Popular LabKit · Recipe
PBS Recipe: How to Make 1X and 10X PBS (pH 7.4)
Phosphate-buffered saline for cell work, ELISA and immunostaining: grams of NaCl, KCl, disodium hydrogen phosphate (any hydrate) and potassium dihydrogen phosphate for 1X and 10X PBS.
Recipe: 1X PBS, pH 7.4, 1000 mL
| Component | Concentration | Amount |
|---|---|---|
| Sodium chloride, NaCl | 137 mM | 8.006 g |
| Potassium chloride, KCl | 2.7 mM | 0.201 g |
| Disodium hydrogen phosphate, Na₂HPO₄ (anhydrous) | 10 mM | 1.420 g |
| Potassium dihydrogen phosphate, KH₂PO₄ | 1.8 mM | 0.245 g |
| Purified water | to 1000 mL |
10X PBS stock and other hydrates
| Component | 1X (per L) | 10X (per L) |
|---|---|---|
| NaCl | 8.006 g | 80.06 g |
| KCl | 0.201 g | 2.013 g |
| Na₂HPO₄ anhydrous | 1.420 g | 14.20 g |
| or Na₂HPO₄·2H₂O | 1.780 g | 17.80 g |
| or Na₂HPO₄·7H₂O | 2.681 g | 26.81 g |
| or Na₂HPO₄·12H₂O | 3.581 g | 35.81 g |
| KH₂PO₄ | 0.245 g | 2.450 g |
Method
- Dissolve the four salts in about 800 mL of purified water.
- Check the pH; it should be close to 7.4. Adjust with HCl or NaOH if needed.
- Make up to 1000 mL. Autoclave (121 °C, 20 min) or filter (0.22 µm) for sterile PBS.
- 10X stock: dilute 100 mL to 1000 mL with water before use. A 10X stock reads about pH 6.8–7.0 and comes to 7.4 on dilution – do not adjust the 10X stock to 7.4.
This is the standard formulation from Cold Spring Harbor Protocols. Some laboratories use Dulbecco’s PBS (DPBS), which differs slightly and may contain calcium and magnesium.
Recipes are calculated with the Popular LabKit buffer model (ionic strength and temperature corrected) and checked against published buffer tables. For a pharmacopoeial test, follow the reagent or buffer recipe in your monograph. Always confirm pH with a calibrated pH meter.
Frequently asked questions
What is the difference between PBS and phosphate buffer?
PBS is a dilute (about 12 mM) phosphate buffer with sodium and potassium chloride added to make it isotonic, for washing cells and diluting antibodies. A plain phosphate buffer has no added salt and is usually more concentrated.
Can I use sodium phosphate dibasic heptahydrate?
Yes – use 2.681 g per litre for 1X, as in the table, or pick the hydrate in the calculator.
Why does my 10X PBS form crystals in the fridge?
Disodium hydrogen phosphate is less soluble when cold. Store 10X PBS at room temperature, or warm and mix before use.