Western blot was performed following a standard protocol. Antibodies were obtained as follow; p53, p21, Chk1, p62, PARP-1, β-actin (Santa Cruz Biotechnology), phospho-p53 (Ser15), phospho-Chk2 (Thr68), phospho-Erk1/2, Erk1/2, phospho-JNK, JNK1/2, phospho-p38, p38, Caspase 3, H2AX (Cell Signaling Technology), phospho-ATM (Ser1981), Chk2, γ-H2AX (Ser139) (Millipore), ATM (Epitomics, Burlingame, CA, USA), and LC3 (Novus Biologicals, Littleton, CO, USA). Chemiluminescence was detected using enhanced chemiluminescence detection reagents.
Phospho p53
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Phospho-p53 is a laboratory reagent used to detect the phosphorylated form of the p53 protein. P53 is a critical tumor suppressor protein that plays a key role in cellular stress response pathways. Phosphorylation of p53 is an important post-translational modification that regulates its activity and stability.
Automatically generated - may contain errors
Lab products found in correlation
β actin, by Santa Cruz Biotechnology (3 mentions)
Phospho p38, by Cell Signaling Technology (2 mentions)
Bcl2 associated x (bax), by Santa Cruz Biotechnology (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Jnk1 2, by Cell Signaling Technology (1 mentions)
Caspase 3, by Cell Signaling Technology (1 mentions)
Phospho jnk, by Cell Signaling Technology (1 mentions)
Phospho chk2 thr68, by Cell Signaling Technology (1 mentions)
Phospho erk1 2, by Cell Signaling Technology (1 mentions)
Erk1 2, by Cell Signaling Technology (1 mentions)
Parp 1, by Santa Cruz Biotechnology (1 mentions)
Antibody against β actin, by Merck Group (1 mentions)
Antibodies against bcl 2, by Santa Cruz Biotechnology (1 mentions)
Ciap2, by Santa Cruz Biotechnology (1 mentions)
Peroxidase labeled donkey anti rabbit and sheep anti mouse immunoglobulin, by Cytiva (1 mentions)
Phospho erk, by Santa Cruz Biotechnology (1 mentions)
Enhanced chemiluminescence ecl kit, by Cytiva (1 mentions)
Phospho akt, by Santa Cruz Biotechnology (1 mentions)
X linked iap xiap, by Santa Cruz Biotechnology (1 mentions)
Antibody against poly adp ribose polymerase parp, by BD (1 mentions)
6 protocols using phospho p53
1
Molecular Signaling Profiling by Western Blot
2
AGS Cell Culture and Protein Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human AGS cells from the American type culture collection (Rockville, MD, USA) were cultured in RPMI 1640 medium (Invitrogen Corp, Carlsbad, CA, USA) supplemented with 10% (v/v) fetal bovine serum (FBS) (GIBCO BRL, Grand Island, NY, USA), 1 mM L-glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin at 37°C in a humidified atmosphere of 95% air and 5% CO2. Antibodies against Bcl-2, Bid, Bcl-xL, phospho p53, p53, BAX, c-IAP-2, X-linked IAP (XIAP), Akt, phospho Akt, ERK, phospho ERK, and procaspase 3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibody against poly (ADP-ribose) polymerase (PARP) was purchased from PharMingen (San Diego, CA, USA). Antibody against β-actin was from Sigma (Beverly, MA, USA). Peroxidase-labeled donkey anti-rabbit and sheep anti- mouse immunoglobulin, and an enhanced chemiluminescence (ECL) kit were purchased from Amersham (Arlington Heights, IL). All other chemicals not specifically cited here were purchased from Sigma Chemical Co. (St. Louis, MO, USA).
3
Western Blotting of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting was carried out as previously described (62 (link)). Primary antibodies to β-actin (Santa Cruz), p62 (CST), SIRT1 (CST), Cas9-hemagglutinin (Cas9-HA; Santa Cruz), PML (CST), XPO1 (CST), p53 (CST), and phospho-p53 (CST) were used.
4
Western Blot Analysis of Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein was isolated using radioimmunoprecipitation assay buffer consisting of 1% (v/v) NP40, 0.1% (w/v) sodium dodecyl sulfate (SDS), 100 μg/mL phenylmethylsulfonyl fluoride, 0.5% (w/v) sodium deoxycholate, protease inhibitors, and phosphatase inhibitors in PBS. Protein concentrations were determined using a BCA Protein Assay Kit (Thermo Fisher Scientific, USA) Proteins were then separated on 10% SDS-polyacrylamide gels and transferred to polyvinylidene fluoride (PVDF) membranes. Membranes were incubated in 5% skim milk with tris-buffered saline Tween (TBST) at room temperature for 1 h and incubated overnight with primary antibodies. The following proteins were probed with the primary antibodies: p16, p21, p53, phospho-p53, caveolin-1, Hsp60, PRDX1-3, and β-actin (Santa Cruz Biotechnology, Dallas, TX, USA). Membranes were washed with TBST and incubated with horseradish peroxidase–conjugated antibody for 1 h. Protein signals were detected using an ECL Detection Kit (BIOFACT, Daejeon, Korea). The signal intensities for each protein band were quantified using the densitometry program, Image J. v1.48. Statistical analyses were performed using Student t test. A p values less than 0.05 were considered statistically significant.
5
Western Blotting Antibodies and Detection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting was carried out as previously described (39 (link)). Primary antibodies to LANA (Abcam, Cambridge, MA), cleaved PARP1 (Cell Signaling Technology [CST]), cleaved caspase 3 (CST), K8 (Santa Cruz), p53 (CST), and phospho-p53 (CST) were used. A monoclonal antibody to ORF65 was previously described (40 (link)).
6
Western Blot Analysis of Heart Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The heart tissue was homogenated in extraction buffer (20 mmol/L Tris HCl, 1 mmol/L Na 3 VO 4 , 5 mmol/L NaF). The heart protein was collected and subjected to 10% or 15% SDS-polyacrylamide gel electrophoresis then transferred to polyvinylidene difluoride membranes, which were blocked for 2 h with 5% non-fat milk in Tris-buffered saline (pH 7.4) containing 0.1% Triton X-100. The membranes were probed overnight at 4 C with the appropriate primary antibody as follows: total-p38, phospho-p38, and phospho-HSP27 (Cell Signaling Technology, Danvers, MA), total-p53, phospho-p53, total-CREB, phospho-CREB, Bax, Bcl2, Cytochrome c, caspase-3 (Santa Cruz Biotechnology, Inc, Santa Cruz, CA). After washing and exposure to horseradish peroxidaseconjugated secondary antibody for 1 h at room temperature, antibody-antigen complexes were visualized by enhanced chemiluminescence assay. Bands corresponding to the protein of interest appeared as dark regions on the developed film. The film images of the Western blots were scanned and were analyzed using Image J (NIH image, National Institutes of Health, Bethesda, MD) analysis software (Tanno et al., 2003) . For quantitation of the proteins of interest, phosphorylated proteins were normalized to total protein expression.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!