Healthy human induced pluripotent stem cell (hiPSC) lines IMR90 and M180 were cultured in mTeSR1 medium on Matrigel-coated plates and were dissociated using ReLeSR. The cells were then transferred onto Reduced Growth Factor (RF)-Matrigel–coated plates for differentiation into hiPSC-derived cardiomyocytes (hiPSC-CM). Differentiation was initiated on day 0 using differentiation medium (RPMI medium [Gibco] supplemented with 1% B27 minus insulin (Gibco) and 6 μmol/L CHIR99201 (Tocris Bioscience) for 2 days. On day 3, the differentiation medium was supplemented with 2.5 μmol/L Wnt-C59 (Tocris Bioscience). The glucose-depletion method was performed on days 11 and 13 by changing of medium to no-glucose RPMI with 1% B27 minus insulin (33 (link)). hiPSC-CM maturation was initiated from day 16 with replating cells onto RF-Matrigel–coated plates and culturing in DMEM containing 5 mmol/L glucose and supplemented with 0.4 mmol/L oleic acid conjugated to BSA, for 1 week (34 (link)). Results from the IMR90 line are presented in Figs. 1 and 2 , with key findings confirmed in the M180 line presented in Supplementary Fig. 1 .
Chir99201
Manufactured by Bio-Techne
CHIR99201 is a small molecule inhibitor that selectively inhibits glycogen synthase kinase-3 (GSK-3). GSK-3 is a serine/threonine protein kinase that plays a crucial role in various cellular processes. CHIR99201 is commonly used in cell culture and stem cell research applications.
Automatically generated - may contain errors
Lab products found in correlation
B 27 minus insulin, by Thermo Fisher Scientific (1 mentions)
Wnt c59, by Bio-Techne (1 mentions)
Rpmi medium, by Thermo Fisher Scientific (1 mentions)
Vu0240551, by Bio-Techne (1 mentions)
B27 supplement, by Thermo Fisher Scientific (1 mentions)
Sodium pyruvate, by Thermo Fisher Scientific (1 mentions)
Glutamax, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
2 mercaptoethanol, by Thermo Fisher Scientific (1 mentions)
Heparin, by Merck Group (1 mentions)
Non essential amino acids solution, by Thermo Fisher Scientific (1 mentions)
Neuropan basal medium, by PAN Biotech (1 mentions)
Recombinant human fgf4, by Thermo Fisher Scientific (1 mentions)
Dmem f12, by PAN Biotech (1 mentions)
Trypsin, by PAN Biotech (1 mentions)
3 protocols using chir99201
1
Differentiation of hiPSCs into Cardiomyocytes
2
Synthesis and Characterization of Compounds
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3
Culturing Mouse Embryonic Stem Cells
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Cell culture mESCs were routinely cultured on 0.1% gelatin (Sigma Aldrich)-coated tissue culture flasks in serum + LIF medium composed of GMEM (ThermoFisher), 10% batch-tested fetal bovine serum (FBS) (Sigma Aldrich), 1x GlutaMAX (ThermoFisher), 1 mM sodium pyruvate (ThermoFisher), 1x non-essential amino acids solution (ThermoFisher), 100 µM 2mercaptoethanol (ThermoFisher) and 10 ng/ml LIF (MPI protein expression facility). Cells were passaged every two to three days using 0.05% Trypsin (PAN Biotech). Basal medium for serum free culture was N2B27, prepared as a 1:1 mixture of DMEM/F12 (PAN Biotech)
and Neuropan basal medium (PAN Biotech) with 0.5% BSA, 1x N2 and 1x B27 supplements (ThermoFisher) and 50 µM 2-mercaptoethanol. For FGF stimulation experiments, short-term serum-free culture was carried out in N2B27 supplemented with 3 µM CHIR99201 (Tocris), 1 µg/ml of Heparin (Sigma) and with or without 10 ng/ml LIF as indicated. Recombinant human FGF4 used was obtained from Peprotech. For live imaging and immunostaining studies, cells were seeded on polymer-bottomed ibidi µ-slides (ibidi) coated with 20 µg/ml fibronectin.
and Neuropan basal medium (PAN Biotech) with 0.5% BSA, 1x N2 and 1x B27 supplements (ThermoFisher) and 50 µM 2-mercaptoethanol. For FGF stimulation experiments, short-term serum-free culture was carried out in N2B27 supplemented with 3 µM CHIR99201 (Tocris), 1 µg/ml of Heparin (Sigma) and with or without 10 ng/ml LIF as indicated. Recombinant human FGF4 used was obtained from Peprotech. For live imaging and immunostaining studies, cells were seeded on polymer-bottomed ibidi µ-slides (ibidi) coated with 20 µg/ml fibronectin.
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