gdnf small molecules and growth factors  (PeproTech)

Journal: Nature protocols

Article Title: Generation of human brain region–specific organoids using a miniaturized spinning bioreactor

doi: 10.1038/nprot.2017.152

Figure Lengend Snippet: Culture media.

Article Snippet: Mouse embryonic fibroblast feeder cells isolated from E13.5 CF1 mouse embryos (Charles River, strain code 023) that have been growth-arrested by γ-irradiation using a Cs-137 irradiator Growth media components and supplements DMEM (Corning, cat. no. 10–013) DMEM/F-12, HEPES (Gibco, cat. no. 11330032) Neurobasal medium (Gibco, cat. no. 21103049) KnockOut Serum Replacement (KOSR; Gibco, cat. no. 10828028) GlutaMAX supplement (Gibco, cat. no. 35050061) MEM non-essential amino acids solution (MEM-NEAA; Gibco, cat. no. 11140050) Penicillin–streptomycin (Pen/Strep; Gibco, cat. no. 15140122) Amphotericin B (Thermo Fisher, cat. no. 15290026 2-Mercaptoethanol (Gibco, cat. no. 21985023) N-2 supplement (Gibco, cat. no. 17502048) B-27 supplement (Gibco, cat. no. 17504044) Matrigel growth factor-reduced basement membrane matrix (Corning, cat. no. 354230) Insulin solution (Sigma-Aldrich, cat. no. I9278–5ML) FBS (Corning, cat. no. 35–010) Small molecules and growth factors A83–01 (Stem Cell Technologies, cat. no. 72022) Dorsomorphin (Stem Cell Technologies, cat. no. 72102) SB-431542 (Stem Cell Technologies, cat. no. 72232) CHIR-99021 (Stem Cell Technologies, cat. no. 72052) Purmorphamine (Stem Cell Technologies, cat. no. 72202) LDN-193189 (Stem Cell Technologies, cat. no. 72146) FGF-2 (Peprotech, cat. no. 100–18B) FGF-8 (Peprotech, cat. no. 100–25A) WNT-3A (R&D Systems, cat. no. 5036-WN-010) SHH (Peprotech, cat. no. 100–45) CNTF (Peprotech, cat. no. 450–13) BDNF (Peprotech, cat. no. 450–02) GDNF (Peprotech, cat. no. 450–10) Ascorbic acid (Sigma-Aldrich, cat. no. 1043003) cAMP (Sigma-Aldrich, cat. no. A9501) Enzymes and other reagents Collagenase, type IV (Thermo Fisher, cat. no. 17104019) BSA (Sigma-Aldrich, cat. no. A9418) Dulbecco’s PBS without calcium and magnesium (DPBS; Corning, cat. no. 21–031) Donkey serum (Millipore, cat. no. S30) 0.1% (wt/vol) Gelatin in water (Stem Cell Technologies, cat. no. 7903) Ethyl alcohol, 95% (vol/vol), 190 proof, pure (Pharmco-Aaper, cat. no. 111000190CSGL) Sterile deionized water DMSO (Sigma-Aldrich, cat. no. 472301) 1-Thioglycerol (Sigma-Aldrich, cat. no. M1753) 10% (vol/vol) Bleach 16% (vol/vol) Formaldehyde, methanol free (Polysciences, cat. no. 18814–10) Sucrose (Sigma-Aldrich, cat. no. S5016) Tissue-freezing medium (General Data, cat. no. TFM-5) Triton X-100 (Sigma-Aldrich, cat. no. T9284) Tween 20 (Sigma-Aldrich, cat. no. P7949) Slide-mounting solution (Fisher Scientific, cat. no. 14-390-5)

Techniques: