α monothioglycerol

Manufactured by Merck Group

Sourced in United States, Canada

α-monothioglycerol is a chemical compound used in various laboratory applications. It serves as a reducing agent and exhibits antioxidant properties. The core function of this product is to facilitate chemical reactions and provide a means to maintain a controlled redox environment in laboratory settings.

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Lab products found in correlation

9 protocols using α monothioglycerol

Clonogenic Assay for Retrovirally Infected Cells

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Retrovirally infected bone marrow cells were plated in duplicates at two concentrations (5 × 10⁴ or 2 × 10⁵ cells per ml) in methylcellulose medium (IMDM (Gibco), 10% fetal bovine serum (FBS), 1% methylcellulose (Fluka), 2% bovine serum albumin (BSA), 100 ng ml⁻¹ stem cell factor (SCF), 50 mM α-monothioglycerol (Sigma), 200 μg ml⁻¹ plasma transferrin, 1 U ml⁻¹ recombinant erythropoietin, and IL-3), with or without neomycin (G148; Gibco) 1 mg ml⁻¹ for 7–10 days. neomycin-resistant colonies were scored and classified^{73 (link)}. For serial replating assays, colony cells were collected, pooled and plated at 5 × 10³ cells ml⁻¹ under the same conditions but without further neomycin selection. Secondary colonies were scored and subjected to tertiary and quaternary replating assays. Colonies were scored using a Leitz Labovert inverted microscope (Leitz Wetzlar).

Smith M.J., Ottoni E., Ishiyama N., Goudreault M., Haman A., Meyer C., Tucholska M., Gasmi-Seabrook G., Menezes S., Laister R.C., Minden M.D., Marschalek R., Gingras A.C., Hoang T, & Ikura M. (2017). Evolution of AF6-RAS association and its implications in mixed-lineage leukemia. Nature Communications, 8, 1099.

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Maintenance of HPC-7 Hematopoietic Progenitor Cells

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The hematopoietic progenitor cell line HPC-7 was kindly gifted by Dr. Britta Will at Albert Einstein College of Medicine. HPC-7 cells were maintained at a density of 2–10 × 10⁵/ml in Iscove’s modified Dulbecco’s medium (Invitrogen) supplemented with 50 ng/ml mouse stem cell factor (Gemini Bio-Products), 1 mM sodium pyruvate (Invitrogen), 6.9 ng/mL α-monothioglycerol (Sigma-Aldrich), 5% bovine calf serum and penicillin‒streptomycin (Invitrogen).

Bera B.S., Thompson T.V., Sosa E., Nomaru H., Reynolds D., Dubin R.A., Maqbool S.B., Zheng D., Morrow B.E., Greally J.M, & Suzuki M. (2023). An optimized approach for multiplexing single-nuclear ATAC-seq using oligonucleotide-conjugated antibodies. Epigenetics & Chromatin, 16, 14.

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Murine Hematopoietic Cell Culture

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Bone marrow mononuclear cells isolated from the femurs and tibiae were cultured for two weeks in Roswell Park Memorial Institute (RPMI) 1640 medium supplemented with 10% FBS and penicillin-streptomycin in the presence of 10 ng/mL recombinant murine IL-3 (PeproTech, Rocky Hill, NJ, USA) and subsequently cultured for two weeks with IL-3 and 10 ng/mL recombinant murine stem cell factor (Peprotech). The 293T cells (Thermo Fisher Scientific) were cultured in Dulbecco modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and penicillin-streptomycin. MEDMC-BRC6 cells [23 (link)] were purchased from RIKEN BRC (Tsukuba, Japan) and cultured in Iscove’s modified Dulbecco’s medium (IMDM; Invitrogen, Thermo Fisher Scientific, Waltham, MA, USA) containing the following: 15% FBS, ITS liquid media supplement (Sigma-Aldrich, St. Louis, MO, USA), 50 mg/mL ascorbic acid (Sigma-Aldrich), 0.45 mM α-monothioglycerol (Sigma-Aldrich), 3 ng/mL IL-3, 30 ng/mL stem cell factor (SCF), 1% penicillin-streptomycin solution and 2 mM l-glutamine (Nacalai Tesque, Kyoto, Japan).

Ohneda K., Ohmori S, & Yamamoto M. (2019). Mouse Tryptase Gene Expression is Coordinately Regulated by GATA1 and GATA2 in Bone Marrow-Derived Mast Cells. International Journal of Molecular Sciences, 20(18), 4603.

