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[Home](https://www.sigmaaldrich.com/US/en)[Protein Concentration & Buffer Exchange](https://www.sigmaaldrich.com/US/en/applications/protein-biology/protein-concentration-and-buffer-exchange)Buffer Reference Center
# Buffer Reference Center
__Section Overview__
- [Choosing the Right Biological Buffer](https://www.sigmaaldrich.com#buffer)
- [What are Buffer Grades?](https://www.sigmaaldrich.com#buffer-grades)
- [Useful pH Ranges of Selected Biological Buffers Chart (25 °C, 0.1 M)](https://www.sigmaaldrich.com#selected-buffers)
- [Tris or Trizma® Buffer Preparation – pH vs. Temperature](https://www.sigmaaldrich.com#trizma)
- [Phosphate Buffer Preparation – 0.2 M solution](https://www.sigmaaldrich.com#phosphate)
- [Citric Acid – Na2HPO4 Buffer Preparation, pH 2.6-7.6](https://www.sigmaaldrich.com#citric)
- [Citric Acid – Sodium Citrate Buffer Preparation, pH 3.0-6.2](https://www.sigmaaldrich.com#citric2)
- [Sodium Acetate – Acetic Acid Buffer Preparation, pH 3.7-5.6](https://www.sigmaaldrich.com#sodiumacetate)
- [Na2HPO4 – NaH2PO4 Buffer Preparation, pH 5.8-8.0 at 25 °C](https://www.sigmaaldrich.com#NA2HPO4)
- [Imidazole (glyoxaline) – HCl Buffer Preparation, pH 6.2-7.8 at 25 °C](https://www.sigmaaldrich.com#imidazole)
- [Sodium Carbonate – Sodium Bicarbonate Buffer Preparation, pH 9.2-10.8](https://www.sigmaaldrich.com#sodiumcarbonate)
- [Buffer Preparation Formulas and Equations](https://www.sigmaaldrich.com#buffer-preparation)
## [](https://www.sigmaaldrich.com)Choosing the Right Biological Buffer
Choose a buffer based on your pH requirements as well as the pKa, a measure of acid strength that accounts for pH, concentration, and temperature. Regulatory or purity needs for your exact application should also be considered. The following tables can help you navigate preparation of many common buffer solutions by pH and pKa.
In addition to the tables below, we've developed several [buffer recipe calculators](https://www.sigmaaldrich.com/US/en/support/calculators-and-apps#buffers) to assist your buffer preparation.
[TBE and TAE for gel electrophoresis](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/gel-electrophoresis/tae-and-tbe-running-buffers-recipe) have slightly more involved recipes. We can help you pick the right one.
## [](https://www.sigmaaldrich.com)What are buffer grades?
Buffer grade indicates the quality and impurity levels appropriate for different uses. We provide six grades of buffers indicated for general lab use, final pharmaceutical formulation and manufacturing, and applications in between that may need trace metal testing or materials of a specified purity.
## [](https://www.sigmaaldrich.com)Useful pH Ranges of Selected Biological Buffers Chart (25 °C, 0.1 M)
BuffersUseful pH RangepKa
(at 20 °C)pKa
(at 25 °C)pKa
(at 37 °C)
[MES](https://www.sigmaaldrich.com/US/en/search/mes?focus=products&page=1&perpage=30&sort=relevance&term=MES&type=product)5.5–6.76.166.105.97
[Bis-Tris](https://www.sigmaaldrich.com/US/en/search/bis-tris?focus=products&page=1&perpage=30&sort=relevance&term=Bis-tris&type=product)5.8–7.2n/a6.506.36
[ADA](https://www.sigmaaldrich.com/US/ensearch/ada?page=1&perpage=30&sort=relevance)6.0–7.26.656.596.46
