The following are recipes for a number of common biological buffers

A good buffer must exhibit the following characteristics [1] :

  1. Water solubility
  2. Chemical stability
  3. High buffering capacity at desired pH
  4. Compatibility with analytical and experimental applications
  5. Compatibility with other solution components

 

GLYCINE–HCL; PH 2.2–3.6, PKA = 2.35

Combine 25 ml 0.2 M glycine and x ml HCl and dilute to 100 ml with DI (Dawson, et al., 1969).

x (ml)

pH

22.0

2.2

16.2

2.4

12.1

2.6

8.4

2.8

5.7

3.0

4.1

3.2

3.2

3.4

2.5

3.6

 

SODIUM ACETATE; PH 3.6–5.6, PKA = 4.76

Combine the following proportions of 0.1N acetic acid and 0.1N sodium acetate (Pearse, 1980).

acetic acid

sodium acetate

pH

185

15

3.6

176

24

3.8

164

36

4.0

147

53

4.2

126

74

4.4

102

98

4.6

80

120

4.8

59

141

5.0

42

158

5.2

29

171

5.4

19

181

5.6

BUFFERED SALINE (PBS, TBS, TNT, PBT)

Buffered saline solutions are used frequently when performing immunolocalization experiments. There are many variations. Presented here are three common formulations (Mishkind, et al., 1987).

PBS 20x stock

TBS

Potassium chloride

4 g

53.6 mM

Potassium chloride 4 g

NaCl

160 g

274 mM

NaCl 160 g

Potassium phosphate monobasic

4 g

29.4 mM

Tris buffer (10 mM, pH 7.5) to 1 liter

Sodium phosphate dibasic (7•H2O) DI

43.2 g

17.5 mM to 1 liter

Use TBS when performing immunocytochemical

experiments on phosphate-sensitive tissues

(photosynthetic tissues typically)

TNT

PBT

NaCl

150 mM

PBS to vol

Tris buffer (100 mM, pH 7.5)

to 1 liter

Tween 20 1% (v/v)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

CACODYLATE BUFFER; PH 5.0–7.4, PKA = 6.27

Sodium cacodylate buffer [Na(CH3)2 AsO2 • 3H2O] is a alternative to Sørensen’s phosphate buffer. It has good pH buffering capacity within the range of pH 5.0–7.4. Cacodylate was introduced for electron microscopy applications by Sabatini et al. (1962) as a method of avoiding adding additional phosphates to sample preparations. Mitochondria and other organelles can be damaged when exposed to the high concentrations of phosphates present in Sørensen’s buffers. Also, cacodylate will not react with aldehyde fixatives as will amine-containing buffers (e.g., Tris). Its efficacy in fixation solutions may be a result of the metabolism-inhibiting effect of the arsenate rather than any special buffering capacity.

Prepare a 0.2 M stock solution of sodium cacodylate in water (4.28 g/100 ml). Add the following amounts of 0.2 M HCl per 100 ml cacodylate stock solution, followed by the addition of DI to a final volume of 400 ml, to obtain 0.05 M cacodylate buffer at the desired pH (Dawes, 1971).

0.2 M HCl

pH

94.0

5.0

90.0

5.2

86.0

5.4

78.4

5.6

69.6

5.8

59.2

6.0

47.6

6.2

36.6

6.4

26.6

6.6

18.6

6.8

12.6

7.0

8.4

7.2

5.4

7.4

 

 

 

 

 

“GOOD” BUFFERS; PKA = 6.15–8.06

Tris–HCl (pKa = 8.06) and maleate (pKa = 6.26) have a working range of pH 5.0–8.6 and may be used successfully to buffer staining solutions (e.g., Toluidine Blue O). Avoid Tris with aldehyde fixatives or osmium tetroxide, however, as the aldehydes reacts with the amino group of Tris, resulting in the loss of buffering capacity. PIPES (pKa = 6.80) is commonly used as a buffer for retention of actin filaments during fixation. Other useful biological buffers include HEPES (pKa = 7.55), MES (pKa = 6.15), and MOPS (pKa = 7.20) (Good, et al., 1966; Perrin and Dempsey, 1974).

 

CITRATE BUFFER; PH 3.0–6.2, PKA = 6.40 Citrate buffer (Gomori, 1955) stock solutions: A: 0.1 M citric acid; B: 0.1 M sodium citrate. Use x ml A + y ml B and dilute to 100 ml with 50 ml DI.