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In Vitro Differentiation of iPSCs

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For in vitro differentiation of iPSCs, the hanging drop method was applied. Hanging drops containing 300 cells in 20 µl Iscove’s modified Dulbecco’s medium plus GlutaMAX™ (Thermo Fisher Scientific) supplemented with 20% FCS, 1× NEAA, 450 µM α-Monothioglycerol (Sigma-Aldrich) were cultured for 2 days. In this time, embryoid bodies had formed and were subsequently transferred into Petri dishes for a 3-day suspension culture period before transfer to gelatinized (0.1%) cell culture dishes for further 5, 15, or 25 days. Medium was exchanged every second or third day. The cells were harvested at the indicated time points, and expression of genes that indicate differentiation into the three germ layers was analyzed.

Gröschel C., Hübscher D., Nolte J., Monecke S., Sasse A., Elsner L., Paulus W., Trenkwalder C., Polić B., Mansouri A., Guan K, & Dressel R. (2017). Efficient Killing of Murine Pluripotent Stem Cells by Natural Killer (NK) Cells Requires Activation by Cytokines and Partly Depends on the Activating NK Receptor NKG2D. Frontiers in Immunology, 8, 870.

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Culture Conditions for Murine Stem Cells

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E14TG2a cells^{38 (link)} and MEDEP-E14 cells^{14 (link)} were obtained from RIKEN Cell Bank. E14TG2a cells were cultured in dishes coated with gelatin (Wako Pure Chemical) and maintained in Glasgow’s minimum essential medium (GMEM, Wako Pure Chemical) supplemented with 10% fetal bovine serum (GE Healthcare), 0.1 mM non-essential amino acids (Wako Pure Chemical), 1 mM sodium pyruvate (Wako Pure Chemical), 1000 U/mL mouse leukemia inhibitory factor (Merck Millipore) and 0.1 mM 2-mercaptoethanol (Wako Pure Chemical). MEDEP-E14 cells were maintained in Iscove’s modified Dulbecco’s medium (IMDM) containing 2 mM L-glutamine (Wako Pure Chemical) supplemented with 15% fetal bovine serum (GE Healthcare), 10 mg/ml bovine insulin, 5.5 mg/ml human transferrin, 5 ng/ml sodium selenite (ITS Liquid Media Supplement, Sigma), 50 mg/ml ascorbic acid (Sigma), 0.45 mM α-monothioglycerol (Sigma), and 3 unit/ml human erythropoietin (Kyowa Hakko Kirin).

Kobayashi N., Kawasaki-Nishi S., Otsuka M., Hisano Y., Yamaguchi A, & Nishi T. (2018). MFSD2B is a sphingosine 1-phosphate transporter in erythroid cells. Scientific Reports, 8, 4969.

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Isolation and Differentiation of CD34+ Cells

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CD34⁺ cells were isolated from peripheral blood using an immunomagnetic technique according to the manufacturer's instructions (#130‐046‐70, Miltenyi Biotec, Bergisch Gladbach, Germany). Briefly, 100 μl FcR blocking reagent and 100 μl CD34 MicroBeads were incubated with 10⁸ cells. The remaining population was cultured at 37°C in 5% CO₂ in Iscove's modified Dulbecco's medium (IMDM; Thermo Fisher Scientific) supplemented with 15% BIT 9500 serum substitute (Stemcell Technologies, Vancouver, Canada), α‐monothioglycerol (Sigma‐Aldrich) and liposomes (phosphatidylcholine, cholesterol and oleic acid; all from Sigma‐Aldrich), in the presence of human recombinant stem cell factor (SCF, 20 ng/ml, Miltenyi Biotec) and human thrombopoeitin (50 nM, Miltenyi Biotec) added once on day 0 to the culture medium, followed by 20 nM thrombopoeitin alone on day 6 with no further SCF addition. For proplatelet formation assays, megakaryocytes were plated on a BSA‐coated surface (chamber slide, Ibidi, Martinsried, Germany) on day 10. On day 13 or 14, the megakaryocytes were fixed using 4% paraformaldehyde and stained for β‐tubulin.

Stoupa A., Adam F., Kariyawasam D., Strassel C., Gawade S., Szinnai G., Kauskot A., Lasne D., Janke C., Natarajan K., Schmitt A., Bole‐Feysot C., Nitschke P., Léger J., Jabot‐Hanin F., Tores F., Michel A., Munnich A., Besmond C., Scharfmann R., Lanza F., Borgel D., Polak M, & Carré A. (2018). TUBB1 mutations cause thyroid dysgenesis associated with abnormal platelet physiology. EMBO Molecular Medicine, 10(12), e9569.

