The mouse hindpaw plantar incision model was created as described previously [36 (link)]. Mice were deeply anesthetized by inhalation of 1.5–2.0% isoflurane (Abbott, Tokyo, Japan) via a nose cone. A 5-mm longitudinal incision was made with a No. 11 blade through the skin and fascia of the plantar foot. The incision was started 2 mm from the proximal edge of the heel and extended toward the toes. The underlying muscle was elevated with a curved forceps leaving the muscle origin and insertion intact. The skin was apposed with a single mattress suture of 8–0 nylon. Morphine (Shionogi & Co. LTD., Japan) was dissolved in phosphate-buffered saline (PBS, pH 7.2), SnPP (Tocris Bioscience, Bristol, UK) was diluted in dimethyl sulfoxide (DMSO), and indomethacin (Nacalai Tesque, Kyoto, Japan) was diluted in Tris buffer (TB, pH8.0). Morphine (3 μg/20 μL or 10 μg/20 μL), naloxone (5 μg/20 μL, Wako, Osaka, Japan), SnPP (400 nmol/20 μL) or indomethacin (50 μg/20 μL) were injected locally to the incisional sites 1 hour after the skin was sutured, and on PODs 1 and 2, or on PODs 5–7. The total amount of solution injected to the hind paws was 20 μL/paw for all experiments. The suture was removed at the end of POD2.
Indomethacin
Manufactured by Nacalai Tesque
Sourced in Japan, United States
Indomethacin is a type of lab equipment used for analytical and research purposes. It is a non-steroidal anti-inflammatory drug (NSAID) that can be used to measure and study inflammatory responses in various experimental models.
Automatically generated - may contain errors
Lab products found in correlation
Indomethacin, by Fujifilm (1 mentions)
Isoflurane, by Abbott (1 mentions)
Kainic acid, by Cayman Chemical (1 mentions)
Fluoro jade c, by Biosensis (1 mentions)
Liraglutide, by Novo Nordisk (1 mentions)
Insulin, by Nacalai Tesque (1 mentions)
Fetal bovine serum (fbs), by GE Healthcare (1 mentions)
3 isobutyl 1 methyl xanthine (ibmx), by Merck Group (1 mentions)
Dulbecco modified eagle medium (dmem), by Merck Group (1 mentions)
Dulbecco modified eagle medium (dmem), by Nacalai Tesque (1 mentions)
Insulin, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Dexamethasone (dex), by Nacalai Tesque (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Insulin, by Thermo Fisher Scientific (1 mentions)
Triiodothyronine t3, by Merck Group (1 mentions)
3 isobutyl 1 methylxanthine ibmx, by Thermo Fisher Scientific (1 mentions)
Dmem f12, by Thermo Fisher Scientific (1 mentions)
Collagenase type 1, by Worthington (1 mentions)
Epilife basal medium, by Thermo Fisher Scientific (1 mentions)
9 protocols using indomethacin
1
Mouse Hindpaw Incision Pain Model
2
Kainic Acid-induced Neurodegeneration Model
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Kainic acid (KA, 78050, Cayman Chemicals, Ann Arbor, MI), indomethacin (IND, 19233-51, Nacalai tesque, Tokyo, Japan), liraglutide (2499410G1021, Novo Nordisk, Bagsværd, Danmark), and sitagliptin phosphate monohydrate (SPM, A4036, ApexBio, Boston, MA, USA) were used for animal treatments. Fluoro Jade C (FJC, TR-100-FJ, Biosensis, CA) was used for staining of degenerating neurons. Pre-stained Protein Marker (02525-35, Nacalai tesque, Tokyo, Japan) was used for Western blots. MK-0524 (DP1 antagonist, 1480, Axon MEDCHEM, Groningen, Netherlands), OC000459 (DP2 antagonist, 1913, Axon MEDCHEM, Groningen, Netherlands) were used as DP selective inhibitors.
