| DMEM/F12 (Life Technologies 11330-032) |
500 mL |
| d-Glucose (Sigma-Aldrich G5400)
|
2.5 g |
| Nicotinamide (Sigma-Aldrich N3376) |
0.61 g |
| Soybean trypsin inhibitor (type 1; Sigma-Aldrich T6522) |
50 mg |
| ITS+ Premix (BD Biosciences 354352) |
2.5 mL |
| Bovine pituitary extract (BD Biosciences 354123; 3 mg/mL in DMEM/F12) |
4.2 mL |
| 3,3′,5-Triiodo-l-thyronine (T3; 50 nm)a |
50 µL |
| Dexamethasone (Sigma-Aldrich D1756; 100 mm in DMSO)
|
5 µL |
| Nu-serum IV (BD Biosciences 355104) |
25 mL |
| Cholera toxin (Sigma-Aldrich C8052; 1 mg/mL in H2O)
|
50 µL |
| Penicillin-streptomycin (100×; Life Technologies 15140-122) |
5 mL |
| Fungizone |
2.5 mL |
| aTo prepare a 50 nm T3 stock solution, dissolve 250 mg of T3 (Sigma-Aldrich T2877) in 10 mL of DMSO and then dilute 1.5 µL of the T3 solution
in 1 mL of DMEM/F12. Store at –20°C.
|
