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Mouse monoclonal anti p53 antibody

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Mouse monoclonal anti-p53 antibody is a laboratory reagent that recognizes the p53 protein. The antibody is derived from a mouse hybridoma cell line and can be used to detect and study the p53 protein in various experimental applications.

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Lab products found in correlation

Bca protein assay, by Thermo Fisher Scientific (3 mentions) Mouse monoclonal anti dnmt3b antibody, by Cell Signaling Technology (2 mentions) Mouse monoclonal anti uxt antibody, by Cell Signaling Technology (2 mentions) Anti β actin mouse monoclonal antibody, by Cell Signaling Technology (2 mentions) Protease inhibitor cocktail, by Roche (2 mentions) Fetal bovine serum (fbs), by Sartorius (1 mentions) Rabbit anti p21 monoclonal antibody, by Cell Signaling Technology (1 mentions) Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Rabbit anti β actin polyclonal antibody, by Santa Cruz Biotechnology (1 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions) Dulbecco s modified eagle s medium dmem, by Merck Group (1 mentions) Hepatocyte medium, by ScienCell (1 mentions) Huh7 cells, by National Collection of Authenticated Cell Cultures (1 mentions) Rabbit polyclonal anti bax antibody, by Santa Cruz Biotechnology (1 mentions) Ecl kit, by Cytiva (1 mentions) Gapdh, by Cell Signaling Technology (1 mentions) Isoprenaline, by Bio-Techne (1 mentions) Ici 118 551, by Bio-Techne (1 mentions) Leukemia inhibitory factor, by Merck Group (1 mentions)

8 protocols using mouse monoclonal anti p53 antibody

1

Immunoblot Analysis of p53 Pathway

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Roswell Park Memorial Institute (RPMI) 1640 medium was purchased from Gibco/Thermo Fisher Scientific, Inc. Fetal bovine serum was obtained from Biological Industries. Mouse anti-p53 monoclonal antibody (catalog no. 2524S), rabbit anti-phospho-p53 polyclonal antibody (catalog no. 2521S), and rabbit anti-p21 monoclonal antibody (catalog no. 2947S) were purchased from Cell Signaling Technology, Inc. Rabbit anti-DLGAP5 polyclonal antibody (catalog no. PA5-82197) was purchased from Invitrogen Company. Mouse anti-actin monoclonal antibody (catalog no. sc-47778) was purchased from Santa Company. Every assay was done in triplicate.
Ke M.J., Ji L.D, & Li Y.X. (2020). Bioinformatics analysis combined with experiments to explore potential prognostic factors for pancreatic cancer. Cancer Cell International, 20, 382.
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2

Cultivation and Characterization of Hepatic Cell Lines

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Huh-7 cells were purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China) and maintained in Dulbecco’s modified Eagle’s medium (DMEM, Sigma) supplemented with 10% fetal bovine serum (Gibco, United States). HepAD38 and HepG2-NTCP cells were obtained from Xiamen University, HepAD38 cells were maintained in DMEM supplemented with 10% FBS and 400 μg/ml G418, and HepG2-NTCP cells were maintained in DMEM supplemented with 10% FBS, 2 μg/ml puromycin and 2 μg/ml Dox. PHHs were purchased from ScienCell and were maintained in hepatocyte medium (ScienCell, Carlsbad, CA, United States). All cells were incubated in a humidified atmosphere at 37°C and 5% CO2. Transfection was performed using Lipofectamine™ 3,000 (Invitrogen, United States) according to the manufacturer’s protocol. Rabbit anti-β-actin polyclonal antibody (100 μg/ml; sc-1, 616-R) and mouse anti-SIRT2 monoclonal antibody (200 μg/ml; sc-28, 298) were purchased from Santa Cruz Biotechnology (Dallas, TX, United States), and mouse anti-p53 monoclonal antibody (72 μg/ml; #48818) was obtained from Cell Signaling Technology (United States). The Human Sirtuin 2 (SIRT2) ELISA Kit was obtained from Abbexa Ltd. (abx383209; Cambridge, United Kingdom).
Wu D.Q., Ding Q.Y., Tao N.N., Tan M., Zhang Y., Li F., Zhou Y.J., Dong M.L., Cheng S.T., Ren F., Chen J, & Ren J.H. (2022). SIRT2 Promotes HBV Transcription and Replication by Targeting Transcription Factor p53 to Increase the Activities of HBV Enhancers and Promoters. Frontiers in Microbiology, 13, 836446.
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3

