P53 fl 393

Manufactured by Santa Cruz Biotechnology

Sourced in United States

The P53 (FL-393) is a lab equipment product offered by Santa Cruz Biotechnology. It is a full-length antibody that recognizes the p53 protein. The p53 protein is a transcription factor that plays a crucial role in regulating cell growth and division.

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Lab products found in correlation

Complete ultra protease inhibitor, by Roche (2 mentions) Iblot, by Thermo Fisher Scientific (2 mentions) β actin clone ac 74, by Merck Group (2 mentions) Nupage bis tris gel, by Thermo Fisher Scientific (2 mentions) P19arf p19arf exon 2, by Rockland Immunochemicals (2 mentions) Hsc70, by Santa Cruz Biotechnology (2 mentions) Gapdh, by Santa Cruz Biotechnology (2 mentions) Mdm2 c 18, by Santa Cruz Biotechnology (1 mentions) Foxo3 75d8, by Cell Signaling Technology (1 mentions) Foxo1 c29h4, by Cell Signaling Technology (1 mentions) α tubulin dm1a, by Merck Group (1 mentions) Alliance 2, by Uvitec (1 mentions) Phospho akt ser473, by Cell Signaling Technology (1 mentions) Anti flag m2 affinity gel, by Merck Group (1 mentions) Raptor, by Santa Cruz Biotechnology (1 mentions) Raptor, by Cell Signaling Technology (1 mentions) Amersham ecl, by Cytiva (1 mentions) γh2ax, by Cell Signaling Technology (1 mentions) Stat3, by Cell Signaling Technology (1 mentions) Pgam5, by Abcam (1 mentions)

Spelling variants of this product

FL-393 (46 citations) Anti-p53 (FL-393) antibody (3 citations) P53 (FL-393; sc-6243, (2 citations) Anti-p53 (DO1 and FL-393) (2 citations)

13 protocols using p53 fl 393

Western Blot Analysis of Cell Signaling Proteins

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Protein extracts were prepared by lysis in RIPA buffer (300 mM NaCl, 2% IGEPAL CA-630, 1% deoxycholic acid, 0.2% SDS, 100 mM Tris-HCl pH 8.0) containing complete ULTRA protease inhibitors (Roche, Basel, Switzerland). Western blots were carried out using 30 μg total protein per sample run on NuPAGE Bis-Tris gels (Life Technologies) and transferred to nitrocellulose membranes with an iBlot (Life Technologies) according to the manufacturer's protocols. Blots were probed with the following antibodies: p53 (FL-393, Santa Cruz Biotechnology, Santa Cruz, CA, USA); p19ARF (p19ARF exon 2, Rockland, Gilbertsville, PA, USA); Mdm2 (C-18, Santa Cruz Biotechnology); FoxO3 (75D8, Cell Signaling Technology); FoxO1 (C29H4, Cell Signaling Technology); and β-actin (clone AC-74, Sigma-Aldrich).

Vandenberg C.J., Motoyama N, & Cory S. (2016). FoxO3 suppresses Myc-driven lymphomagenesis. Cell Death & Disease, 7(1), e2046-.

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Immunoprecipitation and Western Blot Analysis

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For immunoprecipitation (IP), cells were lysed in CHAPS lysis buffer (40 mM Hepes pH7.4, 120 mM NaCl, 2 mM EDTA, 0.3% CHAPS) containing mix of protease inhibitors (Boehringer), plus 5 mM NaF, 10 mM glycerophosphate and 1 mM Na₃VO₄. For IP lysates were pre-incubated with protein G-Agarose (Pierce) and then with the indicated antibody, under gentle rocking at 4 °C overnight. For Western blot (Wb) cells were lysed in RIPA buffer. Membranes were developed using the enhanced chemiluminescence (ECL Amersham) by chemiluminescence imaging system, Alliance 2.7 (UVITEC Cambridge) and quantified by the software Alliance V_1607. Primary antibodies used: MDM4 BL1258 (Bethyl laboratory), MDM4 C82 (Sigma), MDM4 8C6 (Millipore) p53 FL393 (Santa Cruz), α-tubulin DM1A (Sigma), actin C-40 (Sigma), mTOR (Santa Cruz), mTOR (Cell Signaling), anti-FLAG M2 affinity gel (Sigma), phosphoSer473-AKT (Cell Signaling), phosphor-Thr389-S6K (Cell Signaling), Akt (Cell Signaling), S6K1 (Santa Cruz), Raptor (Cell Signaling), Raptor (Santa Cruz).
Fractionation of lysates into heavy membrane and light membrane/cytosolic fractions was performed according to Menon et al. 2014.

Mancini F., Teveroni E., Di Conza G., Monteleone V., Arisi I., Pellegrino M., Buttarelli M., Pieroni L., D’Onofrio M., Urbani A., Pontecorvi A., Mazzone M, & Moretti F. (2017). MDM4 actively restrains cytoplasmic mTORC1 by sensing nutrient availability. Molecular Cancer, 16, 55.