[ACES](https://www.sigmaaldrich.com/US/ensearch/aces?page=1&perpage=30&sort=relevance)6.1–7.56.886.786.54
[PIPES](https://www.sigmaaldrich.com/US/ensearch/pipes?page=1&perpage=30&sort=relevance)6.1–7.56.806.766.66
[MOPSO](https://www.sigmaaldrich.com/US/en/US/en/search/mopso?focus=products&page=1&perpage=30&sort=relevance&term=MOPSO&type=product)6.2–7.6n/a6.906.75
[Bis-Tris Propane](https://www.sigmaaldrich.com/US/en/search/bis-tris-propane?focus=products&page=1&perpage=30&sort=relevance&term=Bis-Tris%20Propane&type=product)6.3–9.5n/a6.8, 9.0n/a
[BES](https://www.sigmaaldrich.com/US/en/search/bes?focus=products&page=1&perpage=30&sort=relevance&term=BES&type=product)6.4–7.87.177.096.90
[MOPS](https://www.sigmaaldrich.com/US/en/search/mops?focus=products&page=1&perpage=30&sort=relevance&term=MOPS&type=product)6.5–7.97.287.207.02
[TES](https://www.sigmaaldrich.com/US/en/search/tes?focus=products&page=1&perpage=30&sort=relevance&term=TES&type=product)6.8–8.27.507.407.16
[HEPES](https://www.sigmaaldrich.com/US/en/search/hepes?focus=products&page=1&perpage=30&sort=relevance&term=HEPES&type=product)6.8–8.27.557.48[](https://www.sigmaaldrich.com/catalog/product/SIGMA/T9269)7.31
[DIPSO](https://www.sigmaaldrich.com/US/en/search/dipso?focus=products&page=1&perpage=30&sort=relevance&term=DIPSO&type=product)7.0–8.2n/a7.607.35
[MOBS](https://www.sigmaaldrich.com/US/en/search/mobs?focus=products&page=1&perpage=30&sort=relevance&term=MOBS&type=product)6.9–8.3n/a7.60n/a
[TAPSO](https://www.sigmaaldrich.com/US/en/US/en/search/tapso?focus=products&page=1&perpage=30&sort=relevance&term=TAPSO&type=product)7.0–8.2n/a7.607.39
[Tris or Trizma®](https://www.sigmaaldrich.com/US/en/search/tris-or-trizma?focus=products&page=1&perpage=30&sort=relevance&term=Tris%20or%20Trizma&type=product)7.0–9.08.208.067.72
[HEPPSO](https://www.sigmaaldrich.com/US/en/search/heppso?focus=products&page=1&perpage=30&sort=relevance&term=HEPPSO&type=product)7.1–8.5n/a7.806.66
[POPSO](https://www.sigmaaldrich.com/US/en/search/popso?focus=products&page=1&perpage=30&sort=relevance&term=POPSO&type=product)7.2–8.5n/a7.807.63
[TEA](https://www.sigmaaldrich.com/US/en/search/tea?focus=products&page=1&perpage=30&sort=relevance&term=TEA&type=product)7.3–8.3n/a7.80n/a
[EPPS](https://www.sigmaaldrich.com/US/en/search/epps?focus=products&page=1&perpage=30&sort=relevance&term=EPPS&type=product)7.3–8.7n/a8.00n/a
[Tricine](https://www.sigmaaldrich.com/US/en/search/tricine?focus=products&page=1&perpage=30&sort=relevance&term=Tricine&type=product)7.4–8.88.168.057.80
[Gly-Gly](https://www.sigmaaldrich.com/US/en/search/gly-gly?focus=products&page=1&perpage=30&sort=relevance&term=Gly-Gly&type=product)7.5–8.9n/a8.20n/a
[Bicine](https://www.sigmaaldrich.com/US/en/search/bicine?focus=products&page=1&perpage=30&sort=relevance&term=Bicine&type=product)7.6–9.08.358.268.04
[HEPBS](https://www.sigmaaldrich.com/US/en/search/hepbs?focus=products&page=1&perpage=30&sort=relevance&term=HEPBS&type=product)7.6–9.0n/a8.30n/a
[TAPS](https://www.sigmaaldrich.com/US/en/search/taps?focus=products&page=1&perpage=30&sort=relevance&term=TAPS&type=product)7.7-9.18.498.408.18