0.1 M citric acid

0.1 M sodium citrate

pH

46.5

3.5

3.0

43.7

6.3

3.2

40.0

10.0

3.4

37.0

13.0

3.6

35.0

15.0

3.8

33.0

17.0

4.0

31.5

18.5

4.2

28.0

22.0

4.4

25.5

24.5

4.6

23.0

27.0

4.8

20.5

29.5

5.0

18.0

32.0

5.2

16.0

34.0

5.4

13.7

36.3

5.6

11.8

38.2

5.8

9.5

41.5

6.0

7.2

42.8

6.2

SØRENSEN’S PHOSPHATE BUFFER; PH 5.8–8.0, PKA = 7.20

Mix appropriate volumes of stock and add an equal volume of distilled water to make a final 0.1 M Sørensen’s phosphate buffer solution (Sørensen, 1909; Gomori, 1955). Keep in mind that high levels of phosphate may be somewhat toxic to plant cells (Sabatini, et al., 1962) and thus Sørensen’s buffer may not be appropriate for some experiments.

Stock solutions: A 0.2 M NaH2PO4 B 0.2 M Na2HPO4

A (ml)

B (ml)

pH

92.0

8.0

5.8

87.7

12.3

6.0

81.5

18.5

6.2

68.5

31.5

6.5

62.5

37.5

6.6

56.5

43.5

6.7

51.0

49.0

6.8

45.0

55.0

6.9

39.0

61.0

7.0

33.0

67.0

7.1

28.0

72.0

7.2

23.0

77.0

7.3

19.0

81.0

7.4

16.0

84.0

7.5

8.5

91.5

7.8

5.3

94.7

8.0

 

 

 

 

PHOSPHATE–CITRATE BUFFER; PH 2.2–8.0, PKA = 7.20/6.40

Add the following to create 100 ml of phosphate/citrate buffer solution. Stock solutions are

0.2 M dibasic sodium phosphate; 0.1 M citric acid (Pearse, 1980).

0.2 M Na2HPO4 (ml)

0.1 M citrate (ml)

pH

5.4

44.6

2.6

7.8

42.2

2.8

10.2

39.8

3.0

12.3

37.7

3.2

14.1

35.9

3.4

16.1

33.9

3.6

17.7

32.3

3.8

19.3

30.7

4.0

20.6

29.4

4.2

22.2

27.8

4.4

23.3

26.7

4.6

24.8

25.2

4.8

25.7

24.3

5.0

26.7

23.3

5.2

27.8

22.2

5.4

29.0

21.0

5.6

30.3

19.7

5.8

32.1

17.9

6.0

33.1

16.9

6.2

34.6

15.4

6.4

36.4

13.6

6.6

40.9

9.1

6.8

43.6

6.5

7.0

 

 

 

 

BARBITAL BUFFER; PH 6.8–9.2, PKA = 7.98

Add the following to create 200 ml of buffered solution. To 50 ml of 0.2 M sodium barbital (Veronal,

41.2 g in 1000 ml) add x ml 0.2 M HCl to create the buffered solution and dilute to 200 ml with DI (Gomori, 1955).

0.2 M HCl (ml)

pH

1.5

9.2

2.5

9.0

4.0

8.8

6.0

8.6

9.0

8.4

12.7

8.2

17.5

8.0

22.5

7.8

27.5

7.6

32.5

7.4

39.0

7.2

43.0

7.0

45.0

6.8

 

 

 

 

TRIS BUFFERS

 

Tris buffers are used commonly in microtechnique applications involving molecular biological procedures. Listed here are a number of common Tris formulations (Maniatis, et al., 1982).

Solution

Preparation

Tris, 1 M stock

Tris base DI Dissolve and adjust pH with the following approximate amount of HCl: pH 7.4 pH 7.6 pH 8.0

121.1 g 800 ml 70 ml 60 ml 42 ml

EDTA, 0.5 M

Disodium ethylene diamine tetraacetate Adjust pH to approx. 8.0 and stir until dissolved

186.1 g

SSC, 20x

NaCl NaCitrate DI Adjust pH to 7.0 with NaOH then add DI to 1 liter

175.3 g 88.2 g 800 ml

SSPE, 20x

NaCl NaH2PO4 • H2O EDTA DI Adjust pH to 7.4 with NaOH then add DI to 1 liter

174 g 27.6 g 7.4 g 800 ml

TE

Tris EDTA Adjust pH using Tris stock solution

10 mM 1 mM

STE (TNE)

Tris NaCl EDTA Adjust pH to 8.0 using Tris stock solution

10 mM 100 mM 1 mM

 

 

 

 

 

 

 

 

 

 

 

GLYCINE– NAOH BUFFER; PH 8.6–10.6, PKA = 9.78

Stock solutions:

0.2 M glycine

0.2 NaOH

Combine 25 ml glycine stock solution with x ml 0.2 M NaOH and dilute with DI to make a 100 ml solution (Pearse, 1980).

