Modified Barth's Saline (MBS) Preparation and Recipe

Table 1. Required components

Component	Amount	Concentration
Hepes (mw: 238.30 g/mol)	11.92 g	0.05002 M
Sodium chloride (mw: 58.44 g/mol)	51.43 g	0.88 M
Potassium Chloride (mw: 74.55 g/mol)	0.75 g	0.01006 M
Sodium bicarbonate (mw: 84.01 g/mol)	2.1 g	0.02499 M
Magnesium Sulfate Heptahydrate (mw: 246.47 g/mol)	2.46 g	0.00998 M

1. Prepare 800 mL of distilled water in a suitable container.
2. Add 11.92 g of Hepes to the solution.
3. Add 51.43 g of Sodium chloride to the solution.
4. Add 0.75 g of Potassium Chloride to the solution.
5. Add 2.1 g of Sodium bicarbonate to the solution.
6. Add 2.46 g of Magnesium Sulfate Heptahydrate to the solution.
7. Adjust the pH of 10x MBS salt solution to 7.8 with 10 M NaOH, and sterilize by autoclaving. Store indefinitely at room temperature.
8. Add distilled water until the volume is 1 L.
9. To prepare 1x MBS, combine 100 mL of 10x MBS salt solution and 7 mL of 0.1 M CaCl2 with 893 mL of autoclaved deionized H2O. Add antibiotics. Store for up to 1-2 mo at 16°C-18°C (the same temperature used for oocyte incubation). Check before use.

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