[AMPD](https://www.sigmaaldrich.com/US/en/US/en/search/ampd?focus=products&page=1&perpage=30&sort=relevance&term=AMPD&type=product)7.8–9.7n/a8.80n/a
[TABS](https://www.sigmaaldrich.com/US/en/search/tabs?focus=products&page=1&perpage=30&sort=relevance&term=TABS&type=product)8.2–9.6n/a8.90n/a
[AMPSO](https://www.sigmaaldrich.com/US/en/US/en/search/ampso?focus=products&page=1&perpage=30&sort=relevance&term=AMPSO&type=product)8.3–9.7n/a9.009.10
[CHES](https://www.sigmaaldrich.com/US/en/search/ches?focus=products&page=1&perpage=30&sort=relevance&term=CHES&type=product)8.6–10.09.559.499.36
[CAPSO](https://www.sigmaaldrich.com/US/en/capso?page=1&perpage=30&sort=relevance)8.9–10.3n/a9.609.43
[AMP](https://www.sigmaaldrich.com/US/en/search/amp?focus=products&page=1&perpage=30&sort=relevance&term=AMP&type=product)9.0–10.5n/a9.70n/a
[CAPS](https://www.sigmaaldrich.com/US/en/search/caps?focus=products&page=1&perpage=30&sort=relevance&term=CAPS&type=product)9.7–11.110.5610.4010.02
[CABS](https://www.sigmaaldrich.com/US/en/search/cabs?focus=products&page=1&perpage=30&sort=relevance&term=CABS&type=product)10.0–11.4n/a10.70n/a
[](https://www.sigmaaldrich.com)
## [](https://www.sigmaaldrich.com)Tris or Trizma® Buffer Preparation Table – pH vs. Temperature
| | | |
|-------------------|-----------|-----------|
| pH at Temperature | | |
| __5 °C__ | __25 °C__ | __37 °C__ |
| 7.76 | 7.20 | 6.91 |
| 7.89 | 7.30 | 7.02 |
| 7.97 | 7.40 | 7.12 |
| 8.07 | 7.50 | 7.22 |
| 8.18 | 7.60 | 7.30 |
| 8.26 | 7.70 | 7.40 |
| 8.37 | 7.80 | 7.52 |
| 8.48 | 7.90 | 7.62 |
| 8.58 | 8.00 | 7.71 |
| 8.68 | 8.10 | 7.80 |
| 8.78 | 8.20 | 7.91 |
| 8.88 | 8.30 | 8.01 |
| 8.98 | 8.40 | 8.10 |
| 9.09 | 8.50 | 8.22 |
| 9.18 | 8.60 | 8.31 |
| 9.28 | 8.70 | 8.42 |
| 9.36 | 8.80 | 8.51 |
| 9.47 | 8.90 | 8.62 |
| 9.56 | 9.00 | 8.70 |
| | | | | |
|-------------------------|------------------|---|---|---|
| g/L for 0.05 M Solution | | | | |
| __Trizma® HCL__ | __Trizma® Base__ | | | |
| 7.02 | 0.67 | | | |
| 6.85 | 0.80 | | | |
| 6.61 | 0.97 | | | |
| 6.35 | 1.18 | | | |
| 6.06 | 1.39 | | | |
| 5.72 | 1.66 | | | |
| 5.32 | 1.97 | | | |
| 4.88 | 2.30 | | | |
| 4.44 | 2.65 | | | |
| 4.02 | 2.97 | | | |
| 3.54 | 3.34 | | | |
| 3.07 | 3.70 | | | |
| 2.64 | 4.03 | | | |
| 2.21 | 4.36 | | | |
| 1.83 | 4.65 | | | |
| 1.50 | 4.90 | | | |
| 1.23 | 5.13 | | | |
| 0.96 | 5.32 | | | |
| 0.76 | 5.47 | | | |
## [](https://www.sigmaaldrich.com)Phosphate Buffer Preparation Table – 0.2 M solution
Potassium
Phosphate Monobasic Anhydrous g/LSodium Phosphate Dibasic Heptahydrate g/L23 °C pH
22.43.495.7
22.084.295.8
21.605.375.9
21.056.606.0
20.408.056.1
19.569.936.2
18.6012.076.3
17.6414.226.4
16.4416.906.5
15.0020.126.6
13.5623.346.7
12.2426.296.8
Potassium Phosphate Monobasic
Anhydrous g/LSodium Phosphate
Dibasic Heptahydrate g/L23 °C pH
10.8029.516.9
9.3632.737.0
7.9235.957.1
6.7238.637.2
5.5241.317.3
4.5643.467.4
3.8445.077.5
3.1246.687.6
2.5248.557.7
2.0449.097.8
1.6849.897.9
1.2750.818.0
## [](https://www.sigmaaldrich.com)Citric Acid – Na2HPO4 Buffer Preparation, pH 2.6–7.61
Citric acid monohydrate, C6H8O7 • H2O, MW 210.14; 0.1 M contains 21.01 g/L. Na2HPO4, MW 141.98; 0.2 M contains 28.40 g/L, or Na2HPO4 • 2H2O, MW 178.05; 0.2 M contains 35.61 g/L.
