Urethane, ethyl carbamate, EC, CAS# 51-79-6; 3-methylcholanthrene, 3-methyl-1,2-dyhydrobenzo[j]aceanthrylene, MCA, CAS# 56-49-5; butylated hydroxytoluene, 2,6-Di-tert-butyl-4-methylphenol, BHT, CAS# 128-37-0; naphthalene, CAS# 91-20-3, and Hoechst33258 nuclear dye (CAS# 23491-45-4), were from Sigma-Aldrich (St. Louis, MO). Bleomycin A2, ((3-{[(2'-{(5S,8S,9S,10R,13S)−15-{6-amino-2- [(1S)−3-amino-1-{[(2S)−2,3-diamino-3-oxopropyl]amino}−3-oxopropyl] −5-methylpyrimidin-4-yl}−13-[{[(2R,3S,4S,5S,6S)−3-{[(2R,3S,4S,5R,6R)−4-(carbamoyloxy)−3,5-dihydroxy-6- (hydroxymethyl) tetrahydro-2H-pyran-2-yl]oxy} −4,5-dihydroxy-6-(hydroxymethyl) tetrahydro-2H-pyran-2-yl]oxy} (1H-imidazol-5-yl)methyl]−9-hydroxy-5-[(1R)−1-hydroxyethyl]−8,10-dimethyl-4,7,12,15-tetraoxo-3,6,11,14-tetraazapentadec-1-yl}−2,4'-bi-1,3-thiazol-4-yl)carbonyl]amino}propyl) (dimethyl)sulfonium; CAS #9041-93-4, was from Calbiochem (Darmstadt, Germany). D-Luciferin potassium salt, (4S)−2-(6-hydroxy-1,3-benzothiazol-2-yl)−4,5-dihydrothiazole-4-carboxylic acid, CAS #2591-17-5, was from Gold Biotechnology (St. Louis, MO).
3 methylcholanthrene
Manufactured by Merck Group
Sourced in United States
3-methylcholanthrene is a chemical compound primarily used in research and laboratory settings. It is a polycyclic aromatic hydrocarbon with a specific molecular structure. This product is commonly utilized for various experimental purposes, but a detailed description of its core function is not available while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Benzo a pyrene, by Merck Group (5 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (4 mentions)
Dexamethasone (dex), by Merck Group (3 mentions)
Rifampicin, by Merck Group (3 mentions)
Aflatoxin b1, by Merck Group (2 mentions)
Phenobarbital, by Merck Group (2 mentions)
4 nitroquinoline 1 oxide, by Merck Group (2 mentions)
N hexane, by Avantor (2 mentions)
Methanol, by Avantor (2 mentions)
Sep pak florisil cartridges, by Waters Corporation (2 mentions)
Dichloromethane dcm, by Avantor (2 mentions)
Acetonitrile acn, by Avantor (2 mentions)
Butylated hydroxytoluene, by Merck Group (1 mentions)
Hoechst33258 nuclear dye, by Merck Group (1 mentions)
2 6 di tert butyl 4 methylphenol bht, by Merck Group (1 mentions)
Naphthalene, by Merck Group (1 mentions)
Florisil cartridge, by Waters Corporation (1 mentions)
Aflatoxin mix, by Merck Group (1 mentions)
Acetonitrile, by Waters Corporation (1 mentions)
Acetic acid, by Merck Group (1 mentions)
21 protocols using 3 methylcholanthrene
1
Biochemical Reagents for Cell Research
2
Quantitative Analysis of Food Contaminants
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The reagents used for the SO2 test were methyl red, 30% hydrogen peroxide solution, hydrochloric acid, 0.01 N sodium hydroxide solution (Factor 1.00, Wako, Tokyo, Japan), ethanol (High-performance liquid chromatography (HPLC) grade, J. T. Baker, Phillipsburg, NJ, USA), and nitrogen gas (purity 99.995%). SO2 was extracted as the test apparatus using a Monier-Williams transformation apparatus and an automatic potentiometer (T50M, Mettler-Toledo, Zurich, Switzerland) was used.