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Nicotine effects on cardiac differentiation

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CCE, a mouse embryonic stem cell line derived from 129/Sv mouse strain, was obtained from ATCC. mESCs were maintained on gelatin-coated plates in feeder-free Iscove's modified Dulbecco's medium (IMDM) (Invitrogen) supplemented with 10% fetal bovine serum (FBS) (ES qualified, Invitrogen), 1% nonessential amino acids, 1% penicillin-streptomycin, 4.5 × 10⁻⁴ M α-monothioglycerol (α-MTG; Sigma), 1,000 U/mL leukemia inhibitory factor (LIF). Cardiac differentiation of mESCs was performed as previously described (Yang et al., 2014 ). In brief, mESCs were developed into EBs (1,250 cells/drop) in a 3D hanging-drop culture for 4 days with LIF-free differentiation medium (IMDM supplemented with 10% FBS, 1% nonessential amino acids, 1% penicillin-streptomycin, and 4.5 × 10⁻⁴ M α-MTG). The early EBs were either plated onto gelatin-coated 12-well culture plates for adherent differentiation or put into a rotary system for 3D microgravity culture with the same differentiation medium for 8 days. EBs were treated with nicotine at a serial concentration (0.01–10 μM) in the presence or absence of 10 μM global nAChR antagonist Hexa during 12 days of differentiation.

Jiang X.Y., Feng Y.L., Ye L.T., Li X.H., Feng J., Zhang M.Z., Shelat H.S., Wassler M., Li Y., Geng Y.J, & Yu X.Y. (2017). Inhibition of Gata4 and Tbx5 by Nicotine-Mediated DNA Methylation in Myocardial Differentiation. Stem Cell Reports, 8(2), 290-304.

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Differentiation of hiPSCs to Platelets

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According to a protocol previously established by Takayama et al. [29 (link)], hiPSC colonies were removed from MEF feeders using a dissociation buffer (0.25% trypsin, 1 mg/ml collagenase IV, 20% KSR, 1 mmol/l CaCl₂ in PBS), transferred onto irradiated C3H10T1/2 cells and differentiated with IMDM medium containing 10 mg/l insulin, 5.5 mg/l transferrin, 6.7 mg/ml selenium, 2 mmol/l L-glutamine, 15% fetal bovine serum (all Gibco), 0.45 μmol/l α-monothioglycerol (Sigma-Aldrich), 50 μg/ml ascorbic acid (Sigma-Aldrich) and 20 ng/ml recombinant human vascular endothelial growth factor (Invitrogen). On day 15, hiPSC-Sacs were disrupted with a cell scraper, crushed with a pipette and passed through a 40 μm cell strainer (BD Falcon). The yielded cells were transferred onto irradiated C3H10T1/2 cells and cultured in the same medium without vascular endothelial growth factor containing 100 ng/ml human TPO (R&D), 50 ng/ml human SCF (R&D), and 25 U/ml heparin (Sigma-Aldrich). Medium was changed every 3 days. According to Takayama et al. [29 (link)], floating cells from days 24 to 30 were collected for platelet and MK analysis.

Orban M., Goedel A., Haas J., Sandrock-Lang K., Gärtner F., Jung C.B., Zieger B., Parrotta E., Kurnik K., Sinnecker D., Wanner G., Laugwitz K.L., Massberg S, & Moretti A. (2015). Functional Comparison of Induced Pluripotent Stem Cell- and Blood-Derived GPIIbIIIa Deficient Platelets. PLoS ONE, 10(1), e0115978.

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Synthesis and Characterization of Metal Chelators

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Triapine, PT, and P44mT were synthesized in our laboratory as previously described [4 (link), 16 (link)]. Dp44mT was from Santa Cruz Biotechnology (Dallas, TX). FeCl₃•6H₂O, FeSO₄•7H₂O, MnCl₂•4H₂O, CoCl₂•4H₂O, NiSO₄•6H₂O, CuCl₂•2H₂O, ZnSO₄•7H₂O, N-acetyl-L-cysteine, α-monothioglycerol, hydroxyurea, chloroquine, cisplatin and catalase (C-1345), were from Sigma-Aldrich (St. Louis, MO). Deferoxamine mesylate and bleomycin were from Cayman Chemical (Ann Arbor, MI). Superoxide dismutase (574594) was from EMD Millipore (Billerica, MA). Dithiothreitol was from Bio-Rad (Hercules, CA).
Triapine and PT were dissolved in anhydrous DMSO at 200 mM. Dp44mT and P44mT were dissolved in DMSO at 10 mM. The stock solutions were further diluted with DMSO and added to cell cultures with the final DMSO concentration being less than 0.05%. cisplatin, which undergoes relatively slow solvolysis in DMSO [26 ], was dissolved in DMSO at 20 mM, immediately aliquoted and stored at −70°C. Thawed aliquots were used one time. bleomycin was freshly prepared in H₂O at 10 mM.

Ishiguro K., Lin Z.P., Penketh P.G., Shyam K., Zhu R., Baumann R.P., Zhu Y.L., Sartorelli A.C., Rutherford T.J, & Ratner E.S. (2014). Distinct mechanisms of cell-kill by triapine and its terminally dimethylated derivative Dp44mT due to a loss or gain of activity of their copper(II) complexes. Biochemical pharmacology, 91(3), 312-322.

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