Phorbol 12 myristate 13 acetate

Manufactured by LC Laboratories

Phorbol 12-myristate 13-acetate is a chemical compound used in laboratory research. It functions as a protein kinase C activator.

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Lab products found in correlation

Rpmi 1640 medium, by American Type Culture Collection (3 mentions) Fetal bovine serum (fbs), by American Type Culture Collection (3 mentions) L glutamine, by American Type Culture Collection (3 mentions) Lncap human prostate cancer cells, by American Type Culture Collection (3 mentions) 20 3h phorbol 12 13 dibutyrate 3h pdbu, by PerkinElmer (2 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (2 mentions) Gst capture kit, by GE Healthcare (1 mentions) Prostratin, by LC Laboratories (1 mentions) Antibiotic antimycotic, by Thermo Fisher Scientific (1 mentions) Phosphatidylcholine pc, by Avanti Polar Lipids (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) 15nh4cl, by Cambridge Isotopes (1 mentions) Rabbit anti gfp antibody, by Cell Signaling Technology (1 mentions) Bicinchoninic acid bca protein assay kit, by Thermo Fisher Scientific (1 mentions) Ht22 cells, by American Type Culture Collection (1 mentions) Rabbit anti na k atpase antibody, by Cell Signaling Technology (1 mentions) Phorbol 12 13 dibutyrate pdbu, by Merck Group (1 mentions) Fetal bovine serum (fbs), by ZenBio (1 mentions) Rabbit anti β actin, by Cell Signaling Technology (1 mentions) Glutathione sepharose 4b, by GE Healthcare (1 mentions)

5 protocols using phorbol 12 myristate 13 acetate

Binding Kinetics of Protein Kinase C Ligands

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[20-³H]Phorbol 12,13-dibutyrate ([³H]PDBu) (17.2 Ci/mmol) was from PerkinElmer Life Sciences (Boston, MA). PDBu, phorbol 12-myristate 13-acetate (PMA), ingenol 3-angelate, and prostratin were from LC Laboratories (Woburn, MA). Sapintoxin D was from Enzo Life Sciences International Inc (Farmingdale, NY). The bryostatin 1 was provided by the Developmental Therapeutics Program, NCI (Frederick, MD). Phosphatidyl-L-serine (PS), phosphatidylcholine (PC), and 1,2-dioctanoylglycerol (DOG) were purchased from Avanti Polar Lipids (Alabaster, AL). LNCaP human prostate cancer cells, fetal bovine serum (FBS), RPMI 1640 medium, and L-glutamine were obtained from the American Type Culture Collection (Manassas, VA). LB broth and LB agar plates used in bacterial culturing were purchased from K-D Medical, Inc. (Columbia, MD). Primers and site-directed mutagenesis kits were from Invitrogen (Life Technologies, Grand Island, NY). For SPR, all experiments were performed with a Biacore 3000 optical biosensor at 25°C. Sensor chip CM-5 with a carboxymethylated dextran matrix, EDC (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide), NHS (N-hydroxysuccinimide), and P20 surfactant, buffers, NBS-EP and HBS-N, and GST-capture kit were obtained from GE Healthcare (Piscataway, NJ).

Kelsey J.S., Geczy T., Lewin N.E., Kedei N., Hill C.S., Selezneva J.S., Valle C.J., Woo W., Gorshkova I, & Blumberg P.M. (2014). Charge density influences C1 domain ligand affinity and membrane interactions. Chembiochem : a European journal of chemical biology, 15(8), 1131-1144.

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THP-1 Macrophage Exposure to Heated Particles

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THP-1 cells (ATCC; TIB-202) were cultured at 37 °C and 5% CO₂ in complete medium, i.e., RPMI-1640 supplemented with 10% fetal bovine serum (FBS), 10 mM Hepes, and 1% antibiotic–antimycotic (Gibco). Cells were subcultured and exposed before reaching confluence. Before particle exposure, THP-1 cells were plated in 96-well plates (100,000 cells/well) in complete medium and differentiated from monocytes to macrophages with 100 nM phorbol 12-myristate 13-acetate (LC Laboratories) for 24 h. Next, cells were washed once with DPBS and exposed for 24 h to increasing concentrations of the particles previously heated at 200 °C for 2 h and then dispersed in culture medium without FBS. Supernatants of cell culture were collected and stored at −80 °C for cytokine assessment.

Pavan C., Santalucia R., Leinardi R., Fabbiani M., Yakoub Y., Uwambayinema F., Ugliengo P., Tomatis M., Martra G., Turci F., Lison D, & Fubini B. (2020). Nearly free surface silanols are the critical molecular moieties that initiate the toxicity of silica particles. Proceedings of the National Academy of Sciences of the United States of America, 117(45), 27836-27846.