Quantitative Western Blot Analysis of Autophagy and Apoptosis Markers in Hippocampal Tissues

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Western blots were analyzed with homogenized and processed hippocampal tissues as we described previously7 (link). Immunoblotting was conducted using the following antibodies: rabbit polyclonal anti-LC3 antibody (Abcam, Cambridge, MA, USA); goat polyclonal anti-cathepsin B antibody (Santa Cruz); goat polyclonal anti-cathepsin D antibody (Santa Cruz); mouse monoclonal anti-p53 antibody (Cell Signaling Technology, Woburn, MA, USA); rabbit polyclonal anti-PUMA antibody (Cell Signaling Technology); rabbit polyclonal anti-Beclin-1 antibody (Cell Signaling Technology); rabbit polyclonal anti-DRAM antibody (Assay Designs & Stressgen Bioreagents, Ann Arbor, MI, USA); and rabbit polyclonal anti-Bax antibody (Santa Cruz, CA, USA). Tissues were incubated in horse radish peroxidase-conjugated secondary antibody (anti-mouse, anti-rabbit or anti-goat IgG; 1:5000; Santa Cruz). After detecting the immunoreactivity with enhanced chemiluminescent autoradiography (ECL kit, Amersham, Arlington Heights, IL, USA) and normalizing to a GAPDH (Cell Signaling Technology) loading control, the levels of protein expression were quantified with Sigma Scan Pro 5 for comparison.
Cui D., Shang H., Zhang X., Jiang W, & Jia X. (2016). Cardiac arrest triggers hippocampal neuronal death through autophagic and apoptotic pathways. Scientific Reports, 6, 27642.
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4

Protein Extraction and Western Blot Analysis

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Cells were lysed in RIPA buffer supplemented with protease inhibitor cocktail (Roche) for 30 min on ice to extract total protein. Protein concentration was determined by BCA protein assay (ThermoScientific). The protein samples were boiled in 1 × sodium dodecyl sulfate buffer for 5 min. Protein in the same amount was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred onto polyvinylidene fluoride membranes. The membranes were blocked with TBST (100 mmol/L Tris-HCl pH7.4, 150 mmol/L NaCl, and 0.05% Tween 20) containing 5% nonfat milk for 1 h, followed by overnight incubation with indicated antibody at 4°C. After being washed three times, the membranes were probed by another matching second antibody. Eventually, it was visualized by enhanced chemiluminescence reagents super signal (ThermoScientific). Mouse monoclonal anti-p53 antibody (1:1,000), mouse monoclonal anti-DNMT3b antibody (1:1,500), mouse monoclonal anti-UXT antibody (1:1,000), and mouse monoclonal anti-β-actin antibody (1:2,000) were purchased from Cell Signaling Technology. ImageJ was used for quantitative analysis of gray ribbon.
Huang Z.F., Tang Y.L., Shen Z.L., Yang K.Y, & Gao K. (2021). UXT, a novel DNMT3b-binding protein, promotes breast cancer progression via negatively modulating lncRNA MEG3/p53 axis. Molecular Therapy Oncolytics, 24, 497-506.
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5

Stem Cell Culture Optimization

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Dulbecco’s Modified Eagle’s Medium (DMEM), KnockOut-Dulbecco’s modified eagle medium (KO-DMEM), fetal bovine serum (FBS), KnockOut Serum Replacement (KSR), β-mercaptoethanol, L-glutamine, non-essential amino acids (NEAA), and GlutaMAX were purchased from GIBCO/Life Science. Leukemia inhibitory factor (LIF) was purchased from Chemicon. Isoprenaline, ICI118551, and SR59230A were purchased from Tocris Bioscience. 5-(and-6)-carboxy-2′,7′-dichlorofluorescin diacetate (carboxy-DCFDA), 8-Hydroxy-2′-deoxyguanosine (8-OH-dG) and N-acetyl-L-cysteine (NAC) were purchased from Sigma-Aldrich. Forskolin and H89 were purchased from Beyotime. HPLC-grade methanol, water, and formic acid were purchased from Merck. Rabbit polyclonal anti-β3 receptor antibody, mouse monoclonal anti-Oct4 antibody, rabbit polyclonal anti-MDM2 antibody, and mouse monoclonal anti-β-Arrestin-1/2 antibody were purchased from Santa Cruz. Rabbit polyclonal anti-β2 receptor antibody was purchased from Abcam. Rabbit monoclonal anti-phospho-histone H2AX antibody, mouse monoclonal anti-p53 antibody, and rabbit polyclonal anti-phospho-MDM2 (Ser166) antibody were purchased from Cell Signalling Technology.
Sun F., Ding X.P., An S.M., Tang Y.B., Yang X.J., Teng L., Zhang C., Shen Y., Chen H.Z, & Zhu L. (2015). Adrenergic DNA damage of embryonic pluripotent cells via β2 receptor signalling. Scientific Reports, 5, 15950.
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6

