Disulfiram

Manufactured by Bio-Techne

Sourced in United Kingdom

Disulfiram is a laboratory reagent used in biochemical and analytical applications. It serves as an inhibitor of the enzyme aldehyde dehydrogenase, which plays a role in the metabolism of alcohol. Disulfiram is utilized in various research and testing procedures to study the effects of alcohol metabolism and related physiological processes.

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Lab products found in correlation

Propidium iodide solution, by Merck Group (2 mentions) Z vad fmk, by InvivoGen (2 mentions) Nigericin, by InvivoGen (2 mentions) Mcc950, by InvivoGen (2 mentions) Poly da dt, by InvivoGen (2 mentions) Acetaldehyde, by Merck Group (2 mentions) D penicillamine, by Bio-Techne (2 mentions) Milli q water, by Merck Group (1 mentions) Reserpine, by Merck Group (1 mentions) Alamarblue reagent, by Thermo Fisher Scientific (1 mentions) Epigallocatechin gallate (egcg), by Selleck Chemicals (1 mentions) Conoidin a, by Cayman Chemical (1 mentions) Vincristine, by Selleck Chemicals (1 mentions) Bactiter glo microbial cell viability assay reagent, by Promega (1 mentions) Bioassay software, by PerkinElmer (1 mentions) Snx 2112, by Selleck Chemicals (1 mentions) Paraformaldehyde (pfa), by Electron Microscopy Sciences (1 mentions) Sytox green, by Thermo Fisher Scientific (1 mentions) Iscove modified dulbecco media (imdm), by Thermo Fisher Scientific (1 mentions) A23187, by Merck Group (1 mentions)

9 protocols using disulfiram

Inflammasome Activation and Inhibition Assay

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E. coli LPS (serotype O:111 B4) was purchased from Enzo Biosciences as a ready-to-use stock solution of 1 mg/ml and used at a working concerntration of 1 μg/ml. Nigericin was bought from Invivogen, dissolved in 100% ethanol to 6.7 mM and used for NLRP3 stimulation at a concentration of 10 μM. Poly(dA:dT) was from Invivogen and dissolved to a stock concentration of 1 mg/ml. Poly(dA:dT) was transfected at a final concentration of 5 μg/ml. MCC950 and zVAD-FMK (both from Invivogen) were resuspended in sterile DMSO to a concentration of 20 mM and used at a final concentration of 10 μM and 20 μM, respectively. Disulfiram was purchased from Tocris Biosciences, dissolved in DMSO to generate a stock solution at 20 mM. During inhibition experiments, Disulfiram was present at a concentration of 50 μM. To inhibit pyroptosis, cells were pre-treated with inhibitors for 30 min and the respective inhibitor was present during signal 2 of inflammasome stimulation. Propidium iodide solution (1 mg/ml) was purchased from MilliporeSigma. Recombinant human M-CSF (CHO expressed, carrier-free) was bought from R&D Systems and dissolved in sterile PBS pH 7.4 (stock concentration 50 μg/ml). YVAD-pNA was from Enzo Biosciences and the stock solution was prepared at 20 mM in DMSO.

Devant P., Cao A, & Kagan J.C. (2021). Evolution-inspired redesign of the LPS receptor caspase-4 into an interleukin-1β converting enzyme. Science immunology, 6(62), eabh3567.

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Inflammasome Activation and Pyroptosis Inhibition

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Ligand and chemical reconstitution E. coli LPS (serotype O:111 B4) was purchased from Enzo Biosciences as a ready-to-use stock solution of 1 mg/ml and used at a working concerntration of 1 µg/ml. Nigericin was bought from Invivogen, dissolved in 100% ethanol to 6.7 mM and used for NLRP3 stimulation at a concentration of 10 µM. Poly(dA:dT) was from Invivogen and dissolved to a stock concentration of 1 mg/ml. Poly(dA:dT) was transfected at a final concentration of 5 µg/ml. MCC950 and zVAD-FMK (both from Invivogen) were resuspended in sterile DMSO to a concentration of 20 mM and used at a final concentration of 10 µM and 20 µM, respectively. Disulfiram was purchased from Tocris Biosciences, dissolved in DMSO to generate a stock solution at 20 mM. During inhibition experiments, Disulfiram was present at a concentration of 50 µM. To inhibit pyroptosis, cells were pre-treated with inhibitors for 30 min and the respective inhibitor was present during signal 2 of inflammasome stimulation. Propidium iodide solution (1 mg/ml) was purchased from MilliporeSigma. Recombinant human M-CSF (CHO expressed, carrier-free) was bought from R&D Systems and dissolved in sterile PBS pH 7.4 (stock concentration 50 µg/ml). YVAD-pNA was from Enzo Biosciences and the stock solution was prepared at 20 mM in DMSO.