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Radioactive Phorbol Ester Binding Assay

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[³H]Phorbol 12, 13-dibutyrate ([³H]PDBu) (13.5 Ci/mmol) was obtained from Perkin Elmer Life Sciences. PDBu and phorbol 12-myristate 13-acetate (PMA) were purchased from LC Laboratories (Woburn, MA). Phosphatidyl-l-serine (PS) and phosphatidylcholine (PC) were from Avanti Polar Lipids (Alabaster, AL). Dimethyl sulfoxide (DMSO) was purchased from Sigma – Aldrich (St. Louis, MO). LNCaP human prostate cancer cells, fetal bovine serum (FBS), RPMI 1640 medium, and l-glutamine were from American Type Culture Collection (Manassas, VA). Reagents used for culturing bacteria (LB Broth, LB agar plates with different selection of antibiotics, etc.) were from K-D Medical, Inc. (Columbia, MD). The oligonucleotide primers used for polymerase chain reaction (PCR) were obtained from Integrated DNA Technologies (Coralville, IA), and site-directed mutagenesis was from Invitrogen.

Czikora A., Pany S., You Y., Saini A.S., Lewin N.E., Mitchell G.A., Abramovitz A., Kedei N., Blumberg P.M, & Das J. (2018). Structural determinants of phorbol ester binding activity of the C1a and C1b domains of protein kinase C theta. Biochimica et biophysica acta. Biomembranes, 1860(5), 1046-1056.

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Lipid and Protein Reagents for Cell Signaling Studies

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1,2-Dioctanoyl-sn-glycerol (DOG) and 2-dihexanoyl-sn-glycero-3-phospho-l-serine (DPS) were obtained from Avanti Polar Lipids (Alabaster, AL). Phorbol 12-myristate 13-acetate (PMA) was purchased from LC Laboratories (Woburn, MA). Phorbol-12,13-dibutyrate (PDBu) was obtained from Sigma-Aldrich (St. Louis, MO). Deuterated dodecylphosphocholine (d₃₈-DPC), DMSO (d₆-DPC), and ¹⁵NH₄Cl were obtained from Cambridge Isotope Laboratories, Inc. (Tewksbury, MA). Protein quantification was conducted using the Bradford protein assay from Bio-Rad (Hercules, CA) and the BCA (bicinchoninic acid) protein assay kit from Thermo Fisher Scientific (Grand Island, NY). Glutathione sepharose 4B was obtained from GE Healthcare Life Sciences (Piscataway, NJ). HT22 cells were purchased from ATCC (Manassas, VA). Fetal bovine serum (FBS) was from ZenBio (Research Triangle Park, NC). Rattus norvegicus Munc13-1 conjugated with green fluorescent protein (GFP) was a generous gift from N. Brose (Max Planck Institute for Experimental Medicine, Gottingen, Germany). The rabbit anti-GFP antibody, rabbit anti-Na/K-ATPase antibody, rabbit anti-β-actin, and HRP (horseradish peroxidase)-conjugated rabbit anti-IgG antibody used for the Western blot analysis were obtained from Cell Signaling (Danvers, MA). All other reagents were purchased from either MilliporeSigma (Burlington, MA) or Thermo Fisher Scientific.

You Y., Katti S., Yu B., Igumenova T.I, & Das J. (2021). Probing the Diacylglycerol Binding Site of Presynaptic Munc13-1. Biochemistry, 60(16), 1286-1298.

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Purification and Labeling of Proteins

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[20-³H]Phorbol 12,13-dibutyrate ([³H]PDBu) (17.2 Ci/mmol) was obtained from PerkinElmer Life Sciences. PDBu and phorbol 12-myristate 13-acetate (PMA) were purchased from LC Laboratories (Woburn, MA). Phosphatidyl-L-serine (PS), phosphatidylcholine (PC), and sphingosine were from Avanti Polar Lipids (Alabaster, AL) and the isopropyl O-D-thiogalactopyranoside (IPTG) was from Sigma (St. Louis, MO). LNCaP human prostate cancer cells, Neuro-2a mouse neuroblastoma cells, HEK-293 cells, fetal bovine serum (FBS), RPMI 1640 medium, and L-glutamine were from the American Type Culture Collection (Manassas, VA). Reagents used for culturing bacteria (LB Broth, LB agar plates with ampicillin, ampicillin solution, etc.) were from K-D Medical, Inc. (Columbia, MD). The primers, the High Fidelity Polymerase, and the ligase used for cloning, the chemically competent MaxEfficiency DH5αT1^R and BL-21(DE3) cells, the DMEM without phenol red used for confocal analysis, the Lipofactamine and Plus reagent, Lipofectamine 3000, and the GST spin purification kit were from Thermo Scientific (Pittsburgh, PA). The QIAquick PCR Purification Kit and the QIAprep Spin Miniprep Kit were from Qiagen (Germantown, MD). The pHTN HaloTag(R) CMV-neo vector and the kit for purification of Halo-tagged proteins were from Promega (Madison, WI).

Das J., Kedei N., Kelsey J.S., You Y., Pany S., Mitchell G.A., Lewin N.E, & Blumberg P.M. (2018). Critical Role of Trp-588 of Presynaptic Munc13-1 for Ligand Binding and Membrane Translocation. Biochemistry, 57(5), 732-741.

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