Western Blot Analysis of Protein Expression

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Cells were lysed in RIPA buffer supplemented with protease inhibitor cocktail (Roche) for 30 min on ice to extract total protein. Protein concentration was determined by BCA protein assay (ThermoScienti c). The protein samples were boiled in 1 × sodium dodecyl sulfate buffer for 5 minutes. Protein in the same amount was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred onto poly-vinylidene uoride membranes. The membranes were blocked with TBST (100 mmol/l Tris-HCl pH7.4, 150 mmol/l NaCl and 0.05%Tween20) containing 5% non-fat milk for 1 h, following incubated with indicated antibody at 4 ℃ overnight. After washed for three times, the membranes were probed by another matching second antibody. Eventually, it was visualized by enhanced chemiluminescence reagents super signal (Thermoscienti c). Mouse monoclonal anti-p53 antibody (1:1000), mouse monoclonal anti-DNMT3b antibody (1:1500), mouse monoclonal anti-UXT antibody (1:1000) and mouse monoclonal anti-β-actin antibody (1:2000) were purchased from Cell Signaling Technology. Image J were used for quantitative analysis of gray ribbon.
Huang Z., Tang Y., Shen Z., Yang K., & Gao K. (2020). UXT, a novel DNMT3b-binding protein, promotes breast cancer progression via negatively modulating lncRNA MEG3/p53 axis.
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7

Immunoprecipitation Assay for p53

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For immunoprecipitation assays (IP)50 , proteins from whole cell lysates (250–500 µg) were incubated overnight at 4 °C with anti-p53 mouse monoclonal antibody (Cell Signaling). The immunoprecipitated proteins recovered by absorption to EZview Red Protein A Affinity Gel (Sigma-Aldrich) were separated by SDS-PAGE, transferred to nitrocellulose membrane and probed for the indicated proteins. Three independent experiments were performed.
De Rosa C., De Rosa V., Tuccillo C., Tirino V., Amato L., Papaccio F., Ciardiello D., Napolitano S., Martini G., Ciardiello F., Morgillo F., Iommelli F, & Della Corte C.M. (2024). ITGB1 and DDR activation as novel mediators in acquired resistance to osimertinib and MEK inhibitors in EGFR-mutant NSCLC. Scientific Reports, 14, 500.
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8

Oligomerization State of p53 in Tissue

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The oligomerization state of p53 in tissue samples was determined using a glutaraldehyde-based cross-linking assay as previously described with some minor modifications (19 (link)). Frozen liver tissue samples (~50 mg) were homogenized in 1 ml cold lysis buffer (20 mM Tris-HCl (pH 7.4), 137 mM NaCl, 1% NP-40, 2 mM EDTA, 10% glycerol, protease/phosphatase inhibitor cocktail (#04-693-116-001 and #04-906-837-001, Roche)) with a IKA Ultra-Turrax homogenizer (#Z404519, Millipore Sigma) for 15 s and centrifuged at 12,000× g at 4 °C for 10 min. The supernatant was depleted of mouse IgG with protein G-agarose (#sc-2002, Santa Cruz Biotechnology). Cross-linking was performed by adding glutaraldehyde (0.1% final concentration; cat. #16120, Electron Microscopy Sciences) to the protein lysate (~2 µg protein/µl lysis buffer), incubating on ice for 15 min, and terminating the reaction with SDS/PAGE protein sample buffer. The protein samples were resolved on 4–20% gradient SDS/PAGE gels (Bio-Rad Laboratories) and immunoblotted with anti-p53 mouse monoclonal antibody (#2524, Cell Signaling Technology). Within each lane, the fraction of p53 immunoreactivity in the oligomer bands (monomer, dimer and tetramer) was determined using Image Lab software (Bio-Rad Laboratories).
Park J.H., Li J., Starost M.F., Liu C., Zhuang J., Chen J., Achatz M.I., Kang J.G., Wang P.Y., Savage S.A, & Hwang P.M. (2018). Mouse homolog of the human TP53 R337H mutation reveals its role in tumorigenesis. Cancer research, 78(18), 5375-5383.
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