SATO Supplement (100×)

• 1. Prepare the following stock solutions (these should be made fresh; do not re-use):

  • Combine 5 mg of progesterone (Sigma-Aldrich P8783) and 200 μL of ethanol to make a progesterone stock solution.

  • Combine 4 mg of sodium selenite (Sigma-Aldrich S5261), 10 μL of 1 n NaOH, and 10 mL of Dulbecco's modified Eagle's medium (DMEM; Gibco/Life Technologies 11960-044) to make a sodium selenite stock solution.

• 2. Combine the following:

  Reagent	Quantity (for 200 mL)	Final concentration (100×)
  BSA (Sigma-Aldrich A4161)	2 g	10 mg/mL
  Transferrin (Sigma-Aldrich T1147)	2 g	10 mg/mL
  Putrescine (Sigma-Aldrich P5780)	320 mg	1.6 mg/mL
  Progesterone stock solution	50 μL	6 μg/mL
  Sodium selenite stock solution	2 mL	4 μg/mL

• 3. Bring to a total volume of 200 mL in DMEM, and then filter-sterilize. Aliquot and store at −20°C.

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