0.2 M NaOH

pH

2.0

8.6

3.0

8.8

4.4

9.0

6.0

9.2

8.4

9.4

11.2

9.6

13.6

9.8

19.3

10.4

22.75

10.6

Source: http://microscopy.berkeley.edu/Resources/instruction/buffers.html

 

 

 

 

1. Hydrochloric Acid-Potassium Chloride Buffer (HCl-KCl); pH Range 1.0 to 2.2

(a) 0.1 M Potassium chloride : 7.45 g/l (M.W.: 74.5)

(b) 0.1 M Hydrochloric acid

Mix 50 ml of potassium chloride and indicated volume of hydrochloric acid.

Mix and adjust the final volume to 100 ml with deionized water. Adjust the

final pH using a sensitive pH meter.

ml of HCl 97 64.5 41.5 26.3 16.6 10.6 6.7

pH 1.0 1.2 1.4 1.6 1.8 2.0 2.2

2. Glycine-HCl Buffer; pH range 2.2 to 3.6

(a) 0.1 M Glycine: 7.5 g/l (M.W.: 75.0)

(b) 0.1 M Hydrochloric acid

Mix 50 ml of glycine and indicated volume of hydrochloric acid. Mix and

adjust the final volume to 100 ml with deionized water. Adjust the final pH

using a sensitive pH meter.

ml of HCl 44.0 32.4 24.2 16.8 11.4 8.2 6.4 5.0

pH 2.2 2.4 2.6 2.8 3.0 3.2 3.4 3.6

3. Citrate Buffer; pH range 3.0 to 6.2

(a) 0.1 M Citric acid: 19.21 g/l (M.W.: 192.1)

(b) 0.1 M Sodium citrate dihydrate: 29.4 g/l (M.W.: 294.0)

Mix citric acid and sodium citrate solutions in the proportions indicated and

adjust the final volume to 100 ml with deionized water. Adjust the final pH

using a sensitive pH meter. The use of pentahydrate salt of sodium citrate is

not recommended.

ml of Citric acid 46.5 40.0 35.0 31.5 25.5 20.5 16.0 11.8 7.2

ml of Sodium citrate 3.5 10.0 15.0 18.5 24.5 29.5 34.0 38.2 42.8

pH 3.0 3.4 3.8 4.2 4.6 5.0 5.4 5.8 6.2

 

 

4. Acetate Buffer; pH range 3.6 to 5.6

(a) 0.1 M Acetic acid (5.8 ml made to 1000 ml)

(b) 0.1 M Sodium acetate; 8.2 g/l (anhydrous; M.W. 82.0) or 13.6 g/l

(trihydrate; M.W. 136.0)

Mix acetic acid and sodium acetate solutions in the proportions indicated and

adjust the final volume to 100 ml with deionized water. Adjust the final pH

using a sensitive pH meter.

ml of Acetic acid 46.3 41.0 30.5 20.0 14.8 10.5 4.8

ml of Sodium acetate 3.7 9.0 19.5 30.0 35.2 39.5 45.2

pH 3.6 4.0 4.4 4.8 5.0 5.2 5.6

5. Citrate-Phosphate Buffer; pH range 2.6 to 7.0

(a) 0.1 M Citric acid; 19.21 g/l (M.W. 192.1)

(b) 0.2 M Dibasic sodium phosphate; 35.6 g/l (dihydrate; M.W. 178.0) or

53.6 g/l (heptahydrate; M.W. 268.0)

Mix citric acid and sodium phosphate solutions in the proportions indicated

and adjust the final volume to 100 ml with deionized water. Adjust the final

pH using a sensitive pH meter.

ml of Citric acid 44.6 39.8 35.9 32.3 29.4 26.7 24.3 22.2 19.7 16.9 13.6 6.5

ml of Sodium

phosphate 5.4 10.2 14.1 17.7 20.6 23.3 25.7 27.8 30.3 33.1 36.4 43.6

pH 2.6 3.0 3.4 3.8 4.2 4.6 5.0 5.4 5.8 6.2 6.6 7.0

 

 

 

 

 

 

 

 

6. Phosphate Buffer; pH range 5.8 to 8.0

(a) 0.1 M Sodium phosphate monobasic; 13.8 g/l (monohydrate,

M.W. 138.0)

(b) 0.1 M Sodium phosphate dibasic; 26.8 g/l (heptahydrate, M.W. 268.0)

Mix sodium phosphate monobasic and dibasic solutions in the proportions

indicated and adjust the final volume to 200 ml with deionized water. Adjust

the final pH using a sensitive pH meter.

ml of Sodium

phosphate, Monobasic 92.0 81.5 73.5 62.5 51.0 39.0 28.0 19.0 13.0 8.5 5.3

ml of Sodium

phosphate, Dibasic 8.0 18.5 26.5 37.5 49.0 61.0 72.0 81.0 87.0 91.5 94.7

pH 5.8 6.2 6.4 6.6 6.8 7.0 7.2 7.4 7.6 7.8 8.0

7. Tris-HCl Buffer, pH range 7.2 to 9.0

(a) 0.1 M Tris(hydroxymethyl)aminomethane; 12.1 g/l (M.W.: 121.0)

(b) 0.1 M Hydrochloric acid

Mix 50 ml of Tris(hydroxymethyl)aminomethane and indicated volume of

hydrochloric acid and adjust the final volume to 200 ml with deionized water.