Devant P., Cao A., & Kagan J.C. (2020). Evolution-inspired dissection of caspase activities enables the redesign of caspase-4 into an LPS sensing interleukin-1 converting enzyme.

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Electrochemical Recordings and Pharmacological Manipulations in Drosophila Larvae

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All chemicals were purchased from Sigma-Aldrich (St. Louis, MO), and solutions were prepared in Milli Q water (Millipore, Billerica, MA) unless noted otherwise. Electrode calibrations were performed in phosphate buffer solution (PBS) (131.25 mM NaCl, 3.0 mM KCl, 10.0 mM NaH₂PO₄, 1.2 mM MgCl₂, 2.0 mM Na₂SO₄, and 1.2 mM CaCl₂) with pH adjusted to 7.4, which was made once a month and stored at 4°C. To make the larval dissection buffer, 11.1 mM glucose and 5.3 mM trehalose were added to the PBS buffer on the day of the experiment. For P2X₂ experiments, ATP solution (0.1 mM) was also prepared daily in the dissection buffer. Ten mM stock solutions of octopamine were prepared in 0.1 M HClO₄ once a month and stored at 4°C. Diluted octopamine solutions for electrode calibration and pressure ejection experiments were prepared daily in PBS buffer.
Disulfiram (Tocris Bioscience, Ellisville, MO) and reserpine (Sigma-Aldrich, St. Louis, MO) stock solutions of 10 mM were prepared in DMSO, and diluted to 400 μM in PBS. Fluoxetine (400 μM) and nisoxetine (80 μM) were prepared in PBS. To add drug to the VNC, 1 mL of the respective solution was added to the Petri dish that contained 3 mL of dissection buffer.

Pyakurel P., Privman E, & Venton B.J. (2016). Fast-scan cyclic voltammetry (FSCV) detection of endogenous octopamine in Drosophila melanogaster ventral nerve cord. ACS chemical neuroscience, 7(8), 1112-1119.

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Morphine Alkaloid Pellet Protocol

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Morphine alkaloid pellets and placebo pellets were obtained from the National Institute on Drug Abuse (Bethesda, MD, United States). Morphine hydrochloride, (Macfarlan Smith, United Kingdom), D-penicillamine, disulfiram (both from Tocris United Kingdom) were all dissolved as appropriate in sterile saline. Acetaldehyde (Sigma Aldrich United Kingdom), was diluted (w/v) into sterile saline, using only glassware.

Hill R., Conibear A., Dewey W., Kelly E, & Henderson G. (2022). Role of Acetaldehyde in Ethanol Reversal of Tolerance to Morphine-Induced Respiratory Depression in Mice. Advances in drug and alcohol research, 1, adar.2021.10143.

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Giardicidal Activity and Cytotoxicity Assay

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The following inhibitors were purchased: Canavanine and metronidazole (Sigma Aldrich); vincristine, epigallocatechin gallate, and SNX-2112 (Selleck Chemicals); disulfiram (Tocris); and conoidin A (Cayman Chemical). For assays of giardicidal activity, serial 1:3 dilutions of these compounds were made in 96-well plates, G. lamblia WB or GS/M trophozoites were added, and cultures were grown for 1–2 days at 37°C under anaerobic conditions [29 (link)]. Parasite cell growth and viability were determined by measuring ATP levels with the BacTiter-Glo microbial cell viability assay reagent (Promega) in a microplate reader. The 50% effective concentration (EC50) was derived from the concentration-response curves using BioAssay software (Cambridge soft). Acute human cytotoxicity was assayed with the human epithelial cell line, HeLa (ATCC CCL-2) [29 (link),30 (link)]. Compounds were serially diluted (1:3) and added to HeLa cell cultures in 96-well plates. Cells were grown for two days, and viable cell numbers were determined using AlamarBlue reagent (Invitrogen). As done for the EC50 calculations, the 50% cytotoxic concentration (CC50) was derived from the normalized concentration-response curves using BioAssay software (Cambridge soft).

Lauwaet T., Miyamoto Y., Ihara S., Le C., Kalisiak J., Korthals K.A., Ghassemian M., Smith D.K., Sharpless K.B., Fokin V.V, & Eckmann L. (2020). Click chemistry-facilitated comprehensive identification of proteins adducted by antimicrobial 5-nitroimidazoles for discovery of alternative drug targets against giardiasis. PLoS Neglected Tropical Diseases, 14(4), e0008224.

