Rabbit anti c ebpβ

Macrophages were lysed with Cell Lytic Buffer (Sigma) and the cell lysate was collected. For p47 ^phox estimation the cell lysate was centrifuged at 600g for 10 min at 4°C to remove unbroken cells and nuclei. The supernatant was then ultracentrifuged at 100,000g for 1hr at 4°C to isolate the membrane fraction. Protein was estimated by Bradford’s reagent and equal amounts of protein sample from each experimental condition was subjected to immunoblot analysis using the following primary antibodies: goat poly anti-MCPIP (1:500), rabbit anti-IRE-1 (1:500), rabbit anti-LC3 II (1:500), rabbit anti-C/EBPβ (1:100), rabbit anti-PPARγ (1:100), rabbit anti-FIZZ1 (1:500), monoclonal anti–p47^phox (1:200), and rabbit anti-Fas (1:1000; Santa Cruz Biotechnology); mouse polyclonal anti-GAPDH (1:1000); rabbit anti-GRP78 (1:500); rabbit anti-BECLin 1(1:1000), goat anti-Arg1 (1:2000; Cell signaling). The immune complexes were detected autoradiographically using appropriate peroxidase-labeled secondary antibodies (Santa Cruz Biotechnology) and enhanced chemiluminescence detection reagent ECL (GE Healthcare). Anti-β-actin and anti-GAPDH antibodies served as loading controls. Specific bands were quantified by densitometry using analytic software (Image J) and expressed as a ratio over loading controls.