Adjust the final pH using a sensitive pH meter.

ml of HCl 44.2 41.4 38.4 32.5 21.9 12.2 5.0

8. Glycine-Sodium Hydroxide, pH 8.6 to 10.6

(a) 0.1 M Glycine; 7.5 g/l (M.W.: 75.0)

(b) 0.1 M Sodium hydroxide; 4.0 g/l (M.W.: 40.0)

Mix 50 ml of glycine and indicated volume of sodium hydroxide solutions and adjust

the final volume to 200 ml with deionized water. Adjust the final pH using a sensitive

pH meter.

ml of Sodium hydroxide 4.0 8.8 16.8 27.2 32.0 38.6 45.5

pH 8.6 9.0 9.4 9.8 10.0 10.4 10.6

 

 

9. Carbonate-Bicarbonate Buffer, pH range 9.2 to 10.6

(a) 0.1 M Sodium carbonate (anhydrous), 10.6 g/l (M.W.: 106.0)

(b) 0.1 M Sodium bicarbonate, 8.4 g/l (M.W.: 84.0)

Mix sodium carbonate and sodium bicarbonate solutions in the proportions indicated

and adjust the final volume to 200 ml with deionized water. Adjust the final pH using

a sensitive pH meter.

ml of Sodium carbonate 4.0 9.5 16.0 22.0 27.5 33.0 38.5 42.5

ml of Sodium bicarbonate 46.0 40.5 34.0 28.0 22.5 17.0 11.5 7.5

pH 9.2 9.4 9.6 9.8 10.0 10.2 10.4 10.6pH 7.2 7.4 7.6 7.8 8.2 8.6 9.0

source: https://www.med.unc.edu/pharm/sondeklab/files/resource-files/protein-purification-handbooks/buffers_calbiochem.pdf

 

Formulas for Qiagen Kit Buffers

Do not autoclave solutions containing ethanol, isopropanol or MOPS; use sterile filtration if necessary.

Buffer AE (elution buffer for genomic DNA preps)

Buffer P1

The buffer and RNaseA can also be ordered from Qiagen separately (catalog numbers 19051 and 19101).

Buffer P2

Buffer P3 (not for spin columns, but for Qiatips, midi, maxi, giga kits)

Buffer DP3 (for Qiagen Directprep 96-well miniprep)

Buffer N3

Buffer PB

Buffer QG

Buffer PE

Buffer QX1 (for solution and binding of agarose gels)

Buffer QXB (for binding of large >3000 bp fragments to columns)

Buffer QBT (equilibration buffer)

Buffer QC (wash buffer)

Buffer QF (elution buffer)

Buffer QN

Buffer FWB2

Buffer B1 (bacterial lysis buffer)

Buffer B2 (bacterial lysis buffer)

Buffer C1 (cell lysis buffer) (store at +4)

Buffer G2 (digestion buffer)

Buffer Y1 (yeast lysis buffer) (store at +4)

Buffer PAA (PAGE gel elution of DNA)

Buffer PNI (purification of oligonucleotides 17 nt and greater)

Source: https://openwetware.org/wiki/Qiagen_Buffers

 

Buffers   western  blottıng

RIPA buffer

RIPA buffer contains the ionic detergent sodium deoxycholate as an active constituent and is particularly useful for nuclear membrane disruption for nuclear extracts. A RIPA buffer gives low background but can denature kinases. It can also disrupt protein-protein interactions and may therefore be problematic for immunoprecipitations and pull-down assays.

50 mM Tris HCl, pH 8.0
150 mM NaCl
​1% NP-40
​0.5% sodium deoxycholate
​0.1% SDS

​The 10% sodium deoxycholate stock solution (5 g into 50 mL) must be protected from light.


NP-40 buffer

20 mM Tris HCl, pH 8.0
137 mM NaCl
10% glycerol
1% NP-40
2 mM EDTA


Cytoskeletal bound proteins extract buffer

10 mM Tris, pH 7.4
100 mM NaCl
1 mM EDTA
1 mM EGTA
1 mM NaF
​20 mM Na4P2O7
2 mM Na3VO4
1% Triton X-100
10% glycerol
0.1% SDS
0.5% deoxycholate


Soluble protein buffer

20 mM Tris-HCl, pH 7.5
1 mM EGTA (Ca2+​ chelator)

Source:http://www.abcam.com/protocols/buffer-and-stock-solutions-for-western-blot