Aflatoxin B1, B2, G1, and G2 mixed solutions (Aflatoxin Mix, Sigma, Burlington, MA, USA) were used as standards for the mycotoxin test. Pididium hydrobromic acid perbromate (PBPB, Sigma, USA) was used as the derivatization reagent; methanol (HPLC grade, J. T. Baker, USA), acetonitrile (HPLC grade, J. T. Baker, USA), acetic acid (HPLC grade, J. T. Baker, USA), and Tween 20 were used as solvents. (Bio-Rad, Hercules, CA, USA); and an immunoaffinity column (AflaTest, Vicam, Manchester, TN, USA) was used.
The standard used for the benzopyrene test was benzopyrene (Sigma, USA). Furthermore, 3-methylcholanthrene (Sigma, USA) was used as an internal standard, and n-hexane (HPLC grade, J. T. Baker, USA), acetonitrile (HPLC grade, J. T. Baker, USA), anhydrous sodium sulfate (HPLC grade, J. T. Baker, USA), dichloromethane (J. T. Baker, USA), and a Florisil cartridge (1 g, Sep-Pak® Waters, Bedford, MA, USA) were used.
Aflatoxin B1, B2, G1, and G2 mixed solutions (Aflatoxin Mix, Sigma, Burlington, MA, USA) were used as standards for the mycotoxin test. Pididium hydrobromic acid perbromate (PBPB, Sigma, USA) was used as the derivatization reagent; methanol (HPLC grade, J. T. Baker, USA), acetonitrile (HPLC grade, J. T. Baker, USA), acetic acid (HPLC grade, J. T. Baker, USA), and Tween 20 were used as solvents. (Bio-Rad, Hercules, CA, USA); and an immunoaffinity column (AflaTest, Vicam, Manchester, TN, USA) was used.
The standard used for the benzopyrene test was benzopyrene (Sigma, USA). Furthermore, 3-methylcholanthrene (Sigma, USA) was used as an internal standard, and n-hexane (HPLC grade, J. T. Baker, USA), acetonitrile (HPLC grade, J. T. Baker, USA), anhydrous sodium sulfate (HPLC grade, J. T. Baker, USA), dichloromethane (J. T. Baker, USA), and a Florisil cartridge (1 g, Sep-Pak® Waters, Bedford, MA, USA) were used.
3
Isolation of Murine Immune Cells and Tumor Induction
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Single cells suspensions from spleen and bone marrow of femurs were prepared. Spleens were carefully passed through a 40 μm cell strainer (BD Biosciences, Heidelberg, Germany), washed twice with PBS/2% FCS and erythrocytes were lysed with hypertonic solution (0.17 M NH4Cl with 20 mM HEPES in H2O) for 10 min at 37°C. For isolation of bone marrow cells epicondyles were cut and diaphyses were flushed with 5 ml 1x PBS. Cells were counted with a haemocytometer (A.Hartenstein, Würzburg, Germany). Peripheral blood for FACS-staining was isolated with EDTA-coated microvettes (Sarstedt, Nümbrecht, Germany).
The BFS1 fibrosarcoma cells were generated in a female C57BL/6N mouse by injection of 1 mg of 3-methylcholanthrene (Sigma, Taufkirchen, Germany) dissolved in 200 μl tricaprylin (Sigma) i.d. in the back of a mouse as described [35 (link)]. Syngeneic tumor cells (BFS1, 2x105 in 50μl PBS) were subcutaneously implanted on the upper part of the loin. The diameters of the tumors were measured in two directions perpendicularly to each other with a caliper.
The BFS1 fibrosarcoma cells were generated in a female C57BL/6N mouse by injection of 1 mg of 3-methylcholanthrene (Sigma, Taufkirchen, Germany) dissolved in 200 μl tricaprylin (Sigma) i.d. in the back of a mouse as described [35 (link)]. Syngeneic tumor cells (BFS1, 2x105 in 50μl PBS) were subcutaneously implanted on the upper part of the loin. The diameters of the tumors were measured in two directions perpendicularly to each other with a caliper.