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Automated Quantification of Neutrophil Extracellular Traps

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WT iNeutrophils, GATA1 KO and primary neutrophils were plated in IMDM (Gibco, 21056023) at a density of 5 x 10⁴ cells per well in ultra-low base 384 well dishes (Aurora, ABD241001A) and stimulated using 50 nM PMA (Sigma-Aldrich, P1585), 25 μg/ml LPS O128:B12, (Sigma-Aldrich, L2887) or 5 μM A23187 (Sigma-Aldrich, C7522) for 3 hours at 37°C. For inhibition studies, cells were pre-treated with 20 μM sotrastaurin (Selleck Chemicals, S2791), 20 μM DPI (Sigma-Aldrich, D2926), 100 μM 4-ABAH (Sigma-Aldrich, A41909) or 20 μM disulfiram (Tocris, 3807) for 1 hour followed by 3 hours of stimulation using 50 nM PMA. After the 3 hour stimulation, a fix/perm/stain solution was added for a final concentration of 2% paraformaldehyde (Electron Microscopy Services, 15710), 0.1% Triton X100 (Sigma-Aldrich, X100-100ML) and 50 nM Sytox Green (Invitrogen, S7020). Nine fields per well were imaged using the Yokogawa CV8000 automated microscope at 20x magnification. Image features were extracted using CellProfiler followed by analysis using custom supervised machine learning software to classify NET versus non-NET nuclei based on nuclei features including size, shape, intensity, etc. Experiments were performed on at least three independent differentiations and three independent donors in at least technical triplicates.

Harper T.C., Oberlick E.M., Smith T.J., Nunes D.E., Bray M.A., Park S., Driscoll C.D., Mowbray S.F, & Antczak C. (2023). GATA1 deletion in human pluripotent stem cells increases differentiation yield and maturity of neutrophils. iScience, 26(10), 107804.

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Morphine Alkaloid Pellet Protocol

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Synthesis and Characterization of Ophiobolin Analogues

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Curcumin, genestein, doxorubicin, and paclitaxel were obtained from Selleckchem (Houston, TX, USA), salinomycin from Cayman Chemicals (Ann Arbor, MI USA), and disulfiram from Tocris Bioscience, (Bristol, UK). OpA was produced by fermentation of the fungus D. gigantea. It was extracted from the fungal culture filtrates, purified, crystallized and identified by ¹H NMR and ESI MS spectra as previously reported^{70 (link)}. The purity of OpA was > 98% as ascertained by ¹H NMR and HPLC analyses.
3-Deoxy OpA was synthesized from ophiobolin I^{71 (link),72 (link)} which was also obtained through fermentation as previously reported^{70 (link)}. A two-step synthetic sequence involving conjugate reduction of the enone which proceeded with high diastereoselectivity (> 19:1 by 600 MHz ¹H NMR) followed by a Ru(IV)-mediated oxidation of the primary alcohol to the aldehyde delivered 3-deoxy OpA. It should be noted that the methyl group at C3 is epimeric with respect to the C3-methyl group in OpA. However, the importance of the C3-hydroxy group and/or the stereochemistry of this methyl group was verified through studies described below and 3-deoxy OpA served as a negative control. Further details are provided in Supplemental Figure 5.

Reisenauer K.N., Tao Y., Das P., Song S., Svatek H., Patel S.D., Mikhail S., Ingros A., Sheesley P., Masi M., Boari A., Evidente A., Kornienko A., Romo D, & Taube J. (2021). Epithelial-mesenchymal transition sensitizes breast cancer cells to cell death via the fungus-derived sesterterpenoid ophiobolin A. Scientific Reports, 11, 10652.

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Cocaine and Isoflavone Interactions in Mice

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Cocaine hydrochloride was obtained from the Spanish Agency of Medicines and Medical Devices (AEMPS, Madrid, Spain), dissolved in sterile saline, and administered at 0.5 mg/kg (intravenously, i.v.). All natural isoflavones (daidzin, daidzein, and genistein) were purchased from LC Laboratories (Woburn, MA) or PhytoLab (Vestenbergsgreuth, Germany), dissolved in 50 μl of Tween 80/0.5% carboxymethylcellulose (CMC) in sterile saline and administered at different doses by intraperitoneal (i.p.) route. Disulfiram was purchased from TOCRIS (Bio-Techne, Minneapolis, MN), dissolved in 50 μl of Tween 80/0.5% CMC in sterile saline and administered at different doses by i.p. route. Phenylephrine (1 mg/kg, subcutaneous (s.c.), not permeable to cross the Blood Brain Barrier), atipamezole (0.4 mg/kg, s.c.), isoproterenol (0.25 mg/kg, i.p.), SKF38393 (0.1 mg/kg, s.c.), quinpirole (0.01 mg/kg, i.p.) and tamoxifen (1 mg/kg, i.p.) were purchased from TOCRIS and dissolved in 50 μl of Tween 80/0.5% CMC in sterile saline.

Martin M., Gutiérrez-Martos M., Cabrera R., Langohr K., Maldonado R., Farre M, & de la Torre R. (2021). Daidzein modulates cocaine-reinforcing effects and cue-induced cocaine reinstatement in CD-1 male mice. Psychopharmacology, 238(7), 1923-1936.

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