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Single component solutions

Organic substances

pK_a and temperature dependence of pH for common buffers
ATP 0.1M
Betaine 5M
Cresol red (Na) 50mM
DTT 1M, 2.2M
dNTP’s 100mM
EDTA 0.5M
EtBr 10mg/ml
Gelatin 2%
Glucose 1; 1.5; 2M
Guanidine HCl 1-8M
HEPES 1M
Imidazol 2M
Paraformaldehyde 37%
PEG 40%
PMSF 100mM
Retinoic acid 10mM
Sucrose 1; 2; 2.5M
Tris Cl 1M
Temperature dependence of pH for TrisCl
Tricine 1M
Triethanolamine 1M
Urea 1-10M

Acids and alkalis

Summary table
NaOH 10M, 1M
KOH 5M
TCA 100%

Detergents

N-Lauroylsarcosine Na 10%
SDS 10%

Organic solvents

Phenol
EthanolEtOH
Preparation of 100% EtOH

Supplement

Densities of some solutions are available on the page "Densities of acids, alkali and organic substances".

Organic substances

pK_a and temperature dependence of pH for common buffers

Buffer	Mw	pK_a 20^oC	Working range	delta pK_a per 10^oC
MES 2-(N-morpholino)ethanesulfonic acid	195.2	6.15	5.5-6.7	-0.110
Bis-Tris bis(2-hydroxyethyl)iminotris(hydroxymethyl)methane	209.2	6.5	5.8-7.2
ADA N-(2-acetamido)-2-imidoacetic acid	190.2	6.60	6.0-7.2	-0.110
PIPES piperasine-N,N'-bis(2-ethanesulfonic acid)	302.4	6.80	6.1-7.5	-0.085
ACES 2-[(2-amino-2-oxoethyl)amino]ethanesulfonic acid	182.2	6.90	6.1-7.5	-0.200
MOPSO 3-(N-morpholino)-2-hydroxypropanesulfonic acid	225.3	6.9	6.2-7.6
Imidazol - HCl	68.08	6.95	6.2-7.8
Bis-Tris Propane 1,3-bis[tris(hydroxymethyl)methylamino]propane	282.3	6.8*	6.3-9.5
BES N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid	213.2	7.15	6.4-7.8	-0.160
MOPS 3-(N-morpholino)propanesulfonic acid	209.3	7.20	6.5-7.9	-0.013
TES N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid	229.2	7.50	6.8-8.2	-0.200
HEPES N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid)	238.3	7.55	6.8-8.2	-0.140
DIPSO 3-[N,N-bis(2-hydroxyethyl)amino]-2-hydroxyprpanesulfonic acid	243.3	7.6	7.0-8.2
TAPSO 3-[N-tris(hydroxymethyl)methylamino]-2-hydroxypropanesulfonic acid	259.3	7.6	7.0-8.2
HEPPSO N-(2-hydroxyethyl)piperazine-N'-(2-hydroxypropanesulfonic acid)	268.3	7.8	7.1-8.5
POPSO piperazine-N,N'-bis(2-hydroxypropanesulfonic acid)	362.4	7.8	7.2-8.5
TEA triethanolamine	149.2	7.8	7.3-8.3
EPPS N-(2-hydroxyethyl)piperazine-N'-(3-propanesulfonic acid)	252.3	8.0	7.3-8.7
Tricine N-tris(hydroxymethyl)methylglycine	179.2	8.15	7.4-8.8	-0.210
Tris (TRIZMA) tris(hydroxymethyl)aminomethane	121.1	8.30	7.0-9.1	-0.310
Bicine N,N-bis(2-hydroxyethyl)glycine	163.2	8.35	7.6-9.0	-0.180
TAPS N-tris(hydroxymethyl)methyl-3-aminopropanesulfonic acid	243.3	8.4	7.7-9.1
Glycylglycine		8.40		-0.280
AMPSO 3-[(1,1-dimethyl-2-hydroxyethyl)amino]-2-hydroxypropanesulfonic acid	227.3	9.0	8.3-9.7
CHES 1-(N-cyclohexylamino)ethanesulfonic acid	207.3	9.3	8.6-10.0
CAPSO 3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid	237.3	9.6	8.9-10.3
AMP 2-amino-2-methyl-1-propanol	89.1	9.7	9.0-10.5
CAPS 3-(cyclohexylamino)-1-propanesulfonic acid	221.3	10.4	9.7-11.1

See also:
/web-links/

Buffer design - online calculations of large list of buffers (Prof. R.Beynon) on the page Java based Molecular Biologist's Workbench EMBL.
Buffer Calculator on the site of LabVelocity. Registration is necessary (free).

Preparation of dNTP's.

The quick protocol for ~100mM stock

dilute all four dNTP's (250mg of each) in 3.676ml of H₂O;
+ 424µl 5M NaOH;
check the pH: ~0.5µl on pH-paper.