x mL 0.1 M-citric acid and y mL 0.2 M-Na2HPO4 mixed
| | | |
|-----|------------------------|------------------|
| pH | x mL 0.1 M-citric acid | y mL 0.2-Na2HPO4 |
| 2.6 | 89.10 | 10.90 |
| 2.8 | 84.15 | 15.85 |
| 3.0 | 79.45 | 20.55 |
| 3.2 | 75.30 | 24.70 |
| 3.4 | 71.50 | 28.50 |
| 3.6 | 67.80 | 32.20 |
| 3.8 | 64.50 | 35.50 |
| 4.0 | 61.45 | 38.55 |
| 4.2 | 58.60 | 41.40 |
| 4.4 | 55.90 | 44.10 |
| 4.6 | 53.25 | 46.75 |
| 4.8 | 50.70 | 49.30 |
| 5.0 | 48.50 | 51.50 |
| 5.2 | 46.40 | 53.60 |
| 5.4 | 44.25 | 55.75 |
| 5.6 | 42.00 | 58.00 |
| 5.8 | 39.55 | 60.45 |
| 6.0 | 36.85 | 63.15 |
| 6.2 | 33.90 | 66.10 |
| 6.4 | 30.75 | 69.25 |
| 6.6 | 27.25 | 72.75 |
| 6.8 | 22.75 | 77.25 |
| 7.0 | 17.65 | 82.35 |
| 7.2 | 13.05 | 86.95 |
| 7.4 | 9.15 | 90.85 |
| 7.6 | 6.35 | 93.65 |
## [](https://www.sigmaaldrich.com)Citric Acid – Sodium Citrate Buffer Preparation, pH 3.0-6.21
Citric acid monohydrate, C6H8O7 • H2O, MW 210.14; 0.1 M contains 21.01 g/L. Trisodium citrate dihydrate, C6H5O7Na3 • 2H2O, MW 294.12; 0.1 M contains 29.41 g/L.
x mL 0.1 M-citric acid and y mL 0.1 M-trisodium citrate mixed
| | | |
|-----|------------------------|------------------------------|
| pH | x mL 0.1 M-citric acid | y mL 0.1 M-trisodium citrate |
| 3.0 | 82.0 | 18.0 |
| 3.2 | 77.5 | 22.5 |
| 3.4 | 73.0 | 27.0 |
| 3.6 | 68.5 | 31.5 |
| 3.8 | 63.5 | 36.5 |
| 4.0 | 59.0 | 41.0 |
| 4.2 | 54.0 | 46.0 |
| 4.4 | 49.5 | 50.5 |
| 4.6 | 44.5 | 55.5 |
| 4.8 | 40.0 | 60.0 |
| 5.0 | 35.0 | 65.0 |
| 5.2 | 30.5 | 69.5 |
| 5.4 | 25.5 | 74.5 |
| 5.6 | 21.0 | 79.0 |
| 5.8 | 16.0 | 84.0 |
| 6.0 | 11.5 | 88.5 |
| 6.2 | 8.0 | 92.0 |
## [](https://www.sigmaaldrich.com)Sodium Acetate – Acetic Acid Buffer PREPARATION, pH 3.7–5.61
Sodium acetate trihydrate, CH3COONa • 3H2O, MW 136.09; 0.2 M contains 27.22 g/L.
x mL 0.2 M-NaOAc and y mL 0.2 M-HOAc mixed
| | | |
|-----------|------------------|-----------------|
| pH, 18 °C | x mL 0.2 M-NaOAc | y mL 0.2 M-HOAc |
| 3.7 | 10.0 | 90.0 |
| 3.8 | 12.0 | 88.0 |
| 4.0 | 18.0 | 82.0 |
| 4.2 | 26.5 | 73.5 |
| 4.4 | 37.0 | 63.0 |
| 4.6 | 49.0 | 51.0 |
| 4.8 | 59.0 | 41.0 |
| 5.0 | 70.0 | 30.0 |
| 5.2 | 79.0 | 21.0 |
| 5.4 | 86.0 | 14.0 |
| 5.6 | 91.0 | 9.0 |
## [](https://www.sigmaaldrich.com)Na2HPO4 – NaH2PO4 Buffer PREPARATION, pH 5.8-8.0 at 25 °C1
Na2HPO4 • 2H2O, MW 178.05; 0.2 M contains 35.61 g/L. Na2HPO4 • 12H2O, MW 358.22; 0.2 M contains 71.64 g/L. NaH2PO4 • H2O, MW 138.01; 0.2 M contains 27.6 g/L. NaH2PO4 • 2H2O, MW 156.03; 0.2 M contains 31.21 g/L.
