Triacsin c

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Triacsin C is a small molecule compound that functions as an inhibitor of long-chain acyl-CoA synthetases. It is commonly used in research applications to study lipid metabolism and signaling pathways.

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Lab products found in correlation

7 protocols using triacsin c

Characterization of LDAH Antibodies

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Custom polyclonal rabbit anti-human and mouse LDAH antibodies were generated at Bethyl Laboratories, and characterized as previously described^{15 (link)}. Bodipy 493/503, LipidTox, and RNAiMAX were from Invitrogen. Fugene HD was purchased from Promega. Isoproterenol hydrochloride, anti-beta actin antibody, anti Flag-M2-HRP, anti Flag-M2-agarose, mouse IgG, oleic acid, 3-isobutyl-1-methylxanthine, insulin, dexamethasone and nocodazole were from Sigma-Aldrich. Triacsin C and anti-ubiquitin antibody were from Santa Cruz Biotechnology. Anti-PLIN1 antibody was from Progen. Anti-beta tubulin and anti-PLIN2 antibodies were from Nobus Biologicals. Anti-PLIN3 was purchased from Fitzgerald. Anti-ATGL and anti-HSL were from Cell Signaling. Alexa Fluor 647-conjugated secondary antibodies and anti-GM130 were from BD Pharmingen. Protein G-agarose and protein-A-agarose were from Thermo Scientific. MG132 was purchased from Calbiochem. Total and free cholesterol were measured using kits from Wako. TAG were determined with Infinity Trglyceride reagents from Thermo Scientific. ¹⁴C-oleic acid was purchased from Perkin Elmer.

Goo Y.H., Son S.H, & Paul A. (2017). Lipid Droplet-Associated Hydrolase Promotes Lipid Droplet Fusion and Enhances ATGL Degradation and Triglyceride Accumulation. Scientific Reports, 7, 2743.

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Visualizing Lipid Ester Synthesis in Huh7 Cells

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For fluorescence microscopic analysis, Huh7 cells precultured with 0.4 mM OA for 12 h were treated with BODIPY 558/568-C₁₂ (Invitrogen) for 1 h to metabolically label newly synthesized lipid esters, rinsed, and cultured for another hour. For the last 2 h of incubation, 1.5 mM hydroxyurea (Sigma-Aldrich) was added to the culture medium with or without 5 µM triacsin C (Santa Cruz Biotechnology, Inc.). After fixation, cells were incubated with BODIPY 493/503 to stain all LDs. For the EM method to visualize newly synthesized lipid esters, cells pretreated with OA for 12 h were cultured with 0.2 mM DHA (Sigma-Aldrich) for 8 h before fixation (Cheng et al., 2009 (link)).

Ohsaki Y., Kawai T., Yoshikawa Y., Cheng J., Jokitalo E, & Fujimoto T. (2016). PML isoform II plays a critical role in nuclear lipid droplet formation. The Journal of Cell Biology, 212(1), 29-38.

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Cell Culture Protocols for Hepatic and Osteosarcoma Cell Lines

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HepG2 (JCRB1054), HEK293 (JCRB9068), and HeLa (JCRB9004), which were obtained from the Japanese Collection of Research Bioresources Cell Bank, McA-RH7777 (ATCC CRL-1601) obtained from American Type Culture Collection, and A549 donated by Dr. Takashi Takahashi (Nagoya University) were cultured in Dulbecco's modification of Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and antibiotics at 37 °C in a humidified atmosphere of 95% air and 5% CO₂. Huh7 from Dr. Eija Jokitalo (University of Helsinki) and U2OS from Dr. Hidemasa Goto (Aichi Cancer Center) were cultured in MEM and McCoy’s medium, respectively, with the same supplementations. In some experiments, OA (Sigma) in complex with fatty acid-free bovine serum albumin (BSA) (Wako) at a molar ratio of 6:1 was added. BAY 13-9952 (Implitapide; Bayer Healthcare), triacsin C (Santa Cruz Biotech), CP-346086, and TM (Sigma) were purchased from respective suppliers.

Sołtysik K., Ohsaki Y., Tatematsu T., Cheng J, & Fujimoto T. (2019). Nuclear lipid droplets derive from a lipoprotein precursor and regulate phosphatidylcholine synthesis. Nature Communications, 10, 473.

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Antibody Sources and Reagents for Lipid Metabolism Research

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Commercial antibodies used are listed in Supplementary Table S2. Antibody against SCD-1 [57 (link)] was a kind gift from Dr. Jean-Baptiste Demoulin, Université Catholique de Louvain, Brussels, Belgium. Anti-human ACSL4 was generously provided by Dr. Stephen Prescott, University of Utah, Salt Lake, USA and Dr. Diana Stafforini, Huntsman Cancer Institute, University of Utah, USA, and used as indicated in [58 (link)]. Triacsin C was purchased from Santa Cruz (Santa Cruz, CA, USA); A939572 was from Biofine International (Biofine International Inc, Vancouver, Canada); Metformin, Phenformin, AICAR, NS-398 and Akt Inhibitor IV were from Sigma-Aldrich, (Sigma-Aldrich, St. Louis, MO, USA).