Accurate protocol for 100mM stocks preparation

add the necessary quantity (see table) of H₂O and Tris base 1M (it is possible to take the volume of salt as ~150µl);

		Mw	V_(H2O)	V_{(Tris base 1M)}	V_(final)
dATP	C₁₀H₁₄N₅O₁₂P₃Na₂x3H₂O	589.2	3.24ml	850 µl	4.24ml
dGTP	C₁₀H₁₃N₅O₁₃P₃Na₃x2H₂O	609.2	3.55ml	400 µl	4.10ml
dCTP	C₉H₁₃N₃O₁₃P₃Na₃x2H₂O	569.1	3.44ml	800 µl	4.39ml
dTTP	C₁₀H₁₄N₂O₁₄P₃Na₃x2H₂O	584.1	3.73ml	400 µl	4.28ml

check the pH: ~0.5µl on pH-paper;
check the quality and concentration (it is useful to take final dilution 1:5000 (~20µM). In this case the optical density will be in the region of A_m~0.3 - the most accurate range for spectrophotometer).
Concentration is c[mM]=k_1:5000xA_m.

Quality:

dATP pH 7.0	A₂₅₀/A₂₆₀=0.80+0.03 A₂₈₀/A₂₆₀=0.12+0.02	dCTP pH 2.0(!)	A₂₅₀/A₂₆₀=0.45+0.03 A₂₈₀/A₂₆₀=2.10+0.15 A₂₉₀/A₂₆₀=1.60+0.10
dGTP pH 7.5	A₂₅₀/A₂₆₀=1.18+0.04 A₂₈₀/A₂₆₀=0.67+0.03 A₂₉₀/A₂₆₀=0.28+0.03	dTTP pH 7.5	A₂₅₀/A₂₆₀=0.65+0.03 A₂₈₀/A₂₆₀=0.73+0.03

Concentration:

	Mw	A_m	pH	E	k (for 1:5000)
dATP[Na₂]	589.2	259	7.0	15.2x10³	328.9
dGTP[Na₃]	609.2	253	7.5	13.7x10³	365.0
dCTP[Na₃]	569.1	280	2.0 (!!!)	13.0x10³	384.6
dTTP[Na₃]	584.1	267	7.5	9.6x10³	520.8

ATP [Na₄]	595.1	259		15.4x10³
CTP [Na₄]	571.1	280		13.0x10³
GTP [Na₄]	611.1	252		13.7x10³
UTP [Na₄]	572.1	262		10.2x10³

Concentration: c[M]=A_max/E; A_max = maximum of absorption.

Paraformaldehyde

PFA (paraformaldehyde) 37% (for histochemistry it should be freshly prepared).

1. mix in the screw-cap tube:

PFA (solid) = 0.37g,
H₂O = 1.0ml
NaOH (1N) = 14µl;

2. solubilize in the boiling water bath (to heat ~1-3'; until pH will drop to ~7.0).

TrisCl: 250ml 1M

V HCl	V HCl	pH
	0	10.44
	1	9.54
	2	9.26
	3	9.08
	3.7	8.98
5ml		8.76
6ml		8.67
	10	8.4
11ml		8.22
	14	8.13
13.5ml		8.06
14ml		8.04
15ml		7.97
	17	7.92
16.5ml		7.82
16.7ml		7.8
17ml		7.77
18.3ml		7.63
18.6ml		7.59
	20	7.75
	23	7.4

Acids and alkalis

Name:	Formula:	Mw	% (w/w)	[M]	g in 1L of subst.	p [g/ml]	ml/L for 1M sol.
Sodium hydroxide	NaOH	40	50%	19.1	763	1.53	52.4
			30.1%	10.0	400	1.329	100
			10%	2.75	111	1.11	363.6
Potassium hydroxide	KOH	56.1	50%	13.5	757	1.52	74.1
			23.06%	5.0	280.6	1.217	200
			10%	1.94	109	1.09	515.5
Ammonium hydroxide	NH₄OH	35.0	28%	14.8	251	0.898	67.6
Acetic acid, glacial	CH₃COOH	60.05	99.5%	17.4	1045	1.05	57.5
Acetic acid	CH₃COOH	60.05	36%	6.27	376	1.045	159.5
Formic acid	HCOOH	46.02	90%	23.4	1080	1.20	42.7
Hydrochloric acid	HCl	36.5	36%	11.6	424	1.18	86.2
Hydrochloric acid	HCl	36.5	10%	2.9	105	1.05	344.8
Nitric acid	HNO₃	63.02	71%	15.99	1008	1.42	62.5
			67%	14.9	938	1.40	67.1
			61%	13.3	837	1.37	75.2
Perchloric acid	HClO₄	100.5	70%	11.65	1172	1.67	85.8
Perchloric acid	HClO₄	100.5	60%	9.2	923	1.54	108.7
Phosphoric acid	H₃PO₄	98.0	85%	18.1	1445	1.71	55.2
Sulfuric acid	H₂SO₄	98.1	96%	18.0	1766	1.84	55.6