x mL 0.2 M-Na2HPO4, y mL 0.2 M-NaH2PO4; diluted to 100 mL with H2O
| | | |
|-----------|--------------------|--------------------|
| pH, 25 °C | x mL 0.2 M-Na2HPO4 | y mL 0.2 M-NaH2PO4 |
| 5.8 | 4.0 | 46.0 |
| 6.0 | 6.15 | 43.85 |
| 6.2 | 9.25 | 40.75 |
| 6.4 | 13.25 | 36.75 |
| 6.6 | 18.75 | 31.25 |
| 6.8 | 24.5 | 25.5 |
| 7.0 | 30.5 | 19.5 |
| 7.2 | 36.0 | 14.0 |
| 7.4 | 40.5 | 9.5 |
| 7.6 | 43.5 | 6.5 |
| 7.8 | 45.75 | 4.25 |
| 8.0 | 47.35 | 2.65 |
## [](https://www.sigmaaldrich.com)Imidazole (glyoxaline) – HCl Buffer PREPARATION, pH 6.2–7.8 at 25 °C1
Imidazole, C3H4N2, MW 68.08
25 mL 0.2 M-imidazole (13.62 g/L), x mL 0.2 M-HCl, diluted to 100 mL with H2O
| | |
|-----------|----------------|
| pH, 25 °C | x mL 0.2 M-HCl |
| 6.2 | 21.45 |
| 6.4 | 19.9 |
| 6.6 | 17.75 |
| 6.8 | 15.2 |
| 7.0 | 12.15 |
| 7.2 | 9.3 |
| 7.4 | 6.8 |
| 7.6 | 4.65 |
| 7.8 | 3.0 |
## [](https://www.sigmaaldrich.com)Sodium Carbonate – Sodium Bicarbonate Buffer PREPARATION, pH 9.2-10.81
Na2CO3 • 10H2O, MW 286.2; 0.1 M contains 28.62 g/L. NaHCO3, MW 84.0; 0.1 M contains 8.40 g/L.
x mL 0.1 M-Na2CO3 and y mL 0.1 M-Na2HCO3 mixed
| | | | |
|-------------------|--------------------|----|----|
| 20 °C | 37 °C | | |
| x mL 0.1 M-Na2CO3 | y mL 0.1 M-Na2HCO3 | | |
| 9.2 | 8.8 | 10 | 90 |
| 9.4 | 9.1 | 20 | 80 |
| 9.5 | 9.4 | 30 | 70 |
| 9.8 | 9.5 | 40 | 60 |
| 9.9 | 9.7 | 50 | 50 |
| 10.1 | 9.9 | 60 | 40 |
| 10.3 | 10.1 | 70 | 30 |
| 10.5 | 10.3 | 80 | 20 |
| 10.8 | 10.6 | 90 | 10 |
## [](https://www.sigmaaldrich.com)[](https://www.sigmaaldrich.com)Buffer Preparation Formulas and Equations
### Percentage by weight (w/v)
(% buffer desired / 100) × final buffer volume (mL) = g of starting material needed.
### Molar Solutions
desired molarity × formula weight × solution final volume (L) = grams needed
### Henderson-Hasselbach Equation
### Reference
1\.
Dawson R, Elliot D, Elliot W, Jones KM. 1986. Data for Biochemical Research. 3rd ed.. Oxford Science Publ..
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- [Tris](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/biological-buffers/tris)
- [HPLC Buffers](https://www.sigmaaldrich.com/US/en/products/analytical-chemistry/analytical-chromatography/hplc-buffers)
- [Phosphate Buffered Saline (PBS)](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/biological-buffers/phosphate-buffer-saline-pbs)
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