2D culture media

Reagent	Final concentration	Amount
DMEM/Ham’s F12	1×	91 mL
HEPES (1M)	10 mM	1 mL
Penicillin-Streptomycin (10,000U/mL)	100 U/mL	1 mL
GlutaMAX (100×)	1×	1 mL
Insulin-Transferrin-Selenium (100×)	1×	1 mL
FBS (100 %)	5 %	5 mL
hEGF (500 μg/mL)	25 ng/mL	5 μL
Bovine pituitary extract (159 mg/mL	30 μg/mL	18.9 μL
Total	n/a	100 mL

2D media can be stored in 10 mL aliquots at −80°C. Once thawed, store at 4°C and use within 1–2 weeks.