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Immunoblotting for DNA Damage Response

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For immunoblots, cell lysates with equal total protein content (2–20 μg) were blotted with antibodies to p53 (FL393), p21 GAPDH, Hsc70, Nek2, and tubulin (all from Santa Cruz Biotechnology); γH2AX (all from Cell Signaling)). All immunoblots were repeated at least two times.

Ghaleb A., Padellan M, & Marchenko N. (2020). Mutant p53 drives the loss of heterozygosity by the upregulation of Nek2 in breast cancer cells. Breast Cancer Research : BCR, 22, 133.

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Antibody Validation for ChIP and Western

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Antibodies used for these studies include AR (N-20, SantaCruz Biotechnologies, for ChIP), AR (441, SantaCruz Biotechnologies, for immunoblotting), AR (PG21, EMD Millipore, for ChIP-re-ChIP), AIB1 (NCOA3, 39797, Active Motif), BRG1 (SMARCA4, G-7, SantaCruz Biotechnologies), β-actin (4967L, Cell Signaling), IRF1 (8478S, Cell Signaling), MEP50 (WDR77, 2823S, Cell Signaling), p300 (C-20, SantaCruz Biotechnologies), p53 (DO-1, SantaCruz Biotechnologies), p53 (FL-393, SantaCruz Biotechnologies), p53 (pAB 421, Calbiochem), PGAM5 (ab126534, abcam), pHistone H2a.z (S139, Cell Signaling), PKN1 (610686, BD Biosciences), STAT3 (12640S, Cell Signaling), TIF2 (NCOA2, 610984, BD Biosciences) and Ki67 (abcam, ab15580).

Liu S., Kumari S., Hu Q., Senapati D., Venkadakrishnan V.B., Wang D., DePriest A.D., Schlanger S.E., Ben-Salem S., Valenzuela M.M., Willard B., Mudambi S., Swetzig W.M., Das G.M., Shourideh M., Koochekpour S., Falzarano S.M., Magi-Galluzzi C., Yadav N., Chen X., Lao C., Wang J., Billaud J.N, & Heemers H.V. (2017). A comprehensive analysis of coregulator recruitment, androgen receptor function and gene expression in prostate cancer. eLife, 6, e28482.

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Western Blot Analysis of Cell Signaling

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Antibodies were purchased from Cell Signaling Technologies (Danvers, MA) unless otherwise indicated. These include mouse IgG against Caspase 8 (1C12), Histone H3 (1G1) (Santa Cruz Biotechnology, Santa Cruz, CA), Cdc-2 (17) (Santa Cruz Biotechnology), and GAPDH (Santa Cruz Biotechnology); rabbit IgG against p21, PARP, Cleaved Caspase 3 (Asp175) (5A1E), LC3A/B (D3U4C), α/β tubulin, p-Cdc-2 (T14/Y15) (Santa Cruz Biotechnology), p53 (FL-393) (Santa Cruz Biotechnology), and p-Histone H3 (Ser10) (Upstate Cell Signaling Solutions, Lake Placid, NY). Secondary antibodies used for immunofluorescence include donkey anti-rabbit IgG for Alexa Fluor 594 (Invitrogen, Waltham, MA). Nuclei were visualized by DAPI (Sigma-Aldrich, St. Louis, MO) staining at 1 µg/ml for 5 min.
Western Blot analyses were carried out as previously described [55 (link)]. Western blots were visualized by chemiluminescence using Bio-Rad ChemiDoc™ Imaging System (Hercules, CA) and specific protein signals were quantified using the Fiji-ImageJ analytical software (NIH, Bethesda, MD).

Graff B.T., Palanivel C., Jenkins C.B., Baranowska-Kortylewicz J, & Yan Y. (2023). Benzimidazole carbamate induces cytotoxicity in breast cancer cells via two distinct cell death mechanisms. Cell Death Discovery, 9, 162.

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Immunoblotting Analysis of Signaling Pathways

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For immunoblots, cell lysates with equal total protein content (2–20 µg) were blotted with antibodies to p53 (FL393), Mdm2, GAPDH, Hsc70 (all from Santa Cruz Biotechnology); Erk1, pErk1/2 (T202/Y204), EGFR, pEGFR (Y845), ERBB2, pERBB2 (Y1221/1222 and pY1248), MET, cleaved PARP, PDGFRα, PDGFRβ, FGFR, AKT, pAKT MdmX, pTK (all from Cell Signaling); HSF1, pHSF1 (S326), Hsp90, Hsp70, Hsp27 (all from Enzo Life Sciences Inc., Farmingdale, NY). All Western blots were repeated at least two times. The phospho-RTK array on primary mammary tumor cells was performed according to the manufacturer’s protocol (Mouse Phospho-RTK Array Kit, R&D Systems).

