Popular LabKit · Recipe
Citrate Buffer pH 6.0: Preparation and Recipe
How to prepare citrate buffer pH 6.0 at 10 mM (the antigen-retrieval buffer), 50 mM or 0.1 M from citric acid and trisodium citrate, with method and tips.
Recipe: 10 mM citrate buffer, pH 6.0, 1000 mL (antigen retrieval)
| Component | Amount |
|---|---|
| Trisodium citrate dihydrate, Na₃C₆H₅O₇·2H₂O | 2.435 g |
| Citric acid monohydrate, C₆H₈O₇·H₂O | 0.361 g |
| Purified water | to 1000 mL |
The widely used alternative for antigen retrieval is to dissolve 2.94 g trisodium citrate dihydrate (10 mM) in water and adjust to pH 6.0 with 1 M HCl, then add 0.5 mL Tween 20 per litre if your protocol asks for it. Both give 10 mM citrate at pH 6.0.
Other concentrations
| Component | 10 mM | 50 mM | 0.1 M |
|---|---|---|---|
| Citric acid monohydrate | 0.361 g | 1.290 g | 2.511 g |
| Trisodium citrate dihydrate | 2.435 g | 12.90 g | 25.90 g |
Method
- Weigh both salts accurately and transfer to a beaker.
- Dissolve in about 800 mL of purified water (80% of the final volume).
- Bring to the temperature of use and check the pH with a calibrated pH meter. Fine-adjust with a few drops of dilute acid or base if needed.
- Transfer to a 1000 mL volumetric flask and make up to the mark. Mix well.
- Label with name, pH, concentration, date and initials. Filter (0.22 µm) or autoclave if it must be sterile.
Citric acid has three pKa values (3.13, 4.76 and 6.40), so citrate buffers cover pH 3.0–6.2. The calculator works out all three equilibria with an ionic-strength correction; at 0.1 M its results agree with the Sigma-Aldrich citrate table within about 0.08 pH (within 0.04 pH above pH 4).
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
Can I make citrate buffer from citric acid and NaOH?
Yes. Dissolve citric acid at the full concentration and titrate to the pH you need with NaOH using a pH meter. The final solution is the same.
What pH range can citrate buffer cover?
About pH 3.0 to 6.2. Below 3 or above 6.2 choose a different buffer, such as McIlvaine citrate–phosphate (pH 2.2–8.0), also in the calculator.
Does citrate buffer interfere with anything?
Citrate binds calcium and other metal ions, so avoid it where free calcium matters.