3
Adipogenic Differentiation Protocol
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Cells were seeded on 6-well tissue culture dishes at a density of 1.0 x106 cells/well, and adipogenic differentiation was initiated by three cycles of an induction/maintenance culture as previously described [33 (link)]. Each cycle consisted of a 3-day culture in induction medium (DMEM (Nacalai Tesque) containing 10% FBS (GE Healthcare), 1 μM dexamethasone (WAKO), 10 μg/ml insulin (Nacalai Tesque), 0.2 mM indomethacin (Nacalai Tesque), and 0.5 mM IBMX (Sigma)), followed by a 3-day culture in maintenance medium (DMEM containing 10% FBS and 10 μg/ml insulin). After an 18-day induction, lipid vacuoles were visualized using Oil Red O staining.
4
Differentiation of Brown Adipocytes
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The interscapular BAT of Sprague-Dawley rats (4 weeks old, male, Tokyo Laboratory Animals Science Co.) was removed and digested in Hanks’ solution (Worthington Biochemical Co., Lakewood, NJ, USA) containing 3.5% bovine blood albumin (Nacalai Tesque, Kyoto, Japan) and 2 mg/mL of collagenase type I (Worthington Biochemical Co.) for 30 min at 37 °C. Brown progenitor adipocytes were obtained by centrifugation at 1000× g for 5 min. Cells were suspended in Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12 Ham (DMEM/F-12; Invitrogen, Waltham, MA, USA) containing 10% fetal bovine serum (Invitrogen) at a density of 800,000 cells/mL and cultured in a 35 mm dish. When the cells were 70–80% confluent, the medium was supplemented with 1 nM triiodothyronine (T3: Sigma-Aldrich, St. Louis, MO, USA), 25 μM insulin (Invitrogen), 0.125 mM indomethacin (Nacalai Tesque), 5 μM dexamethasone (Nacalai Tesque), and 0.5 mM 3-isobutyl-1-methylxanthine (IBMX; Invitrogen). After 48 h, the medium was replaced with 1 nM T3 and 25 μM insulin and the culture was continued. As a control, non-induced cells were cultured in DMEM/F-12 without induction of differentiation. After 72 h, cells were treated with isoproterenol or siRNA.
5
Epithelial Cell Culture and Stimulation
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hOMK were purchased from Cell Research Corp. (Singapore, Republic of Singapore) and cultured in an epithelial culture medium (EpiLife basal medium supplemented with EpiLife Defined Growth Supplement, Thermo Fisher Science, Waltham, MA, USA), at 37 °C in a humidified atmosphere of 95% air and 5% CO2. hOMK were seeded at a density of 1, 3 × 104, 1 × 105, or 1 × 106 cells/well in 96-well plates, 24-well plates, or six-well plates, depending on the experiment, and then stimulated with 5-FU ((0.5 μg/mL or 0.01–10 μg/mL) (Kyowa Kirin, Tokyo, Japan)) and β-cry ((1 × 10−7 M) (CaroteNature, Lupsingen, Switzerland)). In some experiments, the combination of LPS ((1 μg/mL) from P. gingivalis (InvivoGen, San Diego, CA, USA)), RA ((10 nM) (Sigma, St. Louis, MI, USA)) and indomethacin ((10 μg/mL) (Nacalai, Kyoto, Japan)) were also used and dissolved in DMSO.
6
Modulating Inflammation in Acute Colitis
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Acute colitis was induced by 3.5% (w/v) DSS (molecular weight, 5000; Fujifilm, Osaka, Japan) added to the drinking water for 7 days. Seven- to nine-week-old gender-matched mice were included in each group. Body weight was recorded daily. Mice were sacrificed on day 3 or 7, and the colons were collected. To inhibit the NLRP3 inflammasome in vivo, mice received an intraperitoneal injection of 50 mg/kg MCC950 (AdipoGen Life Sciences, San Diego, CA) (or PBS as a control) every other day from one day before to day 7 of DSS administration. To inhibit PGE2 production in vivo, indomethacin (Nacalai Tesque, Kyoto, Japan) was dissolved in ethanol (10 mg/mL) and was added to the drinking water at 1 mg/kg per day, concurrent with DSS treatment.
