Anti p53 antibody

Manufactured by Merck Group

Sourced in Germany

The Anti-p53 antibody is a laboratory reagent used to detect the presence and quantify the levels of the p53 protein in biological samples. The p53 protein is a tumor suppressor that plays a crucial role in regulating cell growth and division. The Anti-p53 antibody is a specific and sensitive tool for researchers investigating p53-related cellular processes and diseases.

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Close spelling variants of this product

Anti-p53 (37 citations) P53 antibody (4 citations) Anti-p53 antibody (OP140, (3 citations) Anti-p53 Ab7 antibody (2 citations) Anti-p53 C-terminal antibody (2 citations)

9 protocols using anti p53 antibody

Molecular Mechanisms of Drug-Induced Cellular Responses

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Alectinib and LY294002 were purchased from LC Laboratories (Woburn, MA, USA) and Merck Ltd. (Darmstadt, Germany), respectively. Hydrogen peroxide, methotrexate, and catalase were purchased from Nacalai Tesque (Kyoto, Japan). GDC‐0068 and rapamycin were purchased from Cayman Chemical (Ann Arbor, MI, USA) and Toronto Research Chemicals Inc. (Toronto, Canada), respectively. Anti‐β‐actin, anti‐Lamin B, anti‐Akt, anti‐p21, and anti‐p53 antibodies and sodium orthovanadate were purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA). To detect murine p53, an anti‐p53 antibody was purchased from Merck Millipore (Darmstadt, Germany). An anti‐NPM1 antibody and anti‐DDX21 antibody were obtained from Novus Biologicals (Centennial, CO, USA). An anti‐RPS7 antibody and anti‐RPL23 antibody were purchased from Abgent (San Diego, CA, USA). An anti‐Flag (M2) antibody, anti‐Fibrillarin antibody, and anti‐EBP2 antibody were purchased from were purchased from Sigma‐Aldrich (St. Louis, MO, USA), Abcam (Cambridge, MA, USA), and ProteinTech (Chicago, IL. USA), respectively. An anti‐RPL5 antibody and anti‐RPL11 antibody were purchased from Bethyl Laboratories (Montgomery, TX, USA). Other antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA).

Uchihara Y., Tago K., Tamura H, & Funakoshi‐Tago M. (2020). EBP2, a novel NPM‐ALK‐interacting protein in the nucleolus, contributes to the proliferation of ALCL cells by regulating tumor suppressor p53. Molecular Oncology, 15(1), 167-194.

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Analyzing DNA Damage Response

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The ATM inhibitor KU-55933 was purchased from Wako Pure Chemicals (Osaka, Japan). TPEN was purchased from Dojindo (Kumamoto, Japan). Anti-p53 antibody was purchased from Merck (Darmstadt, Germany). Anti-ATM, anti-phospho ATM, anti-phospho p53 (Ser 15), and γH2AX antibodies were purchased from Cell Signaling Technology (Danvers, MA). 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Sigma Aldrich (St. Louis, MO). FluoZin-3 AM was purchased from Thermo Fischer Scientific (Waltham, MA).

Hara H., Kobayashi M., Shiiba M., Kamiya T, & Adachi T. (2019). Sublethal treatment with plasma-activated medium induces senescence-like growth arrest of A549 cells: involvement of intracellular mobile zinc. Journal of Clinical Biochemistry and Nutrition, 65(1), 16-22.

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Chromatin Immunoprecipitation of p53

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Target cells were suspended in 1% formaldehyde in PBS, cross-linked for 10 min at room temperature, washed twice with PBS and lysed with SDS lysis buffer (Promega Corporation). At 4°C, the lysates were centrifuged at 1,0000 × g for 10 min; the supernatants were diluted with ChIP dilution buffer (EMD Millipore) and then immunoprecipitated using an anti-p53 antibody (cat. no. ab1101; 1:20) or mouse IgG (cat. no. ab190475; 1:20) (both from Abcam). IgG served as a negative control. The immunoprecipitated DNA was then analyzed by qPCR.

