Serotonin 5-HT2C receptor competition binding experiments were carried out in membranes from HeLa-5-HT2C cells prepared in our group. On the day of assay, membranes were defrosted and resuspended in binding buffer (50 mM Tris-HCl, pH 7.5). Each reaction well of a 96-well plate contained 15 μg of protein, 4 nM [3H]Mesulergine (83.5 Ci/mmol, Perkin-Elmer, Waltham, MA, USA), and different concentrations of the compounds in the range from 0.01 nM to 10µM. Non-specific binding was determined in the presence of 10 μM mianserin (Sigma Aldrich, Spain). The reaction mixture was incubated at 27 °C for 1 h, after which samples were transferred to multiscreen GF/C 96-well plates (Millipore, Spain) pretreated with 0.5% polyethylenimine (PEI, Sigma Aldrich, Spain), filtered, and washed four times with 250 μL wash buffer (50 mM Tris-HCl, pH 6.6). The filters were dried, 35 μL Universol (MP Biomedicals, Spain) per well were added and radioactivity was measured as described above.
Mianserin
Manufactured by Merck Group
Sourced in Spain, Switzerland, Poland, United States
Mianserin is a laboratory product manufactured by Merck Group. It is a heterocyclic compound used in research applications.
Automatically generated - may contain errors
Lab products found in correlation
3h mesulergine, by PerkinElmer (2 mentions)
Multiscreen gf c 96 well plate, by Merck Group (1 mentions)
Citalopram, by Lipomed (1 mentions)
Mazindol, by Lipomed (1 mentions)
Tramadol, by Lipomed (1 mentions)
Venlafaxine, by Lipomed (1 mentions)
Fluoxetine, by Lipomed (1 mentions)
Nisoxetine, by Merck Group (1 mentions)
Pindolol, by Merck Group (1 mentions)
Spiperone, by Merck Group (1 mentions)
3h ketanserin, by PerkinElmer (1 mentions)
Pargyline, by Merck Group (1 mentions)
Hydrocodone, by Lipomed (1 mentions)
Hydromorphone, by Lipomed (1 mentions)
Oxycodone, by Lipomed (1 mentions)
Oxymorphone, by Lipomed (1 mentions)
Codeine, by Lipomed (1 mentions)
3h 8 oh dpat, by PerkinElmer (1 mentions)
Morphine, by Lipomed (1 mentions)
Dextromethorphan, by Lipomed (1 mentions)
4 protocols using mianserin
1
Serotonin 5-HT2C Receptor Binding Assay
2
Radioligand Binding Assay Protocol
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Buprenorphine , citalopram, codeine , dextromethorphan , dihydrocodeine , fentanyl, heroin (diacetylmorphine, diamorphine) , 6‐acetylmorphine (6‐mono‐acetylmorphine), hydrocodone , hydromorphone, mazindol, MDMA, methadone , morphine, oxycodone , oxymorphone , pethidine (meperidine), tramadol, O‐desmethyl‐cis‐tramadol, tapentadol, venlafaxine and fluoxetine were purchased from Lipomed (Arlesheim, Switzerland). Mianserin, nisoxetine, pargyline, pindolol and spiperone were supplied by Sigma‐Aldrich (Buchs, Switzerland). D(S)‐methadone and l(R)‐methadone were obtained from Alsachim (Illkirch Graffenstaden, France). The HPLC purity of all of the substances was >98%. [3H]‐8‐OH‐DPAT, [3H]‐ketanserin and [3H]‐mesulergine were supplied by Perkin‐Elmer.
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3
Quantification of Antidepressants in Biological Samples
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Standards of fluoxetine (FLU) and mianserin (MNS) as well as internal standards (IS, nortryptyline and doxepin) were obtained from Sigma-Aldrich (Poznań, Poland), while paroxetine (PAR) and sertraline (SER) were gifts from the National Drug Research Institute, Warsaw, Poland. All the drugs were of high purity grade (>90%). The standard stock solutions of all compounds were prepared in methanol at concentrations of 1 mg/mL and stored at −20 °C. Working solutions were prepared ex tempore by dilution of the stock solutions with the culture medium. IS working solution (500 ng/mL) was prepared ex tempore by dilution of the stock solution with acetonitrile. The solvents, namely HPLC gradient grade methanol, MS grade acetonitrile (LiChrosolv) and formic acid 98%, were provided by Merck (Darmstadt, Germany). Ultrapure water was obtained from a Millipore water purification system (Milli-Q water). The pH-dependent octanol-water partition coefficients (log D) were calculated with logD predictor (https://disco.chemaxon.com/apps ).
4
Receptor Binding Assays for Natural Compounds
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Recombinant human membrane preparations for M1 (expressed in CHO cells; Lot no. 1706299), 5-HT1A (HEK293 cells; no. 1812363), 5-HT2A (CHO cells; no. 1813092), D1 (L cells; no. 1840059), and D2S (CHO cells; no. 1820984) receptors and MicroScint-O scintillation cocktail were procured from Perkin Elmer (Waltham, MA, USA). [3H] N-methylscopolamine (NMS), [3H] 8-OH-DPAT, [3H] ketanserin, [3H] SCH 23390 and [3H] methylspiperone were purchased from American Radiolabeled Chemicals (St. Louis, MO, USA). Atropine, 5-carboxamidotryptamine (5-CT), ketanserin, SCH 23390 hydrochloride, haloperidol, serotonin hydrochloride, mianserin, human recombinant acetylcholinesterase expressed in HEK 293 cells, acetylthiocholine iodide (ATChI), 5, 5’-dithiobis [2-nitrobenzoic acid] (DTNB) and donepezil hydrochloride were obtained from Sigma-Aldrich (St. Louis, MO, USA). Bacoside A, bacoside A3, bacopaside II, bacopaside X and bacopasaponin C were purchased from Chromadex Inc. (Irvine, CA, USA). Jujubogenin and ebelin lactone were purchased from Shanghai IS Chemical Technology Ltd. (Jinshan, Shanghai, China). Unless stated otherwise, all other reagents of analytical grade were obtained through standard commercial sources.
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