HCT116 cells were washed with ice-cold PBS and lysed on ice in lysis buffer (50 mM HEPES, 150 mM NaCl, 1% Triton X-100) supplemented with protease inhibitor cocktail (Roche). Cell debris was cleared by centrifugation. Whole cell extracts were resolved by 10% SDS-PAGE, transferred onto nitrocellulose membranes, and blotted with antibodies against Sam68 (07-415, Millipore), p53 (HAF1355, R&D), MDM2 (Ab-5, Millipore), p21 (C-19, Santa Cruz), Bax (P-19, Santa Cruz), Puma (4976, Cell Signaling), PARP (F-2, Santa Cruz) and β-actin (A3853, Sigma Aldrich). For immunoprecipitation, anti-Sam68 (07-415, Millipore), anti-p53 (1C12, Cell Signaling), anti-Myc (05-724, Sigma), anti-GFP (Anti-GFP, Roche), mouse or rabbit IgG (Santa Cruz) antibodies were incubated for 2 h with whole cell lysates, and 1 h with 50 μl of 50% slurry protein A/G beads (Sigma) at 4°C. Beads were washed three times with lysis buffer, and bound proteins were recovered by boiling in Laemmli sample buffer.
Bax p 19
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Bax (P-19) is a primary antibody produced by Santa Cruz Biotechnology. It is an immunoglobulin G (IgG) that recognizes the Bax protein. Bax is a pro-apoptotic member of the Bcl-2 family of proteins and plays a key role in the regulation of apoptosis, or programmed cell death.
Automatically generated - may contain errors
Lab products found in correlation
Anti myc, by Merck Group (1 mentions)
Anti p53, by Cell Signaling Technology (1 mentions)
β actin, by Merck Group (1 mentions)
Protease inhibitor cocktail, by Roche (1 mentions)
Parp f 2, by Santa Cruz Biotechnology (1 mentions)
Mouse or rabbit igg, by Santa Cruz Biotechnology (1 mentions)
Ecl plus reagent, by Cytiva (1 mentions)
Sirt1, by Santa Cruz Biotechnology (1 mentions)
P atm ser1981, by Cell Signaling Technology (1 mentions)
Polyacrylamide gel, by Bio-Rad (1 mentions)
Signalfire elite ecl reagent, by Cell Signaling Technology (1 mentions)
β actin, by Santa Cruz Biotechnology (1 mentions)
3 protocols using bax p 19
1
Immunoprecipitation and Western Blot Analysis
2
Western Blot Analysis of Cellular Proteins
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Ten to 50 micrograms of protein were electrophoresed on 8–12% v/v polyacrylamide SDS-PAGE gels. Proteins were electrotransfered onto PVDF membranes. The membranes were subsequently blocked and incubated at room temperature with antibodies (1∶500 #sc-13067 Santa Cruz PGC-1 (H-300), 1∶2000 #ab110304 Abcam Sirt-1, 1∶2000 #sc-99 Santa Cruz p53 (Pab 240), 1∶2000 #06–758 Upstate ac.p53 (Lys 373,382), 1∶1000 #sc-33771 Santa Cruz NRF-1 (H-300), 1∶500 #sc-30963 Santa Cruz mtTFA (E-16)/TFAM/, 1∶1000 #sc-492 Santa Cruz Blc-2 (N-19), 1∶1000 #sc-525 Santa Cruz Bax (P-19), 1∶1000 #sc-27907 Santa Cruz Odf-1 (G-17), 1∶2000 #sc-377305 Santa Cruz LDH-C (H-65), 1: 5000#sc1616 Santa Cruz Actin (I-19). After incubation with primary antibodies, membranes were washed in TBS-Tween-20 and incubated with HRP-conjugated secondary antibodies. After incubation with the secondary antibody, membranes were repeatedly washed. Membranes were incubated with an ECL Plus reagent (RPN 2132, Amersham) and protein bands were visualized on X-ray films. The bands were quantified by ImageJ software (http://imagej.nih.gov/ij/ ) and normalized to beta-actin, which served as an internal control.
3
Western Blot Analysis of DNA Damage Signaling
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RIPA buffer was used for total protein extraction. 20 μg of protein was denatured under reducing conditions and separated on 10% polyacrylamide gels (Bio-Rad Laboratories, Hercules, CA) and transferred to nitrocellulose by voltage gradient transfer. The resulting blots were blocked with 5% (w/v) non-fat dry milk in PBS + 0.1% (v/v) Tween-20. Specific proteins were detected with appropriate antibodies using SignalFireTM Elite ECL Reagent (Cell Signaling Technology). Immunoblotting antibodies were p53 (OP03, Calbiochem, Massachusetts), p-p53 (9284, Cell Signaling Technology, Massachusetts), ATM (2873, Cell Signaling Technology), and p-ATM Ser1981 (13050, Cell Signaling Technology), p21 (C-19, Santa Cruz Biotechnology, California), Bax (P-19, Santa Cruz Biotechnology), Bak (G-23, Santa Cruz Biotechnology), Mdm2 (OP115, Calbiochem), β-actin (I-19, Santa Cruz Biotechnology).
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