Detergents

Detergent	T_melt	Mw [Da]		CMC
Detergent	T_melt	monomer	micelle	%(w/v)	M
Anionic
SDS	206	288	18,000	0.23	8.0 x 10^-3
Cholate	201	430	4,300	0.60	1.4 x 10^-2
Deoxycholate	175	432	4,200	0.21	5.0 x 10^-3
Cationic
C₁₆TAB	230	365	62,00	0.04	1.0 x 10^-3
Amphoteric
LysoPC	-	495	92,000	0.0004	7.0 x 10^-6
CHAPS	157	615	6,150	0.49	1.4 x 10^-3
Zwittergent 3-14	-	365	30,000	0.011	3.0 x 10^-4
Nonionic
Octyl glucoside	105	292	8,000	0.73	2.3 x 10^-2
Digitonin	235	1,229	70,000	-	-
C₁₂E₈	-	542	65,000	0.005	8.7 x 10^-5
Lubrol PX	-	582	64,000	0.006	1.0 x 10^-4
Triton X-100	-	650	90,000	0.021	3.0 x 10^-4
Nonident P-40	-	603	90,000	0.017	3.0 x 10^-4
Tween 80	-	1,310	76,000	0.002	1.2 x 10^-5

Detergent	1	2	3	4	5	6	7	8	9	10	11	12	1 - Strongly denaturing; 2 - Dializable; 3 - Ion exchangeable, unsuitable for ion-exchange chromatography; 4 - Complexes ions; 5 - Strong A₂₈₀; 6 - Assay interference; 7 - Cold precipitates; 8 - High cost; 9 - Ease of purification; 10 - Radiolabeled; 11 - Definite composition; 12 - Auto-oxidation
Anionic
SDS	+	+	+	+	-	-	+	-	+	+	+	-
Cholate	-	+	+	+	-	-	-	-	+	+	+	-
Deoxycholate	-	+	+	+	-	-	+	-	+	+	+	-
Cationic
C₁₆TAB	+	+	+	-	-	-	+	-	+	-	+	-
Amphoteric
LysoPC	+/-	-	-	-	-	-	-	+	+/-	+	+	-
CHAPS	-	+	-	-	-	-	-	+	+	-	+	-
Zwittergent 3-14	+/-	+/-	-	-	-	-	-	+	+	-	+	-
Nonionic
Octyl glucoside	-	+	-	-	-	-	-	+	-	+	+	-
Digitonin	-	-	-	-	-	-	-	+	+	-
C₁₂E₈	-	-	-	+/-	-	-	-	+	-	+	-	+
Lubrol PX	-	-	-	+/-	-	+/-	-	-	-	+	-	+
Triton X-100	-	-	-	+/-	+	+/-	-	-	-	+	-	+
Nonident P-40	-	-	-	+/-	+	+/-	-	-	-	+	-	+
Tween 80	-	-	-	+/-	-	+/-	-	-	-	+	-	+

C₁₆TAB - hexadecyl trimethylammonium bromide;
LysoPC - lysophosphatidylcholine.
* Sodium cholate and sodium deoxycholate are unsoluble at <pH 7.5 or at ionic strength greater then 0.1%. SDS may precipitate below 20^oC;
** Ionic detergents may induce problems with electrophoresis and isoelectric focusing;
*** It is possible to remove by dialysis;
**** Phenol -containing detergents (for example Triton X-100 and NP-40) precipitate during Folin protein assay (but do not interfere with Bradford protein assay).

Organic solvents

Phenol

C₆H₅OH; Mw=94.1g/M
p=1.054, t_m= 43, t_b= 182, pKa=10.0
solubility: 6.8¹⁶H₂O, unlimited ⁶⁶H₂O; unlimited EtOH

Preparation of "acidic" and "neutral" phenol.

a) from the distillation:

distill phenol under H₂O;
adjust water to the volume of about 1/10 of phenol phase;
add 8-hydroxyquinoline to 0.1% (relative to phenol phase) and bMeEtOH (2-mercaptoethanol) to 0.2% (relative to H₂O = 0.02% relative to phenol phase);

---- on this step you obtained the "acidic phenol". Store at -20^oC (~1 year); ----

add about the same volume of 0.2 M Tris-base to the phenol, mix ~0.5-1h;
throw away aqueous phase;
+ 0.1V 0.1M Tris-Cl, pH 8.0
0.2% bMeEtOH
mix ~0.5-1h;
store at 4^oC (in the dark) ~several months.

b) from the good commercial substance:

saturate phenol by the water (add about 1/5 V );
then, according to (a), from the p.2.

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Last modification: 30/07/13

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