7
Detailed Ion Channel Inhibitor Protocol
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All chemicals employed were reagent grade. Amiloride and bumetanide were obtained from Sigma (St. Louis, MO). Sodium butyrate and sodium formate were obtained from Wako Chemicals (Osaka, Japan). HEPES, indomethacin, and ibuprofen were obtained from Nacalai Tesque (Kyoto, Japan). XE991 (10, 10-bis (4-pyridinylmethyl)-9(10H)-anthracenonedihydrochloride) was obtained from Alomone Labs (Israel). Chromanol 293B and bupivacaine were obtained from Tocris Bioscience (Bristol, UK) and Tokyo Chemical Industry (Tokyo, Japan), respectively. Stock solutions of Amiloride (10 mM), XE991 (100 mM), and barium chloride (1 M) were prepared in distilled water. indomethacin (5 mM) and ibuprofen (300 mM) were dissolved in ethanol. bumetanide (100 mM), Chromanol 293B (100 mM), and bupivacaine (300 mM) were dissolved in dimethyl sulfoxide (DMSO). Stock solutions were prepared at a 1000-fold concentration for administration, except for barium chloride at a 200-fold concentration.
8
Agonist-Antagonist Pharmacological Profiling
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The following reagents were all obtained from Sigma-Aldrich (St. Louis, MO, USA) and used
at the indicated final concentrations:
5-HT (10−9–10−5 M); ketanserin tartrate
(10−8–10−7 M); methiothepin maleate
(10−8–10−7 M); His hydrochloride (10−6–10−3M); diphenhydramine hydrochloride (10−7–10−4 M); cimetidine
(10−5 M); Ang II acetate salt (10−9–10−5 M); NA
(10−9–10−5 M); phentolamine mesilate (10−5 M);
propranolol hydrochloride (10−8–10−6 M); BK acetate salt
(10−9–10−6 M); des-Arg9- [Leu8]-BK
(10−5 M); Nω-nitro-L-arginine (L-NNA; 10−4 M);
and sodium nitroprusside (SNP, 10−4 M).
The following reagents were obtained and used at the indicated final concentrations:
HOE140 (10−8–10−6 M; Peptide Institute, Osaka, Japan); indomethacin
(10−5 M; Nacalai tesque, Kyoto, Japan); ACh chloride
(10−9–10−5 M; Daiichi Sankyo, Tokyo, Japan). All drugs were
dissolved in distilled water.
at the indicated final concentrations:
5-HT (10−9–10−5 M); ketanserin tartrate
(10−8–10−7 M); methiothepin maleate
(10−8–10−7 M); His hydrochloride (10−6–10−3M); diphenhydramine hydrochloride (10−7–10−4 M); cimetidine
(10−5 M); Ang II acetate salt (10−9–10−5 M); NA
(10−9–10−5 M); phentolamine mesilate (10−5 M);
propranolol hydrochloride (10−8–10−6 M); BK acetate salt
(10−9–10−6 M); des-Arg9- [Leu8]-BK
(10−5 M); Nω-nitro-L-arginine (L-NNA; 10−4 M);
and sodium nitroprusside (SNP, 10−4 M).
The following reagents were obtained and used at the indicated final concentrations:
HOE140 (10−8–10−6 M; Peptide Institute, Osaka, Japan); indomethacin
(10−5 M; Nacalai tesque, Kyoto, Japan); ACh chloride
(10−9–10−5 M; Daiichi Sankyo, Tokyo, Japan). All drugs were
dissolved in distilled water.
9
Screening of Small Molecule Compounds
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Esmolol hydrochloride, dequalinium dichloride, fosinopril, antimycin A, and oxprenolol hydrochloride were purchased from Santa Cruz Biotechnology. Alexidine dihydrochloride, merbromin, candesartan, amphotericin B, alprenolol hydrochloride, triprolidine hydrochloride, methacycline hydrochloride, cefixime, and ethacrynic acid were purchased from Sigma-Aldrich. Demecarium bromide and cefoperazone were purchased from AK Scientific. Bisoprolol fumarate was purchased from MedChem Express. Benzbromarone, etifenin, cefotetan, and rebamipide were purchased from Tokyo Chemical Industry Co., Ltd. Atractyloside potassium salt was purchased from Toronto Research Chemicals. Furosemide and indomethacin were purchased from Nacalai Tesque. Acemetacin, ketoprofen, bumetanide, colistin sulfate, tranilast, norfloxacin, and doxepin hydrochloride were purchased from LKT Laboratories, Inc.
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