Wan J., Long F., Zhang C, & Liu Y. (2020). miR-181b-p53 negative feedback axis regulates osteosarcoma cell proliferation and invasion. International Journal of Molecular Medicine, 45(6), 1803-1813.

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Immunofluorescence Analysis of Xenograft Models

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Immunofluorescence was performed after drug treatments to the NB cells, following the protocol as described previously (29) . The nuclei were stained with DAPI and localization of p53 was detected using anti-p53 antibody (Merck; 05-224). Expression of CRM1 was visualized by using anti-CRM1 antibody (Cell Signaling Technology; 46249S). Paraffin-embedded xenografts derived from IMR-32 cells were analyzed by hematoxylin and eosin (H&E) and staining was performed with the anti-p53 (Merck; 05-224) and anti-Ki67 antibodies (Abcam; ab16667).

Mitra S., Muralidharan S.V., Di Marco M., Juvvuna P.K., Kosalai S.T., Reischl S., Jachimowicz D., Subhash S., Raimondi I., Kurian L., Huarte M., Kogner P., Fischer M., Johnsen J.I., Mondal T, & Kanduri C. (2021). Subcellular Distribution of p53 by the p53-Responsive lncRNA NBAT1 Determines Chemotherapeutic Response in Neuroblastoma. Cancer research, 81(6).

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Ubiquitination of p53 by MDM2

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U2OS cells were transfected with constructs expressing either Flag-tagged (Flag-Ub) or histidine-tagged (His-Ub) ubiquitin, HA-tagged p53, and MDM2 (not tagged), as indicated. Eighteen hours after transfection, cells were treated with 25 µM MG132 (Calbiochem) for 6 h. Equivalent amounts of clarified cell lysates were immunoprecipitated (as described above) with 1 µg of anti-HA-coupled protein G Sepharose beads (GE Healthcare) followed by immunoblotting with anti-p53 antibody (Sigma, FL393-G) to detect ubiquitinated p53.

Katz C., Low-Calle A.M., Choe J.H., Laptenko O., Tong D., Joseph-Chowdhury J.S., Garofalo F., Zhu Y., Friedler A, & Prives C. (2018). Wild-type and cancer-related p53 proteins are preferentially degraded by MDM2 as dimers rather than tetramers. Genes & Development, 32(5-6), 430-447.

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Western Blot Analysis of Protein Expression

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Cells were lysed on ice with IP lysis buffer (Thermo Scientific, Waltham, MA) containing protease inhibitor cocktail set III (Millipore, Billerica, MA). Total proteins were separated by electrophoresis using Any kD precast polyacrylamide gel (Bio-Rad, Hercules, CA), and transferred onto PVDF membrane. After blocking with 5% skim milk (Thermo Scientific) in TBST buffer, membranes were incubated with the first antibody, respectively: anti-MELK monoclonal antibody (in-house, previously described [8 (link)]), anti-β-actin antibody, anti-p21 antibody, anti-FOXO1 antibody, anti-FOXO3 antibody, anti-pan-AKT antibody, anti-phospho-AKT (Thr308) antibody, anti-phospho-AKT (Ser473) antibody (Cell Signaling, Danvers, MA), and anti-p53 antibody (Sigma-Aldrich). β-actin was used as a loading control.

Matsuda T., Kato T., Kiyotani K., Tarhan Y.E., Saloura V., Chung S., Ueda K., Nakamura Y, & Park J.H. (2017). p53-independent p21 induction by MELK inhibition. Oncotarget, 8(35), 57938-57947.

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Multiparametric Flow Cytometry and Immunohistochemistry

