Green fluorescent protein antibody

H9c2 cells were transfected with an NR4A2 overexpression plasmid or with a negative control plasmid, followed by ischemia treatment. At 60 h after transfection, total protein was collected from cells by using IP lysis buffer (Beyotime, Shanghai, China). After centrifugation at 4 °C, the supernatant was isolated and incubated with protein A/G agarose beads (Beyotime, Shanghai, China) and green fluorescent protein (GFP) antibody (Santa Cruz Biotechnology, Santa Cruz, USA) at 4 °C overnight. The beads were washed 3 times with IP lysis buffer and then eluted with 4× SDS loading buffer. GFP and p53 were detected by western blot assay with their protein-specific antibodies (Cell Signaling Technology, Danfoss, USA).

UNC0638 (#U4885) and BIX-01294 (#B9311) were both purchased from Sigma (St. Louis, MO, USA). The iron chelator DFO was from Novarits (Switzerland). FAC was from JK Chemical (#F5879, Shanghai, China). Kits of the membrane and cytosol protein extraction kit (#P0033), the ROS assay (#S0033) and the cell lysis buffer radio-immunoprecipitation assay (#P0013B) were purchased from Beyotime (Nantong, China). Monoclonal antibodies specific for G9a (1:1,000; #3306), YY1 (1:1,000; #2185), HDAC1 (1:1,000; #34589), HDAC2 (1:1,000; #57156), Histone H3 (1:2,000; #4499), K9 dimethylated Histone H3 (1:2,000; #4658), β-actin (1:5,000; #8457) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (1:5,000; #5174) were purchased from Cell Signaling Inc. (Danvers, MA, USA). The other antibodies were as follows: HEPH antibody (1:500; #sc-365365, Sigma), green fluorescent protein (GFP) antibody (1:1,000; #sc-69779, Santa Cruz Biotechnology), Alexa Fluor 488 goat anti-mouse IgG (H + L) antibody (1:200; #A-11001, Life Technologies, OR, USA) and Alexa Fluor 594 goat anti-rabbit IgG (H + L) antibody (1:200; # A-11012, Life Technologies).