TrypLE
Mouse embryonic fibroblasts (MEFs) (Gibco)
PBS-/- (Gibco)
PBS+/+(Gibco)
Accumax (Fisher #NC0443331)
Rock inhibitor (R&D #1254)
Cell culture media
MEF Media
100x MEM NEAA (Life Tech #11140-050)
FBS (Thermo Sci #SH30071.03)
1x DMEM (Life Tech #11995-065)
hESC Media
100x Glutamax (Life Tech #35050-061)
100x MEM-NEAA (Life Tech #11140-050)
100x Beta-mercaptoethanol (Millipore #ES-007-E)
KSR (Life Tech #10828-028)
DMEM/F-12 (Life Tech #11320-033)
FGF-2 -10ng/ml (R&D 233-FB-025/C)
Day 0 Media
hESC media
Activin A- 10ng/ml (R&D 338-AC/CF)
HeregulinB -10 ng/ml (Peprotech 100-03)
Day 1 Media
100x Glutamax (Life Tech #35050-061)
1x RPMI 1640 (Mediatech #15-040-CM)
FBS- 0.2% (Corning/Fisher - #35-011-CV)
ITS-1:5,000 (Gibco)
Activin A-100 ng/ml (338-AC/CF)
Human WNT3a- 50 ng/ml (R&D -5036-WN-010/CF).
Day 2 Media
100x Glutamax (Life Tech #35050-061)
1x RPMI 1640 (Mediatech #15-040-CM)
FBS- 0.2% (Corning/Fisher - #35-011-CV)
ITS- 1:2,000 (Gibco)
Activin A -100 ng/ml (338-AC/CF)
Day 3 Media
100x Glutamax (Life Tech #35050-061)
1x RPMI 1640 (Mediatech #15-040-CM)
FBS- 0.2% (Corning/Fisher - #35-011-CV)
ITS- 1:1,000 (Gibco)
TGFbi IV- 2.5 μM (CalBioChem #616454)
KGF- 25 ng/ml (R&D- 251-KG-050/CF)
Day 4‐5 Media
100x Glutamax (Life Tech #35050-061)
1x RPMI 1640 (Mediatech #15-040-CM)
FBS- 0.4% (Corning/Fisher - #35-011-CV)
ITS -1:1,000
KGF-25 ng/ml (R&D- #251-KG-050/CF)
Day 6‐7 Media
100x Glutamax (Life Tech #35050-061)
1x DMEM (Life Tech #11960-051)
B27- 1:100 (Gibco)
TTNPB - 3 nM (Sigma-Aldrich#T3757)
Day 8 Media
100x Glutamax (Life Tech #35050-061)
1x DMEM (Life Tech #11960-051)
B27-1:100 (Gibco)
TTNPB - 3 nM (Sigma-Aldrich#T3757)
EGF- 50 ng/ml (R&D #236-EG)
Day 9-11 Media
100x Glutamax (Life Tech #35050-061)
1x DMEM (Life Tech #11960-051)
B27- 1:100 (Gibco)
EGF- 50 ng/ml (R&D #236-EG)
KGF- 50 ng/ml (R&D- #251-KG-050/CF)
Day 12‐20 Media
100x Glutamax (Life Tech #35050-061)
1x DMEM (Life Tech #11960-051)
B27- 1:100 (Gibco)
NEAA- 1:100 (Gibco)
ALKi II- 10 μm (Cayman Chemical #14794)
LDN‐193189 - 500 nM (Fisher #04-0074-10)
Xxi - 1 μm (Millipore #565789)
T3- 1 μM (Sigma-Aldrich #T6397)
Vitamin C - 0.5 mM (Sigma-Aldrich#49752)
N-acetyl Cysteine- 1 mM (Sigma-Aldrich)
Zinc sulfate- 10 μM (Sigma-Aldrich)
Heparin sulfate- 10 μg/ml (Sigma-Aldrich#H3149)
Day 20-27 Media
100x Glutamax (Life Tech #35050-061)
CMRL (Life Tech #11530)
1:100 B27 (or 10% FBS) (Gibco)
1:100 NEAA (Gibco)
ALKi II -10 μm (Cayman Chemical #14794)
Vitamin C -0.5 mM (Sigma-Aldrich#49752)
T3- 1 μM (Sigma-Aldrich #T6397)
N-acetyl Cysteine -1 mM (Sigma-Aldrich)
Zinc sulfate- 10 μM (Sigma-Aldrich)
Heparin sulfate - 10 μg/ml (Sigma-Aldrich#H3149)