4
3D Spheroid Compound Screening Assay
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Acetaminophen, 3-methylcholanthrene, and aflatoxin B1 were purchased from Sigma-Aldrich. The compounds were dissolved in dimethyl sulfoxide (DMSO, Sigma-Aldrich) and diluted with culture medium. This preparation was carried out immediately before use every time. The final concentration of DMSO was less than 0.1%. After culture day 6, 3D spheroids were exposed to each compound, and culture medium containing 300 μM D-luciferin and compound was refreshed every 2 or 3 days. Bioluminescence from the 3D spheroids was measured nondestructively for 5 s in each well using Phelios (ATTO). Reproducibility of the luminescence measurement during the experimental period was confirmed with the standard LED plate TRIANT (WSL-0001, ATTO).
5
Generation of NCF1* Sarcoma Mouse Model
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NCF1m1J mutant mice (NCF1*) NCF1* mice were previously backcrossed into C57Bl/10.Q/rhd mice. They were genotyped as previously described and crossed to generate heterozygous NCF1*/+ and homozygous NCF1*/* mutant mice [22 (link)]. To induce sarcoma tumorigenesis, mice (approximately 20 g, 8–16 weeks old) were treated by intramuscularly injection of 0.025 ml of an emulsion consisting of 3-methylcholanthrene (Sigma-Aldrich)-Corn oil (5mg/ml) in the right hind leg. Mice were observed weekly for the development of intramuscular tumor development from 70–160 days. Tumors were harvested and mice sacrificed as described below. From harvested tumors, cell lines were established using typical mechanical dissociation and Collagenase IV/DNAase/Trypsine treatment followed by single cell suspension generation by passage through filter prior to seeding into T-25 flasks with complete RPMI medium containing 10% FBS 10mM nonessential amino acids, 50 units/ml penicillin, 50 μg/ml streptomycin.
6
Cyst-enriched Organoid Differentiation and Induction Assay
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Conventional organoids/cyst-enriched organoids were dissociated into single cells using TrypLE Select Enzyme (10×). After a filtration by 70 µm cell strainer and centrifugation, the cell pellet was resuspended in cyst-enriched medium and adjusted to 3.5 × 105 cells/mL. 200 µL of cell suspension was seeded on Transwell (Corning, Cat#3378 or 3470) coated with Matrigel diluted 50-fold with cold-PBS at 37 °C for 2 h, and medium was added to the basal compartment. On day 2 or 3, the medium was changed to the differentiation medium which consisted of basal medium containing 1 × B-27 Supplement, 1 mM N-acetyl-cysteine, 10 nM Gastrin, 100 ng/ml Noggin, 0.5 µg/ml R-spondin 1, 500 nM A83-01, 10 µM Y-27632, 3 mM VPA (Abcam). The medium was replaced every 2 or 3 days. For the induction assay, differentiation medium containing 50 µM β-naphtoflavone (Sigma), 5 µM 3-methylcholanthrene (Sigma), or 100 nM 1α, 25-dihydroxyvitamin D3 (Sigma) was exposed for 48 h. CYP1A activity and CYP3A activity were analyzed by P450-Glo Assays according the manufacturer’s protocol (luciferin-CEE for CYP1A, luciferin-IPA for CYP3A, Promega). The monolayer integrity was assessed by TEER. The TEER value was measured using Millicell ERS-2 system (Merck).
7
Ligand Screening for Nuclear Receptors
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Compound 1 (CITCO, (6-(4-chlorophenyl)imidazo [2,1-b][1,3]thiazole-5-carbaldehyde-O-(3,4-dichloro-benzyl)oxime)),
rifampicin (rif), TCPOBOP, and PK11195 were obtained from Sigma-Aldrich
(St. Louis, Missouri, United States, now Merck), which is now known
as Merck (Darmstadt, Germany). Phenobarbital (Luminal 200 mg/mL injection)
was manufactured by Desitin Pharma spol. s.r.o. (Prague, Czech Republic).
Ligands for nuclear receptors (GW3965, thyroxin, obeticholic acid,
dexamethasone, fenofibrate, GW501516, rosiglitazone, 3-methylcholanthrene,
calcitriol, testosterone, and estradiol) were purchased from Sigma-Aldrich
(now Merck). The prototype ligands were used at 100 nM (dexamethasone,
calcitriol), 1 μM (thyroxin), or 10 μM concentrations.
