Hif 1α antibody nb100 134

Chromatin immunoprecipitation (ChIP) was performed according to previous reports9 (link)42 (link). Cells were cross-linked for 10 min using 1% paraformaldehyde and then subjected to fragmentation using Sonifier (Branson, Dansbury, USA). Samples were immunoprecipitated with the HIF-1α antibody (NB100-134, Novus Biologicals). ChIP samples were also quantified with RT-PCR using specific primer pairs. All mouse primer sequences for PCR are listed in Supplementary Table 2.

Mouse lungs were lysed in ice‐cold lysis buffer, mixed with a commercial sample buffer (Invitrogen), and heated at 95°C for 5 min. Samples were then electrophoresed on SDS–polyacrylamide gels, after which the gels were transferred to PVDF membranes. Blots were incubated overnight at 4°C with various primary antibodies. We used the HIF‐1α antibody (NB100‐134) from Novus Biologicals, LLC. The dilution is 1:500, and the molecular weight is 93 kDa. Primary antibody was followed by appropriate secondary antibodies. After washing, the signal was detected using a Super Signal chemiluminescent substrate Western Blotting reagent (Pierce Biotechnology) with the chemiluminescent‐sensitive film.¹⁴