Yallowitz A., Ghaleb A., Garcia L., Alexandrova E.M, & Marchenko N. (2018). Heat shock factor 1 confers resistance to lapatinib in ERBB2-positive breast cancer cells. Cell Death & Disease, 9(6), 621.

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Standardized Immunohistochemistry Protocol

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Standardized immunohistochemical staining were performed on murine FFPE tissues. The following primary antibodies were used and listed in detail in Supplementary Table 1: p53 FL393 (Santa Cruz); cyclinD1, pan-Cytokeratin, and α-smooth muscle actin/SMA (all Abcam); phospho-HSF1 (Bioss); and pRB and Hsp70 (both Cell Signaling). The ImmPRESS™ Peroxidase polymer reagent based on 3, 3-diaminobenzidine (DAB, Vectorlabs), or Alexa Fluor®488-coupled and Alexa Fluor®647-coupled secondary antibodies (immunofluorescence) were used as detection systems. Hematoxylin (DAB) or DAPI (immunofluorescence) were used as counterstains. Stained sections were analyzed by microscopy (AxioScope, Zeiss) with ZENblue software V3.0 (Zeiss). Grading for phospho-HSF1 and p53 was as follows: p53^high, >25% with intense nuclear staining; Hsp70^high, >50% with high intensity.

Isermann T., Şener Ö.Ç., Stender A., Klemke L., Winkler N., Neesse A., Li J., Wegwitz F., Moll U.M, & Schulz-Heddergott R. (2021). Suppression of HSF1 activity by wildtype p53 creates a driving force for p53 loss-of-heterozygosity. Nature Communications, 12, 4019.

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Western Blot Protein Analysis

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Cell lysates were prepared as described (17 (link)). Primary antibodies were MDM2: 4H26L4 (Invitrogen, Carlsbad, CA; 700555) 1:300, MYCN: NCMII100 (Santa Cruz) 1:200, p53: FL393 (Santa Cruz) 1:300, p21: C-19 (Santa Cruz) 1:100; α-tubulin: B 5-1-2 (Sigma) 1:5000; and HRP-conjugated secondary antibodies (Santa Cruz). Densitometry was performed with ImageJ.

Tran H.N., Singh H.P., Guo W., Cambier L., Riggan L., Shackleford G.M., Thornton M.E., Grubbs B.H., Erdreich-Epstein A., Qi D.L, & Cobrinik D. (2020). Reciprocal Induction of MDM2 and MYCN in Neural and Neuroendocrine Cancers. Frontiers in Oncology, 10, 563156.

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Immunoprecipitation and Western Blot Analysis

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Primary antibodies were used at a concentration of approximately 1 ug primary antibody per 200 μl protein extract for immunoprecipitation and Western blotting as described elsewhere [21] (link). Antibodies used were INSM1 (A-8) (1:1000), Menin (C-19) (1:1000), Actin (1:2000), Cyclin D1 (HD11) (1:300), p27 (F-8) (1:500), p53 (FL-393) (1:2000), Hes1 (E:5) (1:500), Notch 3 (A-6) (1:800); all purchased from Santa-Cruz Biotechnology (USA). Antibodies against phosphor-ERK (1:1000), phosphor-Akt (S473) (D-96) (1:1000), Akt (1:1000) (1:1000) were purchased from Cell Signaling (Japan). Secondary antibodies were obtained from GE Healthcare (UK).

Capodanno Y., Chen Y., Schrader J., Tomosugi M., Sumi S., Yokoyama A., Hiraoka N, & Ohki R. (2021). Cross-talk among MEN1, p53 and Notch regulates the proliferation of pancreatic neuroendocrine tumor cells by modulating INSM1 expression and subcellular localization. Neoplasia (New York, N.Y.), 23(9), 979-992.

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Immunohistochemical Analysis of Cell Signaling Pathways

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Flag-Tag (F3165, 1:1000) and actin (A2066, 1:1000) were obtained from Sigma. BrdU (B44, 1:200) and Ki67 antibodies (556528, 1:300) were obtained from BD Pharmingen (San Jose, CA, USA). Caspase-2 (11B4, 1:500) was obtained from Alexis (Farmingdale, NY, USA). Mdm2 N-terminal (IF2, 1:200) and p53 (DO-1, 1:2000) were obtained from Calbiochem (Billerica, MA, USA). p21 (F-5, 1:100), p53 (FL393, 1:1000) and Mdm2 (SMP14, 1:1000) were obtained from Santa Cruz (Dallas, TX, USA). Cleaved Caspase-3 (9661, 1:500), phospho-Chk2 (T68) (2661, 1:1000), and Parp (9532, 1:1000) were obtained from Cell Signaling Technology (Danvers, MA, USA).

Terry M.R., Arya R., Mukhopadhyay A., Berrett K.C., Clair P.M., Witt B., Salama M.E., Bhutkar A, & Oliver T.G. (2014). Caspase-2 impacts lung tumorigenesis and chemotherapy response in vivo. Cell Death and Differentiation, 22(5), 719-730.

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