Sánchez-Martínez R., Cruz-Gil S., de Cedrón M.G., Álvarez-Fernández M., Vargas T., Molina S., García B., Herranz J., Moreno-Rubio J., Reglero G., Pérez-Moreno M., Feliu J., Malumbres M, & de Molina A.R. (2015). A link between lipid metabolism and epithelial-mesenchymal transition provides a target for colon cancer therapy. Oncotarget, 6(36), 38719-38736.

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Cell Culture Conditions and Reagents

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U2OS, Huh7, and A549 cells were kindly donated by Dr. Hidemasa Goto (Aichi Cancer Center, Nagoya, Japan), Dr. Eija Jokitalo (University of Helsinki, Helsinki, Finland), and Dr. Takashi Takahashi (Nagoya University, Nagoya, Japan), respectively. Cells were cultured in DMEM (U2OS and A549) or minimum essential medium (Huh7) supplemented with 10% FBS and antibiotics at 37°C in a humidified atmosphere of 95% air and 5% CO₂. In some experiments, OA (Sigma) in complex with fatty acid–free BSA (Wako) at a molar ratio of 6:1 was added. BAY13-9952 (Implitapide; Bayer Healthcare), triacsin C (Santa Cruz Biotechnology), tunicamycin, and Torin1 (Sigma) were purchased from the indicated suppliers.

Sołtysik K., Ohsaki Y., Tatematsu T., Cheng J., Maeda A., Morita S.Y, & Fujimoto T. (2020). Nuclear lipid droplets form in the inner nuclear membrane in a seipin-independent manner. The Journal of Cell Biology, 220(1), e202005026.

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Lipid and Cell Signaling Pathways Analysis

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Oleic, palmitic, palmitoleic, linoleic, and methyl oleic acids, 5-bromo-2’-deoxyuridine (BrdU), etomoxir (Sigma), CAY10587, bisindolylmaleimide-1 (Cayman Chemical, Ann Arbor, MI), U0126 (Cell Signaling Technology, Danvers, MA), PKCζ myristolated pseudo substrate (Enzo Life Science, Plymouth Meeting, PA), triacsin C (Santa Cruz Biotechnology, Dallas, Santa Cruz, CA), GO-6976 (Tocris Bioscience, Bristol, UK) were purchased commercially.

Brelje T.C., Bhagroo N.V., Stout L.E, & Sorenson R.L. (2017). Prolactin and oleic acid synergistically stimulate β-cell proliferation and growth in rat islets. Islets, 9(4), e1330234.

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Quantifying Palmitate Incorporation in Lipids

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20E4cells/mL cells were grown in RPMI complete media and supplemented with a mix containing 0.2mM nonradiolabeled palmitate (conjugated to BSA 6:1), additional 10uM BSA, 0.15uCi [1-¹⁴C] palmitic acid (mixed at 37° for 30 min). Cells were treated with 10uM Triacsin C (Santa Cruz, in DMSO) for 2 h prior to fatty acid treatment. After 6 h fatty acid treatment, cells were collected via centrifugation for 5min 1000xg and washed twice in cold PBS containing 0.5% BSA following by one wash in PBS without BSA. Lipids were extracted from cell pellets with addition of 3:2 hexane:isopropanol and 20min gentle shaking, followed by centrifugation 5min 5000xg. Lipid extracts were dried under air stream and neutral lipids were separated by TLC using hexane:diethyl ether:acetic acid (80:20:1). TLC plates were exposed to a phosphor imaging cassette and revealed by a phosphorimaging (Typhoon FLA 7000). Incorporation of radiolabeled palmitic acid into lipid classes (cholesterol esters, triglycerides, diacylglycerides and phospholipids) was measured as band intensity on TLC using Image J software. Remaining cell pellets were dried and proteins were solubilized using 0.3N NaOH in 0.1% SDS. Band intensity was normalized to protein concentration.

Piccolis M., Bond L.M., Kampmann M., Pulimeno P., Chitraju C., Jayson C.B., Vaites L.P., Boland S., Lai Z.W., Gabriel K.R., Elliott S.D., Paulo J.A., Harper J.W., Weissman J.S., Walther T.C, & Farese RV J.r. (2019). Probing the Global Cellular Responses to Lipotoxicity Caused by Saturated Fatty Acids. Molecular cell, 74(1), 32-44.e8.

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