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For multi-parametric flow cytometry and DEPArray, primary antibodies were obtained from the following sources: FITC-CD45 (#304054; 1:200), FITC-CD34 (#343504; 1:200), FITC-CD105 (#323204; 1:200), FITC-CD90 (#328108; 1:200), FITC-CD73 (#344016; 1:200), FITC HLA-A/B/C antibody (#311404; 1:200), PerCP/Cy5.5-CD146 (#342014; 1:100), PE-Human NG2/MCSP (#FAB2585P, 1:100), BV421-Ki67 (#350506; 1:100), were obtained from Biolegend, anti-Melan-A antibody (# AC12-0297-03; 1:200) from Abcore, and FITC-Anti-S100 (#ab76749; 1:50) was purchased from Abcam.
For immunohistochemistry, anti-human, anti-Melan-A antibody (# ab51061; 1:100), anti-tenascin-C (# ab108930, 1:100) and anti-p21 (#ab188224, 1:100) were purchased from Abcam, HLA-ABC (#565292; 1:100) from BD Biosciences, USP7 (#GTX125894; 1:200) from Genetex, and PTEN antibody (# sc-7974; 1:20) was purchased from Santa Cruz Biotechnology, anti-p53 antibody (#SAB4503021, 1:100) was purchased from Sigma, CCP110 (#12780-I-AP, 1:100) antibody was obtained from Proteintech. Anti-mouse, USP7 (#26948-1-AP, 1:200) and PTEN (#603000-1-Ig, 1:200) antibodies were purchased from Proteintech, Alexafluor-conjugated anti-mouse, anti-rabbit secondary IgG antibodies used for immunofluorescence staining (1:500 dilution) were obtained from Cell Signaling Technology. USP7 inhibitors P5091 (#SML0770) and P22077 (#2301) were purchased from Biovision.

Vishnoi M., Boral D., Liu H., Sprouse M.L., Yin W., Goswami-Sewell D., Tetzlaff M.T., Davies M.A., Glitza Oliva I.C, & Marchetti D. (2018). Targeting USP7 identifies a metastasis-competent state within bone marrow-resident melanoma CTCs. Cancer research, 78(18), 5349-5362.

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Immunofluorescent Detection of p53 in Prostate Tissues

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For detection of p53 protein via IF staining analysis, prostate tissues of subset groups were fixed in 10% formalin for 48 h, embedded in paraffin block, sliced into 4 µm thick sections, and mounted on a glass slide. Sections were then deparaffinized with xylene, rehydrated with different concentrations of EtOH, and pretreated with blocking buffer containing 10% goat serum in 1× PBS solution, for 30 min at room temperature. The pretreated sections were subsequently incubated with anti-p53 antibody (Sigma-Aldrich Co.), diluted 1:300 in blocking buffer. After thorough washing in 1× PBS solution, the slide sections were incubated with goat FITC-labeled anti-rabbit IgG serum for 1 h, washed thrice in 1× PBS for 3 min each, and mounted with vector shield mounting medium. Finally, the green fluorescence in stained cells was observed at 400× magnification via fluorescence microscopy (Eclipse TX100, Nikon, Tokyo, Japan).

Park J.J., Kim J.E., Jeon Y., Lee M.R., Choi J.Y., Song B.R., Park J.W., Kang M.J., Choi H.J., Bae S.J., Lee H., Kang B.C, & Hwang D.Y. (2020). Deletion of NKX3.1 via CRISPR/Cas9 Induces Prostatic Intraepithelial Neoplasia in C57BL/6 Mice. Technology in Cancer Research & Treatment, 19, 1533033820964425.

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Western Blot Analysis of Cell Signaling Proteins

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Anti-Bcl-xl antibody was purchased from Thermo Fisher Scientific (Cat. No. 66020–1-IG). Anti-Bcl-2 antibody was purchased from Thermo Fisher Scientific (Cat. No. MA5–11757). Anti-GAPDH antibody was purchased from Sigma-Aldrich (Cat. No. G8795). Anti-p53 antibody was purchased from Sigma-Aldrich (Cat. No. P6874). Anti-p21 antibody was purchased from Cell Signaling Technology (Cat. No. 2947T). All primary antibodies were used at a suggested dilution in 5% non-fat milk in Phosphate-buffered saline with 0.1% Tween-20 (PBST) buffer for western blot assay. All secondary antibodies were used at a 1:4000 dilution in 5% non-fat milk in Phosphate-buffered saline with 0.1% Tween-20 (PBST) buffer for western blot assay.

Chang M., Gao F., Chen J., Gnawali G, & Wang W. (2022). MDM2-BCL-XL PROTACs enable degradation of BCL-XL and stabilization of p53. Acta materia medica, 1(3), 333-342.

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