining with rabbit polyclonal anti-HNE antibody (Alpha Diagnostic International, San Antonio, TX) at 1:500 dilution. Oxidative stress-induced DNA damage was measured by the presence of 8-hydroxy-2′-deoxyguanosine (8-oxo-DG) with mouse monoclonal anti-8-oxo-DG at 1:200 dilution (Rockland Immunochemicals Inc., Limerick, PA), as described previously by others.^{40 (link),41 (link)}