Goat anti rabbit secondary antibody conjugated with hrp

Cells were lysed in RIPA buffer (Pierce, Rockford, IL) supplemented with protease inhibitor cocktail and phosphatase inhibitors (Sigma). Total proteins were separated in 10% SDS-PAGE pre-cast gels (Bio-Rad) and transferred onto Nitrocellulose membranes (Millipore Corp., Bedford, MA). Membranes were first probed with Arginase 1 (Arg1) (1:500, BD Biosciences, San Jose, CA), Ym1 (1:500, StemCell Technologies, Vancouver, British Columbia, Canada), STAT6 (1:1000, Cell Signaling, Danvers, MA), phosphor-STAT6 (1:1000, Cell Signaling), C/EBPβ (1:1000, Santa Cruz Biotechnology, Inc. Santa Cruz, CA), or β-actin (1:1000, Sigma) antibodies, followed by goat anti-rabbit secondary antibody conjugated with HRP (1:5000, Millipore Corporation, Billerica, MA) or goat anti-mouse secondary antibody conjugated with HRP (1:3000, Millipore Corporation). The Protein detection was performed using Pierce ECL Western Blotting Substrate (Pierce).

Cells were lysed in RIPA buffer (Pierce, Rockford, IL) supplemented with protease inhibitor cocktail and phosphatase inhibitors (Sigma). Total cellular extracts (30 µg) were separated in 10% SDS-PAGE pre-cast gels (Bio-rad) and transferred onto Nitrocellulose membranes (Millipore Corp., Bedford, MA). Membranes were first probed with HIF-1α (1 ug/ml, Sigma) and β-actin (1:1000, Sigma) antibodies, followed by goat anti-rabbit secondary antibody conjugated with HRP (1:5000, Millipore). The Protein detection was performed using Pierce ECL Western Blotting Substrate (Pierce).