The compounds were dissolved in DMSO, and the final concentration
of DMSO in the entire reaction mixture or cultivation media was 0.1%.
rifampicin (rif), TCPOBOP, and PK11195 were obtained from Sigma-Aldrich
(St. Louis, Missouri, United States, now Merck), which is now known
as Merck (Darmstadt, Germany). Phenobarbital (Luminal 200 mg/mL injection)
was manufactured by Desitin Pharma spol. s.r.o. (Prague, Czech Republic).
Ligands for nuclear receptors (GW3965, thyroxin, obeticholic acid,
dexamethasone, fenofibrate, GW501516, rosiglitazone, 3-methylcholanthrene,
calcitriol, testosterone, and estradiol) were purchased from Sigma-Aldrich
(now Merck). The prototype ligands were used at 100 nM (dexamethasone,
calcitriol), 1 μM (thyroxin), or 10 μM concentrations.
The compounds were dissolved in DMSO, and the final concentration
of DMSO in the entire reaction mixture or cultivation media was 0.1%.
8
Optimizing Cell Culture Conditions
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Dulbecco’s modified Eagle’s medium (DMEM), phosphate-buffered saline (PBS), Hank’s buffered salt solution, Earle’s balanced salt solution, penicillin, streptomycin, human plasma fibronectin, and trypsin-ethylenediaminetetraacetic acid were purchased from Invitrogen Life Technologies, Carlsbad, CA. Type IV collagenase, HEPES (4-[2-hydroxyethyl] piperazine-1-ethanesulfonic acid), glucagon, calcium chloride, hydrocortisone, sodium dodecyl sulfate, hydrogen peroxide, glutaraldehyde, dicumarol, 3-methylcholanthrene, calf thymus DNA, chitosan, and hyaluronic acid (HA) was purchased from Sigma-Aldrich, St. Louis, MO. All other chemicals were purchased from Fisher Scientific, Waltham, MA, unless otherwise noted.
9
CYP1A induction in mouse liver
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Young adult male C3H/HeN mice (n = 5–6 per group) were treated with the AhR inducer 3-methylcholanthrene (Sigma; dissolved in corn oil) by a single i.p. injection of 10, 25 or 50 mg/kg body weight 48 h prior to sacrifice. Mice were killed between 9 and 11 a.m. to avoid circadian variations; livers were excised, transferred to Carnoy's fixative and subsequently embedded in paraffin. Tissue slices of 5 μm thickness were stained for CYP1A as recently described (61 (link)) using a rabbit antiserum at 1:500 dilution (gift of Dr R. Wolf, University of Dundee, UK) and a horseradish peroxidase-conjugated secondary antibody (1:100; Dako, Glostrup, Denmark) with the substrates 3-amino-9-ethylcarbazole/H2O2. Mice had access to tap water and standard chow ad libitum. All animals received humane care and protocols complied with institutional guidelines. Width of CYP1A-positive zones was assessed using an AxioImager light microscope and AxioVision software (Zeiss, Oberkochen, Germany).
10
Organoid-Based Cytochrome P450 Induction
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Standard multicellular organoids (1,200 cells/ organoid) were formed in round bottomed 96 well plates. For comparison, 2D cultures were formed by plating 105 cells (in the same proportions used to form the organoids) on matrigel coated flat bottomed 96 well plates to form a confluent layer. Drug toxicity in the organoids and monolayer cultures was then assessed by inducing cytochrome P450 activity using a mixture of rifampicin (25mM), 3-methylcholanthrene (3.78μg/mL), and phenobarbital (58.0μg/mL) (Sigma) in HCM medium (Lonza), inducing the cells for 24 hours. Then diazepam was added (2.5μg/mL) in HCM medium for 24 hours. Diazepam metabolites temazepam, nordiazepam, and oxazepam were measured in the cell supernatant using a Quantum Discovery Max triple quadrupole mass spectrometer (Thermo/Fisher)
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